WELCOME TO THIS NIEHS WORKSOP THE WORKSHOP AND TO THE OFFICE, EVERYONE WHO SUPPORTED THE RESEARCH THAT WENT INTO THE CASE THAT YOU ARE GOING TO HEAR ABOUT DURING THIS PARTICULAR WORKSHOP. BUT I WOULD LIKE TO INTRODUCE DR. PAUL COATS THE DR. AT N.I.H. TO GIVE US THE OPENING WELCOME. WELCOME. >> THANK YOU. >> GOOD MORNING. ON BEHALF OF DIETARY SUPPLEMENTS AND FOLKS DEDICATED TO THIS ISSUE I WOULD LIKE TO EXTEND MY WELCOME AS WELL. I'M SETTING THE STAGE, THAT'S ALL. WITH LUCK YOU HAVE ALREADY HEARD THIS SO IF YOU ARE STARTING TO DOZE, IT'S OK. JUST A REMINDER ABOUT THE REGULATORY DEFINITION FOR DIETARY SUPPLEMENT. IT'S A PRODUCT INTENDED TO SUPPLEMENT THE DIET. MAKES SENSE. CONTAINS A VITAMIN, MINERAL, AMINO ACID, DIETARY SUBSTANCE AND THE CLASS OF PRODUCTS THAT ARE CALLED HERBS OR BOTANICALS. JUST AS A REMINDER, IT'S THE RULES FOR FOODS AND NOT DRUGS THAT APPLY TO DIETARY SUPPLEMENTS. SO AN INGREDIENT IS PRESUMED TO BE SAFE BASED ON ITS HISTORY OF USE IN HUMANS AND THAT SOMETIMES IS A STRETCH. A REMINDER THAT DIETARY SUPPLEMENT USE IS REALLY VERY WIDE SPREAD IN THE UNITED STATES BASED ON DATA FROM THE NATIONAL HEALTH AND NUTRITION EXAMINATION SURVEY. OVER A TEN YEAR PERIOD. QUITE A STABLE NUMBER, IF YOU LOOK AT THESE FOUR LINES, THE BLUE ONE, SECOND FROM THE TOP IS THE ALL ADULTS NUMBER. AND OVER A TEN-YEAR PERIOD, ABOUT 50% OF AMERICAN ADULTS USE DIETARY SUPPLEMENTS, ON A VERY REGULAR BASIS. THIS IS DAILY, OR FREQUENTLY. THE LINE IS ABOVE IT, REPRESENTS WOMEN. THE LINE BELOW IT REPRESENTS MEN. SO WOMEN TEND TO BE USERS MORE THAN MEN ARE. BUT EVEN CHILDREN ABOUT A THIRD OF THE POPULATION, A LITTLE LESS THAN A THIRD USES DIETARY SUPPLEMENTS ON A REGULAR BASIS. SO THIS IS PUBLIC HEALTH, IN MY VIEW. OUR OFFICE WAS CREATED AS PART OF THE DIETARY SUPPLEMENT HEALTH AND EDUCATION ACT. AND WE FRAMED OUR MISSION IN STRENGTHENING KNOWLEDGE AND UNDERSTANDING OF DIETARY SUPPLEMENTS IN FOUR MAJOR AREAS, TO EVALUATE SCIENTIFIC INFORMATION, TO DISSEMINATE THE RESULTS OF THAT RESEARCH AND EDUCATE THE PUBLIC TO FOSTER AND ENHANCE THE HEALTH OF THE PUBLIC. THIS CLOSELY MIRRORS THE N.I.H. MISSION STATEMENT AS WELL. I'M GOING TO RUN THROUGH A COUPLE THINGS THAT HAVE GOTTEN OUR ATTENTION OVER THE YEARS. AS WE HAVE BECOME MORE AND MORE ENGAGED IN THE DETAILED STUDY OF DIETARY SUPPLEMENTS, THAT INCLUDES BOTANICALS. SEVERAL ISSUES HAVE ARISEN FOR US THAT WE PAY CLOSE ATTENTION TO. IN TERMS OF METHODS THAT EVALUATE AS INTERVENTIONS, RESEARCH PROTOCOLS, USUALLY CLINICAL TRIALS. THESE ISSUES HAVE ARISEN FOR US. THE NOTION OF THE GENERALIZABILITY OF RESEARCH FUNDINGS FROM THE POPULATIONS THAT ARE UNDER STUDIED. IF WE HAVE ONLY STUDIED YOUNG MEN IT'S HARD TO MAKE A GENERALIZATION TO OLDER WOMEN. THIS IS ALSO TRUE FOR THE SECOND BULLET. WE HAVE BECOME MUCH MORE INTERESTED IN THE NOTION THERE MAY BE PEOPLE WHOSE RESPONSES ARE PROGRAMMED FOR ONE REASON OR ANOTHER. PERHAPS GENETICALLY. SO IT WOULD BE VERY USEFUL IF WE COULD FIND WAYS TO DISTINGUISH RESPONDERS FROM NON-RESPONDERS. BECAUSE NOT EVERYBODY RESPONDS THE SAME WAY. IF YOU ARE LOOKING TO PROVE THERE'S A BENEFIT, YOU MIGHT BE HAVING TO LOOK FOR THE PEOPLE WHOM HARMS ARE POSSIBLE AS WELL. THE NOTIONS, AND THESE ARE COMMON TO A LOT OF KINDS OF INTERVENTIONS NOT JUST DIETARY SUPPLEMENTS BUT WHEN DO YOU INTRODUCE THEM. OVER WHAT PERIOD OF TIME DO YOU TAKE THEM. WHAT END POINTS DO YOU LOOK AT. WHAT DOSES OR SERVING SIZES? A LOT OF THIS HAS MADE US REALIZE IN THE FIELD OF DIETARY SUPPLEMENTS WE HAVE TO MOVE A LITTLE FURTHER BACK AND ENGAGE MORE SERIOUSLY IN EARLIER PHASE STUDIES AND THAT GOES STRAIGHT TO CLINICAL TRIALS. SOME OF THE SAME ISSUES ARE PRETTY, I WOULD SAY, UNIQUE TO DIETARY SUPPLEMENTS, YOU COULD PROVE ME WRONG. AND SOME IN PARTICULAR ARE QUESTIONS THAT ARE RAISED ABOUT BOTANICAL DIETARY SUPPLEMENTS, NOTION OF IDENTIFICATION, CHARACTERIZATION. IF YOU ARE USING THEM IN RESEARCH STUDIES YOU HAVE A RELIABLE SOURCE OF MATERIAL. IF YOU HAPPEN TO BE A CONSUMER YOU HAVE SOME CONFIDENCE THAT PRODUCT FROM TIME TO TIME REMAINS ABOUT THE SAME. I WANTED TO GIVE YOU AN EXAMPLE OF WHERE WE TOOK A STAND COMPWIET A NUMBER OF YEARS AGO. WE RECEIVED CONGRESSIONAL LANGUAGE IN THE EARLY 2000'S IN DIETARY SUPPLEMENTS CONTAINING PRODUCTS. THIS IS THE ONLY DATA SLIDE I HAVE. WHAT WE DID TO BEGIN THAT PROCESS IN COLLABORATION WITH THE NATIONAL CENTRE FOR COMPLEMENTARY AND ALTERNATIVE MEDICINE THEN, NOW NATIONAL CENTRE FOR COMPLIMENTARY AND INTEGRATED HEALTH IS TO CONVENE A SYSTEMATIC REVIEW USING THE EVIDENCE-BASED PRACTICE CENTRE AT RAND IN SOUTHERN CALIFORNIA. UNDER THE AEGIS OF RESEARCH AND QUALITY AND WE CALLED ON THEM TO REVIEW ALL OF THE AVAILABLE LITERATURE. PUBLISHED AND UNPUBLISHED AND THAT WAS PRETTY IMPORTANT TOO. ON THE USE OF E EPHEDRA ON WEIGHT MANAGEMENT AND PERFORMANCE ENHANCEMENT. WE WANTED THEM TO GIVE US THE EVIDENCE FOR EFFICACY AND SAFETY. IN TERMS OF WEIGHT LOSS THERE WAS A MODEST EFFECT OF EPHEDRA CONTAINING PRODUCTS LESS THAN SIX MONTHS. THAT WAS THE STANDARD BY WHICH MOST INTERVENTIONS FOR WEIGHT LOSS WORK EVALUATED USUALLY LESS THAN SIX MONTHS. NO EVIDENCE ON EFFECT ONAGE -- ATHLETIC PERFORMANCE, WHICH WAS A SURPRISE, IT WAS WIDELY TOUTED FOR BEING FOR WEIGHT LOSS AND ATHLETIC PERFORMANCE. THERE WERE SIDE EFFECTS. AN ARRAY OF SERIOUS SIDE EFFECTS, ADVERSE EVENTS FILED WITH THE F.D.A. AND MANUFACTURER. ALL OF THIS INFORMATION WAS COMPILED IN THE EVIDENCE REPORT, ULTIMATELY PUBLISHED IN JAMA IN 2002. -- 2003. A STEP THAT OCCURRED NOT LONG AFTERWARD IS THE FOOD AND DRUG ADMINISTRATION FELT ENOUGH INFORMATION HAD BEEN GATHERED TO DEMONSTRATE THAT THERE WAS A CONCERN WITH EPHEDRA AND THEY PULLED EPHEDRA CONTAINING PRODUCTS FROM THE MARKETPLACE. THAT WAS FOR US THE BIGGEST CHALLENGE. WE HAVE DONE A LOT OF REVIEWS SINCE THEN AND NONE HAVE BEEN AS EXCITING AS WAS EPHEDRA. WE HAVE A LONG HISTORY OF BEING INTERESTED IN ISSUES RELATED TO SAFETY OF DIETARY SUPPLEMENTS. I CALL TO YOUR ATTENTION TWO FAIRLY RECENT WORKSHOPS. N.I.D.D.K.'S DRUG INDUCED LIVER INFORMATION NETWORK. AND WE SPONSORED A WORKSHOP ALONG WITH THE AMERICAN ASSOCIATION FOR THE STUDY OF LIVER DISEASES ON LIVER INJURY FROM HERB AND HERBAL AND DIETARY SUPPLEMENTS. I WOULDN'T HAVE CALLED IT THAT, ACTUALLY. I THINK WE WOULD HAVE PREFERRED TO CALL IT HERBAL DIETARY SUPPLEMENTS. THERE IS NO DISTINCTION BETWEEN HERBAL AND DIETARY SUPPLEMENTS, IN MY VIEW. ALONG WITH NATIONAL TOXICOLOGY PROGRAM LAST YEAR ALSO EVALUATING ASSESSING THE EVIDENCE OF POTENTIAL HEALTH EFFECTS AT HIGH INTAKES OF FOLIC ACID. THIS IS WHERE WE GET A LOT OF INFORMATION. I'M VERY MUCH LOOKING FORWARD TO THE OUT COMES OF THE WORKSHOP TODAY. AND THIS MAY WELL LEAD TO SOME WORK THAT WE CONDUCT AND SPONSOR IN THE OFFICE OF DIETARY SUPPLEMENTS. SO THANK YOU VERY MUCH. [APPLAUSE] >> THANK YOU, PAUL. I WOULD LIKE TO CALL ON DR. LINDA BERNEHAUM. DIRECTOR OF THE NATIONAL TOXICOLOGY PROGRAM TO GIVE US SOME OPENING REMARKS. >> GOOD MORNING, EVERYONE. I THINK WE HAVE HAD A REALLY GOOD INTRODUCTION SO FAR FROM PAUL COATS, WHICH I APPRECIATE BECAUSE IT'S ALLOWED ME TO GO QUITE RAPIDLY AND WE CAN GET INTO THE MEAT OF THE NEXT TWO DAYS AND I'M EXCITED I WILL BE ABLE TO STAND HERE LEARNING WITH YOU AND DOING SOME SCIENCE. IF ANY OF YOU ARE NOT FAMILIAR WITH THE N.T.P. A BRIEF REMINDER OF WHO WE ARE. A CROSS-AGENCY EFFORT IN THE DEPARTMENT OF HEALTH AND HUMAN SERVICES HEADQUARTERED AT N.I.H. AND COMPONENTS OF F.D.A. NATIONAL CENTRE FOR TOX LOGICAL RESEARCH IN ARKANSAS AND N.I.A.S. PART OF THE C.D.C. N.T.P. HAS IS ONE OF THE INTRA MURAL DIVISIONS AS WELL. WHAT IT'S EFFORT IS TO CONDUCT RESEARCH ON COMPOUNDS NOMINATED TO US. YOU MAY SAY NOMINATED BY WHO, THE ANSWER IS ANY AND EVERYONE. WE GET NOMINEES FROM SISTER FEDERAL AGENCIES, WE GET NOMINEES FROM THE PUBLIC AND DIFFERENT STAKEHOLDER GROUPS IN ORDER TO CONDUCT THE DIFFERENT KINDS OF EVALUATION AND TESTING ACTIVITIES THAT WE DO. WE HAVE OVER THE YEARS SINCE IT'S FOUNDING, IT WAS FOUND IN 1978 BY JOSEPH CALIFANO. SINCE THAT TIME OVER THOUSANDS OF AGENTS HAVE BEEN EVALUATED AND COMPREHENSIVE TOX STUDIES FROM GENE TOX ALL THE WAY TO THE TWO YEAR GOLD STANDARD BY LAB RATS AND MICE AND WE COULD HAVE A WHOLE DISCUSSION ABOUT WHETHER THE GOLD STANDARD FOR THE 20th CENTURY IS WHAT WE WANT TO BE DOING IN THE 21st CENTURY. THOSE GOLD STANDARD STUDIES ARE DONE UNDER G.L.P. COMPLIANCE AND FOLLOWING THE GUIDELINES THAT ARE REQUIRED, OFTEN SUPPLEMENTS THROUGH THE STUDIES. IN ADDITION TO THE TESTING ACTIVITIES AND TEST DEVELOPMENT ACTIVITIES WE ARE MANDATED TO DO DETAILED ANALYSIS ACTIVITIES. SO WE ARE LEGISLATIVELY MANDATED IN THE PUBLIC SERVICE HEALTH ACT TO PRODUCE A REPORT ON CARCINOGENS WE NOW ATTEMPT TO DO EVERY OTHER YEAR. STAY TUNED BECAUSE THE 14th REPORT SHOULD BE COMING OUT TOWARD THE END OF THE YEAR. WE HAVE AN OFFICE WE FORMED A NUMBER OF YEARS AGO CALLED OFFICE OF HEALTH ASSESSMENT AND TRANSLATION. THIS IS THE OFFICE THAT IS CONDUCTING MANY OF THE VERY DETAILED SYSTEMATIC REVIEWS OF A VARIETY OF SUBSTANCES AND APPROACHES. THEY ISSUED A REVIEW ON EFFECTS OF LOW LEVEL LEAD. DEMONSTRATING THERE'S REALLY NO SAFE LEVEL BECAUSE THERE ARE EFFECTS ON CHILDREN AND ADULTS FIVE MICRO GRAMS. CHEMO THERIC APPROACH DURING PREGNANCY AND THE THIRD IS TO RUN THAT INTERAGENCY COORDINATING COMMITTEE FOR THE VALIDATION OF ALTERNATIVE TEST MESSAGE WHICH INVOLVES 15 DIFFERENT FEDERAL PARTNER AND OVER SEEN BY THE N.T.P. EVALUATION CENTRE FOR TOX METHODS AND YES WE HAVE LOTS OF ACRONYMS AS ANYONE IN THE FEDERAL GOVERNMENT DOES. WHY IS N.T.P. INTERESTED IN BOTANICALS? MANY THINK N.T.P. IS JUST LOOKING AT CHEMICALS. IF YOU LOOK AT THE SEVERAL THOUSAND SUBSTANCES OR PROCEDURES OR PROCESSES WE HAVE TESTED YOU COULD SEE IT'S MANY MORE THAN JUST INDIVIDUAL CHEMICALS. WE DEVELOP AND APPLY TOOLS OF MODERN TOXICOLOGY, MODERN BIOLOGY. WE ARE LEADERS IN THE WHOLE TOX 21 EFFORT TO DEVELOP HIGH THROUGHPUT AND MID THROUGHPUT DIFFERENT APPROACHES TO EVALUATE COMPOUNDS OR APPROACHES, SUBSTANCES MORE RAPIDLY. NOW PAUL JUST SHOWED DATA SHOWING 50% OF ADULTS USE NATURAL PRODUCTS, SUCH AS BOTANICAL SUPPLEMENTS ON A DAILY BASIS. THE NUMBERS I HAVE ARE A LITTLE LOWER AND IT MAY DEPEND ON SOME DEFINITIONS BEING USED. BUT THE POINT IS WITH A POPULATION OF 300-350 MILLION EVEN 18% OF ADULTS IS A LOT AND 4% OF CHILDREN AND KNOWING CHILDREN ARE A MORE SUSCEPTIBLE POPULATION, I THINK THAT'S SOMETHING WE WITH WE SHOULD BE THINKING ABOUT. IN 1994, IN, I DON'T KNOW ABOUT ANYONE ELSE BUT ALLERGIES ARE REALLY BAD RIGHT NOW. MANDATED REGULATION OF DIETARY SUPPLEMENTS BY F.D.A. BUT THEY DO NOT MANDATE SAFETY TESTING. IT'S DONE ACCORDING TO GOOD MANUFACTURING PRACTICES. SINCE DIETARY SUPPLEMENTS AS PAUL SAID ARE CONSIDERED TO BE FOOD. MOST OF THESE HAVE BEEN IN USE, I WOULD SAY SOME PROBABLY ALMOST AS LONG AS MAN HAS WALKED THE FACE OF THE EARTH. THEY ARE CONSIDERED GENERALLY RECOGNIZED AS SAFE. DO NOT, NOT REQUIRED TO TEST THEM. WE HAVE RECEIVED NOMINATIONS. AS I SAID WE COULD GET THEM FROM ANYWHERE, BUT WE HAVE GOT A LOT OF NOMINATIONS FROM THESE DIETARY SUPPLEMENTS, BOTH FROM PUBLIC AND DIFFERENT FEDERAL AGENCIES. BUT YOU KNOW YOU JUST HAVE TO PICK UP "U.S.A. TODAY" OR "NEW YORK TIMES", WASHINGTON JOURNAL, OR WASHINGTON POST. WHAT IS THAT SHEET WHEN YOU CHECK OUT? "THE DAILY INQUIRER", SOMETHING LIKE THAT. WHAT YOU WILL FIND ALL THE TIME ARE QUESTIONS, BOTH RECOMMENDATIONS AND CONCERNS ABOUT DIFFERENT SUPPLEMENTS. THEY ARE VERY MUCH IN THE NEWS. PAUL MENTIONED EPHEDRA WHICH WE KNOW HAS SERIOUS ADVERSE HEALTH CONSEQUENCES BUT ANY TIME THEY CAN PROVE IT WILL HELP YOU LOSE WEIGHT CUSTOMERS ARE LINING UP IN ORDER TO PURCHASE IT. THEY THINK THAT'S A LOT EASIER THAN EXERCISE IN WATCHING WHAT THEY EAT. BUT I THINK WE ARE BEGINNING TO UNDERSTAND AND THERE HAVE BEEN A NUMBER OF REALLY SERIOUS HEALTH EVENTS, FOR EXAMPLE. MANY OF YOU UNDERSTAND TRYPTOPHANE, THAT LEAD NOT ONLY TO ILLNESS BUT DEATH AND IT INCREASES OUR LEVEL OF UNDERSTANDING. N.T.P. HAS BEEN INTERESTED IN THE SAFETY OF SUPPLEMENTS FOR A LONG TIME. ALMOST 20 YEARS AGO WE SPONSORED A WORKSHOP THAT REALLY LOOKED AT, NEEDS TO LOOK AT POTENTIAL TOXICITY, ASSOCIATED WITH HIGH OR PROLONGED USE. MOST WHO TAKE SUPPLEMENTS IT'S NOT JUST A ONE-TIME THING. OFTEN THEY TAKE IT EVERY SINGLE DAY FOR YEARS AND YEARS. THERE ARE REAL ISSUES WHAT ARE IN THE PRODUCTS YOU USE. WE NEED TO INCREASE OUR CONSUMER EDUCATION THROUGH WHAT'S AVAILABLE IN THE PACKAGES PEOPLE BUY. WE KNOW THERE ARE HERB DRUGS AND HERB-HERB INTERACTIONS. NEVER MIND OVER THE COUNTER AND PRESCRIBED PHARMACEUTICALS, NEVER MIND OTHER THINGS IN YOUR DIET THAT COULD ALTER YOUR RESPONSIVENESS. AND RESPONDERS AND NON-RESPONDERS. WE NEED TO UNDERSTAND WHO ARE THE SENSITIVE POPULATIONS TO SOME OF THESE DIFFERENT EFFECTS. THOSE ARE THE RECOMMENDATIONS 20 YEARS AGO. I WOULD SAY MANY ARE STILL RELEVANT AND WE WILL BE ADDRESSING SOME OF THEM TODAY. ONE OF THE THINGS I THINK WE NEED TO BE THINKING ABOUT IS YOUR MEDICINE WOMAN AND YOUR SHAMAN FOR THOUSANDS OF YEARS HAVE DEALT WITH HERBS AND BOTANICAL SUPPLEMENTS. BUT THEY OVER TRIAL AND ERROR UNDERSTOOD WHAT THE INHERENT TOXICITY WAS AND IT WAS A REAL RESTRICTED POPULATION THAT WOULD BE EXPOSED TO THESE THINGS, AS OPPOSED TO WHAT WE HAVE NOW WHERE ANYONE COULD GO IN AND PURCHASE THINGS OVER THE COUNTER. IF YOU HAVEN'T SEEN ONE OF OUR FACT SHEETS, THIS IS JUST KIND OF A VERY BRIEF OVERVIEW. IT'S GOOD FOR THE PUBLIC. YOU CAN DOWNLOAD IT FROM THE N.T.P. AND N.I.E.H. WEBSITE WHICH TALKS ABOUT BOTANICAL ACTIVITIES AND THINGS WE HAVE ACTUALLY STUDIED. YOU COULD SEE THE LAST ON THE RIGHT IS E EPHEDRA WHICH CLEARLY DAMAGED THE HEART MUSCLE OF THE RATS AND EVIDENCE FROM HUMAN STUDIES. ALOE VERA, WHICH USING IT ON YOUR SKIN MAY NOT BE TERRIBLY HAZARDOUS BECAUSE IT MAY NOT BE ALL ABSORBED BUT IT LEAD TO TUMORS IN MALE AND FEMALE RATS. AND THERE'S EVIDENCE WHEN YOU ARE EXPOSED TO SUNLIGHT AS WELL AS ALOE VERA WE HAVE THOSE INTERACTIONS. BUT I REMEMBER IN AFRICA ON A VACATION AND OUR GUIDE WAS TELLING US ABOUT WHEN HE HAD A BOUT OF MALARIA HE BOILED ALOE VERA ROOT AND DRANK IT BECAUSE IT WOULD HELP HIM GET THROUGH THAT MALARIA EPISODE BUT IT WAS SO BITTER, HIS KIDS HE TOOK TO THE DOCTOR WHEN THEY HAD THEIR MALARIA. HERBS IN FAIRLY HIGH USE IN THE POPULATION ARE BEING SHOWN TO AT LEAST CAUSE CANCER IN ONE OR BOTH SPECIES OF THE TWO-YEAR RODENT STUDIES. SO WE HAVE A LOT OF ONGOING WORK AS FAR AS BOTANICALS. THE BLACK CO HOSH EXTRACT, CONCERN FROM THE ANIMAL STUDIES AND WE ARE ACTUALLY IN A CLINICAL STUDY RIGHT NOW WHERE WE ARE RECRUITING WOMEN WHO TAKE BLACK COHOSH USUALLY FOR MENSTRUAL SYMPTOMS AND WE ARE DOING ANALYSES TO SEE IF WE SEE RESPONSES IN THE BLOOD AS WE SEE IN THE ANIMALS, BIO MARKERS WHICH MIGHT INDICATE POTENTIAL FOR ADVERSE EFFECT IN THE POPULATION. I HAVE TO SAY WITH BLACK COHOSH, YOU GO TO THE DRUGSTORE AND WHAT YOU BUY TODAY IS NOT WHAT YOU BUY TOMORROW. THERE'S JUST A WHOLE LIST OF SOME OF THE DIFFERENT ONE'S WE ARE LOOKING AT. I THINK GREEN TEA EXTRACT IS ESPECIALLY INTERESTING BECAUSE IT'S PROMOTED TO DO ALL KINDS OF DIFFERENT EFFECTS. ANTI CARCINOGENIC. N.T.P. IS HEADQUARTERS AT N.I.E.H.S. MANY OF YOU MAY BE FAMILIAR WITH THE PLAN WE ARE REALLY LIVING ALL OF OUR DIVISION, DIVISION OF N.T.P. INTRA MURAL AND EXTRA MURAL ARE PUTTING THE WORK WE ARE DOING IN CONTEXT OF THE STRATEGIC PLAN AND MIXTURES RESEARCH AND WHAT ARE WE DEALING WITH WHEN DEALING WITH BOTANICAL SUBSTANCE, MIXTURES, ADDRESSES OUR GOAL FOR NONE OF US ARE EXPOSED TO ONE AT A TIME. I THINK WE COULD USE OUR BOTANICALS PROGRAM TO ADDRESS ISSUES HOW DO WE DEAL WITH COMPLEX MIXTURE, ADDRESS DIFFERENT SIMILARITY AND WHAT WE LEARN IS GOING TO ALLOW US TO TACKLE REALLY CHALLENGING PROBLEMS SUCH AS DIFFERENT COMMERCIAL FORMULATIONS AS WELL AS ENVIRONMENTAL CONTAMINANT MIXTURES. SO WITH THAT I WANT TO SAY THANK YOU, AND I'M REALLY LOOKING FORWARD TO A REALLY EXCITING TWO DAYS OF SCIENCE. THANK YOU. >> THANK YOU LIN. LINDA. I WOULD LIKE TO CALL CYNTHIA RIDER. THE DRIVING FORCE. I WANT TO GIVE HER A ROUND OF APPLAUSE PUTTING THIS ONE ON. CYNTHIA WILL GIVE US THE MARCHING ORDERS FOR THE WORKSHOP. >> THANK YOU FOR BEING HERE. THANKS TO LINDA AND PAUL FOR GREAT INTRODUCTIONS, THAT'S PERFECT. BEFORE WE GET INTO TALKING ABOUT SOME LOGISTICS AND INTRODUCING SOME TOPICS IN THE WORKSHOP WE REALLY WANTED TO IDENTIFY WHAT ARE THE KEY CHALLENGES WE WANT TO FOCUS ON. WE THOUGHT IDENTIFYING ACTIVE CONSTITUENTS, DIFFERENT BOTANICALS WERE REALLY CROSS CUTTING THEMES THAT HIT BOTH HAZARD CHARACTERIZATION, TO GIVE YOU A LITTLE OUTLINE OF HOW THINGS ARE GOING TO GO. OUR FIRST SESSION IS TO PROVIDE PERSPECTIVES ON THE CHALLENGES ASSOCIATED WITH BREAULT TAN CALS. -- BOTANICALS. EACH OF THESE WILL BE SIMILAR FORMAT WITH SPEAKERS FOLLOWED BY A PANEL DISCUSSION. FIRST TOPIC ONE WILL BE ON DETERMINING EQUIVALENCE, AND TWO IDENTIFYING ACTIVE CONSTITUENTS AND TOPIC THREE WILL COVER BEST PRACTICES PANEL DISCUSSION WILL HAVE A MODERATOR, THE PANELS WILL BE COMPOSED OF SPEAKERS AND OTHER EXPERTS AND QUESTIONS CAN COME FROM PREPARED QUESTIONS, QUESTIONS FROM THE AUDIENCE WILL BE GREAT AND WEBCAST PARTICIPANTS CAN EMAIL QUESTIONS. WE ARE MONITORING THAT. THE OVER ARCHING GOAL IS TO UNIFORM RESEARCH ON BOTANICAL SAFETY. WE HOPE TO COMMUNICATE THE CURRENT SCIENCE AND KEY TOPIC AREAS FEEDBACK AND IDENTIFY GAPS IN RESEARCH NEEDS SOME LOGISTICS IF YOU WOULD PLEASE SILENCE YOUR MOBILE DEVICES PLEASE USE MICROPHONES SO IN PERSON AND WEB AUDIENCE CAN HEAR YOUR QUESTIONS, UNFORTUNATELY THERE'S NO FOOD OR BEVERAGE BUT THERE ARE PRODUCTS AVAILABLE, OPEN FROM 7-3 AS WELL AS ACROSS THE STREET AT NATUR CENTRE FROM 6:30 TO 2:30. IF YOU HAVE FOLLOW-UP QUESTIONS MARY WOLF IS HERE TO LIAS FOR ANYONE WHO WOULD LIKE TO TALK FURTHER ABOUT THESE TOPICS AND N.T.P. THERE'S A LOT OF PEOPLE TO THANK. THIS HAS BEEN A PROCESS. THE BOTANICAL WORKSHOP PLANNING COMMITTEE. JOSEPH FROM N.I.H.O.D.S., PAUL HOWARD, REPRESENTED BY SUSAN KARLSSON, SUZANNE FITZPATRICK AND LEA ROSENFELD. BIG TEAM OF N.I.E.S. FOLKS WE CARAVANED HERE TO ATTEND. WE WOULD REALLY LIKE TO THANK THE OFFICE OF DIETARY SUPPLEMENTS FOR FUNDING OUR CASE STUDY DEVELOPMENT. WE WILL TALK IN LESSON ONE. ALL HAD A PART IN PUTTING TOGETHER THE CASE STUDIES. I WOULD LIKE TO THANK POST TRAINEES. KRISTAN RYAN AND KELLY, KRISTEN IS NO LONGER A POST DOC. THANKS TO THEM THEY WILL BE ANSWERING QUESTIONS AND PROVIDING SUPPORT I WOULD LIKE TO THANK OUR SUPPORT STAFF. FOR ALL THINK HAVE DONE TO PUT TOGETHER THE WORKSHOP AND LIFTER HILL CONTACTS MELISSA INCLUDING PAT. A BIG THANKS TO ALL THE SPEAKERS, PANELISTS AND MODERATORS PARTICIPATING IN THE MEETING AND ALL THE MEETING ATTENDEES IN PERSON AND WEBCAST. WE HOPE YOU ENJOY THE WORKSHOP. THIS IS GREAT, WE ARE STILL ON TIME. GREAT WHEN YOU ARE MODERATOR MAKING SURE EVERYONE IS ON TIME. IN ORDER TO DO THAT FOR THE FOLKS ON OUR SESSION WE HAVE THESE CARDS. WHEN YOU ARE GETTING CLOSE I WILL GIVE YOU THREE MINUTES AND WHEN YOU ARE REALLY CLOSE I WILL GIVE YOU A WRAP UP. JUST TO MAKE SURE WE GET THROUGH THE DAY. PERSPECTIVE ON CHALLENGES ASSOCIATED WITH BOTANICALS AND I WOULD LIKE TO INTRODUCE DR. JOE BETS FROM O.E.S., JOE IS PROBABLY A FAMILIAR SITE. HE IS THE DIRECTOR OF DOCTOR SUPPLEMENTS AND MATERIALS MATERIALS AT O.D.S., HAS A LONG HISTORY IN DEALING WITH COMPLEXITY OF BOTANICALS THIS. SHOULD BE A VERY GOOD OPENING SESSION. GOOD MORNING, EVERYBODY. WE WILL WIZ THROUGH THIS PRESENTATION AND HOPEFULLY SET THE STAGE FOR THE REST OF THE WEEK. WE WILL GO STRAIGHT TO SOME REGULATORY STUFF. WHAT ARE DIETARY SUPPLEMENT PRODUCTS MADE OF? THESE ARE DEFINITION THAT'S COME OUT OF THE PREAMBLE TO THE GOOD PRACTICES FOR GOOD MANUFACTURING FOR DIETARY SUBSTANCES. FOR ANYTHING THAT'S COME IN CONTACT WITH YOUR PRODUCT *. THESE INCLUDE USING INSOLVENTS LATER REMOVED FROM THE PRODUCT BEFORE YOU MANUFACTURE IT. WE WILL COVER SOME THINGS TALK THERE MAY BE SOMETHING LEFT EVEN THOUGH IT'S NOT INTENDED TO BE IN YOUR FINISH PRODUCT. IT WON'T BE ON A LABEL, THESE ARE THE SOURCE OF THINGS YOU HAVE TO THINK ABOUT. INGREDIENTS ARE ESSENTIALLY, THEY ARE BROKEN DOWN INTO TWO CATEGORIES, DIETARY INGREDIENTS, EXCIPIENTS, BINDERS, COLOURING AGENTS MATERIALS USED TO COAT YOUR TABLET. THE SHELL YOUR CAPSULE IS MADE OF, ETC.. DIETARY. YOU COULD SEE THERE'S A BUNCH OF THINGS THAT ARE DIETARY INGREDIENTS ANY PRODUCT OTHER THAN TOBACCO THAT CONTAINS THOSE, DR. COATES GAVE A LITTLE OF THESE EARLIER. DR. HOP WILL TALK HOPEFULLY ABOUT PRODUCT INTEGRITY AND HOW IMPORTANT THAT IS TO DOING N.I.H. SPONSORED RESEARCH ON DIETARY SUBSTANCE, BOTANICALS IN GENERAL. THE FIRST THING TO NOTE IS PHYTO CHEMISTRY IS AVAILABLE. THE AMOUNTS COULD VARY FROM BATCH TO BATCH OF HERB. FROM THE BASED ON GEOGRAPHY, CLIMATE, ENVIRONMENTAL STRESS, DROUGHTS, EXCESS RAIN IN AN AREA WHERE THE THINGS ARE GROWN AND THERE ARE NUMEROUS, NAMELESS VARIABILITY IN PLANT CHEMISTRY THAT WE ARE ONLY STARTING TO BEGIN TO CAPTURE. IT'S DIFFICULTY TO MEASURE AND MAINTAIN. COMPANIES SET THEIR OWN SPECIFICATION. THEY AREN'T REQUIRED TO USE MANDATORY SPECIFICATIONS. AND DIFFERENT COMPANIES MAY SET DIFFERENT SPECS. THE PROCESS BY WHICH THE MATERIAL IS MANUFACTURED DEFINES THE PRODUCT. WE TALKED ABOUT INTEGRITY, NEEDED FOR RAW MATERIALS NEED TO KNOW WHAT PLANTS, THERE ARE OTHER ISSUES PEOPLE MAY OR MAY NOT CONSIDER. PURITY, QUALITY, COMPOSITION AND STRENGTH OF THE MATERIAL THAT GOES INTO YOUR PRODUCT. WE TALK ABOUT IDENTITY AND THINGS GET MORE COMPLICATED. GOLDEN SEAL ON YOUR UPPER LEFT THERE IS FAIRLY EASY TO IDENTIFY WHEN YOU HAVE IN TACT LEAVES AND FLOWERS. IT'S STILL FAIRLY EASY TO IDENTIFY WHEN YOU PULL UP FRUITS AND RYE SOMES BUT WHEN YOU GRIND IT UP AND MAKE IT IN A POWDER, ALL THOSE FEATURES ARE LOST THEN YOU ARE DEPENDING ON MICRO SKOPY AND CHEMISTRY AND BY THE TIME YOU GET TO THE BROWN GOO IN THE CENTER THAT'S AN EXTRACT YOU ARE RELYING ON CHEMISTRY RATHER THAN BOTANICAL MORPHOLOGY. WE SWITCH TO PURITY. YOU WANT TO MAKE SURE YOU HAVE THE TARGET SPECIES AND PLANT PART ONLY. YOU ARE REQUIRED TO IDENTIFY THE SPECIES YOU ARE USING IN THE MATERIAL BUT ALSO THE PLANT PART IT WAS MADE OUT OF. WE HAVE SEEN ADULTATION OF GINSING ROOT. MICROPIA THERE ARE RECALLS AS WELL AS ARTICLES, PESTICIDES FOR DIETARY SUPPLEMENTS. MICRO TOXINS, F.D.A. 20 PART PER BILLION DOESN'T REALLY PUT A NEW MER CAL LIMIT ON THE MICRO TOXINS IN NATURE. ONE HAS TO KEEP AN EYE OUT FOR OTHER TOXINS. THEY ARE NATURALLY OCCURRING IN BUTTERBUR, THEY ARE PRESENT IN A NUMBER OF OTHER MEDICINAL. THERE ARE GUIDELINES, METHODS, TOLERANCES YOU DON'T WANT CADMIUM, LEAD, SINCE THEY GROW IN THE GROUND AND SOME OF THESE PLANTS ARE ACCUMULATORS OF TOXIC ELEMENTS THIS IS SOMETHING YOU HAVE TO KEEP IN MIND. AND THEN FINALLY UNDER CLEAR AND SYNTHETIC SUBSTANCES DR. BIRNBALM'S SLIDE. IT'S EASY TO THINK OF IMPURITY, TOXIC ELEMENTS AND PESTICIDES BUT THERE'S ALSO A PURITY ISSUE IN TIMES OF THE PLANT MATERIAL THAT'S IN YOUR PRODUCT. A SPECIFICATION FROM THE JAPANESE PHARMACOPEDIA, SAYS THAT THE PURITY FOR THE EPHEDRA, WOODY STEMS SHOULD NOT EXCEED 5% AND EPHEDRA HERB DOESN'T CONTAIN STEMS. THEY ARE JUST GRASS. LAWN GRASS OR OTHER FOREIGN MATTER. THERE'S ALSO OTHER, THE PLANT TOXINS, THE NATURALLY OCCURRING TOXINS OR OTHER THINGS YOU HAVE TO START THINKING ABOUT INTRODUCED CONTAMINANTS, SO PURELY THEY ARE A CONCERN, THIS ARTICLE PUBLISHED IN 2014 THEY WERE DISCOVERED RELATIVELY LOW LEVELS IN THINGS LIKE GREEN TEA, CHAMOMILE, FENNEL AND MINUTE. SOME, OTHERS ARE SOLD AS CONVENTIONAL TEAS AND TEA BAGS. IT'S SUSPECTED MOST OF THESE ALKALOIDS GROW AS WEEDS AND FIELDS HARVESTED, BOTANICKLY HARVESTED. WHEN YOU MECHANICALLY HARVEST A FIELD OF CHAMOMILE, YOU MAY BE HARVESTING IT ALONG WITH CHAMOMILE AND THAT ENDS UP IN YOUR PRODUCT. MOST MATERIALS IN COMMERCE ARE EXTRACTS. WE REMEMBER THE BOILER PLATE, RATHER THAN BIO MASS SO INSTEAD OF PLANT MATERIAL MOST OF THE THINGS IN COMMERCE ARE EXTRACTS. THERE IS A SPECIFIC TERMINOLOGY ASSOCIATED WITH EXTRACTS, LIQUID EXTRACTS ARE WHAT YOU WOULD EXPECT. YOU TAKE SOME GOLDEN SEAL AND MIX IT WITH ETHANOL AND SOME WATER AND YOU END UP WITH A SOLUTION THAT'S GOT ETHANOL AND WATER SOLUBLE EXTRACTABLES. WHEN YOU REMOVE THAT SOLVENT YOU END WITH THE GUMMY MATERIAL CALLED A NATIVE EXTRACT AND THEN THE TERMINOLOGY, AND THIS GETS CONFUSING. THE INDUSTRY USES THE TERM EXTRACT TO DESCRIBE THE NATIVE EXTRACT AFTER IT'S BEEN SPRAY DRIED ONTO A CARRIER. SOME COMMON CARRIERS ARE MALL TO DEX TRINH METHO CELLOSE. WHEN YOU ARE DEALING WITH THESE INGREDIENTS YOU HAVE TO KNOW THE MATERIAL YOU ARE DEALING WITH. THIS IS TRUE WHETHER YOU ARE MANUFACTURER OR WHETHER IN AN F.D.A. SITUATION DOING FORENSIC ANALYSIS YOU HAVE TO KNOW WHAT THE MATERIAL IS MADE OUT OF. IT SHOULD BE PART OF ANY SPECIFICATION SET IN YOUR G.M.P. YOU AREN'T NECESSARILY RECEIVING St. JOHN'S WORST YOU ARE RECEIVING AN EXTRACT OF St. JOHNS WORT SPRAY DRIED ON SOMETHING AND THAT'S IMPORTANT TO KNOW. THESE ARE EXTRACTS IN OLD T.C.M. AND OTHER SORTS OF TONES REFERRED TO AS TRADITIONAL. THEY INCLUDE THINGS LIKE MILLING, OR GRINDING. THEY ARE REALLY OLD SCHOOL HYDRO ALCOHOLIC SOLVENTS, THINGS LIKE WINE, EVEN VINEGAR IN SOME CASES FILED BY FILTRATION OR DECANTATION. CONCENTRATION, OR DRYING, LETTING IT EVAPORATE. AND THESE ARE MORE OR LESS REPRESENTATIVE OF THE NATIVE PLANT. SO WHATEVER YOU CAN GET OUT AND WATER AND DILUTE ALCOHOL IS WHAT YOU WILL GET OUT OF THESE EXTRACTS. THE MORE MODERN EXTRACTS ARE STANDARDIZED EXTRACTS. HAS DIFFERENT TYPES OF MODERN EXTRACTS. STANDARDIZED IS AN EXTRACT MANUFACTURED. I PUT THE WORD MANIPULATED TO LET YOU KNOW PEOPLE ARE MESSING AROUND WITH THE COMPOUNDS TO GET THE SAME AMOUNTS TO CONTAIN A DEFINED AMOUNT OF COMPOUND OR GROUP OF COMPONENTS CALLED MARKER COMPOUNDS. MANIPULATION DONE THROUGH SELECTIVE EXTRACTION, ETC.. THIS ASSURES BATCH TO BATCH REPRODUCABILITY. THERE'S NORMALIZED EXTRACTS WHICH ARE SIMILAR INSTEAD OF KNOWN ACTIVE, FOR EXAMPLE BELLA DONNA EXTRACT WE WOULD STANDARDIZE TO ATROPINE. YOU THINK YOU HAVE A HANDLE BUT YOU AREN'T QUITE SURE. QUANTIFIED EXTRACT WHERE YOU MEASURE JUST BECAUSE YOU CAN. NOT NECESSARILY BECAUSE WE HAVE ANY SUSPICION ABOUT THE BIOLOGICAL ACTIVITY. THE INTERESTING PART THERE ARE TWO CATEGORIES WHERE YOU DON'T 100% KNOW WHAT THE ACTIVE BIOLOGICAL EXTRACT IS. THERE ARE SOME PLANTS WE KNOW THE ACTIVES AND PHARMACOLOGY OF THOSE COMPOUNDS, ATROPINE, ETC. BUT THERE ARE A LARGE NUMBER OF PLANTS WE SIMPLY DON'T KNOW WHAT THE BIOLOGICALLY ACTIVE CONSTITUENTS ARE. AT ANY RATE THE EUROPEAN UNION TREATS IT AS THE PHARMACEUTICAL INGREDIENT RATHER THAN ANY PARTICULAR COMPOUND OR SET OF COMPOUNDS. MARKER COMPOUNDS IS THE TERM BROUGHT UP. ONE OR MORE NATURALLY OCCURRING CONSTITUENTS OF THE PLANT. CHEMO TAXONOMY WHERE UNIQUE COMPOUNDS WERE USED TO IDENTIFY OR ESTABLISH PLANT IDENTITY IN PHILO GENETIC RELATIONSHIPS. SOME COMPOUNDS UBIQUITOUS, FLAVANOIDS. St. JOHN'S WORT. BY LOOKING FOR THESE UNIQUE COMPOUNDS YOU COULD TRY TO GET A HANDLE ON THE IDENTITY. THERE'S A CLASSIC REFERENCE WHICH I BELIEVE IS AVAILABLE IN ENGLISH TRANSLATION NOW BY RICHARD HEADNOUR. NOT JUST ABOUT BIOLOGICAL ACTIVITY. THEY ARE ABOUT IDENTITY AND CONSISTENCY OF FINISHED PRODUCTS. WE TALKED ABOUT STRENGTH EARLIER. WE EXPRESSED THE AMOUNT OF THE UNIT OF HERB USED FOR UNIT OF EXTRACT. YOU WANT TO TRY TO REPRESENT THE ACTUAL AMOUNT OF HERB THAT VOLUME. 30 MILLILITERS OF EXTRACT REPRESENTS. THERE ARE 1:1 DILUTIONS, 50:1 ETC. THAT TENDS TO BE USED AS A PHARMACOLOGICKAL TERM. IT'S EASIER TO GET THE STRENGTH, YOU TELL HOW MUCH HERB WAS USED VERSUS HOW MUCH SOLVENT. POTENCY COULD BE RELEVANT IN STANDARDIZED EXTRACT BECAUSE THERE IT'S KNOWN. WE TALK ABOUT VARIATION IN THE PLANT. A LOT OF THESE MATERIALS ARE PROPRIETARY. DOZENS OF COMPANIES THAT MAKE THE EXTRACTS AND THEY ALL HAVE THEIR PROPRIETARY PROPERTIES. COFFEE. THERE ARE DIFFERENCES IN HOW THE PRODUCTS ARE PRODUCED, STARTING RIGHT AT THE BEGINNING. SO HOW ONE DRIES OR ROASTS THE COFFEE BEANS WILL HAVE AN EFFECT ON THE CHEMISTRY OF THE COFFEE BEANS. THE COMPOSITION. HOW THEY ARE GROUND OR MILLED WHEN YOU DO YOUR EXTRACTION. HOW EFFICIENT THE EXTRACTION PROCESS IS FOR SOME COMPOUNDS AND OTHERS. HE ESPRESSO GRINDS, COURSE GRINDS MEDIUM GRINDS ETC.. WE CAN TALK ABOUT STRENGTH AND WHEN YOU START YOUR KRUPPS IN THE MORNING, I PUT THREE BIG SCOOPS OF COFFEE IN THE CONTAINER, IN THE FILTER. MY WIFE CAN'T DRINK THAT COFFEE BECAUSE IT'S TOO STRONG FOR HER SO SHE PUTS ONE SCOOP FOR THE SAME AMOUNT OF WATER. IMAGINE IF YOU COULD CHOOSE TO EXTRACT WITH SOMETHING BESIDES HOT WATER. COFFEE USES HOT WATER. WE ARE TALKING ABOUT EXTRACTS MADE WITH ETHANOL, ACETONE OR CO2. YOU COULD IMAGINE THE CHEMISTRY OF THE FINISHED EXTRACT WILL BE CONSIDERABLY DIFFERENT CONSIDERING WHAT SOLVENT YOU USE TO MAKE YOUR EXTRACT. IT'S INSTANT COFFEE, THE LITTLE CRYSTALS OF INSTANT COFFEE. THAT'S A FLOWABLE PRODUCT. NATURAL PRODUCT TEND TO BE GUMMY AND HARD TO MANIPULATE UNLESS YOU HAVE A CARRIER OR WAY OF MANIPULATING SO YOU GET A NICE DRY POWER FLOWABLE. YOU CAN'T MAKE CAPLETS OUT CH THE GOOEY GUMMY STUFF. WE GO FROM GENERAL EXAMPLE TO A MORE SPECIFIC EXAMPLE. GINKGO. A PROFESSOR WORKED FOR A WHILE AT ENDINA AND AGREED TO SHARE SOME SLIDES WITH ME. WE TAKE GINGKO. THOSE ARE BEAUTIFUL LEAVES. EASY TO IDENTIFY EVEN IN THE LEAF STAGE. THE CHEMISTRY OF GINGKO. THEY MEASURE THE CONTENT, FLAF NOL GLYCE SIDE. GINKOLIC ACID YOU WANT TO KEEP AT A MINIMUM. YOU MEASURE AND ADJUST AT LEAST TEN DIFFERENT COMPOUNDS. IT TURNS OUT THAT THOSE NUMBERS, THOSE SPECIFIC NUMBERS DON'T OCCUR WHEN THE GINGKO LOOKS LIKE THAT. THEY OCCUR WHEN THE GINGKO LOOKS LIKE THAT. SO IN THE FALL JUST BEFORE THE GINGKO LEAVES FALL OFF THE TREE IS WHEN THOSE PHYTO CHEMICALS HAVE THOSE RATIOS AND AMOUNTS. THERE YOU COULD SEE THE VARIATION. IN THE SPRING THE FLAVANOID GLYCOSIDES LOOK LIKE THIS. YOU COULD DO PATTERN RECOGNITION. METRIC ANALYSES. THIS IS JUST A GENERAL PROTON AN MORE SPECTRUM OF WHOLE GINGKO EXTRACTS. SO THERE ARE VARIOUS BATCHES OF THE ENDINA BRANDED GINGKO ABOUT A BALOBA. THERE ARE DISTINCTIONSES BETWEEN THE DIFFERENT EXTRACTS. SO IF YOU DO SOME PATTERN RECOGNITION, SOME ARTIFICIAL INTELLIGENCE EVALUATION YOU COULD SEE REAL DIFFERENCES. THERE'S NO ATTEMPT TO TRY TO IDENTIFY COMPOUNDS WE ARE JUST LOOKING AT PATTERNS, FINGERPRINTS. IF YOU TAKE THOSE COMMERCIAL PRODUCTS ALL MADE BY ONE MANUFACTURER YOU COULD SEE THEY CLUSTER IN ONE AREA. WHEREAS GINGKO PRODUCTS FALLOUT SIDE OF THAT CLUSTER. YOU COULD SEE THE RED SQUARES, THE RED BOXES ON THE SLIDE. IT'S COMPLEX. IT'S HARD. IT'S NOT IMPOSSIBLE. IT'S JUST HARD AND EXPENSIVE TO TRY TO CHARACTERIZE THESE MATERIALS AS MUCH AS YOU REALLY WANT TO. HOW DO YOU COMPARE EXTRACTS, HOPEFULLY WE WILL GET SOME ANSWERS DURING THE COURSE OF THE DAYS. YOU NEED TO KNOW HOW THE MATERIAL YOU ARE STUDYING IS MADE AND HOW THE OTHER STUFF IS MADE, IN A WORLD OF PROPRIETARY EXTRACTS WHERE LITTLE OF THIS INFORMATION PATENTS IS AVAILABLE IN THE PEER-VIEW LITERATURE THAT'S REALLY HARD. IT MAY INVOLVE TRADE SECRETS AND LAWYERS. YOU HAVE TO BE PREPARED TO BE DISAPPOINTED OR ASSIGN CONFIDENCALITY AGREEMENTS OR BOTH IF YOU ARE GOING TO WORK WITH THESE MATERIALS. HOPEFULLY WHAT WE WILL BE GETTING TO LATER TODAY AND TOMORROW IS HIGHER ORDER METHODS TO CHARACTERIZE A MATERIAL. THIS MAY INCLUDE ENLISTING CHEMISTS, BIO ASSAY AND STATISTICIANS AS CO AUTHORS. AGAIN YOU MAY NOT GET TO A SATISFACTORY ANSWER WITHOUT A FAIR AMOUNT OF TIME AND ENERGY AND RESOURCES BEING SPENT. THIS IS TO SET THE STAGE, HOPEFULLY THE FIRE DRILL DIDN'T PUSH US TOO FAR BEHIND. I HOPE THE NEXT COUPLE DAYS WILL BE USEFUL AND INFORMATIVE. SO THANK YOU VERY MUCH. >> IF THERE ARE A COUPLE QUESTIONS WE DO HAVE TIME, I WANT TO MAKE SURE EVERYBODY HAS A CHANCE FOR AT LEAST ONE QUESTION. HERE IN THE FRONT. BE SURE TO USE YOUR MIC. >> ARE BIO MARKER ARE-IVE BIO ACTIVE? >> THEY AREN'T ALWAYS. WE WOULD LIKE TO GUESS THEY ARE BUT THAT'S NOT ALWAYS PROVEN. >> THE FIRE DRILL TOOK US ABOUT 15 MINUTES BEHIND. WE WILL TAKE FIVE MINUTES OFF THE BREAK AND TAKE TEN MINUTES OFF FOR LUNCH, WE HAD AN HOUR FOR LUNCH, THERE'S LUNCH DOWNSTAIRS OVER THE ROAD, 50 MINUTES WILL BE MORE THAN ENOUGH, TRUST ME. THE NEXT SPEAKER, DR. CARA WELCH FROM THE U.S. F.D.A. ONE OF THE SENIOR ADVISORS IN THE OFFICE OF DIETARY SUPPLEMENTS PROGRAM. SHE WILL TELL US ABOUT THE F.D.A. REGULATORY LANDSCAPE. >> GOOD MORNING, EVERYONE. SO OFFICE OF DIETARY SUPPLEMENTS PROGRAM IS EXCITED ABOUT THIS MEETING. THERE'S ALWAYS RESEARCH THAT NEEDS TO BE DONE. WE TRY TO DO AS MUCH AS WE CAN ON OUR END BUT AS MUCH AS WE CAN TO AID THE RESEARCH BEING DONE IN OTHER AREAS. WE ARE LOOKING FORWARD TO THE NEXT COUPLE DAYS AS WELL. IF YOU ALL HAVE BEEN FOLLOWING THE NEWS OR AT LEAST F.D.A. NEWS WE DON'T ALWAYS GET INTO MAINSTREAM MEDIA BUT F.D.A. CREATED THE OFF OF DIETARY SUPPLEMENTS PROGRAM. IN LATE DECEMBER OF 2015, I THINK IT ACTUALLY BECAME OFFICIAL. IT WAS TIME BECAUSE OF THE MASSIVE GROWTH WITHIN THE INDUSTRY. VARIOUS NUMBERS HAVE BEEN THROWN AROUND AS MUCH OF A GROWTH OF TEN TIMES SINCE DESHEA. $40 BILLION INDUSTRY $40 MILLION INDUSTRY TO A $40 BILLION. WE ARE RESPONDING TO THE GROWTH AND PUTTING US IN THIS POSITION OF AN OFFICE AT LEAST PUTS US IN A BETTER PLATFORM TO ADVOCATE FOR RESOURCES FOR ACTIVITY. FOR MONEY. ALSO DIETARY SUPPLEMENTS PROGRAM WE ARE THE PROGRAM OFFICE FOR DIETARY SUPPLEMENTS. WE ARE THE REGULATORY CENTER WITHIN THE F.D.A. FOR ACTIONS REGARDING DIETARY SUPPLEMENTS. HOWEVER, WE WORK WITHIN THE CENTER FOR FOOD SAFETY AND APPLIED NUTRITION. VARIOUS OFFICES COME TOGETHER TO DO PRETTY MUCH EVERYTHING. WE ALSO WORK WITH OTHER CENTERS AND THE OFFICE OF REGULATORY AFFAIRS WHICH IS A MASSIVE AREA OF F.D.A. ALL OF OUR DISTRICT OFFICES AND INVESTIGATORS, AND LABS THAT MATTER OBVIOUSLY PLAYS A HUGE ROLE IN THE REGULATION OF DIETARY SUPPLEMENTS AS WELL GIVEN THEY ARE OUR INVESTIGATORY FORCE. ONE THING IS STRATEGIC REVIEW. WE NEED TO RE-REVIEW. ARE WE USING OUR AUTHORITY, RESOURCES TO THE BEST OF THEIR ABILITY. IS THERE SOMETHING WE DON'T NEED TO FOCUS ON ANY MORE. WHERE DO WE WANT TO MOVE OVER THE NEXT FIVE YEARS THESE ARE OUR CONSTANT PRIORITIES. PROTECTION OF PUBLIC HEALTH. WE WILL TALK ABOUT COMPETING PRIORITIES GENERALLY LEANS HEAVY ON PROTECTION OF PLK HEALTH. REMOVE IT FROM THE MARKET TO ADDRESS IT, TO ADJUST MANUFACTURING IF NEED BE, TO ADJUST JUST TO RESEARCH THE PRODUCT OF COMMERCE. RESEARCH WHAT IT SHOULD BE. WE NEED TO ENSURE PRODUCT IDENTITY AND WE HAVE BEEN WORKING ON THE NEXT FEW YEARS, SIGNIFICANTLY OVER THE LAST YEAR. WE HAVE PUT OUT A NUMBER OF ACTIONS, WARNING LETTER, BLITZES SO TO SPEAK ON CERTAIN INGREDIENTS WE BELIEVE SHOULDN'T BE IN DIETARY SUPPLEMENTS FOR ONE REASON OR ANOTHER. RESEARCH TO ENSURE WHAT THE LABEL SAYS ON THE PRODUCT IS ACTUALLY WHAT THE PRODUCT CONTAINS AND WHAT CAN WE DO TO ENSURE THAT. BECAUSE IT'S A QUESTION I'M ALWAYS ASKED I JUST PUT IT ON THE SLIDES NOW. IT'S A PRIORITY FOR US TO PUBLISH AS SOON AS POSSIBLE. ENFORCING G.M.P. COMPLIANCE. FLEXIBILITY TO GET INTO A FIRM, FLEXIBILITY AND SEE HOW THEY CAN DO WHAT THEY ARE DOING. SETTING OUT, DO THEY KNOW WHAT THEY ARE MAKING, SPECIFICATIONS, PROCEDURES, DO THEY HAVE RECORDS IN PLACE BEFORE THEY START MANUFACTURING PRODUCT. ARE THEY MANUFACTURING TO THOSE SPECIFICATIONS. ARE THEY KEEPING IT TO DEMONSTRATE IT. KEEPING RESERVE PRODUCT IF A PRODUCT CROPS UP THEY CAN TEST THAT PRODUCT AND ENSURE ANY ISSUE THAT'S CAME UP RESULTS OF THAT PRODUCT OR ANYTHING ELSE. ENFORCING G.M.P. COMPLIANCE IS A HUGE AREA AND I WILL FINISH OFF MY TALK ON THAT. ANOTHER CONSISTENT SLIDE I GIVE ARE THE PROBLEMS WE ARE SEEING IN INDUSTRY AND CERTAIN AREAS OF MANUFACTURING PRACTICES. FIRST AND FOREMOST PROTECTING THE PUBLIC HEALTH. THIS IS OUR TOP PRIORITY. THIS IS THE REASON THAT WE WILL ADJUST OUR PRIORITIES ON THE FLY. SOME OF THE RESEARCH F.D.A. HAS BEEN WORKING ON TAKING IN VITRO RESULTS HOW CAN WE LINK, VERY MUCH THE NEXT DAY AND A HALF IS VERY IMPORTANT TO US. HOW COULD WE BREAK DOWN BOTANICAL RESEARCH, HOW DO WE KNOW THE RESULTS AND ACCURATELY USE THOSE RESULTS. LIVER TOXICKITY FOR ADDITIONAL PROBLEMS. IT'S THE RESULT OF AN AREA ALREADY MENTIONED A COUPLE TIMES BUT STIMULANTS, HIDDEN INGREDIENTS IN PRODUCTS. WHAT COULD BE A LEGITIMATE BOTANICAL EXTRACT COULD BE, MAYBE THEY ARE NATURALLY OCCURRING STIM ULANTS OR MAYBE NATURALLY ADDED IN. WE HAVE BEEN WORKING ON OTHER AREAS. YOU MAY HAVE SEEN SOME EFFORTS ON OUR END TO REMOVE BOTANICAL DIETARY INGREDIENTS FOR BETTER OR WORSE. CRATEM. IT'S A NEW DIETARY INGREDIENT FOR WHICH THERE IS NO NOTIFICATION, IF IT SHOWS UP IN A DIETARY SUPPLEMENTS, IT IS AN ADULTERATED DIETARY SUPPLEMENTS. WE HAVE ISSUED AN IMPORT ALERT THAT'S BEEN IN PLACE FOR OVER TWO YEARS NOW. AND THAT IMPORT ALERT ANYTHING COMING INTO THE COUNTRY AS DIETARY SUPPLEMENTS CONTAINING PRODUCT OR BULK INGREDIENT WE WILL FLAG IT AND PUT IT ON OUR IMPORT ALERT TO DATE, THIS WAS ABOUT A MONTH AGO NOW, THERE WERE 101 FIRMS LISTED ON THAT IMPORT ALERT. OVER TWO YEARS THE NUMBER OF PRODUCTS WE HAVE BEEN ABLE TO OBTAIN WITHOUT PHYSICAL EXAMINATIONS. CRATUM IS NOT AN ILLEGAL DRUG. THE SECTION OF THE ACT WE REFERENCE WOULD BE SECTION 402-F1B. WE USE THAT TERM OFTEN. I APOLOGIZE IF I REVERT TO THAT. WE HAVE ALSO TAKEN SOME ACTION DOMESTICALLY. WE DISCOVERED THAT PRODUCT IS GETTING IN. TO THE BEST OF OUR ABILITY WE AREN'T STOPPING ALL OF IT. WE ARE FOCUSING OUR EFFORTS DOMESTICALLY. IN JANUARY 2016 WE DID A SEIZURE ABOUT $400,000 WORTH OF MATERIAL IN ILLINOIS. AND THERE WAS A NICE AMOUNT OF PRESS ON THAT. IT'S IMPORTANT TO GET THAT OFF THE MARKET AND GET OFF THE PRODUCT VERY CLEAR. PURE POWDERED CAFFEINE. I'M TALKING ABOUT BULK AMOUNTS. PURE CAFFEINE POWDER SOLD DIRECTLY TO CONSUMERS *. THE PRODUCTS WERE LISTED AS DIETARY SUPPLEMENTS, AND LABELLED TO, THE SERVING SIZE WAS SUPPOSED TO BE 1/16 OF A TEASPOON. ACCORDING TO OUR RESEARCH AND DETERMINATION THE AMOUNT OF FACE AMOUNT AND TOXIC AMOUNT WAS VERY SMALL SO WE DID SEND OUT SOME WARNING LETTERS SAYING WE BELIEVE IT PRESENTED SIGNIFICANT RISK OF ILLNESS OR INJURY TO CONSUMERS. THAT SPECIFIC LANGUAGE, EARLIER I SAID 402-F1A. SAME SECTION WE USE FOR EPHEDRA ALPHA. WE BELIEVE THE MAJORITY OF PRODUCT HAS BEEN PULLED OFF THE MARKET BUT THERE ARE PEOPLE WHO WILL TRY TO SELL ANYTHING AND THE INTERNET ISN'T ALWAYS OUR FRIEND ON THAT END. SUFFICE IT TO SAY WE DON'T CONSIDER BULK PURE POWDERED CAFFEINE TO BE A SAFE DIETARY SUPPLEMENTS. ALSO FOCUSING ON ENSURING PRODUCT INTEGRITY. WE HAVE PUT A LOT OF EFFORT INTO WARNING LETTERS. WE PUT OUT ON SPECIFIC INGREDIENTS BUT THERE'S ANOTHER END TO ENSURING PRODUCT INTEGRITY. WHEN YOU LABEL ON THE PRODUCT IS IT ACTUALLY THAT INGREDIENT AND WHEN YOU LABOR A CERTAIN AMOUNT, STANDARDIZED TOOL AMOUNT ARE YOU PROVIDING AS MUCH OF THAT PRODUCT. THIS ISN'T ALWAYS THE EASIEST WORK ON OUR PART. WE MAY NOT HAVE THE APPROPRIATE METH DOLG. WE ARE DOING OUR BEST ON THAT. ORIGINALLY PRESENTED AT A CONFERENCE, LOOKING BOTH PRESENTED AT A CONFERENCE. BOTANICAL INTEGRITY, LOOKING AT SPECIFIC STANDARDIZED EXTRACTS AND ARE THEY IN THE PRODUCT IN THE AMOUNT THEY SHOULD BE. WE HAVE LOOKED AT PROBUY OTIC INTEGRITY. NOT THE FOCUS ON THIS PRESENTATION OR NEXT DAY AND A HALF BUT PROBUY OTIC IS A HUGE SOURCE OF THE DIETARY SUPPLEMENTS INDUSTRY. WE HAVE DONE RESEARCH LOOKING AT PROBUY OTIC, SPECIFICALLY WHAT THEY LABEL ON THE PRODUCT IS THAT MICROBIALS THAT ARE IN THE PRODUCT. WE ARE INTERESTED IN ALL THINGS PROBIOTICS AND WILL CONTINUE TO MOVE RESEARCH FORWARD ON THAT END. AS I MENTIONED BEFORE, SOME OF THE LETTERS THAT WE HAVE ISSUED ON SPECIFIC INGREDIENTS OVER THE LAST YEAR WE HAVE BEEN QUITE ACTIVE. THE FIRST ONE ARE A COUPLE N.D.I.'S OR NEW DIETARY INGREDIENTS. THEY DIDN'T HAVE THE REQUIRED NOTIFICATION ON FILE. D.M.B.A. OR A.M.P. CITRATE WHATEVER YOU WANT TO CALL IT. VERY SIMILAR STRUCTURE TO D.M.A.A. WE ARE STRUGGLING WITH THAT FROM TIME TO TIME. PURPORTED INGREDIENT WHILE THAT IS NOT ESTABLISHED YES OR NO THERE ARE NO NOTIFICATION ON FILE. THERE ARE SEVERAL PRODUCTS THAT HAD AS A DIETARY INGREDIENT. 14 DOESN'T ENCOMPASS THE ENTIRE INDUSTRY. BUT SOMETHING IS BETTER THAN NOTHING. WE NEED TO PUT THE INDUSTRY ON NOTICE THIS ISN'T SOMETHING THAT SHOULD BE USED WITHOUT NOTIFICATION. THE OTHER IS STRAIGHT BOTANICALS. DIETARY INGREDIENT. THERE WAS NO NOTIFICATION ON FILE. A NUMBER OF FOLKS USING IT IN THEIR DIETARY SUPPLEMENTS. SO SIX WARNING LETTERS WERE ISSUED MARCH 2016 LAST MONTH. WE ARE PUTTING IT OUT THERE SO THE INDUSTRY IS AWARE, THEY NEED TO MOVE ONTO THE NEXT BOTANICAL DU JOUR FOR BETTER OR WORSE. THESE AREN'T DIETARY INGREDIENTS IF THEY AREN'T THEY CAN'T BE NEW DIETARY INGREDIENTS. B.M.T.B.A. IS A SIMILAR WAY TO TALK ABOUT BETA META METHYL MIN. THERE WAS NO NOTIFICATION ON FILE. THEY WOULD HAVE RECEIVED A RESPONSE APRIL 2015 WAS A BUSY MONTH. PICAMILLEN. DOMINATION OF NIACIN AND GABBA. IT DIDN'T FIT THE DEFINITION NOR PART OF CONVENTIONAL FOOD DIET IN IT SELF. WE ISSUED FIVE WARNING LETTERS IN 2015 TO FIRMS LABELLING AS A DIETARY INGREDIENT. ACCORDING TO OUR RESEARCH MAJORITY HAS BEEN PULLED. WE AREN'T ALWAYS 100% SUCCESSFUL BUT WE TRY TO KEEP CHIPPING AWAY. THERE ARE SEVERAL SEN NIF RIN. WE ISSUED SEVEN WARNING LETTERS IN MARCH OF 2016, LATE LAST MONTH. WE WILL SEE WHAT THE INDUSTRY RESPONSE TO THAT IS AS WELL. PROBABLY IMPORTANT TO THE LAST COUPLE SLIDES, N.D.I.'S AND OTHER INGREDIENTS NOT DIETARY INGREDIENTS WOULD BE N.D.I. GUIDANCE. WE UNDERSTAND IT'S IMPORTANT. WE ISSUED A DRAFT GUIDANCE ON N.D.I.'S THE REQUIREMENT TO SUBMENT NOTIFICATION FOR JULY 2011. COMING UP ON FIVE YEARS AGO. IT WAS NOT, WE DID SEE WE WILL REVISE AND PUBLISH ANOTHER DRAFT GUIDANCE. IT'S OUR OPPORTUNITY TO CLARIFY THINGS THAT WEREN'T CLEAR TO PROVIDE EXTRA EXPLANATION, IF NEED BE TO ADJUST OUR POSITION. AND WE ARE WORKING ON THAT. LIKE I SAID BEFORE, IT'S A TOP PRIORITY. WE UNDERSTAND THE IMPORTANCE OF IT, IDEALLY FIRMS WILL READ OUR CURRENT DRAFT GUIDANCE AS WELL. THE UPCOMING REVISED DRAFT GUIDANCE AND TAKE NOTE OF THINGS THAT FIT THE DEFINITION AND DO NOT, THAT DO NEED NOTIFICATION ON FILE. HOW DOES ONE SUBMIT A NOTEFICATION. WHAT ARE THE REQUIREMENTS NEEDED FOR A NOTEFICATION AND WHAT DO WE THINK CONSTITUTES A SOLID NOTIFICATION ALL OF THAT IS IMPORTANT. UNIQUE NOTIFICATIONS. I SHOULD SAY NOTIFICATIONS ON UNIQUE N.D.I.'S IS HOW IT SHOULD BE WORDED. 610? -- 610 IN 21 YEARS IS NOT A VERY LARGE PERCENTAGE. WE BELIEVE THERE SHOULD BE MORE AND THERE ARE MORE NOTIFICATIONS THAT HAVE BEEN SUBMITTED. MORE FIRMS HAVE RESUBMITTED SHOULD THEY RECEIVE A RESPONSE THEY DIDN'T APPRECIATE. THE ACTUAL NUMBER IS AROUND 900 BUT LOOKING AT THE UNIQUE NUMBERS IT'S ABOUT 610. WHAT WE ARE SEEING IS THAT, NICELY IN GREEN, GREEN MEANS GO, A.K.L. WHAT WE CALL AN ACKNOWLEDGMENT. ABOUT 24% OF THE 610 HAVE RECEIVED AN ACKNOWLEDGMENT. AN ACKNOWLEDGMENT IS AS GOOD OF A THUMB'S UP AS YOU ARE EVER GOING TO GET FROM US. IT'S NOT AN APPROVAL SYSTEM. IT'S A NOTEFICATION SYSTEM. WE WILL RESPOND BACK, WE ACKNOWLEDGE YOUR SUBMISSION. WE HAVE NO FURTHER OBJECTIONS, THAT'S AN A.K.L. AN I.A.L. IS AN INADEQUATE LETTER. ABOUT 33%. INADEQUATE MEANS WE ACKNOWLEDGE YOU SUBMITTED A NOTEFICATION BUT WE HAVE SOME QUESTIONS OR CONCERNS. WE BELIEVE YOUR CONCLUSION THAT A PRODUCT CONTAINING IT IS REASONBLY EXPECTED TO BE SAFE IS INADEQUATE IN ONE WAY OR THE OTHER. MEANS THEY DIDN'T PROVIDE ALL THE REQUIRED INFORMATION, LISTED IN THE REGULATION. IF THEY DON'T HAVE THAT, THEY WILL RECEIVE BACK A RESPONSE. YOUR SUBMISSION WAS INCOMPLETE. AS FAR AS WE ARE CONCERNED THAT NOTIFICATION NEVER HAPPENED. AND THEN AN N.D.L. WHICH IS NOT A DIETARY INGREDIENT OR DIETARY SUPPLEMENTS LETTER. THAT'S A RESPONSE BACK THAT SHOULD SOMEONE HAVE SUBMITTED A NOTEFICATION ON PICAMILLEN THEY WOULD HAVE RECEIVED A N.D.L. AND NOTIFY THAT WAS NOT A DIETARY INGREDIENT OR POSSIBLY PUT INTO A TOPICAL OR INJECTABLE SO IT DOESN'T FIT THE DEFINITION OF A DIETARY SUPPLEMENTS. THERE ARE A NUMBER OF REASONS ONE MIGHT RECEIVE THAT RESPONSE. ENFORCING G.M.P. COMPLIANCE. NORMALLY I SPEND MUCH MORE TIME ON THIS, HOWEVER THIS ISN'T A G.M.P. CONFERENCE. SO I WON'T BE FOCUSING ON THIS. HOWEVER, I DO LIKE TO PROVIDE THE NUMBERS FOR THOSE THAT ARE INTERESTED. WE HAVE BEEN INCREASING OUR NUMBER OF INSPECTIONS OF DIETARY SUPPLEMENTS FACILITIES. 2011 TO 2015, THE NUMBERS UP THERE. FAIRLY SETTLED ON THOSE NUMBERS. ALWAYS SOME SHIFTING AROUND BUT I BELIEVE THOSE ARE FINAL NUMBERS. THE COLUMNS NEXT TO IT ARE O.A.I. OFFICIAL ACTION INDICATED. N.A.I. IS NO ACTION. WHAT WE REALLY NEED TO SEE IS DROPPING IN THE ACTION OF O.A.I. UNFORTUNATELY, IT GOES UP, IT GOES DOWN BUT STAYING AROUND 20-30%. IN OUR TERMS IT'S TOO HIGH. AND WE ARE FOLLOWING UP WITH RESULTING ACTIONS. SO WE SEE A NUMBER OF WARNING LETTERS, W.L. WE TRY TO STICK WITH, WE ARE HAVING ANYWHERE FROM 70-80. 2014 WAS AN ABERRATION. WE ARE SEEING AN INCREASE IN INJUNCTION. AN INJUNCTION WAS WHEN THE GOVERNMENT MOVES TO ENJOIN THE FIRM, LABOR INTENSIVE ACTION. OR SEIZURE. IF WE HAVE A PROBLEM WITH THE PRODUCT, AND NEED TO SEIZE THE PRODUCT WE WILL MOVE TO A SEIZURE. THIS IS THE SLIDE OF IMPORTANCE. TOP VIOLATIONS FOR 21 CFI REGULATION REQUIREMENT FOR G.M.P.'S ARE REALLY, I THINK I HAVE 11 THINGS LISTED THERE. THE TOP 11. WHAT WE ARE SEEING IS A PROBLEM WITH PRODUCT SPECIFICATIONS, MASTER MANUFACTURING RECORDS WITH DIETARY INGREDIENT IDENTITY AND WITH BATCH RECORDS AND THEN WITH QUALITY CONTROL PROCEDURES. THOSE ARE THE TOP FIVE I JUST LISTED. THEY ARE EITHER COMPLETELY AB -- ABSENT OR VERY INADEQUATE. WITHOUT THIS, YOU CANNOT PUT OUT A QUALITY PRODUCT. YOU DON'T KNOW WHAT YOU ARE SETTING UP TO MAKE. YOU AREN'T KEEPING A RECORD OF IT. AND YOU DON'T HAVE THE QUALITY CONTROL IN PLACE TO TEST IT AS YOU MOVE ALONG. WITH THAT, I HAVE MY WARM UP, MY WRAP UP SIGN. SO THANK YOU ALL. NOT SURE WE HAVE TIME FOR QUESTIONS BUT I'M AVAILABLE. I WILL BE AROUND FOR THE BREAK AND LUNCH. THANK YOU. WE WILL TAKE A BREAK NOW SO KARA WILL BE AVAILABLE IF YOU WANT TO ASK SPECIFIC QUESTIONS. I WILL PULL FIVE MINUTES FROM THE BREAK. SO EVERYONE RECONVENE AND WE WILL RESTART AFTER THE BREAK AT 10:40. I WOULD LIKE TO WELCOME DR. CRAIG HOPP. YOU CAN TALK TO CRAIG. CRAIG IS GOING TO TALK ABOUT THE QUEST FOR RIGOR AND REPRODUCABILITY IN BOTANICALS RESEARCH. THANK YOU, CRAIG. >> THANK YOU. JUST FOR CLARIFICATION. I'M THROUGH N.C.C.H., SIDE OF THINGS. MY COLLEAGUE IS IN THE AUDIENCE HERE SOMEWHERE, I BELIEVE, THE COUNTERPART THAT HANDLES IT FROM O.D.S.'S SIDE. I WOULD LIKE TO START BY THANKING CYNTHIA FOR THE INVITATION TO SPEAK HERE TODAY AND THANK N.I.E.H.S. FOR PUTTING THIS TOGETHER. A TOPIC NEAR AND DEAR TO OUR HEARTS AT N.C.C.I.H. WHAT I HOPE TO TALK TO YOU ABOUT, HOW WE APPROACH THIS QUEST, AS I CALL IT FOR RIGOR AND REPRODUCABILITY IN BOTANICAL RESEARCH. BY WAY OF BACKGROUND, WE WERE KNOWN AS NICAN, SUPPORTED A NUMBER OF LARGE PHASE 3 CLINICAL TRIALS TO ASSESS THE EFFICACY OF A VARIETY OF DIETARY SUPPLEMENTS, PRIMARILY BOTANICALS FOR A VARIETY OF INDICATIONS FOR WHICH THEY WERE COMMONLY USED AND ALMOST UNIVERSALLY THOSE TRIALS FAILED TO DEMONSTRATE THE EFFICACY THAT HAD BEEN HYPOTHESIZED FOR THOSE PRODUCTS. LET'S GIVE LESSONS LEARNED ABOUT WHAT'S COME OUT OF THAT. IF YOU CAN'T UNDERSTAND THE SLIDE TOO WELL, THIS IS St. JOHN'S WORT, PUBLISHD IN 2002, THE PAPER IN JAMA, COMPARED WITH ZOLOFT AND PLACEBO. REALLY NO SEPARATION BETWEEN THE THREE LINES. YOU COULD SAY St. JOHN'S WORT IS AS EFFECTIVE AS SY TRY LEAN. MIDDLE LINE IS PLACEBO AND BOTTOM LINE IS SIR TRA LEAN. THIS IS AN EXAMPLE BLACK COHOSH USED FOR MENOPAUSAL SYMPTOMS. RED CLOVER WITH HORMONE REPLACEMENT THERAPIES, SPECIFICALLY ESTROGENS AS WELL AS PLACEBO. IT WAS VERY EFFECTIVE BUT FOR REASONS, SOMETHING PEOPLE DON'T LIKE TO DO ANY MORE. BLACK COHOSH, RED CLOVER, THE TAKE HOME MESSAGE WAS NEITHER OF THESE THINGS WAS MORE EFFECTIVE THAN PLACEBO BUT THERE WERE SIGNIFICANT REDUCTION IN SYMPTOMS THEY DIDN'T EXCEED OR IMPROVE UPON THE LEVELS SEEN WITH PLACEBO. SEEN WITH ECHINACEA, USED WITH PREVENTION OR REDUCE DURATION OF COLDS, THERE WAS NO PERCEIVED BENEFIT WITH THAT PRODUCT. SO THIS GOT US ALL TO THINKING, WHAT DO WE DO NOW? WHAT'S THE TAKE-HOME MESSAGE HERE? WELL FIRST, THESE STUDIES FAILED TO REPORT PRIOR REPORT BENEFIT. WE DIDN'T OPEN THESE STUDIES BECAUSE IT SEEMED LIKE A GOOD IDEA. THERE WAS FAIR AMOUNT OF LITERATURE THERE WAS POTENTIAL FOR BENEFIT FOR EACH OF THESE PRODUCTS FOR THE STUDIES WE DID. WE WERE UNABLE TO REPLICATE THE STUDIES THAT HAD BEEN REPORTED PREVIOUSLY. SECONDLY,ALMOST THE MOMENT THESE STUDIES WERE PUBLISHED WE WERE CRITICIZED AND FROM DIFFERENT ANGLES ABOUT THE DESIGN OF THE STUDY. THEY WERE VERY RIGOROUSLY CONDUCTED, VERY WELL CONTROLLED AND VERY WELL RUN. BUT THEY WERE ALSO NOT BASED ON A LOT OF PRECLINICAL PRE-K DATA OR DOSE RANGING DATA. WE WERE CRITICIZED WE WEREN'T USING THE RIGHTD PRODUCT, DOSE, FORMULATION, POPULATION, FOR SOME DRIT -- CRITICISMS WE WERE UNABLE TO DEFEND OURSELVES RIGOROUSLY. THINGS ARE STUDIED IN VEE TROE, ANIMALS. PHASE ONE, PHASE TWO, PHASE THREE, THE NORMAL PROTOCOL REALLY WANT FOALED THROUGH. GOOD REASONS FOR THAT, ACTIVELY USED BY MILLIONS AND WE WANTED TO KNOW WHETHER OR NOT THERE'S WAS ANY BENEFIT. WHAT WE LEARNED WE CAN'T SHORT CIRCUIT THE NORMAL PROCESS THAT PEOPLE TAKE. THE OTHER LESSON WE LEARNED IS BECAUSE OF THE CRITICISMS WE COULDN'T REALLY VIGOROUSLY DEFEND WE WERE NOT ABLE TO MAKE DEFINITIVE CONCLUSIONS ABOUT THESE THINGS. I WILL EMPHASIZE THAT N.C.I.H. ARE NOT CRITICAL OF A CLINICAL FINDING. ONLY IF WE CAN SAY IT DEFINITIVELY. BECAUSE OF THE QUESTIONS THAT AROSE ABOUT THIS, CALLING INTO QUESTION, THE RIGOR WITH WHICH THE STUDIES UPON WHICH THEY WERE BASED WE COULDN'T MAKE DEFINITIVE CONCLUSIONS WE DON'T KNOW FOR SURE IF THEY HAVE A BENEFIT. STILL QUESTIONS WE WERE UNI ABLE TO ANSWER. PLACEBO RESPONSES ARE VERY HIGH. PARTICULARLY FOR STUDIES THAT INVOLVE SUBJECTIVE END POINTS LIKE PAIN OR MENOPAUSAL SYMPTOMS. WE REALLY SEEK TO BRETTER DESIGN OUR STUDIES IN THE FUTURE TO IMPROVE UPON THAT. AS WE WERE COMING TO LEARN THESE LESSONS AND TEN YEARS AGO OR SO, PEOPLE WERE CRITICIZING THAT NOTHING YOU DO SEEMS TO WORK, THERE WAS A WASTE OF MONEY WE WERE POINTING OUT THIS IS NOT UNUSUAL FOR PHASE THREE TRIALS TO NOT SORT OF PAN OUT THE WAY YOU WANT THEM TO. YOU ARE UNABLE TO DEFINITIVELY SHOW A BENEFIT. ABOUT THE TIME, THIS PAPER CAME OUT IN 2011. FROM BAYER, THE GERMAN VERSION OF THAT. THEY WERE LOOKING AT WHAT THEY HAD DONE IN HOUSE. CAN WE DEVELOP AROUND THIS TARGET, THE TAKE-HOME MESSAGE EITHER MEN'S HEALTH, THEY LOOK AT A NUMBER OF DOZENS OF DRUG TARGETS ABOUT TWO THIRDS THEY COULD NOT REPLICATE THE STUDIES THAT WERE REPORTED IN THE LITERATURE. POINTING TO ISSUE, ABOUT A YEAR AFTER THAT, THEY RUNG THE BELL AND N.I.H. SORT OF QUICKLY ANSWERED THE CALL, IF YOU WILL. THIS WAS A PAPER THAT CAME OUT OF A BUNCH OF COLLEAGUES AT N.I.D.S., SILVERBERG CHAMPIONED THIS CAUSE CALLING FOR TRANSPARENCY IN THE REPORTING OF RESEARCH, ESPECIALLY PRE-CLINICAL RESEARCH TO IMPROVE ON THE, FOR N.I.H. TO MAXIMIZE OUR RESEARCH DOLLARS. WE DIDN'T WANT TO WASTE TAX PAYER MONEY BUT WE WERE COMING INTO THE SAME SCENARIO THAT WE WERE UNABLE TO REPLICATE OR DEMONSTRATE THINGS WE EXPECTED THERE TO BE BENEFIT FOR. AFTER MUCH MEETINGS AND WORKSHOPS AND INTERNAL DISCUSSIONS HAD AMONG SENIOR LEADERS AT N.I.H. AND NEXT THING YOU KNOW A RIGOR AND REPRODUCABILITY POLICY. THIS IS THE WEBSITE. THERE'S LOTS OF ASPECTS TO THIS ABOUT WANTING TO MAXIMIZE THE IMPACT OF N.I.H. RESEARCH FUNDING SO SOME OF THE TENETS OF THIS POLICY, THIS IS FOR GRANT APPLICATIONS IS THAT THE APPLICANT MUST SHOW THE SCIENTIFIC PREMISE MUST DEMONSTRATE RIGOROUS AND UNBIASED RESULT. THEY HAVE TO HAVE AUTHENTIFICATION. IT'S VERY PERNTMENT TO THE DISCUSSION TODAY. A LITTLE MORE ABOUT THAT REQUIREMENT. I GUESS REGULATIONS THAT YOU WILL OFFER N.I.H. THE PROTOCOL POLICY STATES THEY MAY OR MAY NOT BE GENERATED AND MAY DIFFER FROM LABORATORIES OVER TIME. MAY HAVE QUALITIES AND OR QUALIFICATIONS OF RESEARCH DATA. DOES THIS SOUND LIKE BOTANICAL RESEARCH TO YOU. IT DOES TO ME. THOUGH IT WASN'T WRITTEN WITH BOTANICALS IN MIND. IT'S CERTAINLY EXACTLY THE SORT OF THINGS WE STRUGGLE WITH ANYBODY THAT'S INVOLVED IN BOTANICAL RESEARCH STRUGGLES THEY DIFFER FROM PLACE TO PLACE. VERY IMPORTANT TO THE RESEARCH. THEY ARE GOING TO INFLUENCE THE DATA. THESE INCLUDE CELL LINES, ANTIBODIES AND OTHER BIOLOGICKS. N.I.H. TENDS TO FOCUS MORE ON BIOLOGY THAN CHEMISTRY BUT CERTAINLY RECOGNIZE THE IMPORTANCE CHEMISTRY HAS FOR THE STUDIES. THIS IS SOMETHING THAT'S COME INTO PLAY IN THE LAST YEAR OR SO AT N.I.H. BUT N.C.C.I.H. I THINK WE HAVE BEEN AHEAD OF THE CURVE ON THAT BY QUITE A BIT IN SOME CASES. I WILL TALK FIRST ABOUT THE CLINICAL RESEARCH ON BOTANICALS AND THINGS WE ARE FOCUSED ON CURRENTLY FOR THE STUDY WE DO. I MENTIONED WE DO A LOT OF VERY LARGE PHASE THREE STUDIES THAT DIDN'T PAN OUT. OUR CONCLUSION WE MAY HAVE JUMPED INTO THOSE PREMATURELY. IN THE CLINICAL SETTING, WE STIPULATE TRIALS MUST BE DESIGNED SO WHATEVER YOU FIND WHETHER POSITIVE OR NEGATIVE WILL PROVIDE HIGH SCIENTIFIC QUALITY INFORMATION THAT WILL BE SUPPORT DECISIONS ABOUT GOING FORWARD OR NOT TO GO FORWARD. WE AREN'T DOING THAT FULL SCALE CLINICAL TRIAL NOW. THIS CONCEPT OF BIOLOGICAL SIGNATURES IS SOMETHING WE FOCUS ON QUITE A BIT THESE DAYS. WE USE THE TERM BIOLOGICAL SIGNATURE BECAUSE FOR BOTANICAL PRODUCTS WE REALIZE THERE'S NOT A SINGLE TARGET THAT WILL BE USED BY THAT COMPLEX INTERVENTION. A WEB, IF YOU WILL, OR A NETWORK OF BIOLOGICAL RESPONSES THAT RESULT FROM THAT. SO THAT'S ON THE CLINICAL SIDE. AS A CENTER, AS A POLICY ACROSS A CENTER WE HAVE THIS POLICY THAT WAS ORIGINALLY CALLED BIOLOGICALLY ACTIVE AGENTS USED IN COMPLIMENTARY AND ALTERNATIVE MEDICINE, AND PLACEBO MATERIALS. THIS HAS BEEN SHORTENED TO BE CALLED PRODUCT INTEGRITY POLICY. AS FATE WOULD HAVE IT THAT POLICY WAS PUBLISHED ALMOST 11 YEARS AGO TO THE DAY OF THIS MEETING APRIL 29th OF 2005. THE KEY TENET WE ASKED INVESTIGATORS TO PROVIDE NICAN THAT THE QUALITY OF THE PRODUCTS WAS SUFFICIENT TO ENSURE IT COULD BE REPRODUCED. THIS CONCEPT OF REPRODUCABILITY IN BOTANICAL RESEARCH IS SOMETHING THAT N.I.E.S. AND O.A.H. ENSURED IMPORTANCE OF GETTING AHEAD OF THE DATA WHEN IT COMES TO REPRODUCABILITY IN BOTANICAL PRODUCTS. IT'S BEEN REVISED SINCE IT'S ORIGINAL ITERATION WE RECOGNIZE THERE'S A SPECTRUM OF RESEARCH THAT N.C.I.H. FUNDS MULTI COMPLEX HERB ALIFORM LAS ALL THE WAY TO HIGHLY REFINED RESEARCH ON ISOLATED CONSTITUENTS, BE THEY I.C.B.G., YOU NAME IT. REQUIREMENTS, THE POLICY, WHAT IT BASICALLY REQUIRES IS THAT THEY HAVE A SERIES OF QUESTIONS THEY HAVE TO ADDRESS. BEFORE WE WILL FUND THEM TO DO THE RESEARCH. THEY HAVE TO TELL US WHAT IS THE, WHAT IS THE BINOMIAL IF IT'S A COMPLEX PRODUCT. WHAT IS THE MANUFACTURING PROCESS, FINGERPRINT, IF YOU WILL OF THAT PRODUCT. THEY HAVE TO HAVE INDEPENDENT ANALYSIS. NO JUST SHOWING ME THE C. OF A. FROM THE SUPPLIER AND CALLING THAT GOOD. WE REQUIRE THEM IN SOME WAY, SHAPE OR FORM INDEPENDENTLY VERIFY AND WE DON'T STIPULATE HOW THEY DO THAT. IT'S NOT DONE ON A CASE-BY-CASE BASIS BUT HAVE TO CORROBORATE THE DATA PRESENTED IN ANY CERTIFICATE OF ANALYSIS PROVIDED BY THE MANUFACTURER. ON THE OTHER SIDE IF IT'S A VERY REFINED PRODUCT. IF THEY ARE STUDYING RES VER TROLL WE DON'T CARE WHERE THE GRAPES WERE GROWN REALLY. WE WANT TO KNOW HOW PURE IS IT. IS IT AS PURE AS YOU THINK IT IS. THAT'S NOT ALWAYS THE CASE AS WE ARE AWARE OF. STILL REQUIRING THIS FOCUS ON INDEPENDENT ANALYSIS. OR THEY CAN FIND OFTEN TIMES A THIRD PARTY GROUP THAT COULD DO THAT. IN ADDITION TO A SPECTRUM OF COMPLEXITY OF THE PRODUCT THERE'S ALSO A SPECTRUM OF COMPLEXITY OF THE RESEARCH. THIS I'VE SEEN, N.C.I.H. FUNDS IN VITRO RESEARCH TO INVOLVING HUMANS AND OUR PRODUCT INTEGRITY POLICY REFLECTS THAT. WE DON'T REQUIRE PEOPLE DOING IN VITRO RESEARCH TO GET A LETTER FROM THE SUPPLIER SAYING THEY WOULD SUPPORT I.N.V. WE TRY TO STAGE THINGS AND REQUIRE CERTAIN TYPES OF INFORMATION AS THE RESEARCH PROGRESSES. WE HOPE INVESTIGATORS AS THEY ARE DOING IN VITRO RESEARCH ARE THINKING AHEAD WHAT THEY MIGHT NEED FOR THE NEXT STAGE IF THEY COME BACK TO FUND THAT WORK. THIS SHOWS HOW WE ESCALATE THE INFORMATION THAT WE ASK FOR THE INVESTIGATORS TO PROVIDE US. OFTEN TIMES THEY HAVE TO GO TO THE MANUFACTURER TO GET THIS INFORMATION AND OFTEN TIMES IT'S NOT ALWAYS STRAIGHT FORWARD TO GET THAT DEPENDING ON THE CIRCUMSTANCES OF COURSE. NOW I WOULD LIKE TO GIVE A COUPLE EXAMPLES HOW THIS POLICY IN ACTION HAS REALLY HELPED N.C.I.H. TO HELP THE RESEARCH AND REPRODUCABILITY WE FUND. THE POLICY WAS IN PLACE I THINK A YEAR BEFORE IT REALLY SORT OF SHOWED IT'S MERITS. THIS PLANT CALLED DULSE MARRA. THERE WAS A RESEARCHER OUT OF THE STATE OF FLORIDA LOOKING AT A SOUTH AMERICAN PLANT FOR A THERAPEUTIC, THEY CALLED IT DULCAMARA. THEY GAVE THEM SOLANUM DULCAMARA BUT THAT WASN'T WHAT THEY WANTED. IT WAS A COMMON NAME FOR A PLANT THEY REALLY MET CALANCHO. BECAUSE OF THE POLICY WE WERE ABLE TO CATCH THIS MISTAKE. BEFORE IT GOT INTEGRATED INTO THE RESEARCH. IMAGINE WHAT THE PAPER THAT CAME OUT OF THIS WOULD BE UNINTERPRETABLE. PROBABLY NOT REPRODUCIBLE. AGAIN THE POLICY WE HAD IN PLACE VERY QUICKLY CONFIRMS OUR RATIONAL FOR PUTTING IN PLACE. MORE RECENTLY WE FUNDED RESEARCH ON COMMON PLANTS SUCH AS CINNAMON. THERE'S CASSIUS CINNAMON. THEY HAVE DIFFERENT LEVELS OF COUMARIN SOMETHING YOU DON'T WANT. IT'S MORE PREVALENT IN CASSIUS. WHAT WE FOUND WHEN PEOPLE WERE LOOKING AT SIN MUN CINNAMON. WE STARTED TO USE A LITTLE D.N.A. ANALYSIS I WON'T GET INTO THE PRO'S AND CON'S AT THE MOMENT BUT FOR THIS CONTEXT IT WAS QUITE HELPFUL. WE COULD NOT FIND A POWDERED CINNAMON PRODUCT THAT WAS A SINGLE SPECIES. ALMOST ALWAYS BLENDS OF MULTIPLE SPECIES. THAT'S A LITTLE BIT PROBLEMATIC. TRYING TO SAY CINNAMON HAS BIOLOGICAL EFFECTS WE WOULD LIKE TO PIN IT TO A PARTICULAR SPECIES. HOWEVER IF YOU GET CINNAMON STICK THAT'S MUCH MORE DIFFICULT TO BLEND A CINNAMON STICK. BECAUSE OF THIS POLICY WE BECAME AWARE OF THIS PRODUCT AND HAVE BEEN ABLE TO ADVISE PEOPLE ACCORDINGLY. ANOTHER EXAMPLE IS GREEN TEA. ACTUALLY I THINK IN THE LAST WEEK A REPORT FROM N.T.P. ON GREEN TEA. THE ISSUE WE HAVE IS WHAT DO THEY MEAN WHEN THEY SAY GREEN TEA. THEY ARE TALKING ABOUT E.C.G.C. OR CATECHINS. SOME OF THESE PRODUCTS ARE HIGHLY CONCENTRATED AND WHEN YOU DO THE ANALYSIS OF THE PRODUCT, THE INDEPENDENT ANALYSIS IT'S REALLY NOT GREEN TEA AT ALL. AND JOE TOUCHED UPON THIS. IT'S A CONCENTRATED COLLECTION OF A SMALL NUMBER OF CATECHINS. SO THIS, AGAIN, THIS IS ALSO AN EXAMPLE OF GREEN TEA WE AGREE IS CAMELIA SIN NEN SIS. THIS IS SUCH A WIDELY CULTIVATED PRODUCT IT COULD BE VERY CHALLENGING AND WE HAVE BEEN UNABLE TO GET SOMETHING THAT LOOKS LIKE A GENETICALLY HOMOGENOUS EXAMPLE. WHEN YOU REPORT THIS DATA ACCURATELY. IF YOU ARE TALKING ABOUT IN YOUR APPLICATION YOU WILL USE GREEN TEA AND TESTED AS AN ANTIOXIDANT. WHAT YOU ARE STUDYING IS E.C.G.C. REPORT ON E.C.G.C. DON'T REPORT ON THE ACTIVITY OF GREEN TEA WHEN THAT'S REALLY NOT WHAT YOU ARE STUDYING. ALL RIGHT, RIGHT ON CUE. I GET MY WRAP UP SINGLE AND TAKE-HOME MESSAGES SLIDE SO EVERYTHING IS WORKING OUT GREAT. THEY REALLY TWO-FOLD. CERTAINLY ANYBODY INVOLVED IN RESEARCH UNDERSTANDS HOW DIFFICULT IT IS TO CONTROL FOR ALL THE VARIABLES IN PLACE WHEN YOU ARE TRYING TO DO BIOMEDICAL RESEARCH. WHAT I WOULD LIKE TO POINT OUT AND WHAT I THINK EVERYBODY IS WELL AWARE OF. WHEN YOU ADD BOTANICAL PRODUCTS TO THAT, YOU ADD A LETTER OF COMPLEXITY ABOUT EIGHT-FEET DEEP. YOU HAVE AN ENORMOUS NUMBER OF ADDITIONAL VARIABLES YOU HAVE TO CONTROL. THIS MAKES RESEARCH OF BOTANICALS EXTREMELY DIFFICULT FOR MANY TO DO. ESPECIALLY THOSE NOT WELL VERSED IN THESE CHALLENGES. THEY ARE MOLECULAR BIOLOGISTS. THEY DON'T UNDERSTAND THE COMPLEXITIS. THE REASON THE POLICY IS IN PLACE. TAKE HOME MESSAGE NUMBER ONE, TAKE HOME MESSAGE NUMBER TWO, I HOPE I HAVE SHOWN NICAM AND N.I.A.H. HAVE POLICIS IN PLACE TO HELP MAXIMIZE THE RIGOR AND REPRODUCABILITY OF THE RESEARCH WE FUND. WITH THAT, I WILL WRAP UP AND THANK YOU FOR YOUR ATTENTION. PL*D [APPLAUSE] >> THANK YOU, CRAIG. SO THE FINAL, WE'VE GOT, SECOND PERSON IN THE SECOND HALF OF THE SESSION IS DUFFY. COUNCIL OF RESPONSIBLE NUTRITION AND LICENSED NATUROPATHIC DOCTOR AS WELL. >> THANK YOU VERY MUCH, NIGEL. IT'S REALLY GREAT TO BE HERE. AND REALLY APPRECIATE BEING PART OF THIS CONVERSATION. I THINK WE ARE CONTINUING THE WHIRLWIND TOUR WE HAVE HEARD FROM F.D.A. FROM OFFICE OF DIETARY SUPPLEMENTS FROM N.C.I.A.H. AND NOW WE WILL HEAR FROM N.I.H. I WORK FOR A TRADE ASSOCIATION HERE IN WASHINGTON D.C. THAT REPRESENTS BOTH MANUFACTURERS OF THE FINISHED PRODUCT AS WELL AS SUPPLIERS OF THE RAW MATERIAL. THIS WOULD INCLUDE BOTANICAL RAW MATERIALS. TODAY I'M HOPING TO SORT OF HIGHLIGHT SOME OF THE ROLES OF THE INDUSTRY AND THE SAFETY OF DIETARY SUPPLEMENTS. OUR ROLES AND RESPONSIBILITY. WE MANUFACTURER THESE PRODUCTS. I WANT TO TALK ABOUT WHAT THE LARGER, THE DISTINCTION BETWEEN PLANTS AND DRUGS AND HOW THESE INHERENT DIFFERENCES IMPACT SAFETY. WE ARE LOOKING AT SAFETY OBSERVATIONS. WE TEND TO HAVE WORLD EXPERTS IN TOXICOLOGY. ALL OF THE ANALYTICAL EXPERTS. WE TEND TO HAVE LESBO S -- LESS BOTANICAL EXPERTS. THAT LEADS US, WE GET SO FAR DOWN THIS PATH AND THEN WE REALIZE 20 YEARS LATER WE ARE WORKING WITH COMPLICATED STUFF. IF WE REALLY INCLUDE THESE OTHER SHAREHOLDERS WE WILL HAVE THAT WILL BENEFIT US. MY FINAL PART OF OUR TALK WE WILL LOOK AT THE ALOE VERA. IT'S FAIR TO SAY BEFORE THE DRAFT REPORT ON ALOE VERA, THE INDUSTRY WAS NOT ENTIRELY AWARE OF THIS INITIATIVE. AT LEAST NOT ENGAGED IN. THERE WERE SOME OUT LIARS WHO SAW THE WORK OF THE N.T.P. WHO GOTTEN GAUGED AND PROVIDED COMMENTS AND WENT TO MEETINGS BUT IT WASN'T UNTIL WE SAW THE IMPACT. I WILL SHARE A LITTLE OF WHAT WE LEARNED FROM THAT. I THINK IT'S BEEN COVERED FAIRLY WELL, THIS CONCEPT DIETARY SUPPLEMENTS ARE REGULATED IN THE CATEGORY OF FOOD. IT'S SO IMPORTANT, AS WE ATTEND THESE MEETINGS AND HAVE OTHER FOLKS LOOKING AT THE CATEGORY, WE WILL SEE A MIGRATION -- HARVEST CONDITIONS. TO A LOT OF PEOPLE FROM THE DRUG SIDE OF THE EQUATION, THAT'S JUST A PROBLEM. IT DOESN'T MAKE SENSE. SCIENTISTS LIKE THINGS BLACK AND WHITE. THEY LIKE THINGS REPRODUCIBLE. THEY LIKE THINGS IN CONTROLLED SETTINGS AND THEY LIKE TO CONTROL VARIABLES. HERBALIST AND PEOPLE WHO USE PLANTS REALLY EMBRACE THAT INFLUENCE OF NATURE. THAT INFLUENCE OF THE WEATHER AND PROCESSING AT CERTAIN TIMES, YOU ALREADY HAVE TWO PARADIGMS SOMEWHAT STRUGGLING WITH ONE ANOTHER. I THINK WHAT THAT LENDS ITSELF TOWARD WE REALLY NEED NEW METHODS WE NEED TO BRING THE WORLDS TOGETHER AND DECIDE HOW DO WE PROPERLY ASSESS THE SAFETY. I COMMENT THE GROUP TODAY AND VARIOUS STAKEHOLDERS FOR TAKING A SERIOUS LOOK AT THAT QUESTION. WHILE WE ARE REGULATED AS A CATEGORY OF FOOD. WE ARE ALSO REGULATED DIFFERENT THAN LIKE A CAN OF SOUP OR TOMATO OR SOMETHING LIKE THAT. I PULLED THIS QUOTE WHERE IT SAYS THE FEDERAL GOVERNMENT SHALL TAKE SWIFT ACTION THAT ARE UNSAFE OR UNADULTERATED. HOWEVER THE FEDERAL GOVERNMENT SHALL NOT TAKE ANY ACTION TO OPPOSE ANY REGULATORY BARRIERS LIMITING SAFE PRODUCTS TO CONSUMERS. THEY WERE A DELICATE BALANCE BETWEEN ACCESSIBILITY AND SAFETY. PEOPLE DID NOT WANT TO MAKE THESE PRODUCTS LIKE DRUGS. YOUR VITAMIN C, FISH OIL, ECHINACEA. DO YOU HAVE TO GO TO YOUR DOCTOR. DOES IT REALLY NEED PHASE 1, 2, OR 3 STUDIES. THE ANSWER WAS NO. WE SHOULD BALANCE QUALITY AND SAFETY. THIS IS A VERY BROAD STROKE IN OUR PARADIGM FOR SAFETY. WE DON'T HAVE PRE-MARKET APPROVAL. WHEN YOU SEE IN THE NEWS SUPPLEMENTS ARE UNREGULATED REALLY WHAT PEOPLE ARE REFERRING TO WE AREN'T REGULATED EXACTLY THE SAME AS DRUGS. IT'S NOT THAT WE AREN'T REGULATED. CARA WELCH REPRESENTS, LOTS OF PEOPLE AT F.D.A. ARE PAID TO REGULATE THIS PRODUCT CATEGORY. HOW DO WE REGULATE IT? I USE THE ANALOGY OF THE STOOL, THREE-PRONGED STRUCTURE. THE FIRST IS INGREDIENT SELECTION. IF PRESUMED OR SHOWN TO BE SAFE, THAT'S THE FIRST STEP. USE THE INGREDIENTS WE KNOW ARE SAFE. THE OLD DIETARY INGREDIENTS OR GRANDFATHERED INGREDIENTS. IF YOU HAVE SOMETHING NEW YOU ARE REQUIRED TO SUBMIT SAFETY -- TO F.D.A. JUST LIKE IN GENERAL FOOD YOU HAVE TO USE A FOOD OR FOOD ADDITIVE. INGREDIENTS ALLOWED TO GO INTO CONVENTIONAL FOOD ARE HIGHLY REGULATED, IF YOU WILL. THE NEXT STEP IS MANUFACTURING STANDARDS, THERE'S BEEN SEVERAL REFERENCES TO THE GOOD PRACTICES AND THESE ESSENTIALLY MANDATE YOU IDENTIFY YOUR MATERIAL, 100% IDENTITY TESTING. YOU HAVE MANUFACTURING RECORDS. YOU SET SPECIFICATIONS, SO YOU, AS A MANUFACTURER, YOU KNOW WHAT YOUR ACTIVE, WELL YOUR VARIOUS CHEMICAL. YOU SET LIMITS FOR CONTAMINANTS, IF YOU KNOW THERE ARE CONTAMINANTS NATURALLY OCCURRING, YOU ARE TESTING, FILTERING FOR AND MAKE SURE YOU DON'T RELEASE TO THE PUBLIC ANYTHING CONSIDERED ADULTERATED AND WE HAVE A POST-MARKET SURVEILLANCE SYSTEM. OUR ADVERSE EVENT REPORTING AND RECORD KEEPING REQUIREMENT. A VERY IMPORTANT COMPONENT OF ANY SAFETY STRUCTURE AND THIS IS IDENTICAL. THE EXACT SAME RULES AND REGULATIONS APPLY TO OVER THE COUNTER DRUGS AS FOR DIETARY SUPPLEMENTS. I HOPE THIS PROVIDES SOME PERSPECTIVE FOR SOME SCIENTISTS NOT AS FAMILIAR WITH OUR REGULATORY STRUCTURE. ADULTERATED. YOU WILL SEE THE TERM ADULTERATED IS USED. WHEN A PRODUCT IS ADULTERATED NOW F.D.A. HAS A VARIETY OF TOOLS IN ITS TOOL BOX TO ACT ON THAT PRODUCT. VERY AGGRESSIVELY OR JUST SENDING A WARNING LETTER. IT CAN BE ADULTERATED WITH A SIGNIFICANT RISK OF ILLNESS OR INJURY AND THAT'S IN THE CONDITION OF USE. AND CARA MENTIONED POWDERED CAFFEINE. CAFFEINE ITSELF IS NOT UNSAFE. IT'S BEEN THROUGH LOTS OF TOX LOGICAL PROFILES AND TESTING HOWEVER WHEN IN PURE CRYSTAL FORM AND LEAVING IT TO CONSUMERS TO USE SPOONFULS, F.D.A. COULD CALL THAT ADULTERATED. IF YOU INTRODUCE, AND YOU DON'T DO PROPER 75 DAY. IT'S ADULTERATED. CARA WELCH SHOWED. AGAIN YOU DON'T DO THAT, ADULTERATED F.D.A. CAN TAKE ACTION. IMMINENT HAZARD TO PUBLIC HEALTH OR SAFETY. THIS IS ONE WHERE IT'S LIKE THEY HAVE A CRISIS ON THEIR HANDS. THEY HAVE RECALL AUTHORITY. ADMINISTRATIVE DETENTION AUTHORITY. AND THEN LOWER DOWN. IF YOUR FACILITY IS NOT IN COMPLIANCE WITH G.M.P.'S, THE PRODUCTS THAT COME OUT YOUR DOOR ARE CONSIDERED ADULTERATED AND F.D.A. CAN ACT ON THOSE PRODUCTS. MORE ON THE INGREDIENT SAFETY. I MENTIONED OLD INGREDIENTS WHEN CONGRESS GOT TOGETHER. THEY GOT THEIR ACT TOGETHER. THEY PUT THE SHEA, AMENDED COSMETIC FOOD ACT. THEY WEREN'T GOING TO ASK TO DO RETROACTIVE SAFETY INSPECTIONS ON THESE. IT HAPPENS IN LOTS OF REGULATED INDUSTRIES WITH OVER THE COUNTER DRUGS WHEN LAWS PUT IN PLACE YOU DON'T WANT TO IMPOSE AN UNNECESSARY BURDEN ON THOSE RR IN THE MARCH KRET PLACE. IN 1994 IF THE PRODUCT IS ON THE MARKET. IF YOU ARE NEW AND THAT INCLUDES AN OLD INGREDIENT YOU MODIFIED SIGNIFICANTLY YOU HAVE TO PROVIDE WITH SAFETY DOSSIER. THIS IS WHAT WE CALL THE NEW DIETARY INGREDIENT NOTIFICATION. A FAIRLY COMPREHENSIVE DOCUMENT. IT'S WHAT WAS OUTLINED IN THE DRAFT GUIDANCE WHICH WE ARE WAITING ON THE REVISED VERSION. IT REALLY SPELLS OUT TO INDUSTRY, IF YOU WANT TO INNOVATE AND INTRODUCE NEW PRODUCTS THIS IS WHAT WE EXPECT WITH REGARD TO THE EVIDENCE FOR SAFETY. YOU MIGHT HAVE LOTS OF EVIDENCE PEOPLE HAVE BEEN CONSUMING THIS FOR A LONG TIME. AND THAT'S TOTALLY LEGITIMATE EVIDENCE TO INCLUDE. HOWEVER, YOU MIGHT HAVE EVIDENCE THAT THEY CONSUMED A LITTLE BIT OF THIS AND YOU WANT TO PROVIDE MORE IN YOUR DIETARY SUPPLEMENT. NOW YOU HAVE TO PROVIDE EVIDENCE WHETHER CLINICAL, TOX LOGICAL DATA TO FILL THAT INFORMATION GAP. AT THE END OF THE DAY, F.D.A. NEEDS TO BE SATISFIED AND THAT'S WHERE DR. WELCH PUT UP THE PIE GRAPH. YOU WANT ONE OF THOSE GOOD DAY LETTERS, THAT YOUR DOSSIER REASONBLY ESTABLISHES THE INGREDIENT AS SAFE. ONE OF THE MESSAGES TO THE AUDIENCE, DIETARY SUPPLEMENT SAFETY IS A TOP PRIORITY FOR THE INDUSTRY. THOSE WHO HAVE BEEN IN THE INDUSTRY FOR THE LONG HAUL. THOSE ARE SELLING GINGKO AND WHEY PROTEIN AND FISH OIL, OUR CONSUMER IS OUR GREATEST ASET. ASSET. WE ARE VERY MUCH INTERESTED IN PROVIDING SAFE PRODUCTS. WHEN THE EPHEDRA DEBACKLE WAS TAKING PLACE, WE KNOW IT WAS FORMULATED IN PRODUCTS FOR WEIGHT LOSS BUT WE HAD THIS OTHER BOTANICAL COMMUNITY THAT SAID WE USE IT FOR BRONCHITIS IN DIFFERENT AMOUNTS. VERY DIFFERENT THAN WEIGHT LOSS. WE DON'T WANT TO LOSE THIS BOTANICAL. CHINESE DOCTORS, THEY REALIZE WE NEED TO ACT, WE NEED TO GIVE UP THIS INGREDIENT. IT'S NOT SAFELY BEING FORMULATED AND OUR CONSUMER IS OUR BEST ASSET. WE HAD TO GIVE UP AN INGREDIENT WHICH ARGUABLY SHOULD BE STILL USED IF FORMULATED FOR BRONCHITIS. WHAT EVIDENCE THE INDUSTRY REALLY SUPPORTS? SAFETY AND PUTTING CONSUMERS FIRST. THE INDUSTRY WAS THE NUMBER ONE ADVOCATE FOR RELEASING THE GOOD MANUFACTURING PRACTICES. SO DESHEA WAS PATENTED IN '94 AND THEY SAID THEY WOULD WRITE GOOD MANUFACTURING PRACTICES BUT IT TOOK A LONG TIME UNTIL 2007 TO PUT THAT DOCUMENT TOGETHER. THE INDUSTRY WAS A HUGE SUPPORTER. THEY SAW HOW IMPORTANT PUTTING THESE STANDARDS IN PLACE WAS. IN 2006, THE INDUSTRY WAS A MAJOR SUPPORTER OF THE ADVERSE EVENT REPORTING SYSTEM. POST-MARKET SURVEILLANCE. WE WERE FOR THAT, WE THOUGHT IT WAS A VALUABLE PIECE TO OUR SAFETY PARADIGM. JUST IN 2012 MY ORGANIZATION MAJORLY SPEAR HEADED DASKA IT GIVES D.E.A., THE DRUG ENFORCEMENT AGENCY INCREASED AUTHORITY TO ACT ON QUESTIONABLE STEROID INGREDIENT. PRIOR THEY HAD TO DO EXTENSIVE ANALYTIC WORK BEFORE THEY COULD MAKE SOMETHING ILLEGAL. FOOD SAFETY MODERNIZATION ACT, SIMILARLY WE SAW THE VALUE IN OUR SUPPLY CHAIN INTEGRITY THAT IMPACTS OUR INGREDIENT SUPPLIERS AND WE HAVE A VARIETY OF SELF REGULATORY INITIATIVES WE IMPOSE ON OUR OWN MEMBERS. THEY TEND TO BE LABELLING. AROUND CAFFEINE THAT INCLUDES TRANSPARENCY AND LABELLING. QUITE A BIT OF LEGAL OBLIGATION, HAVE YOU TO PRODUCE PRODUCTS THAT DON'T PRESENT A SIGNIFICANT RISK. WHEN YOU HAVE SOMEONE SAYING HEY, SELL POWDERED CAFFEINE, YOU HAVE TO DO AN ASSESSMENT AND DETERMINE YOURSELF, IS THIS AN UNREASONABLE RISK? NOTHING THAT PRESENTS AN IMMINENT HAZARD. NOTHING THAT COULD CONTAIN A POISONOUS INGREDIENT. NEW OR OLD DIETARY INGREDIENTS OR GENERALLY RECOGNIZED AS SAFE IN THE FOOD SUPPLY. YOU MUST BE OPERATING OUT OF A G.M.P. COMPLIANT FACILITY AND ADHERE TO THE ADVERSE EVENT REPORTING LAW. NOW, HOW ARE THESE CATEGORIES DIFFERENT? I THINK ONE OF THE THINGS THAT'S UNDER APPRECIATED WE ARE FOOD, SO RIGHT OUT OF THE GATES THERE'S NO THERAPEUTIC BENEFIT. WE CAN ONLY SUPPORT HEALTHY. WE ARE NOT DRUGS. SO ALREADY WE CAN'T DO A BENEFIT-RISK EQUATION. A LOT OF TIMES IF THERE'S A RISK IN A PRODUCT, EVEN IF IT'S SMALL, LIKE YOU SEE CONSUMER REPORTS, CHAMOMILE CAN CAUSAL ERGIES AND DANGER IN THOSE PRODUCTS ON YOUR SHELF. WAIT A MINUTE. CHAMOMILE HELPS A LOT OF PEOPLE AND ONLY PEOPLE WITH HAY FEVER ALLERGIES. JUST BECAUSE SOME WITH HAY FEVER MAY REACT TO CHAMOMILE PEOPLE GET WORKED UP THAT IT'S AN UNSAFE DIETARY SUPPLEMENT. THE SAME STORY GOES WITH A LOT OF PRODUCTS. OBVIOUSLY WITH DRUGS YOU HAVE A BENEFIT-RISK EVALUATION. THIS IS AN OUTCOME WITH A VERY LONG CONVERSATION. QUESTION A MANUFACTURE ESTABLISHES IT'S OWN SPECIFICATION. YES THIS IS HOW I MAKE AND SELL GINGKO AND THE GUY OVER HERE CAN SET DIFFERENT ONE'S AND THE TWO PRODUCTS WILL BE DIFFERENT. I KNOW THAT'S A HUGE CHALLENGE FOR THE PUBLIC SAFETY COMMUNITY. WE HAVE TO ACCEPT REGULATIONS ALLOW FOR THIS FLEXIBILITY. AND, TO MAKE MATTERS WORSE, YOU HAVE BATCH-TO-BATCH VARIABILITY. I GET GINGKO FROM THE SAME GUY EVERY YEAR. WE HAVE A GREAT SUPPLY CHAIN INTEGRITY. I HAVE SEED-TO-SHELF CONTROL HOWEVER IT RAINED A TON THIS YEAR AT HIS FARM. AND MY CHEMISTRY IS COMING OUT DIFFERENT. STILL MEETS MY SPECIFICATION. I CAN STILL SELL THIS LEGALLY BUT IT'S A LITTLE DIFFERENT THAN THE LAST BOTTLE. BUT GUESS WHAT. SO IS THE SOUP YOU ARE BUYING, A LITTLE DIFFERENT THAN THE LAST CUP OF SOUP. NATURE HAS VARIABILITY. WE MUST BE OK WITH THAT. UNLIKE DRUGS, NEW TO NATURE, SYNTHETIC, COMPOSITION, WELL CHARACTERIZED VERY LITTLE BATCH-TO-BATCH VARIABILITY. HERBS, LONG HISTORY OF SAFE USE. THIS IS CHALLENGING FOR MANY SAFETY ASSESSMENT PEOPLE. HISTORY OF SAFE USE DOESN'T MEAN SAFETY. I HEAR IT ALL THE TIME. BUT WE MUST HONOR IN TRADITIONAL CHINESE MEDICINE, WESTERN CHINESE METE MEDICINE. THESE FOLKS HAVE USED THESE PRODUCTS AND KNOW A LOT ABOUT THEM, THERE'S WISDOM THAT COULD LEND ITSELF TO OUR SAFETY ASSESSMENTS. WE HAVE A HUGE PUBLIC HEALTH CHALLENGE. HOW DO WE EVALUATE THE SAFETY OF THESE PRODUCTS, COMPLEX MIXTURES, VARIETY AMONG COMPANIES AND ANIMAL MODELS MAY NOT BE PREDICTIVE. I KNOW THERE'S A DESIRE TO USE WHAT'S IN THE INDUSTRY, WHAT'S IN COMMERCE, WE REALLY THINK THAT'S IMPORTANT. AT THE SAME TIME WE HAVE MON GRAPHS, WE HAVE U.S.P., AMERICAN HERBAL, ALL OF THESE SPECIFICATIONS MANUFACTURERS COULD USE, IF WE FOCUS N.T.P. STUDY ON ESTABLISHED MONO GRAPH HERB THERE'S POTENTIAL TOXICKITY. WE KNOW THE VERSION OF GINGKO WE NEED TO BE CAUTIOUS ABOUT. OR IT COULD COME OUT THERE'S NO ISSUES AND NOW YOU CREATED A SAFE HARBOR FOR INDUSTRY TO SAY I WILL MANUFACTURE TO THOSE SPECIFICATIONS BECAUSE I KNOW THOSE HAVE BEEN ASSESSED IN IN VITRO STUDIES. REALLY QUICK ON ALOE VER RO, WE KNOW THE STUDY CAME OUT IN 2011. THIS IS WHERE THE INDUSTRY GETS THE RUB. TWO YEARS LATER CONSUMER GROUPS START CALLING ALOE VERA UNSAFE, DON'T CAKE IT. -- TAKE T. THEY DOWN PLAYED THE DIFFERENCE AND IT WASN'T MENTIONED IN THE FIRST TALK. THE ALOE VERA TAKEN TO SHOW CANCER-CAUSING EFFECTS WAS NOT THE ALOE VERA PEOPLE CONSUMED LONG TERM. THAT WAS THE ALOE VERA THAT HAD THE GREEN OUTER LEAF ON IT, VERY HIGH IN THE ALOE A. INDUSTRY MIGRATED OVER YEARS BY KNOWING ABOUT THE PLANT THEY FILTERED THAT OUT. BUT WHEN C.F.P.I. COMES OUT AND TELLS CONSUMERS TO STOP TAKING ALOE, THAT SUBTLELY GETS LOST. SALES DROP. THEY RECONFIRM THE N.T.P. FINDINGS AND NOW 2015, ALOE VERA AT LEAST WHOLE LEAF HAS TO BE LISTED BY THE STATE OF CALIFORNIA AS CAUSING CANCER. MAJOR IMPACT ON THE INDUSTRY. I KNOW THERE'S A SUBTLETY. YOU CAN STILL SELL FILTERED ALOE. THE DAMAGE HAS BEEN DONE. HINDSIGHT IS 20/20. THEY PUT TOGETHER THE MONO GRAPH TO REFLECT WHAT IS SOLD IN INDUSTRY. HOW DO YOU VERIFY PURITY. HOW DO YOU IDENTIFY THE PLANT. HOW DO YOU MAKE SURE IT DOESN'T HAVE TOO MUCH ALOE IN IT. IF THE N.T.P. REPORT IS ON THIS VERSION OF ALOE, I THINK YOU WOULD SEE INDUSTRY REALLY EMBRACING MAKING SURE ALL PRODUCTS MET THESE STANDARDS. ? IN SUMMARY, HOW DO WE REGULATE THE INDUSTRY ENSURE SAFE PRODUCTS BY ONLY USING ALLOWABLE INGREDIENTS. I KNOW I'M REFERRING TO A RESPONSIBLE INDUSTRY. YOU READ ABOUT OUT LIARS IN THE NEWS BUT HERE THERE IS A CORE FOCUS GROUP OF COMPANIES THAT HAVE BEEN IN THIS INDUSTRY FOR A LONG TIME. AND WE ARE EQUALLY ENRAGED BY THE BEHAVIOR OF THE BAD ACTORS AND WE ARE HERE TO MAKE THOSE NEXT STEPS TO GET THROUGH THIS PERIOD SO WE CAN CONTINUE TO GROW AS AN INDUSTRY AND BE PART OF PEOPLE'S HEALTH. WE SET MANUFACTURING STANDARDS G.M.P.'S WE HAVE POST MARKET SURVEILLANCE. SAFETY IS A TOP PRIORITY FOR EVERYBODY, INCLUDING RESPONSIBLE INDUSTRY. THESE N.T.P. STUDIES ONE OF THE MOST GRACIOUS GOVERNMENT AGENCIES VERY GOOD ABOUT OUR COMMENTS. WE STEPPED IN ON THE ALOE WHEN THEY WERE JUST CALLING IT ALOE VERA. WE REALLY HAVE TO USE THE TERMINOLOGY TO REFLECT USING WHOLE LEAF UNFILTERED BECAUSE THERE'S A WHOLE OTHER PRODUCT THAT HAS NOTHING TO DO WITH THIS STUDY. C.R.N. THIS IS A SHOUT OUT TO N.C.I.A.H. WE NOTICED IN THEIR DRAFT STRATEGIC PLAN FOR THE NEXT FIVE YEARS THEY ARE LOOKING AT THE DEVELOPMENT OF BIO AND FRAMATIC. THE INDUSTRY NEEDS TO MOVE IN A DIRECTION WE ARE ADAPTING TO THESE COMPLEX MIXTURES NOT TRYING TO SHOVE THE ROUND PEG IN THE SQUARE HOLE AND I'M EXCITED TO LEARN HOW THAT'S GOING. SMALL SUGGESTION IN THE CURRENT STATE, CONTINUE THESE TWO YEARS MOUSE AND RAT STUDIES THEY FOCUS ON WELL CHARACTERIZED THINGS WITH MONO GRAPHS WELL ESTABLISHED AND IF SOMEONE WILL PROVIDE THAT MATERIAL TO YOU WE CAN HELP YOU GET IT MADE TO THOSE SPECIFICATIONS SO YOU KNOW WHAT ARE YOU TESTING. AND I BELIEVE IT HAS GREAT VALUE BECAUSE THEN AT THE END WHEN THESE STUDIES COME OUT WE DON'T HAVE TO ARGUE ABOUT PICKING THE WRONG PRODUCT OR MATERIAL. WE COULD ALL BE OWNING THE FACT WE CHOSE FOR SOME REASON TO USE THIS FORM OF GINGKO OR ALLOW. -- ALOE. THANK YOU VERY MUCH. I HOPE I WAS ABLE TO ADD TO THE CONVERSATION. >> THANK YOU. IT'S GOOD TO HAVE A CONVERSATION. THE FINAL SPEAKER IN THIS MORNING'S SESSION IS HELEN, LIAISON DIETARY SUPPLEMENTS U.S. PHARMACOPEIA. THANK YOU VERY MUCH. >> IT'S MID MORNING AND I KNOW IT'S NEARLY LUNCH TIME AND MY SLIDES ARE NOT FUNCTIONING FOR SOME REASON. I DON'T KNOW IF SOMEONE CAN HELP ME HERE. WOW, THIS IS BAD. I WANTED TO TALK TO YOU ABOUT BOTANICAL QUALITY STANDARDS AND HOW IT COULD BE USED TO IDENTIFY PROPER MATERIALS. WE HAVE HEARD A LOT ABOUT THE NEED TO USE MATERIAL WHEN DOING RESEARCH AND WE HAVE SEEN QUESTIONS COME UP BECAUSE OF THIS. FOR THOSE WHO ARE AT OXFORD THIS IS A REPEAT PERFORMANCE SO, ENJOY. AGAIN TO TALK ABOUT THE STANDARD. HAS BOTANICAL. IN THE U.S.C. I REALIZE -- WE HAVE (INAUDIBLE) AND WE ALSO HAVE CONTINUUM. WHAT IS CONFUSING FOR PEOPLE IT'S NOT CLEAR WHERE IT COMES FROM BUT WHATEVER YOU HAVE IN DIETARY SUPPLEMENTS CONTINUUM IS WHAT IS USUALLY IN THE U.S.D.N.F. THE COMPENDIUM GIVES A COLLECTION OF ALL THE BOTANICALS QUALITY STANDARDS, TOGETHER WITH THE OTHER MATERIALS LIKE GUIDANCE MATERIALS AND ANY INFORMATION RELEVANT. WHERE YOU CAN FIND ALL THIS INFORMATION COLLECTED TOGETHER. WE HAVE THE COMPENDIUM, THIS WE USUALLY ONLY AVAILABLE ONLINE. IN OTHER WORDS, IT'S FREE. SO IT MEANS ONCE YOU REGISTER AT THE U.S.C. WEBSITE YOU ARE ABLE TO ASSESS THIS AND IT'S AVAILABLE FOR ANYBODY WHO HAS INTERNET, ACCESS TO THE INTERNET. BASICALLY A MON GRAPH IS ONLY ONE OF THE STANDARDS FOR BOTANICALS YOU FIND WITHIN. BUT THERE ARE GENERAL CHAPTERS THAT DESCRIBE CERTAIN FEATURES OF CERTAIN TESTS THAT NEED TO BE DONE AND THIS GENERAL CHAPTER ALSO SOMETHING THAT HAS TO BE FAMILIAR WITH, AS YOU GO THROUGH A MONO GRAPH YOU FIND SPECIFIC TEST IN SPECIFIC GENERAL CHAPTER AND THEREFORE, IN ORDER TO BE ABLE TO SAY THAT YOUR MATERIAL CONFORMS TO THE STANDARD ONE HAS TO BE FAMILIAR WITH THIS TEST AND BE ABLE TO PERFORM THEM AND MEET THE ACCEPTANCE CRITERIA. WHAT I WANTED TO TALK ABOUT TODAY IS SEE TO IDENTIFY MATERIAL AND WHEN WE TALK ABOUT, WE ARE NOT ONLY REFERRING TO THE TEXT IN THE MONOGRAM BUT ALSO THAT YOU NEED TO MEET ALL THE OTHER REQUIREMENTS WITHIN THE MONOGRAM IN ORDER TO CLAIM YOU MEET THE STANDARD AND IN ORDER TO CLAIM THAT YOU HAVE A GOOD QUALITY MATERIAL. THE FIRST THING AS I MENTIONED HAVE DIFFERENT COMPENDIUM. FARM CO PIA AND NATIONAL FORMULARY RY BUT THEY USUALLY HAVE BOTANICAL QUALITY STANDARDS. BASICALLY IT'S THE SAME, HOPE WE WILL BE ABLE TO SHOW YOU THAT WHEN THE COMPUTERS COOPERATE. THE TITLE OF THE BOTANICAL WILL USUALLY BE THE NAME, WHICH IS ACCORDING TO THE REGULATION, IN ADDITION MAKE SURE WE HAVE PLANT PART INDICATED DIFFERENT FACTORS OF THE PLAN MAY HAVE, IT'S IMPORTANT, DIFFERENT PART OF THE MATERIAL YOU ARE USING. THAT HAS ALREADY COME OUT, IN FACT, FOR EX THE PREVIOUS SPEAKER TALKING ABOUT ALOE. TALK ABOUT THE INNER PART OF THE LEAF. PROBABLY VERY DIFFERENT AND IF YOU HAVE THE ROOT IT SHOULD BE A DIFFERENT MATTER ALL TOGETHER. IT'S IMPORTANT TO HAVE PART OF THE PLAN AS PART OF THE MONO GRAPH. WE HAVE A DIFFERENT GUIDELINE. THIS MONOGRAPH USED IN DIFFERENT COUNTRIES OF THE WORLD. WE WANT PEOPLE TO BE ABLE TO UNDERSTAND EACH OTHER WHEN TALKING ABOUT THEIR MATERIALS. WE ARE GETTING CLOSE, SEEING SOMETHING HERE. AFTER THE TITLE THE NEXT THING YOU IDENTIFY WE SEE IN THE MONOGRAPH IS INVESTIGATION. WILL USUALLY GIVE YOU A PURE DEFINITION OF WHAT THE MONO GRAPH IS TALKING ABOUT, IN ADDITION TO GIVING YOU THE NAME OF THE MATERIAL. BINOMIAL AND THE PLANT PART, IT WILL ALSO GIVE YOU A MINIMUM LABEL OF CHEMICAL CO STIT WENTZ THAT WOULD BE REQUIRED TO MEET THE STANDARD. THAT BECOMES AN IMPORTANT PART OF THE MON GRAPH. SUBSEQUENT YOU COME ACROSS A TEST TELL YOU THE MATERIAL AND FOR THE TEST VERY IMPORTANTLY THE U.S.P. DOES A LOT OF WORK AROUND THIS AS YOU MAY ALREADY KNOW, U.S.P. STANDARDS, ACTUALLY REGION BY EXPERTS, BY THE U.S.P. STAFF, WE HAVE EXPERTS, SOME ARE IN THE ROOM RIGHT NOW WHO ARE VERY KNOWLEDGEABLE IN THIS AREA. TO MAKE SURE WE KNOW THE DANGER IN A PARTICULAR BOTANICAL SO IN THAT WAY WE END UP WITH GOOD QUALITY MATERIAL, SO AT THE END OF THE DAY YOU ARE ABLE TO SAY, ACTIVE CONSTITUENTS OR COULD ACTUALLY BE A MARKER TO GIVE YOU WHAT THE QUALITY OF THE MATERIAL IS. NORMALLY WE DON'T ONLY DO ONE. SO YOU REASONABLE ONLY LOOKING AT THE MICROSCOPIC FEATURES BUT LOOK AT FRENCH -- THANK YOU SO MUCH. ALL RIGHT. I WILL RUSH QUICKLY, ALREADY TALKING ABOUT MOST OF THIS ANYWAY. BASICALLY THAT WAS THE MISSION OF THE U.S.B. MANY OF YOU ARE FAMILIAR WITH, THESE ARE THE BOOKS I WAS TALKING ABOUT. ALSO HAVE BOTANICAL. THOSE ARE STANDARDS FOR FOOD COMMODITIES, THE ONE ON THE RIGHT. THE GREEN ONE IS THE COMPENDIUM WHICH CONTAINS MONOGRAMS AND MATERIALS ABOUT THAT SUPPLEMENTS AND THE EXTREME RIGHT YOU HAVE, AVAILABLE ONLY ONLINE. WE LOW WE HAVE THE STANDARD PART OF THE REFERENCE MATERIALS YOU HAVE PART OF THE STANDARDS AND OTHER PARTS OF THE OFFERS. I TALKED ABOUT ALL THESE. IN THE CASE THIS IS CONSISTENT WITH G.M.P. MONO GRAPH FOR BOTANICALS, MOST PLACES YOU HAVE A SET OF MONO GRAPHS, A FAMILY OF MONO GRAPHS. BASICALLY WE HAVE THE MATERIAL MON KNOW GRAPH PLANT AND PLANT EXTRACT MONO GRAPH AND THE NAMES WILL BE AS INDICATED THERE WE ARE TRYING TO HAVE NOMENCLATURE. THE DEFINITION AGAIN GIVES YOU CHARACTERISTICS. PART OF THE PLANT IMPORTANTLY, ACCORDING, IF IT'S A SUPPLEMENT. WE HAVE THE COMPOUND OR COULD BE JUST ANOTHER COMPOUND THAT HELPS TO DETERMINE. ALSO GONE THROUGH ALL OF THIS AS WELL. WHAT I WAS TALKING ABOUT IN THIS CASE TALKING ABOUT -- DEFINITION CONSIST WITH DRIED FRUIT AND BODY OF LUCIUM. HERE IS AN EXAMPLE I HAVE THE TITLE INDICATED THERE. FOR TESTING FOR IDENTIFICATION WE HAVE DIFFERENT METHODS SO HERE WE HAVE ACTUALLY ALSO GIVE AN IDEA OF WHAT BECAUSE WE UNDERSTAND THERE COULD BE VARIATION, COUPLED WITH THE PROCEDURE. WHICH IS WELL DESCRIBED AND GIVES YOU AN OUTLINE OF WHAT IT WILL LOOK LIKE IN TERMS OF PEEKS THAT WILL BE AVAILABLE. WE DON'T JUST LOOK AT THE NUMBER OF PEEKS BUT AVAILABILITY OF MATERIALS IN THOSE PEEKS. IN ADDITION WE HAVE THE COMPOSITION. WE EXPECT TO HAVE LESS THAN 0.3 -- ACID. THIS IS JUST A CONTINUATION OF WHAT THE MONO GRAPH WOULD LOOK LIKE AND GIVE A CLEAR CRITERIA. HOPE YOU ARE ABLE TO SEE THAT. WE HAVE HERE WHAT YOU CALL SPECIFIC TESTS. WE KNOW THAT IT CONTAINS (INAUDIBLE) ALSO SACCHARIDES, THOSE ARE GIVEN SOME SPECIFIC TEST FOR THAT MONO GRAPH. THE OTHER THING WE ALSO HAVE. VERY IMPORTANT, THIS FORMS ANOTHER KIND OF TEST SPECIFIC TO THE CHEMICALS, YOU WILL NOT ONLY LOOK AT MICROSCOPIC FEATURES BUT ALSO MICROSCOPIC FEATURES. THIS FORMS PART OF THE TESTING TECHNIQUES THAT WE HAVE. IN ADDITION TO THAT WE ALSO HAVE OTHER TESTS THAT REFER TO SPECIFIC EXAMPLES, WE HAVE TEST FOR TOTAL ASH AND SOLUBLE EXTRACT AND SO ON AND SO FORTH AND FOR THIS ONE ALSO HAS TO ASSIGN THE DIFFERENT CHAPTERS AND BE ABLE TO CONFORM AS WELL. AT THE END OF THE MON GRAPH YOU USUALLY FIND ADDITIONAL REQUIREMENTS. WE HAVE DETERMINED TO HAVE THAT KIND OF A LABEL. THE MATERIALS THAT ARE LISTED. WE HAVE AT THE END OF THE MON GRAPH DIFFERENT MATERIALS AND AMONG THE DIFFERENT MATERIALS WE HAVE THE DIFFERENT ACIDS BUT ALSO WE HAVE THE -- IN ADDITION -- ALONGSIDE YOUR OWN MATERIALS THAT WAY YOU ARE ABLE TO SEE WHETHER YOU ARE CONFORMING TO WHATEVER THE MONO GRAPH SAYS. HOW THE DIFFERENT TECHNIQUES APPLY TO THE DIFFERENT MATERIALS. USUALLY WE HAVE A GOOD PORTION BEING ABLE TO IDENTIFY USING MICROSCOPIC FEATURES AND MICROSCOPY. YOU FIND THAT THE TEST FOR MICRO SCOPY. BY THE TIME YOU REACH DRY PLANT EXTRACT THERE'S HARDLY ANY MICROSCOPIC THINGS THAT WILL BE HELPFUL. AND YOU NEED TO USE THE PHYTO CHEMICAL TEST. THAT WILL BE REFLECTED IN THE MONOGRAPHS. I'M NOT GOING TO SHOW BUT AT LEAST I'VE SHOWN WHAT A MONOGRAPH OF U.S.B. LOOKS LIKE. OF COURSE THIS PARTICULAR MONOGRAPH IS SPECIFIC TO THE FRUIT AND BODY. WE COULD HAVE MONOGRAPHS FOR THIS. HAVING THIS INFORMATION HELPS WITH THE CHEMICAL TESTING TO BE ABLE TO IDENTIFY THE MATERIALS. HERE WE HAVE FOR THE POWDER, YOU WILL BE LOOKING AT DIFFERENT FEATURES. THIS IS ALL THAT YOU COULD BE ABLE TO DO, I BELIEVE USING ONLY THIS INFORMATION IT WOULD BE DIFFICULT TO IDENTIFY THE MATERIALS SO AT THIS POINT YOU BECOME MORE DEPENDENT ON THE CHEMICAL TEST AND HERE WHAT WE ARE SHOWING. THAT ARE FOUND. YOU COULD SEE WHAT YOU WOULD BE LOOKING AT. WE NOT ONLY DESCRIBE, WE HAVE HERE MATERIAL THAT HAS VERY LITTLE OF THIS. BUT YET IT STILL CONFORMS SO DESCRIBING THE WHOLE OF THE PROFILE IS VERY IMPORTANT. THIS IS THE 366. THIS IS UNDER WHITE LIGHT. AND THIS IS THE PROFILE. YOU COULD SEE WELL SEPARATED PEAKS. ALL OF THIS IS ADDED UP. AS YOU COULD SEE IDENTIFICATION OF THE MATERIAL. IDENTITY TEST ALONE IS NOT SUFFICIENT FOR YOU TO BE ABLE TO COMPLETELY IDENTIFY THE MATERIALS. EVEN IF WE HAVE, WE KNOW, SAY FOR EXAMPLE A LOT HAS BEEN SAID ABOUT D.N.A. TESTING, YOU WOULD BE ABLE TO TEST, KNOW YOU ACTUALLY HAVE THE GROUP RATE OR THE MATERIAL YOU ARE TALKING ABOUT BUT AT THE SAME TIME WHAT YOUR INTERESTS WOULD BE IS PHYTO CHEMICAL IMAGERY. IN FACT PROBABLY MUCH MORE IMPORTANT. I WILL NOT, BEFORE I FINISH, I HAVE TO MENTION WE HAVE CONSIDERED ALSO THE ISSUE OF D.N.A. TESTING WHICH WE KNOW ISN'T GOING AWAY BUT AT THIS POINT, IT WAS PUBLISHED IN THE P.F. FOR COMMENTS AND THE MATERIAL, THIS WAS PROVIDED TO A CHAPTER WHICH IS NOW AVAILABLE IN THE ARTICLES OF BOTANICAL ORIGIN, CHAPTER 563. HOWEVER (INAUDIBLE) WHERE IT WILL FIT. AS WE KNOW IT WOULD JUST BE TEST TO ONE OF THE OTHER TESTS FOR IDENTITY. AT THIS MOMENT THEY ARE CONSIDERING WHETHER TO PUBLISH AS A CHAPTER IN THE U.A.C. OR IN THE U.S.B. ITSELF. HERE JUST TO DEMONSTRATE HOW D.N.A. BECOMES NOT VERY USEFUL AS YOU USE TO PLANT MATERIALS HERE YOU HAVE GOOD QUALITY D.N.A. AS YOU GO TO THE EXTRACT. YOU MAY STILL HAVE D.N.A. BUT NOT VERY GOOD QUALITY AS SHOWN HERE, BY THE TIME YOU REACH EXTRACT YOU HARDLY HAVE ANY D.N.A. TO SAY YOU WILL USE D.N.A. FOR IDENTITY IS NOT SOMETHING THAT WOULD REALLY PASS ON IN ADDITION THEY PROVIDE TO REALLY ENABLE MATERIAL YOU HAVE CONFORM TO GOOD QUALITY BUT DOING THE IDENTITY TEST IS NOT ENOUGH ONE WOULD NEED TO DO THE TEST FOR COMPLETION AS WELL AND THERE ARE ALSO OTHER TESTS WITHIN THE MONOGRAPH THAT I DIDN'T DELVE SO MUCH INTO BECAUSE OF TIME. BUT WE ALSO HAVE SPECIFICATIONS FOR IMPURITIES FOR BIO -- TO MAKE SURE YOU DON'T HAVE ORGANISMS WITHIN YOUR MATERIAL. AND ALL OF THESE HAVE TO BE MET TO BE ABLE TO CLAIM THEY HAVE MATERIAL THAT CONFORMS TO THE U.S.B. STANDARDS AND ALSO ONE HAS TO BE FAMILIAR WITH GENERAL CHAPTERS TO PERFORM SOME OF THE TESTS THAT ARE NOT FULLY DESCRIBED. ? IN ADDITION TO THAT WE HAVE MATERIAL THAT ARE KEY TO PERFORMING THE TEST. BUT ALSO FOR COMPARISON TO BE ABLE TO KNOW YOUR MATERIAL IS THE RIGHT MATERIAL. SO THAT IN ORDER TO MEET THE MEETING U.S.B. STANDARD TO CONFORM WITH ALL OF THE MONOGRAPHS. LASTLY I JUST WANT TO MENTION OUR U.S.B. STANDARD I THINK WE MENTION, NOTE BY THE U.S.B. STAFF BUT BY A GROUP OF EXPERTS THEY BENEFIT AN ART FORM IN THE AREAS THAT TOXICOLOGIES, DEVELOP WITH THESE STANDARDS. THANK YOU VERY MUCH. AND SORRY ABOUT THE PROBLEMS. >> THAT'S HOW YOU DEFINE GRACE UNDER PRESSURE RIGHT THERE. ANOTHER ROUND OF APPLAUSE. WE ARE ACTUALLY ONLY 20 MINUTES BEHIND AND GIVEN WE HAD A 15 MINUTE FIRE DRILL WE AREN'T TOO BAD. APPLAUSE, THANK ALL THE SPEAKERS FOR THIS MORNING'S SESSION FOR KEEPING ON TIME. AND YOU GOT THE MESSAGE WE WILL TAKE A 50 MINUTE LUNCH BREAK. WE WILL TRY TO RECONVENE AT 12:40. THAT'S WHEN WE ARE GOING TO TRY TO RECONVENE. GOING TO START WITH OUR AFTERNOON SESSION. WE HAVE GOT EIGHT SPEAKERS THIS AFTERNOON SO IT'S JAM PACKED. AND THEY'RE ALL GOING TO ADDRESS SOME ASPECT OF TOPIC NUMBER 1 WHICH IS DETERMINING PHYTOEQUIVALENCE AMONG BOTANICAL DIETARY SUPPLEMENTS. WE WILL HAVE ONE BREAK IN THE AFTERNOON, WE'LL STILL TRY TO HAVE IT AROUND THE 2:30, SO WE MAY SHIFT SPEAKERS A LITTLE BIT. AND AFTER OUR FINAL SPEAKER WE'LL CONVENE A PANEL DISCUSSION EXPLORING THREE BASIC AREAS, MINIMUM DATA NEEDS FOR DETERMINING PHYTOEQUIVALENCE, THE USEFULNESS OF CHEMICAL STRUCTURAL FEATURES AND OTHER CHARACTERISTICS. AND THE PROS AND CONS OF DIFFERENT STATISTICAL APPROACHES IN ANALYZING THE DATA PHYTOEQUIVALENCE. AS WELL AS ANYTHING ELSE THAT MAY ARISE IN THE AFTERNOON SESSION. WITH THAT WE'LL START IN WITH OUR FIRST SPEAKER. CYNTHIA RIDER, SHE DOESN'T NEED MUCH OF AN INTRODUCTION. SHE'S RESPONSIBLE FOR THIS TWO DAY WORKSHOP. SHE'S A TOXICOLOGIST SPECIALIZING IN THE COMPLEX TOXICOLOGY OF MIXTURES FROM THE NTP AND SHE WILL PRESENT CASE STUDIES ON PHYTOEQUIVALENCE TO EXPLORE THE QUESTION OF HOW SIMILAR IS SIMILAR ENOUGH. >> SO WHILE I'M GETTING MY CORRECT SLIDES UP HERE, I WILL TELL YOU MY BACKGROUND AS KRISTINE SAID WAS -- IS IN MIXTURES TOXICOLOGY, NOT NECESSARILY IN BOTANICALS. SO I LEARNED THE TERM PHYTOEQUIVALENCE FROM JOE BECK WHO CORRECTED MY USE OF THE TERM SUFFICIENT SIMILARITY WHICH COMES FROM THE MIXTURES WORLD. SO BUT THE THING I LIKE ABOUT SUFFICIENT SIMILARITY IS IT ALLOWS FOR SOME WIGGLE ROOM SO IT'S NOT EXACTLY EQUIVALENT BUT WHAT IS SIMILAR ENOUGH? SO AS MANY SPEAKERS HAVE DISCUSSED THROUGHOUT THE DAY, BOTANICALS ARE COMPLEX MIXTURES, THEY HAVE MANY CONSTITUENTS, MULTIPLE ACTIVE CONSTITUENTS, AND THESE CAN DIFFER DEPENDING ON IF YOU'RE INTERESTED IN THE PHARMACOLOGICAL PROPERTIES OR THE TOXICOLOGICAL ACTIVITY. POTENTIAL FOR INTERACTIONS AND THERE'S OFTEN LARGE UNIDENTIFIED FRACTION. IN ADDITION TO THE INHERENT COMPLEXITY OF THESE BOTANICALS, THERE'S ALSO A LOT OF VARIABILITY IN THE MARKETPLACE. THIS CAN COME AS WE DISCUSS FROM DIFFERENCES IN RAW MATERIAL, DIFFERENCES IN PROCESSING MANUFACTURING AND ADULTATION AND COMBINATION. SO THIS IS THE GENERAL ISSUES WE DEAL WITH BUT FROM NTP PERSPECTIVE WE HAVE SEVERAL CHALLENGES. OUR PRIMARY ARE IN TEST ARTICLE SELECTION AND IN RELATING ANIMAL DOSES TO HUMAN INTAKE. SECONDARILY WE WORK ABOUT EXTRAPOLATING OUR FINDINGS TO OTHER BOTANICALS, IDENTIFYING ACTIVE CONSTITUENT AND DRUG BOTANICAL INTERACTIONS, ALL DISCUSSED THROUGHOUT THE WORKSHOP SO TOMORROW WE HAVE A SIGNATURES AS WELL AS ON ADME, THAT WILL BE TOMORROW BUT FOR TODAY, WE'LL TALK HOW THESE ARE INTERRELATED ISSUES, THEY DON'T EXIST IN ISOLATION. SO IN ORDER TO SELECT THE APPROPRIATE TEST ARTICLE OR TO RELATE ANIMAL DOSES TO HUMAN INTAKE, IT'S IMPORTANT NOTE SOME THINGS ABOUT ACTIVE CONSTITUENTS. ADDITIONAL WHETHER I TEST ARTICLE SELECTION IS FLIP SIDE EXTRAPOLATING FINDINGS TO OTHER BOTANICALS SO THESE ARE ALSO RELATED. SO THE FIRST THING WE'LL TALK ABOUT IN THIS SESSION IS BOTH OF THESE REQUIRE COMPARING ACROSS MULTIPLE LOTS OF BOTANICALS. TO TELL YOU ABOUT OUR TRADITIONAL APPROACH, HOW WE TYPICALLY LOOK AT TEST ARTICLE SELECTION, WE EVALUATE MULTIPLE LOTS FROM VARIOUS SUPPLIERS TO FIND A SINGLE REPRESENTATIVE TEST ARTICLE. AND BY REPRESENTATIVE WE HAVE TO CONSIDER, DOES IT HAVE GREATEST EXPOSURE POTENTIAL? DO WE LOOK FOR SOMETHING THAT HAS THE GREATEST AVAILABILITY P IN THE MARKETPLACE? OR SOMETHING THAT'S THE MOST LIKE A REFERENCE STANDARD. OR HIGHEST LEVEL ACTIVE INGREDIENT SO MOST POTENT. HERE WE'RE LOOKING FOR WORST CASE SCENARIO. WE TRY TO KEEP ALL THESE THINGS IN MIND WHEN SELECTING TEST ARTICLES BUT PRETRITO GET AS CLOSE AS POSSIBLE TO THE REFERENCE STANDARD. WE HAVE MULTIPLE METHODS TO GO ABOUT LOOKING A ACROSS THE DIFFERENT TEST ARTICLE POTENTIAL TEST ARTICLES. WE HAVE UNTARGETTED CHEMISTRY WHERE WE COMPARE ACROSS CROW MAT GRAMS AND TARGETED CHEMISTRY TO QUANTIFY CONCENTRATIONS OF MARKER COMPOUNDS. SO THE FLIP SIDE OF THAT AS I SAID IS HOW REPRESENTATIVE IS THE TEST ARTICLE. WE SELECT ONE TEST ARTICLE TO MOVE FORWARD WITH, WE TEST, VERY ROBUST BIOASSAYS AND REPORT FINDINGS. WE WANT TO KNOW HOW THIS REFERENCE TEST ARTICLE MATCHES ALL THE THINGS THAT COULD EXIST ON THE MARKETPLACE. THINK ABOUT THE DIFFERENCES IN RAW MATERIALS, DIFFERENT PRODUCTS AVAILABLE AND DIFFERENCES IN PROCESSING, THERE'S AN INFINITE NUMBER OF THINGS THAT WE CAN COMPARE TO. SO IN ORDER TO MORE SYSTEMATICALLY TALK ABOUT THIS ISSUE, WE HAVE BEEN DEVELOPING CASE STUDIES AND WE AGAIN THANK ODS FOR THEIR CONTRIBUTION FUNDING CASE STUDY DEVELOPMENT. OUR GOAL WITH THE CASE STUDIES IS TO WORK THROUGH THE PROBLEM OF DETERMINING PHYTOEQUIVALENCE OR SUFFICIENT SIMILARITY WITH MULTIPLE EXAMPLES. AND IN EACH OF THESE EXAMPLES WE WANT TO COMPARE DIFFERENCE APPROACHES FOR DETERMINING SUFFICIENT SIMILARITY USING CHEMICAL SIMILARITY, BIOLOGICAL SIMILARITY, SUPERVISED APPROACHES THAT REQUIRE SCIENTIFIC JUDGMENT AS WELL AS UNSUPERVISED APPROACHES THAT ARE MORE DATA DRIVEN. THROUGHOUT THAT WE WANT TO IDENTIFY KNOWLEDGE GAPS. SO THE THREE CASE STUDY BOTANICALS THAT WE WANT TO LOOK AT ARE GINKGO BILOBA EXTRACT, HE CAN NAY SHAH EXTRACT. WE WANTED TO GET A DIVERSITY OF PROFILES. SO GINKGO BILOBA EXTRACT IS ON ONE SIDE OF THE SPECTRUM. HERE WE HAVE A RELATIVELY LARGE IDENTIFY FRACTION, CHEMISTRY IS WELL DEFINED. THERE'S MARKER CONSTITUENTS, HIGH CONFIDENCE THEY'RE ASSOCIATED WITH BIOLOGICAL ACTIVITY. WE FOUND CYTOTOXICITY WAS MAJOR FINDING. FOR BLACK COHOSH EXTRACT, THIS LARGE UNIDENTIFIED FRACTION WE HAVE LOWER CONFIDENCE THAT MARKER CONSTITUENTS ARE ASSOCIATED WITH TOXICITY. AND THE KEY END POINT WE WANT TO LOOK AT IS GENOTOXICITY. AT THE OTHER END HAND WE HAVE ECHINACEA PURPUREA. A LARGE UNIDENTIFIED FRACTION N. ADDITION IT HAS WEAK ACTIVITY. SO SO FAR IN NTP STUDIES WE FOUND ENHANCED IMMUNE RESPONSE AS THE MAJOR FINING. SO FAR. SO WHAT DO WE HAVE WITH CASE STUDIES? HERE WE LISTED OUT WHAT WE'RE GATHERING SO THE CHEMISTRY, THE LOSS WE PROCURED, WHAT TYPE OF CHEMISTRY HAS BEEN DONE AND WHAT TYPE OF IN VIVO STUDY SO I WON'T GO THROUGH THIS IN DETAIL, JUST WANT TO SAY THAT WE WON'T HAVE TIME THIS AFTERNOON TO COVER ALL THREE CASE STUDIES SO WE'LL FOCUS CAN ATTENTION MOSTLY ON GINKGO BILOBA EXTRACT. AND ONE TALK WILL BE SPECIFIC TO TO BLACK COHOSH EXTRACT. BUT WE PLAN TO CONTINUE ASSESSING WHAT WE HAVE DONE SO FAR AND PUBLISHING THE DATA IN THE PEER REVIEWED LITERATURE SO YOU WILL HEAR MORE ABOUT THE MR. CASEY: STUDIES MOVING FORWARD BUT THE -- CASE STUDIES MOVING FORWARD BUT THE MAIN IS ON GINKGO BILOBA EXTRACT SO I'LL GO THROUGH BACKGROUND AND TO TALK ABOUT EXACTLY WHAT WE HAVE DONE WITH GIN GINKBO. 'S A POPULAR DIETARY BOTANICAL SUPPLEMENT OFTEN USED TO PROMOTE CIRCULATION AND BRAIN FUNCTION. IT WAS SELECTED FOR STUDY AT NP BASED ON LACK OF TOXICITY AND CARS KNOW GENICITY DATA IN THE LITERATURE. WELL CHARACTERIZED CHEMICAL PROFILE. THE MARKER CONSTITUENTS ARE THOUGHT TO BE ASSOCIATED WITH BIOLOGICAL ACTIVITY. STANDARDIZED GINKGO BILOBA EXTRACT SHOULD CONTAIN 24% FLAVINOL GLYCOSIDES, 6% TERPENE LACK TO BEES AND LESS THAN 5 PPM GENKGOLIC ACIDS SO EGB 761 BY SCHWABE IS SELECT FORD THE CLINICAL TRIALS, AND WE HAVE HIGH CONFIDENCE IN THE QUALITY OF THAT PARTICULAR PRODUCT. SO WHEN NTP BEGAN WORK WITH GINKGO BILOBA, WE HAD NOT YET DEINVOLVED OUR TEST ARTICLE SELECTION PROCESS TO WHERE IT IS TODAY. WE WANTED TO GET SCHAWE EBV 761 AND WE WERE UNABLE TO PROCURE BOTH SO WE ASKED COLLEAGUES IN THE INDUSTRY WHAT THEY RECOMMENDED AND WE FOUND SHANGHAI JEAN LING GINKO AND COMPARED IT USING CROW MAT GRAMS AND WE SET QUALITATIVELY THIS LOOKED SIMILAR AND MOVED FORWARD WITH STUDIES OF GINKGO BILOBA SO WE DID IN VIVO AND IN VITRO GENOTOXICITY STUDIES THREE MONTH TOXICITY STUDIES IN THE FISHER RATS AND B 6C 3F 1 MICE, IN RATS AND MICE AS WOMEN THE FIND WHERE IS TOXICITY INCLUDED THE LIVER, NOSE AND THYROID GLANDS AND WE HAVE CARS KNOW GENICITY IN RATS AND CLEAR EVIDENCE OF CARS KNOW GENICITY IN MICE IN THESE STUDIES SO WE'LL FOCUS CAN IN ON THE LIVER AS A TARGET. WHAT WE SEE IN THREE MONTH, INCREASED LIVER WEIGHT, SEVERAL NON-NEOPLASTIC LESIONS IN THE TWO YEAR AGAIN NON-NEOPLASTIC LESIONS IN THE RAT AS WELL AS LIGHT INCREASE IN HEPATOCELLULAR CARCINOMA THAT WAS EQUIVOCAL. THEN WE HAD CARS KNOW GENICITY IN MICE AND LIVER, ALSO A SUITE OF NON-NEOPLASTIC LESIONS. THEN SENT TISSUE FROM THE TWO YEAR MOUSE STUDY TO COLLEAGUES AT NCTR AND THEY FOUND THAT THE MAJOR PATHWAYS IDENTIFIED WITH GINKGO BILOBA TREATMENT WERE DRUG METABOLIZING ENZYME, NECESSARY TWO MEDIATED OXIDATIVE STRESS PATHWAYS AND MIC GENE CENTERED NETWORK IN CELL CYCLE CELLULAR MOVEMENT AND CANCER. OTHER COLLEAGUES DID SOME MECHANISTIC WORK, THEY FOUND IN VIVO MOUSE STUDIES THAT CAR IS POTENTIALLY PXR INVOLVED IN THE HEPATIC RESPONSE. ALSO PEOPLE HAVE DONE WORK IN HUMAN HEPATOCYTES AND G2 CELLS AND FOUND CAR AND AHR ACTIVATION WERE PRESENT WITH GINKGO BILOBA TREATMENT. SO NOW MOVING TO WHAT WE WANTED TO DO WITH CASE STUDIES. OUR FIRST PHASE WAS PROCUREMENT. SO WE WANTED TO GET OUR HANDS ON AS MANY GINKGO BILOBA SAMPLE AS POSSIBLE. WE HAD ONE TEST ARTICLE THAT SERVED AS A REFERENCE BUT WE HAD UNDER THREE STORAGE CONDITIONS SO WE HAD AMBIENT STORAGE FOLLOWED BY MINUS 20-DEGREES THEN ANOTHER ONE THAT WAS STORED IN AMBIENT CONDITIONS FOR MOST OF THE PERIOD OF TIME THAT WE HAD IT. HEN WE HAD ONE THAT WAS PUT IMMEDIATELY INTO THE FROZEN ARCHIVE. WE ALSO PROCURED 20 LOTS OF GINKGO BILOBA FROM DIFFERENT SUPPLIERS SO SOMETIMES MULTIPLE LOTS ARE FROM ONE SUPPLIER BUT WE HAD ONE OR MORE LOTS FROM A RANGE OF SUPPLIERS OF GINKGO BILOBA. WE ALSO GOT TWO STANDARD REFERENCE MATERIALS FROM NIST. FOUR FORMULATIONS CONTAINING EGB 761 AND 12 MARKER CONSTITUENTS REPRESENTING TERPENE LACTONES FLAVOR NOL GLYCOSIDE AND TWO -- FLAVOR NOL AS WELL AS GENGCOTOXIN SO THE FIRST PHASE OF THE WORKS, AND JIM HARNLY WILL TALK IN DETAIL BUT WE DID SOME UNTARGETTED CHEMISTRY SO WITH EACH LOT WE RAN THROUGH HPLC ELSD AND THIS IS JUST AN EXAMPLE OF THE CROW MAT GRAMS. THERE'S LOTS OF PEEKS IN TERPENE AREA AND WE HAD OTHER LOTS, QUALITATIVELY BUT THERE'S SOME LOTS I WILL HIGHLIGHT WHERE YOU SEE A COMPLETELY FLAT SLIDE IN THE AREA OR MAYBE ONE OR TWO PEAKS. THEY LOOK DISTINCTLY UNLIKE THE STANDARD REFERENCE MATERIAL. SO AS I SAID JIM WILL TALK MORE ABOUT THE ANALYSIS OF THIS DATA. BUT I WANT TO MENTION SO SIX OF THE LOTS THAT DID -- THAT HAD A NOTABLE ABSENCE OF PEAKS IN KEY AREAS WEREN'T EVALUATED FOR CONCENTRATION OF THE 12 MARKER CONSTITUENTS SO THESE DIDN'T GO ON TO TARGETED TESTING. WE MEASURED THE 12 MARKET COMPOUNDS, AND THIS WAS A PRINCIPLE COMPONENT ANALYSIS WITH RDAC PUT TOGETHER OF THE RELATIONSHIP BETWEEN THESE. SO WHEN WE GO D&O PRINCIPLE COMPONENT ANALYSIS WE FIND THIS SCATTERING OF GINKO LOTS. WE USE THIS IN ORDER TO SELECT TEST ARTICLES FOR FIVE DAY IN VIVO STUDY AND SCOTT WILL TALK MORE ABOUT THE RESULTS FROM THAT STUDY AS WILL CHRIS GENNINGS LATER IN THE SESSION. BUT SHOW YOU HERE IS OUR GIN CO, THE NTP TEST ARTICLE WITH RELATIVELY HIGH TERPENE LACK TONE LEVELS. TO STANDARDIZED EXTRACT SO TERPENE LACTONES 1/2 NOL GLYCOSIDES AND LOW ACIDS THEN DIGENT GIN CO-. ROUNDED OUT ARE SET SO TO GO THROUGH WHAT WE HAVE IN TERMS OF BUY LODGE DACAL ENDS POINTS EVALUATED WITH GINGKO, BIOLOGICAL END POINTS. MEDIUM AND LONG TERM STUFF CAN DIS AND WE MEASURED A HOST OF END POINTS FROM FROM HISTOPATHOLOGY AND 41 TISSUES, CLINICAL HISTORY HEMATOLOGY SO WE CAN ONLY DO THAT WITH ONE LOT. WE CAN'T TEST MULTIPLE LOTS IN THESE TWO YEAR ASSAIL SAYS THAT TAKE A LONG TIME, VERY EXPENSIVE SO WE WANT TO LINK THOSE TO SHORTER TERM ASSAYS. WE INCORPORATED THE FIVE DAY RAT STUDY AND LOOKED AT LIVER WEIGHT AND GENE EXPRESSION IN THE LIVER BECAUSE THESE WERE IDENTIFIED AS IMPORTANT DURING OUR LONG TERM STUDIES. WE ALSO WANTED TO INCORPORATE HIGH THROUGH PUT ASSAYS. SO WE USE HUMAN PRIMARY HEPATOCYTES WHICH STEVE FERGUSON WILL TALK ABOUT IN A LITTLE WHITE. WE HAD END POINTS THAT WE THOUGHT SHOULD BE EFFECTED BY GINGKO KNOWING WHAT WE KNOW ABOUT ITS MECHANISM OF ACTION. HERE WE CAN TEST ALL LOTS, ALL STANDARD REFERENCE MATERIALS AS WELL AS THE PRODUCT FORMULATIONS AND MARKER CONSTITUENTS FOR TOTAL OF 41 SAMPLE. WHAT WE HOPE TO DO WITH THIS IS HOPE TO RELATE THESE SHORT-THE TERM AND HIGH THROUGH PUT END POINTS BACK TO OUR KNOWN TARGETS IN THE LONG TERM TESTS. AND THEN IN THE REVERSE WE HOPE TO FOLLOW THIS REFERENCE LOT THROUGH THESE DIFFERENT ASSAYS SO THAT WE CAN SEE HOW IT EXISTS IN THIS UNIVERSE OF SAMPLES THAT WE'RE TESTING. WE STARTED WITH ALL THE AVAILABLE GINKGO BILOBA IN THE MARKET PLEASE AND SELECT A SUBSET TO EVALUATE THOSE FOR POTENTIAL TEST ARTICLE USE. WE DO SOME LIMITED CHEMICAL ANALYSIS TO START OFF WITH, WE SELECT OUR TEST ARTICLE, WE HAVE THAT ONE TEST ARTICLE THAT WE THOROUGHLY CHARACTERIZE IN TERMS OF DOING COMPLETE CHEMICAL CHARACTERIZATIONS, HAZARD CHARACTER RATION AND ADME/TK WORK. SO YOU CAN SEE THIS PROCUREMENT OF SAMPLES AND LIMITED CHEMICAL ANALYSIS IS VERY IMPORTANT IN THIS PROCESS. THE QUESTION WE'RE REALLY ASKING IS WHETHER THAT'S ENOUGH. IS THAT OKAY TO TEST SELECT TEST ARTICLE THAT WAY, DOES THAT WORK? SO THIS CASE STUDY PROJECT IS REALLY TRYING TO ROOK AT AER SPECTRUM OF SAMPLES LIKE IN VITRO AND SHORT TERM IN VIVO ASSAYS, CHEMICAL ANALYSIS ACROSS A WIDER SAMPLE OF TEST ARTICLES AND STATISTICAL METHODS TO COMPARE WHAT WE'RE FINDING. COMPARE ACROSS THE DIFFERENT SAMPLES. SO TO GIVE YOU AN OVERVIEW WHAT WE'RE GOING TO TALK ABOUT IN THIS SESSION, GLENN RICE WILL START OFF GIVING BACKGROUND ON SUFFICIENT SIMILARITY FROM A RISK ASSESSMENT PERSPECTIVE. WE WILL SHOW US HOW WE THINK ABOUT SUFFICIENT SIMILARITY USING EXAMPLES OUTSIDE THE BOTANICAL WORLD. THEN JAMES HARNLY WILL TALK UNTARGETED CHEMISTRY USING DATA GENERATED IN THE GINKGO BILOBA EXTRACT CASE STUDY. KERRI WILL TALK TARGETED CHEMISTRY AND THROUGH AUTHENTICATION PROCESS. STEVE FERGUSON WILL FOLLOW WITH IN VITRO HEPATOCYTE DATA FOR GINKGO BILOBA EXTRACT. FURTHER CASE STUDY. STEPHANIE THERE L TALK ABOUT IN VITRO BLACK NUCLEUS DATA WITH BLACK COHOSH TO CONTRAST WITH THE GINGKO EXAMPLE. THEN CHRIS WILL BRING THE CHEMISTRY AND BIOLOGY ABOUT SUPERVISED APPROACH FOR COMBINING THIS DATA. TO DETERMINE SUFFICIENT SIMILARITY AND WE'LL HEAR FROM SCOTT AURBACH ON A MORE DATA DRIVEN APPROACH THAT DOESN'T HAVE AS MUCH SUPERVISION PERHAPS FOR COMBINING CHEMISTRY AND BIOLOGY TO LOOK AT THE SUFFICIENT SIMILARITY QUESTION. SO THROUGHOUT THIS AFTERNOON SESSION I HOPE WE CAN THINK ABOUT QUESTIONS TO TALK ABOUT DURING THE PANEL DISCUSSION. CHRISTINE MENTIONED HIGHLIGHTS TO THE DISCUSSION SO WE'RE INTERESTED IN FINDING OUT MORE ABOUT WHAT KINDS OF CHEMICAL AND BIOLOGICAL DATA ARE MOST USEFUL FOR DETERMINING SUFFICIENT SIMILARITIES. HOW PREDICTIVE OF BIOLOGICAL RESPONSE IS CHEMICAL SIMILARITY. SO IS THAT FIRST STAGE WHERE WE JUST USE THE LIMITED CHEMISTRY AND IS THAT ENOUGH TO HAVE CONFIDENCE IN OUR SELECTED TEST ARTICLES. WHAT CHARACTERISTICS OF THE BOTANICAL IN QUESTION DETERMINE WHETHER OR NOT IT IS A GOOD CANDIDATE FOR PHYTOEQUIVALENCE ASSESSMENT, DO WE HAVE TO KNOW BASELINE AMOUNT OF INFORMATION ABOUT ACTIVE CONSTITUENTS AND CHEMISTRY OF THESE IN ORDER TO MOVE FORWARD WITH THESE METHODS OR CAN WE USE THEM REGARDLESS OF WHAT WE KNOW. WHAT ARE PROS AND CONS FOR EVALUATING SIMILARITY. HERE WE TALK ABOUT SUPERVISED VERSUS DATA DRIVEN APPROACHES. WITH THAT I WOULD LIKE TO THANK EVERYBODY WHO WORKED ON THIS PROJECT DEVELOPING THE CASE STUDIES, LEAD CHEMIST AT NTP WORKED ON EVERY SINGLE BOTANICAL WE WORKED ON. BRAD COLLINS HAD INVOLVEMENT IN THE GINGKO PARTICULARLY. OR CONTRACT LABS MRI AND MATTEL GENERATE AD LOT OF THIS DATA, OUR STUDY SCIENTISTS FOR EACHANECIA AND AT THIS TEN LIVE STONE IS THE STUDY FOR BLACK COHOSH EXTRACT. IN VITRO WORK PERFORMED BY STEVE POST-DOC AND STEPHANIE LED THE BLACK COHOSH EFFORT WITH ILS. SCOTT AURBACH AND MOLLY VALLANT AND BATTELL AND THANKS FOR JAMES HARNLY AND CHRIS GENERAL BINGES AND THANKS TO ODS FOR FUNDING OF THE STAYS CASTE STUDY DEVELOP. WHICH IS VERY HELPFUL. WITH THAT I THANK YOU FOR YOUR ATTENTION. IF YOU HAVE ANY QUESTIONS. [APPLAUSE] HE CAN NAY SHAH CO-HOSH >> OUR NEXT SPEAKER IS DR. GLENN RICE, ENVIRONMENTAL HEALTH SCIENTIST AT US EPA OFFICE OF RESEARCH AND DEVELOPMENT WITHIN NATIONAL CENTER FOR ENVIRONMENTAL ASSESSMENT. HE'S AN EXPERT IN BOTH COMPONENT BASED AND WHOLE MIXTURE RISK ASSESSMENT AND HAS DONE PIONEERING WORK TO DEVELOP CASE STUDY USING WATER DISINFECTION BY-PRODUCTS. THIS AFTERNOON HE WILL TALK ABOUT THE MIXTURE RISK ASSESSMENT CONSIDERING SUFFICIENT SIMILARITY. >> A NUMBER OF CO-AUTHORS, THIS PROJECT EMERGED FROM SOME COLLABORATIVE WORK. ONE OF MY CO-AUTHORS IS IN THE ROOM AS WELL SO ANY MISTAKES OR HARD QUESTIONS GO TO CHRIS. SOME OF THE OTHER THINGS I COME UP WITH TODAY EMERGE FROM DISCUSSIONS WITH JASON LAMBERT AND THIS ON AOP REALLY COME FROM THOSE DISCUSSIONS. BECAUSE I'M FROM EPA THE VIEWS EXPRESSED IN THIS PRESENTATION IN NO WAY REFLECT ON THE AGENCY OR ITS POLICIES. AS KRISTINE POINTED OUT, WHAT I HOPE TO TALK ABOUT TODAY IS AN OVERVIEW REAL QUICKLY OF CHEMICAL MIXTURES METHODS AND FOCUS IT ON SUFFICIENT SIMILARITY WHICH IS ALLUDED TO BY CYNTHIA IN THE PREVIOUS TALK. AND THEN WE WILL TALK PRINCIPLE COMPONENTS ANALYSIS AND HOW IT'S USED TO EVALUATE SUFFICIENT SIMILARITY, DO THAT FAIRLY HIGH LEVEL. DR. BETS SHOWED A PCA SLIDE IN HIS PRESENTATION EARLIER TODAY AS WELL. USED IT IN A SIMILAR WAY. THIS IS A GENERAL APPROACH FOR MIXTURES RISKS IN THE ENVIRONMENT THAT WE USE AT EPA. WE HAVE COMPONENT APPROACHES SIMILAR MIXTURE COMPONENTS WE'LL FOCUS ON AND MIXTURES OF CONCERN. THIS IS REALLY DATA DRIVEN THE WAY THE EPA GUIDELINES WERE LAID OUT, THE CHOICE OF THE APPROACH IS REALLY GOING TO DEPEND ON THE AVAILABLE DOSE RESPONSE AND EXPOSURE DATA. THERE IS INCREASING PREFERENCE BY RISK ASSESSORS. WE PREFER A HIXTURE OF CONCERN, IF WE CAN'T WE USE A SURROGATE SIMILAR MIXTURE AND IF WE HAVE TO, WE EEL USE COMPONENT APPROACHES. THERE'S FEW TOX STUDIES CERTAINLY IN THE ENVIRONMENTAL MIXTURES WORLD THAT I COME FROM. THAT STUDY DOSE RESPONSE ASSESSMENTS FOR WHOLE MIXTURES IN ADDITION TO TYPICAL RESOURCES REQUIRED FOR A SINGLE CHEMICAL ASSAY, MIXTURE STUDIES REQUIRE A LOT MORE RESOURCES GENERALLY. ALSO JUST SOME OF THE PREVIOUS DISCUSSIONS WITH SOME OF THE SPEAKERS HAD, YOU SEE FOR THE BOTANICALS I DON'T THINK COMPONENTS ARE PRACTICAL AT THIS TIME. SOME OF THE COMPONENTS AREN'T WELL CHARACTERIZED, THERE SEEMS TO BE A NUMBER OF UNCERTAINTY REGARDING WHICH COMPONENTS ARE BIOLOGICALLY ACTIVE IN THE AMONG SOME OF THE BOTANICALS ET CETERA. SO IT PUSHED THIS TOWARDS THIS IDEA OF SUFFICIENT SIMILARITY. SO WHAT DO WE MEAN BY THAT? THIS IS TAKEN PRIMARILY FROM US EPA 2000 GUIDANCE ON CHEMICAL MIXTURES. AND SUFFICIENTLY SIMILAR REFERS TO THIS IDEA THAT THE TOX COLOGIC CONSEQUENCES OF THE EXPOSURE TO TWO MIXTURES ARE NEARLY IDENTICAL. SO JUST KIND OF A QUICK FLOW DIAGRAM HOW THAT WORK, YOU HAVE A MIXTURE OF CONCERN, THE FIRST QUESTION YOU WOULD ASK YOURSELF, IS IT FEASIBLE TO DEVELOP A DOSE RESPONSE DATA ON THAT MIXTURE OF CONCERN. THE ANSWER IS YES. DEVELOP IT USING THE RISK ASSESS MENT. IF NO, WE WANT TO UNDERSTAND IS THE MIXTURE INFORMATION ABOUT THERE'S NO MIXTURE OF INFORMATION KNOW INFORMATION ABOUT MIXTURE OF CONCERN, YOU GET INFORMATION ON THE CHEMICAL -- SOME OTHER SIMILAR CHEMICALS AND IF THE ANSWER IS YES WE USE THOSE AS SURROGATES FOR THE DOSE RESPONSE DATA IF THE ANSWER IS NO WE USE COMPONENT DATA. SO ONE THING CYNTHIA ASKED ME TO THINK THROUGH AND TALKED WITH THE GROUP OF AUTHORS WHAT CONSTITUTES USEFUL DATA TO EVALUATE SUFFICIENT SIMILARITY. THINKING PRIMARILY ABOUT ENVIRONMENTAL DATA BUT I THINK THESE WILL APPLY AT LEAST IN SOME ASPECT TO THE TYPE OF INFORMATION YOU'RE CONCERNED ABOUT WITH BOTANICALS. THE FIRST TYPE OF DATA IS JUST A CHEMICAL COMPOSITION. CLEARLY YOU WANT TO KNOW THE MAJOR CHEMICALS, CHEMICAL CLASSES, UNDERSTAND THE DIFFERENT PROPORTIONS OF THE COMPONENTS IN THE DIFFERENT MIXTURES THAT YOU HAVE IN THE ENVIRONMENT VERSUS THOSE THAT YOU TEST. AND YOU WILL PREPARE THE MIXTURES IN DIFFERENT WAYS, FAIRLY COMPLEX MIXTURES LIKE THIS INFECTIOUS BY-PRODUCTS FROM DRINKING WATER TREATMENT. PREPARATION METHODS HAVE SIGNATURE AFFECTS ON THE MIXTURE ITSELF AND YOU NEED TO BEGIN TO THINK ABOUT THOSE AS YOU PREPARE ENVIRONMENTAL MIXTURES WHICH MIXTURE OF CONCERN TO THE ONES WE TEST. THEN THERE'S THIS IDEA OF THE UNIDENTIFIEDED FRACTION, AND DOES IT CONTRIBUTE TO THE TOXICITY OF THE CHEMICALS. ALSO THE NUMBER OF DIFFERENT TYPES OF TOXICITY DATA. THIS IS WHERE SOME OF THE CONVERSATIONS THAT I HAD WITH DR. LAMBERT AND OTHERS HAVE KIND OF BEGUN TO GET ME THINKING ABOUT WHAT WE NEED. SO FIRST TO RUN -- DO SUFFICIENT SIMILARITY WE NEED DOSE RESPONSE DATA FOR THE WHOLE TESTED MIXTURE. THAT'S A REQUIREMENT. YOU NEED TO HAVE THAT TO USE IT. IDEALLY WE WANT TO UNDERSTAND THE RELATIONSHIPS BETWEEN DOSES TESTED AND THE ENVIRONMENTAL A EXPOSURE DOSES. SO ROUTES OF EXPOSURE, MAGNITUDE OF DOSE, ET CETERA. WE ALSO WANT TO UNDERSTAND HOW USEFUL TOXICITY DATA TESTS MIGHT BE. ARE QUESTION W DEALING WITH PRECURSOR AFFECT, PERTURBATION FOR MIXTURE CAN AND WE WANT TO UNDERSTAND THOSE RELATIONSHIPS BETWEEN THOSE TYPES OF OUTCOMES AND THE APICAL END POINT OF CONCERN, WHAT'S THE OUTCOME WE'RE CONCERNED ABOUT PEOPLE. ARE WE DEALING WITH WITH IN VIVO DATA, IN VITRO, IN SILICO DATA, HOW GOOD IS THE LAB SPECIES WE DEAL WITH. HOW RELEVANT IS THAT THE HUMAN WE'RE CONCERNED ABOUT, ET CETERA. THE OTHER THING TO KNOW, AND GIVES ADDITIONAL CONFIDENCE IS THIS IDEA DO WE UNDERSTAND WHAT THE KEY OR ACTIVE INGREDIENT OR COMPONENT IS IN THE MIXTURE. DO WE KNOW WHAT -- AT LEAST THAT WE HAVE SOME HYPOTHESIS ABOUT THE COMPONENTS THAT ARE ACTUALLY ELICITING THE TOXICITY IN ASSAY AND CAN WE CONFIRM THAT WITH INDIVIDUAL CHEMICAL TESTS. FINALLY THINK ABOUT STATISTICAL CRITERIA AND PART OF THE EVALUATION YOU WANT TO ESTABLISH TRICKERY TIERIA FOR JUDGING WHETHER THE MIXTURES DIFFERENCE OR NOT. AND AT LEAST IN THE STUDY WE DID WITH CHRIS A NUMBER OF YEARS AGO WE DID THIS AND IT WORKED EXTREMELY WELL. AND CLEARLY THIS CAN EVOLVE WITH SUCCESSSIVE EMPERIMENTS AND CAN BECOME AN AN ITERATIVE PROCESS. YOU WANT IT TRANSPARENT, PRACTICAL, APPLICABLE TO TYPES OF TOXICITY STUDIES YOU MIGHT USE. YOU WANT REPRODUCIBLE AND ALSO VERY RELIABLE. WHAT DO WE NORMALLY HAVE, CYNTHIA ALLUDED TO, WE HAVE A FULL WHOLE MIXTURE STUDIES TESTED IN VIVO OR IN VITRO AND RESPONSES ARE AT THE ORGANISM MALL OR ORGAN LEVEL. TYPICALLY WITH APICAL OUTCOMES. THIS IS STARTING TO CHANGE AS WE GET A LOT OF ADDITIONAL INFORMATION FROM HIGH THROUGH PUT PLATFORMS SO THE LIMITATIONS OF TRADITIONAL DATA ARE FEW MIXTURES GET TESTED, THERE'S CHEMICAL AND TOXICITY DIFFERENCES AMONG THE MIXTURES NOT WELL KNOWN OR UNDERSTAND AND THE RELEVANCE OF TOXICITY TESTS TO THE APICAL OUTCOMES SO YOU WANT TO USE IMMUNOGENICITY INFORMATION AS SURROGATE FOR CANCER WHICH IS A STUDY WE PUBLISHED 15 YEARS AGO NOW. BUT AGAIN, IT'S THE LIMITATION OF THE TYPE OF PATIENT YOU HAVE, WHEREAS THE HIGH THROUGH PUT PLATFORM OR TOXICOGENOMICS, BIOINFORMATICS START TO PROVIDE AND DOUSE US WITH JUST ABUNDANCE OF ADDITIONAL DATA ON THE MIXTURES AND ON COMPONENTS. BUT I THINK THE USES OF THOSE DATA REQUIRE SOME THOUGHT AND CAUTION. WE HAVE TO BE CAREFUL INTERPRETING THE RELATIONSHIP BETWEEN MEASURED PERTURBATIONS AND THE APICAL OUTCOMES OF CONCERN. AND HOW THOSE RELATIONSHIPS INFORM RISK BY THE MIXTURE AND ALSO QUESTIONS OF SUFFICIENT SIMILARITY. SO THIS IS FROM JERRY HANDILY'S PAPER ON KEY FEATURES OF ADVERSE OUTCOME PATHWAYS. YOU CAN SEE A NUMBER OF DIFFERENT EVENTS IN THIS CONCEPTUAL DIAGRAM FROM MACRO MOLECULAR INTERACTIONS, CELLULAR RESPONSES LEADING TO ORGAN RESPONSES, ORGANIST MALL RESPONSES AND POPULATION RESPONSE, AOPs ARE DEVELOPED, THIS IS NOT THE END POINT POPULATION RESPONSES ARE NOT AN END POINT WE DEAL WITH A GREAT DEAL IN HUMAN HEALTH RISK ASSESSMENT. BUT COUPLE OF KEY POINTS EMERGE. FIRST THESE ALL THESE RESPONSES TO MIXTURES HAVE A BIOLOGICAL UNDERPINNING. AND THESE EVENTS THAT COMPRISE CHEMICAL AND BIOLOGICAL INTERACTIONS OCCUR IN THIS BROADER CONTEXT OF BIOLOGICAL SYSTEM WHICH HAVE VARIANCE THEMSELVES. WE MAY NOT KNOW OR COMPLETELY UNDERSTAND ALL THESE DIFFERENT EVENTS OR THEIR IMPLICATIONS IN PRESENTING THINGS IN RA LINEAR FASHION THAT MAY NOT BE LINEAR IN REAL LIFE BUT AOP, THIS IS A USEFUL C KNOWLEDGE WE HAVE CONCERNS THE LINKAGE BETWEEN MOLECULAR INITIATING EVENT AND ADVERSE OUTCOME AND DIFFERENT BIOLOGICAL LEVELS OF ORGANIZATION. THIS IS TRAINED IN A SEQUENTIAL WAY BUT AGAIN, THINGS PROBABLY DON'T HAPPEN THAT SEQUENTIALLY. THINK ABOUT MIXTURES RISK RECEPTION, WHAT DO WE CAN DO WITH THIS INFORMATION. THINKING HIGH LEVEL. AOP DEPICT PLAUSIBLE CONNECTIONS BETWEEN MOLECULAR INITIATING EVENT AND ADVERSE OUTCOME PATHWAY. WE HAVE PRETTY GOOD UNDERSTANDING LEVEL OF BIOLOGICAL ORGANIZATION AVAILABLE TO BE TESTED. WE CAN OBTAIN INFORMATION TO CHARACTERIZE DOSE RESPONSE OR SIMILARITY, AMONG MIXTURES AT THE VARIOUS LEVELS. I THINK THIS IS ASSESSING THE CONFIDENCE BETWEEN RESPONSE WE LOOK AT AND THE APICAL OUTCOME WE'RE MORE CONCERNED ABOUT FROM SUFFICIENT SIMILARITY. SO WE MIGHT USE DATA THAT ARE FURTHER UPSTREAM, TRYING TO ASSESS SIMILARITY OF OUTCOME, VERY FAR DOWNSTREAM. THE OTHER POINT TO MAKE IS IT FORCES YOU TO MAKE A EXPLICIT CHARACTERIZATIONS OF THESE RELATIONSHIPS. BETWEEN THE RESPONSE IN THE ASSESSMENT TO ESTIMATE SUFFICIENT SIMILARITY IN THIS CASE AND THE PLAUSIBLE END POINT. BETWEEN EACH STEPS WE'RE SHOWING PREVIOUS SLIDES YOU GOT KEY EVENT RESPONSE RELATIONSHIP SOME SHAPES OF KEY EVENTS MAY NOT BE MONO TONIC. SO THERE'S A LOT OF COMPLEXITIES BUT THERE'S A COMPLEXITIES THAT ARE TYPICALLY GLOSSED OVER AND ASSESSMENTS. I THINK NEW DATA STREAMS FORCE US TO CONTEND WITH THAT. I HOPE IT MAKES FOR BETTER RISK ASSESSMENT. ONE QUICK EXAMPLE, THIS IS WORK FROM MCDONALD ET ALL 2004, ONE OF THE SIGN TESTS WERE ASSOCIATED WITH LOOKING AT EXHAUST SAMPLES FROM DIFFERENT ENGINES AND LOOKING AT SUFFICIENT SIMILARITY EVALUATED BASED ON COME POUNDS RELATIVE TONGSISTY INFORMATION BUT THEY DID HAVE BOTH CHEMICAL TOXICITY INFORMATION AVAILABLE FOR THE WHOLE MIXTURE. PRINCIPLE COMPONENTS ANALYSIS, AS WELL AS MULTI-VARIANT REGRESSION. INVESTIGATE STATISTICAL RELATIONSHIPS AMONG THE PARTICULATE AND SEMIVOLATILE ORGANIZE CHEMICAL CONSTITUENTS OF GASOLINE AND DIESEL VEHICLE EXHAUST. FOUR DIFFERENT GASOLINE EXHAUST SAMPLES WHETHER GAS WAS COLLECTED ROOM TEMPERATURE, WHERE CARS WERE 33-DEGREES FAHRENHEIT AND NORMAL VERSUS HIGH OMITTING CARS THAT EMITTED WHITE GATS VERSUS HIGH EMITTING CARS WITH BLACK GAS. SIMILARLY FOR DIESEL EXHAUST SAMPLES THEY HAD NORMAL CARS AT ROOM TEMPERATURE, AT 30-DEGREES AND ALSO HAD ONE THAT WAS HIGH AS WELL. SO THEY HAVE SEVEN SAMPLES THAT ARE VERY COMPLEX VOLATILE AS WELL AS PARTICULATE, DRAWING FROM THEIR SAMPLE. THEY'RE USING A PRINCIPLE COMPONENTS ANALYSIS APPROACH. FOR CLUSTER ANALYSIS, IT TRIES TO CAPTURE THE VARIANTS OF THE DATA SET AND REDUCE DIMENSIONALITY OF THE DATA WITHOUT A LOT OF E DETAILS. BUT EACH TRANSFORMED VARIABLE IS A LINEAR COMBINATION OF THE ORIGINAL VARIABLES. AND IT'S CORRELATED WITH THE OTHERS. EACH PRINCIPLE COMPONENT REDUCES VARIABILITY THROUGH TRANSFORMATIONS IN THE DATA SET. SO I WILL IGNORE THE CHEMISTRY DATA IN THE INTEREST OF TIME, JUST SHOW THEY HAD THEY HAD 11 TESTS FOUR TYPES OF LONG TOXICITY TESTING DATA THEY USED. THAT HAD INFORMATION POTENCY ESTIMATES, CYTOTOXICITY, PARENCHYMAL CHANGE AND GENERAL TOXIC POTENCY ESTIMATES THEY USED FROM THE LUNG, YOU CAN SEE THE DIFFERENT TYPES OF TISSUES THEY ASSESS. THEY ALSO HAD BACTERIAL MUTAGENICITY DATA USED AS WELL. SO SUBSEQUENT SLIDES HE WILL STRAIT SEVEN EXHAUST SAMPLES DEMONSTRATE GROUP DIFFERENTLY WHETHER TOXICITY DATA OR MUTAGENICITY DATA, NOT SHOWING THE CHEMISTRY DATA BUT THE SAME BASIC PATTERN. HERE IS PRINCIPLE COMPONENTS ANALYSIS FOR THE LUNG TOXICITY DATA AND YOU CAN SEE WHERE THERE'S THREE DIFFERENT GROUPS OF DATA. BIGGER ANALYSIS WHICH SHOW STATISTICAL AS ASPECTS TO THIS. I'M GOING TO GLOSS OVER FOR TODAY. THREE DIFFERENT GROUPS OF DATA YOU MIGHT GROUP THESE TOGETHER BASED ON PRINCIPLE COMPONENTS ANALYSIS OF THESE LUNG TOXICITIES, 11 LUNG TOXICITY ASSAYS. FOUR DIFFERENT MUTAGENICITY ASSAYS, WE SEE FOUR DIFFERENT GROUPS, BASED ON THESE DIFFERENT MUTAGENIC RESPONSES TO THE DIFFERENT SAMPLES AND YOU CAN SEE THAT THESE ARE DIFFERENT GROUPING THAN PREVIOUS SLIDE SO SIMILAR IN SOME CONTEXT BUT DIFFERENT IN OTHER CONTEXT. THIS IS AN IMPORTANT POINT THINKING SUFFICIENT SIMILARITY. SO PCA APPROACH, TAKES A RELATIVELY RICH SET OF CHEMICAL AND BIOLOGICAL MEASURES BY ENVIRONMENTAL SAMPLES AND THE GROUPING DEPENDS ON TOXICOLOGIC TESTS YOU'RE ANALYZING. SO WE SEE THESE DIFFERENT GROUPS THAT I JUST ABBREVIATED HERE, WIND UP IN VERY DIFFERENT GROUPS, LOOK AT HD, PAIRS UP WITH BG, IN OUR FIRST EXAMPLE, WHEREAS THE SECOND ONE YOU CAN SEE WHILE THESE ARE DIFFERENT GROUP TWO OTHER TYPES OF EXHAUST SAMPLES WIND UP IN THE SAME GROUPS WITH THEM SO I THINK THE POINT IS, I'M TRYING TO -- MADE HERE IS TA THERE'S LOGICAL SIMILARITIES DEPEPPING ON TYPE OF TEST SO WHAT THE APICAL CONCERN HERE IS DEPENDENT STRONGLY ON TYPE OF TEST SO UNDERSTANDING RELATIONSHIP BETWEEN WHAT YOU TEST AND THE OUTCOME BACKS A REALLY IMPORTANT CONSIDERATION FOR SUFFICIENT SIMILARITY. WELL, WE HAVE MULTIPLE TYPES OF ASSAYS THAT ARE SIMILAR, IT WINDS UP IN THAT 11 TOXICITY TEST ASSAYS THEY USED FOR LUNG TOXICITY. STATISTICALLY MORE HEAVILY IN ANALYSIS OF SUFFICIENT SIMILARITY. AGAIN, YOU SORT OF USE EXPERT TOX COLOGIC INFORMATION. THAT'S A LOT OF TOXICOLOGISTS IN THE ROOM TO BRING THE BEAR ON THOSE TESTS USED TO EVALUATE SUFFICIENT SIMILARITY. THERE'S A NUMBER OF FORMAL INFERENCE THINGS I IGNORED HERE IN THIS PARTICULAR STUDY BUT CLEARLY YOU WANT TO BE VERY ROBUST WITH YOUR STATISTICAL METHODS USED AND YOU WANT TO ADD ADDITIONAL TESTS TO THAT TO ROUND OUT OTHER SAMPLES TO SEE HOW SIMILAR THOSE FIT WITH THE SAMPLES HERE. JUST TO TEST YOUR OVERALL STATISTIC OF PREDICTED OUTCOMES. PRINCIPLE COMPONENTS, THE REALLY USEFUL APPROACH IN THE CONTEST -- CONTEXT HERE FOR BOTANICALS BUT THEY ARE REALLY THE TRANSFORMED REAL WORLD VALUES AND MEASURES SO IT'S NOT ANYTHING MAGICAL, THERE'S AN EXAMPLE WE USE FROM THE DVP WORLD WHERE YOU CAN'T SHOW THE MEASURES REPORTED ARE CONSISTENT WITH OUR UNDERSTANDING OF THE TOXICOLOGY OF THE INFECTION BY-PRODUCTS. WE WANT TO EVALUATE WHICH REAL WORLD MEASURES, WHETHER TOXICITY, TOOK CITIES DRIVING ANALYSIS AND SEEK BIOLOGICAL EXPLANATION. TO REINFORCE WHAT THE STATISTICS ARE SHOWING YOU. THE TESTS YOU EVALUATE DEPENDS ON HEALTHES OF CONCERN FOR THE MIXTURE AND THERE'S GOING TO BE SOME JUDGMENT HERE. DON'T THINK THERE'S A SITUATION WE WON'T INVOKE HOPEFULLY THAT'S A STRUCTURED ELICITED STRUCTURE FOR A GROUP OR SINGLE TOXICOLOGIST. FROM GIVEN EVALUATING RESPONSE AND ENVIRONMENTAL MIX CHUS AND MANY THAT ARE OUT THERE, I THINK IS SUFFICIENT SIMILARITIES POTENTIALLY USEFUL APPROACH, FOR ESTIMATES RISK ASSOCIATED WITH IN THIS CASE IN MY CASE ENVIRONMENTAL MIXTURES BUT I THINK IN CASE PRESENTED TO THE GROUP HERE, POTENTIALLY SOME OF THE BOTANICALS, USE BIOSTATISTICAL INFORMATION I SHOWED YOU A SHORT OVERVIEW EXAMPLE FOR ASSESSING SIMILARITIES BETWEEN DIFFERENT GROUPS OF MIXTURE, THEY'RE QUITE COMPLEX, I THINK IT'S ESSENTIAL TO EXAMINE CHEMICAL DATA TOT TOXICOLOGICAL TESTS UTILIZED. I DIDN'T SHOW THAT BUT THAT'S GOING TO INCLUDE THE DOSE RANGES TESTED. THE DIFFERENT HOW YOU THINK PEOPLE ARE BEING EXPOSED TO THE MIXTURES AND THE ENVIRONMENT, AGAIN, WHICH IS BIG OF A DEAL WITH BOTANICALS, I HEARD THE DIFFERENCE BETWEEN POTENTIALLY EXPOSURE WITH THE ALLO VERA, WHETHER INHALATION DOSE VERSUS DERMAL DOSE. WANT TO MAKE SURE YOU HAVE A PROCEDURE THAT'S REPEATABLE, USING VISUALIZE, AGAIN IT'S POTENTIALLY MORE POWERFUL THINKING AOP CONSTRUCTS THAT FORCE TO CONFRONT UNCERTAINTIES IN THE RELATIONSHIP BETWEEN TOXICITY TESTS THAT WE CAN RUN AND THE OUTPOINTS -- OUTPUT -- END EFFECTS AND OUTCOMES WE'RE CONCERNED ABOUT. ACKNOWLEDGE A NUMBER OF FOLKS THAT I WORKED WITH OVER THE YEARS ON DIFFERENT TYPE OF WHOLE MIXTURE STUDIES AND SUFFICIENT SIMILARITY, AND SAY THANK YOU VERY MUCH. FOR YOUR TIME AND ATTENTION. [APPLAUSE] >> OUR NEXT SPEAKER IS DR. JAMES HARNLY, HE'S A RESEARCH CHEMIST AT USDA FOOD COMPOSITION AND METHODS DEVELOPMENT LABORATORY WHERE HE'S CURRENTLY DEVELOPING METHODS FOR IDENTIFYING AND AUTHENTICATING BOTANICAL SUPPLEMENTS USING INNOVATIVE METHODS, TODAY HE WILL DESCRIBE CHARACTERIZATIONS OF BOTANICAL MATERIALS USING CHEMO METRIC METHODS. >> THANK YOU FOR THE INVITATION TO BE HERE TODAY. YOU CAN SEE I'M WITH THE U.S. DEPARTMENT OF AGRICULTURE BUT THE DATA CAME FROM CYNTHIA RIDER IN NIEHS AND OTHER RESEARCHERS THERE, THEIR NAMES AREN'T ON HERE, THIS IS AN EARLIER VERSION OF THE TITLE AND IT GOT OMITTED. WHAT I WOULD LIKE TO DO IS TAKE THREE STEPS. ONE A GENERAL DESCRIPTION OF UNTARGETED ANALYSIS, THE OTHER IS GOING TO BE A SECOND IS A PERSPECTIVE ON SIMILARITY AND THIRD IS THE STATISTICS ON THE GINKGO BILOBA FOR CASE STUDY. SO TYPES OF CHARACTERIZATIONS, YOU HEARD A NUMBER OF TERMS WE USE DIFFERENT TERMS FOOD CHEMISTRY AND IN NATURAL PRODUCTS. BUT WHAT COME ACROSS, THESE ARE GOING TO HAVE SIMILAR MATHEMATICAL SOLUTIONS. SO SIMILARITY, AUTHENTICATION. TAXONOMIC EXACTNESS, IDENTIFICATION, ADULTERATION, PHYTOEQUIVALENCE, REGARDLESS WHAT YOU CALL IT IS PERSPECTIVE, THESE ARE ALL THE SAME PROBLEM AND BEST ADDRESSED WITH THE SAME APPROACH. ON TARGET METHODS WITH MULTI-VARIANT ANALYSIS. SO AN UNTARGETED ANALYSIS, WE USE AN ENTIRE CHROMATOGRAM OR SPECTRUM, AND WE CALL IT A FINGERPRINT BECAUSE THERE'S NO ATTEMPT TO IDENTIFY COMPONENTS, WE DON'T CARE WHAT THOSE ARE. BUT EVERY DATA POINT IS A POTENTIAL MARKER ESPECIALLY LOOKING FOR ADULTERATION. IF YOU SAY I LOOK AT THREE YOU LEAVE YOURSELF OPEN TO UNEXPECTED SIGNALS LATER. NONE OF THESE POINTS ALREADY ARBITRARILY DISCARDED SOME ARE NOISE BUT WE'LL USE EVERYTHING TO BEGIN WITH. THE MODELING THE SIMPLE. ONE CLASS, CLASSIFICATION, THE ONLY INFORMATION YOU REALLY HAVE IS ON YOUR AUTHENTIC MATERIALS. SO YOU WILL BUILD YOUR MODEL AROUND THAT AUTHENTIC MATERIAL. ALL THE INFORMATION -- NO OTHER INFORMATION IS REQUIRED. YOU'RE GOING TO COMPARE REFERENCE SAMPLES AND YOU WILL USE THE REFERENCE SAMPLES AN CONFIRM THE TEST SAM P BILL HAS NO UNEXPECTED COMPONENTS AND THAT EXPECTED COMPONENTS -- EXCUSE ME MAKE SURE EXPECTED COMPONENTS ARE THERE AND UNEXPECTED COMPONENTS ARE NOT. PETER SOL AT FDA TOLD ME YEARS AGO THAT AUTHENTICATION IS A SESAME STREET QUESTION. WHICH ONE IS DIFFERENT? OR MORE IN LINE WITH WHAT WE'RE TRYING TO DO. IS THIS ONE, AT THE BOTTOM, THIS ONE THE SAME AS THE OTHERS. IF YOU'RE -- SOME OF YOU IN THE ROOM REMEMBER THIS LITTLE DID DIONUSES ME STREET, IT WAS ENTERTAINING. SO CONCEPTUALLY SIMILARITY IS SIMPLE. NO PROBLEMS BUILD A MODEL WITH FINGERPRINTS FROM THE REFERENCE MATERIALS SET YOUR LIMITS HOW MUCH DEVIATION YOU TOLERATE THEN YOU COME PYRE THE TEST SAMPLE AND SAY IS IT IN THE MODEL OR OUT? VERY SIMPLE. THE HARD PART IS SPECIFYING AND COLLECTING THE REFERENCE SAMPLES FOR THE MODEL. YOU WANT THOSE TO ENCOMPASS AS MUCH NATURAL OCCURRING SOURCE OF VARIANTS AS POSSIBLE. COLLECTING YOUR SAMPLES. TWO, SPECIFYING THE METHOD TO ENCOMPASS AS MANY SPECIFIC BOTANICAL FEATURES OR RESPECTS AS POSSIBLE. HENCE, WE USE COMPLETE SPECTOR, COMPLETE CHROMATOGRAMS, NOTHING IS THROWN OUT. SO WE USE FINGERPRINTS SO THE CARTOON RIGHT HAND SIDE SHOWS YOU THEY COME FROM DIRECT ANALYSIS OF SOLIDS, EXTRACTS, NO SEPARATIONS OR YOU CAN TAKE THE EXTRACT YOU CAN SEPARATE CHROMATOGRAM, ANY SORT OF SEPARATION PROCESS. THE FINGERPRINT YOU'RE LEFT WITH IS A COMPLEX COLLECTION OF DATA. THE ONLY WAY YOU CAN MAKE SENSE OF IT IS THROUGH STATISTICAL OR CHEMOME TREK ANALYSIS TO GAIN KNOWLEDGE. ANY SPECTROSCOPIC METHOD CAN BE USED. ANYTHING THAT GIVES A PATTERN. I HAVE SHOWN THIS ON THE NEXT SLIDE WHETHER CHROMATOGRAM, UV SPECTRA, NMR SPECTRA, MASS SPECTRA. YOU'RE GETTING A A PATTERN AND YOU WILL LIFT THE TEST SAMPLE PATTERN AND SAY DOES IT MATCH OR NOT MATCH MY PATTERN? SO YOU CAN USE ANY TECHNIQUE YOU WANT THAT GIVES A PATTERN. LY TALK STRICTLY CHEMICAL ANALYSIS, CHEMICAL PATTERNS TODAY FROM CHROMATOGRAPHIC OR SPECTRA SCOPIC ANALYSIS. TOUGHEST IS COLLECTING THE REFERENCE P SAMPLES. THEY CAN BE VOUCHERED, DEPENDS YOUR PURPOSE WHAT ARE YOU TRYING TO DO. YOU PREFER AS MANY AS POSSIBLE. YOU WANT TO REPRESENT THE MATERIAL OF INTEREST AND VARIANTS. JOE DID AN EXAMPLE, A GOOD JOB EARLIER DESCRIBING WHERE THE VARIATION CAN RISE, SPECIES, ENVIRONMENTAL CONDITIONS PROCESSING, ANYTHING DIFFERENT, PRODUCE A DIFFERENT CHEMICAL COMPOSITION TO YOUR SAMPLE. SOME CASES THE NAME MAY NOT BE OF COURSE. WHEN I TALK ABOUT GINGKO LATER YOU WILL SEE, BUT YOU HAVE TO BE CAREFUL TO COMPARE APPLES TO APPLES. IF SAMPLES ARE PROCESS DIFFERENTLY EXCIPIENT IS USED, THEY'RE NOT -- IT'S NOT APPROPRIATE TO USE REFERENCE MATERIALS OR SAMPLES NOT PREPARED THE SAME WAY, AS YOUR TEST SAMPLE. I WOULD LIKE A BRIEFION OVERVIEW, BASED ON PREVIOUS WORK WE DID. BOTANICALS, I WILL LOOK AT ECHINACEA PANEX THREE GENUS, WITHIN THOSE, YOU CAN -- I WILL SHOW YOU THE DISTRIBUTION THEN LOOK AT JUST THE ECHINACEA. WE HAVE TWO DIFFERENT SPECIES AND PLANT PARTS AND WITHIN THE PLANT PARTS IN SPECIES YOU HAVE RAW ENCEPHTHROUGHS. SO DO PKA ANALYSIS, -- PCA ANALYSIS LOOKING AT FLOW INJECTION MASS SPEC SO WE TAKE 1500 DATA POINTS, PUT THEM INTO THE PCA PROGRAM, THIS IS WHAT YOU GET. SO YOU CAN SEE VERY NICELY SEPARATE YOUR PANEX AND ECHINACEA. THIS ISN'T SURPRISING WE KNOW THERE'S DIFFERENT PLANTS, CHEMISTRY SHOULD BE DIFFERENT, THIS SAYS IN FACT IT IS DIFFERENT. IF YOU TAKE ECHINACEA, WE HAVE TWO SPECIES, AND WE HAVE TWO PLANT PARTS, AERIAL AND ROOTS. SO EAR IS ECHINACEA AND GUSTAFOLA ROOTS. YOU CAN SEE THE OPEN RED SQUARES. ECHINACEA AERIAL IS THE CLOSED RED INDICTMENT AND THEN THE BLACK CROSSES ARE THE ECHINACEA PUPRORIA ROOT SO THEY CLUSTER DIFFERENTLY. LOOKING AT THE SUPPLEMENTS ASSOCIATED WITH EACH, YOU CAN SEE A GREENISH BLUE, THESE ARE THE SUPPLEMENTS FOR THE SAFOLIA ROOT. THE GREEN ARE TRIANGLES FOR THE PUR,UREA ROOT. AND PURPLISH ICONS ARE FOR PURPUREA AERIAL. SO THIS IS VISUALLY DIFFERENT. STATISTICALLY WHERE DO THEY SIT? ONE LAST POINT, TAKING JUST THE PURPUREA DATA YOU CAN SEE LOOKING AT THE SAMPLES AND THE SUPPLEMENTS YOU WOULD SAY THEY'RE PROBABLY DIFFERENT. AND SURE ENOUGH, THEY ARE. THIS IS WHAT'S CALLED SIMKA, WE BUILD A MODEL AROUND OUR PURPUREA AERIAL, THE SOLID RED ICONS. THEN WE SET A 95% CAN CONFIDENCE LIMIT AND SAY ARE THE OTHERS SIMILAR OR DIFFERENT BASED ON THEIR LOADINGS. YOU CAN SAY BOTH PURPUREA AERIAL SOLID SUPPLEMENT AND A LIQUID SUPPLEMENT ARE STATISTICALLY DIFFERENT. SO THE PCA PLOTS DON'T GIVE YOU A FEEL FOR THE STATISTICS INVOLVED. THESE ARE THE RESULTS FROM THE NESTED POOLED ANALYSIS OF VARY I CAN'T WANTS. NESTED IN THE FIRST SLIDE, THE DISTRIBUTION AND GOOD STATISTICIAN WOULD SHUTTER BECAUSE THEY'RE NOT EVEN NUMBERS IN EACH CATEGORIES. IT IS POOLED BECAUSE WE TAKE VARIANTS OVER THE SPECTRA OR CHROMATOGRAM. THE MOST INTERESTING VALUE TO SUM UP HERE IS WE HAVE 100% FOR GRAND MEAN RESIDUALS. 60% COMES BETWEEN SPECIES. 33% IS WITHIN THE SPECIES. THIS BREAKS DOWN, 79% AND 18% ECHINACEA. IF YOU GO OVER AND LOOK AT THE VARIANTS BETWEEN PLANT PARTS AND SPECIES, THAT'S 9%, THE DIFFERENCE BETWEEN THE PLANT PARTS AND THE SUPPLEMENTS IS ONLY ONE PERCENT. THE TOTAL AT THE BOTTOM 3% ANALYTICAL VARIANTS. EACH PROBABILITY ARE HIGHLY SIGNIFICANT IN OTHER WORDS PROBABILITY MEAN OF THOSE DATA ARE EQUAL IS HIGHLY UNLIKELY. THERE'S SIGNIFICANTLY DIFFERENT. HOW CAN WE SUM THIS UP? SIMILARITY. THE GENUSES ARE DIFFERENCE. SPECIES WITHIN THE GENES ARE DIFFERENT. YES. PLANT PARTS WITHIN THE THOUGH SPECIES ARE DIFFERENCE, YES. THE SUPPLEMENTS ARE PLANT PARTS, STATISTICALLY. GENERAL COMPOSITION, SAME RESULT. SUPPLEMENTS, ARE MOST SIMILAR TO THE PLANT PARTS. IF YOU LOOK AT THE SUPPLEMENT PCA SUPPLEMENTS IDENTIFY MORE CLOSELY WITH THE PLANT PART OR THE SPECIES. SUPPLEMENTS ARE SIMILAR TO GENUS. YES. LOOK AT THE PATTERNS, ON PCA, YOU WOULD SAY THESE ARE PART OF THE GENUS SPECIES THAT YOU EXPECT. EFFICACIOUS ON THIS THIS SYMPOSIUM, EQUALLY SAFE? WE DON'T KNOW. NMR MASS SPEC, HIGH RESOLUTION MASS SPEC, GIVE US NICE DATA. WE CAN TELL YOU THINGS ABOUT THE SAMPLE BUT NOTHING ABOUT SAFETY OR EFFICACY. THAT'S THE JOB FOR THE REST OF YOU. SO THE AUTHENTICITY OF GINKGO BILOBA IS -- THAT'S A COP OUT. SO WE LOOKED AT 26 SUPPLEMENTS TWO NIST REST RMs AND TEN PURE STANDARDS. UNHYDROIZED AND HYDROLYZED, WE USE HPLC, I HEARD CYNTHIA -- TO CHANGE MY SLIDE. UNHYDROLYZED IS EXTRACTED, OBVIOUSLY WE TAKE SOLID MATERIALS, TO THE GET THE LIQUID TO MAKE THE LIQUID, ETHANOL WATER MIXTURE. IF YOU HYDROLYZE IT AND TREAT WITH ACID, YOU BREAK DOWN USING WATER AND ETHER BONDS. HERE ARE THE SAMPLES WE RAN. NOTICE THIS WILL BE IMPORTANT LATER. U AN V ARE THE NEST REFERENCE MATERIALS, ONE AND ONE F ARE FROZEN MATERIAL AND THE NIEHS MATERIAL THEY PURCHASED. FROM SO APPLYING CHEMOMETRICS IS NOT NECESSARILY A STRAIGHT FORWARD JOB. IN A PERFECT WORLD, IF YOU PUT ALL THE DATA IN, FROM THE UNHYDROLYZED SAMPLES YOU HAVE CLOUTERS HYDROLYZED, UNHYDROLYZED. THAT MEANS ALL AGREED FOR THE UNHYDROLYZED AND ALL AGREED FOR THE HYDROLIZE. IN FACT THAT DIDN'T HAPPEN. WHEN YOU PLOT THEM AT ONE TIME ON PCA, YOU GET GARBAGE. IF YOU CONSIDER ONLY THE HYDROLYZED SAMPLES, TO SIMPLIFY, AND I HAD TO GO BACK AND LOOK AT THE CHROMATOGRAMS, IT DIDN'T WORK JUST TO LOOK AT THE PCA. SO I DROPPED ALL THE ONE COMPONENT SAMPLES. YOU HEARD CYNTHIA TALK ABOUT THIS, YOU HAVE BASELINE SHIFTS HOW THE DATA PLOT ON PCA BECAUSE YOU HAVE TO COMPARE APPLES TO APPLES AND THE BASELINE WAS NOT THE SAME. SO DROP THE FIRST 4.3 MINUTES SO THE FLAT PACED PART GOES OUT, THE SOLVENT PEAK GOES OUT WHICH IS HIGHLY VARIABLE. WE DROPPED THE LAST FIVE MINUTES BECAUSE THERE WAS NO DATA OUT THERE. WE CAN REDUCE NUMBER OF DATA POINTS. THEN WE DRIVE ADVERTISE. I DRIVETIZED. THIS GETS RID OF THE HUMP IN THE BASELINE. SO ONCE WE GOT RID OF THE -- DRIVETIZE, I DRIVETIZE, YOU SAW THE RETENTION TIMES WITH WERE NOT WELL ALIGN SOD WITH THE DERIVATIVE YOU HAVE YOU HAVE POSITIVE AND NEGATIVE GOING LOW WHEN THEY T THE SLIDE OFFSET GIVES YOU POSITIVE AND NEGATIVE LOBES OVERLAPPING, AGAIN YOU GET GARBAGE OR PLOTTING ON THE ON SITE SIDE OF THE PCA. RELYING ON RETENTION TIMES. THEN WITH THESE CHANGES PRE-PROCESSING WE GOT CLUSTERS THAT MAKE SENSE. THEN I WENT BACK AND PROCESS THROUGH THE HYDROLYZED SAMPLES USING THE SAME STEPS. THIS IS HOW I DID IT. HOW I'M GOING TO PRESENT THE DATA IS GOING TO BE IN A MORE LOGICAL WAY. SO YOU CAN UNDERSTAND TRANSFORMATIONS. THESE ARE UNHYDROLYZED GINGKO SAMPLES. WE HAVE ROUGHLY FIVE CLUSTERS. TAKE MY WORDS FOR IT. WHEN YOU HAVE THIS ON THIS SCREEN IN A PROGRAM AND YOU ROTATE THE ACT SEIZE, YOU CAN SEE VERY DISTINCTLY FIVE CLUSTERS WITH A STATIC PRESENTATION LIKE THIS, IT'S MORE DIFFICULT AND YOU SAY MAYBE THEY SHOULDN'T BE IN THE CLUSTER. ANYWAY, SO WE GET FIVE GENERAL CLUSTERS. IF WE GO BACK AND LOOK AT THE CHROMATOGRAPHIC FINGERPRINTS WE FINDS THERE'S A BASIS FOR THESE CLUSTERS. YOU CAN SEE FOR CLUSTERS A B C D AND E, THERE ARE DISTINCT DIFFERENCES BETWEEN THE CHROMATOGRAMS. I PUT IN THREE VERTICAL LINES HERE, THIS IS CAMPORAL, THIS IS UNIDENTIFIED PEAK. I WILL SHOW YOU CAMP ROLL STANDARDS IN A MINUTE BUT YOU CAN SEE DIFFERENT DEGREES OR NUMBERS OF COMPONENTS, DIFFERENT POSITIONS OF COMPONENTS, DIFFERENT MAGNITUDES OF COMPONENTS. I SHOULD EXPLAIN THAT THIS DATA IS ALL NORMALIZED. SO I TOOK THE DATA, THE RAW INTENSITIES AFTER PRE-POSSESSING. NORMALIZED THOSE SO EVERY DATA POINT IS SQUARED, YOU SET SOME EQUAL TO ONE AND USE NORMALIZATION FACTOR TO MULTIPLY EVERY DATA POINT. IF THE SIGNALS ARE LOWER YOU GET MORE NOISE, IF THEY'RE MULTIPLYING BY LARGER FACTOR, OR YOU HAVE LARGE SIGNALS, IT'S QUIETER. SO THEY ARE DIFFERENT. LET'S LOOK AT CLUSTER A. SIMPLE -- SAMPLES UV WXYZ, 1 AND 1F THAT ARE SLIGHTLY DIFFERENT BUT CLUSTER CLOSE TOGETHER SO INSETS 1 AND 2 SHOW DIFFERENCES BETWEEN U AND V REFERENCE MATERIAL AND SAMPLES. AND W SAMPLE W IS SLIGHTLY DIFFERENT IN 3 AND 4, WE GO BACK AND LOOK AT THE LABEL AND W CONTAINS GO TO COLON SO IT HAD AN ADDITIONAL COMPONENT WHICH SHOWS UP, IT WASN'T MUCH BECAUSE OBVIOUSLY YOU PLOTTED SIMILAR. SO THESE THREE SAMPLES, YOU HAVE THE TWO -- EXCUSE ME. THE EIGHT SAMPLES YOU HAVE THE UV, UNV FROM THE NIST, WXYZ WERE FROM THE SAME VENDOR, FOUR LABELS FOUR SAMPLES. AND THEN ONE AND ONE F ARE THE REFERENCE MATERIALS OR THE MATERIALS THAT NIEHS WAS USING. I SAID I WOULD SHOW YOU ONE PEAK SAMPLES, CYNTHIA TALKED ABOUT THESE TWO A B C AND F. YOU CAN SEE THEY HAVE ONLY SINGLE PEAK HERE IN THE MIDDLE AND LOOK DOWN, HERE IS ROOTEN AND IT CORRESPONDS NICELY, THESE HAVE A SINGLE COMPONENT, THEY PUT ROUTEN INTO THE MATERIAL AND SELLING AS GINKGO BILOBA. SAMPLE G HAS GINGKO A, GINGKO -- GIN COLIC ACID, IT'S 15 AND 17 SEE AT THE BOTTOM NICELY. JUST THE GIN COLIC ACID AND SAMPLE H HAD NOTHING. SO NORMALIZED THERE'S NOTHING THERE, IT WILL BLOW UP THE NOISE SO THE SQUARE ENDS UP ONE. WHEN YOU HYDROLYZE THE SAMPLES THIS IS WHAT YOU SEE. I LEFT THE ORIGINAL COLORS FROM THE UNHYDROLYZED THERE SO YOU CAN SEE HOW THEY SHIFT AROUND. YOU CAN SEE THAT THE 1 AND 1F SHIFTED FROM WHAT WAS PREVIOUSLY THE FIRST GROUP. AND YOU SEE TWO APPEAR TO COMBINE AND ONE SPLIT OFF HERE. SO IT GAVE YOU A TOTALLY UNEXPECTED OR UNANTICIPATED CHANGE WHEN YOU HYDROLYZE THEM. I WANT TO SHOW YOU CLUSTER A A FIRST, UV XY ANDZ, I DROPPED W, WITH THE GO TO, BUT YOU CAN SEE WHAT THE UNHYDROLYZED SAMPLE LOOKED LIKE. UPON HYDROLYSIS YOU'RE LEFT WITH CAMPEROL AND ISORAMNATIN SO YOU HAVE GONE FROM GLYCOSIDES FORMS OF THE FLAVINOIDS TO THE GLYCONES YOU STRIPPED OFF THE SUGARS. THE HYDROLYZE FROM 1 AND 1F YOU CAN SEE THEY SIMILARLY GO TO CAMPEROL BUT THEY HAVE ALSO GOT A STRONG GLOBAL IDE PEAK NOT PRESENT IN THE OTHER MATERIAL, IT WAS THERE BUT MUCH REDUCED CONCENTRATIONS. THEN DOWN BELOW, THESE ARE STANDARDS WE USE TO IDENTIFY THESE PEAKS. GLOBAL IDE, GINGKO IDE JBE, ISORAMNATIN AND GIN COLIC ACID, 15 AND 17. GOING BACK TO THE PREVIOUS CLUSTER, AND LOOK AT WHAT'S HAPPENING TO SOME OF THESE VALUES, I WILL TAKE SELECT SAMPLES, ONE OUT OF HERE U, J, SO FORTH AND SHOW HOW THEIR PROFILES LOOK. AND YOU CAN SEE AS EXPECTED CURSETEN AND CAMPEROL ARE STRONG, SAMPLE J HAVE NO IDEA WHAT THIS IS. BUT IT'S CERTAINLY NOT SIMILAR TO UK AND THE REST OF THEM. YOU CAN SEE FOR THE MOST PART CURSEITIN AND CAMPEROL ARE THE MAIN COME POINTS, THERE'S ISO RAMNATIN IN THERE AND PEAK OUT HERE. SO WE DON'T KNOW WHAT THAT IS. WE HAVE TO DO METABALOMICS. THIS IS THE ORIGINAL UNHYDROLYZED SAMPLE. YOU CAN SEE WITHOUT HYDROLYSIS, CLUSTERS B, C, D AND E HAVE CURSETIN AND CAMPEROL PRESENT AND MAYBE IN SOME CASES A LITTLE BIT OF ISORAMNATIN. WHY WOULD A NATURAL UNHYDROLYZED SAMPLE HAVE AN AGLYCONEIA? IT SHOULDN'T. IT SAYS AS JOE POINTED OUT, THESE ARE THE EXTRACTS, STANDARDIZED E TRACTS, WE KNOW THE CASE FOR GINKGO BILOBA AND THEY HAVE BEEN MANIPULATED. HIGHLY MANIPULATED AS YOU CAN SEE. THESE COMPONENTS SHOULDN'T BE THERE. AS WE DID FOR ECHINACEA AND PANAX, AND GINGKO. IT'S SIMILAR, NOTHING CHANGED. DO A PCA PLOT YOU WOULD SAY ? I SUPPLEMENTS WOULD BE ASSOCIATED WITH CLUSTERED CLOSELY TO RAW GINKGO BILOBA. WE KNOW THIS, THAT THE SUPPLEMENTS ARE DIFFERENT FROM PREVIOUS STUDIES BY BUT IF YOU PLOT FOUR GENUSES YOU CAN SEE SUPPLEMENTS WITH GINKGO BILOBA. ARE ALL COMPONENTS THE SAME? NO. ARE ALL SAW PREMS EFFICACIOUS AND SAFE? WE DON'T KNOW. ALL SUPPLEMENTS EFFICACIOUS AND SAFE? JOB SECURITY FOR THE REST OF YOU. SUMMARY, AUTHENTICATION, DETECTION OF ADULTERATION IS A SIMPLE CONCEPT, COLLECT AUTHENTIC SAMPLES, BUILD A MODEL SET STATISTICAL LIMIT AND COMPARE THE TEST SAMPLE. SIMILARITY IS A RELATIVE TERM, YOU CAN EXAMINE IN TERMS OF STATISTICS. , GENERAL COMPOSITION, SPECIFIC COMPONENTS OR EFFICACY SAFETY. CROW MATTED GRAPHIC FINGERPRINTING ARE COMP MEN TEAR AND NECESSARY FOR THE INTERPRETATION OF THE DATA. YOU CAN'T PLOT PCA AND SAY THAT'S THE RESULT. YOU HAVE TO GO BACK AND CONFIRM THE DATA WITH THE RAW -- CONFIRM PCA PLOTS WITH THE RAW DATA. AS I HAVE SHOWN, MOST OF THE GINKGO BILOBA SUPPLEMENTS ARE ADULTERATED, IE LACK OF SIMILAR COMPONENTS IN CONCENTRATION RATIOS. THAT ONE MANUFACTURER VENDOR IN THE NIEHS SAMPLES WAS THE ONLY ONE THAT APPEARED NOT TO HAVE ADDED GINGKO LIEDS, CURSEECIN CAMP EROL. SO THEY MEET INDUSTRY SPECK BUT SAMPLES HAVE BEEN SPIKED AS I POINTED OUT. SO I WOULD LIKE TO ACKNOWLEDGE RESEARCH SUPPORT FROM THE RESEARCH SERVICE, INTERAGENCY AGREEMENT WITH THE OFFICE OF DIETARY SUPPLEMENTS, HERE AT NIH. WITH THAT, THANK YOU VERY MUCH. [APPLAUSE] >> OUR NEXT SPEAKER IS DR. KERRI LEVANSELER, DIRECTOR OF NSF INTERNATIONAL CHEMISTRY LABORATORY. SHE'S AN EXPERT IN THE APPLICATION OF VARIOUS INSTRUMENTAL TECHNIQUES FOR ANALYSIS OF A WIDE RANGE OF ANOLYTES INCLUDING DIETARY SUPPLEMENT CONSTITUENTS. AND HER TALK WILL EXPLORE TARGETED ANALYSIS OF HERBS, MARKERS, ACTIVES, AND NATURAL TOXINS AND MORE. KERRI. >> THANK YOU VERY MUCH. AT NSF INTERNATIONAL WE'RE A THIRDS PARTY CERTIFYING ORGANIZATION SO THE STANDARD FOR DIETARY SUPPLEMENT AND USING THAT STANDARD, WE WILL APPROVE DIETARY SUPPLEMENTS, TO CARRY OUR MARK, THAT INVOLVES TARGETED ANALYSIS, CONTAMINANT TESTING, AUDITING, MANUFACTURING FACILITY. SO IT'S THROUGH THAT STANDARD THAT -- AND TESTING WE DO TO SUPPORT THAT, WE BRING THAT EXPERIENCE TO THIS AUDIENCE. WHAT IS IN PLANTS? I KIND OF -- I'M SURPRISED THAT PEOPLE SOMETIMES THINK THERE CAN BE JUST A TEST FOR THE PLANT ITSELF. LIKE -- I THREE TO THINK OF WELL IF YOU HAD A CARROT, WHAT WOULD BE IN THE CARROT? THERE'S A LOT OF STUFF. THE WATER MAKES UP A LOT, CARBOHYDRATES, DIFFERENT FORMS OF FIBER. SORRY ABOUT THAT. AND A SLEW OF OTHER PHYTOCHEMICALS SO WE TALKED HOW WE HAVE IMMENSE DIVERSITY HERE. THEN PEOPLE MENTION HOW IMPORTANT THE PLANTED PART IS. THAT PART OF THE PLANT MAKE AS A HUGE DIFFERENCE, WHETHER WE'RE TALKING ABOUT THE SEED, TENNING TO BE MAYBE MORE FAT LIKE YOU GET YOUR PUMPKIN SEEDS HAVE MORE FAT TO THEM. YOUR ROOT, LEAF, FLOWER, OR AERIAL PART OR -- IT CERTAINLY MAKES A HUGE DIFFERENCE IN THE CHEMISTRY DIFFERENCES. SO WHEN WE COME TO SELECTIVE TESTING WHAT WE CHOOSE AS OUR TEST DEPENDS ON WHAT WE'RE TRYING TO DETERMINE. A LOT IS SPOKEN ABOUT IDENTITY TESTING, MAKING SURE WE HAVE THE LIGHT SPECIES. CONFIRMING THE RIGHT PLANT PART, WE TALKED ABOUT ACTIVE CONSTITUENTS BUT AS DISCUSSED SOMETIMES WE'RE NOT SURE WHAT COMPONENT IN THE PLANTS ARE ASSOCIATED WITH THE EFFICACY OR BIOACTIVITY. THE MARKER COMPOUNDS, SOMETIMES DO ALIGN WITH WHAT'S CONSIDERED ACTIVE BUT SOMETIMES THEY HAVE BEEN COMPOUNDS THAT APPEAR UNIQUE FOR THAT PLANT AND MAYBE JUST EASIER TO MEASURE. WHETHER IT'S RELATED TO HERBAL EFFECTS, WE'RE NOT SURE. NATURAL TOXINS, BECAUSE IT'S NATURAL DOESN'T MEAN IT'S GOOD FOR US. AND THEN ALL THE SLEW OF CONTAMINANT AND ADULTERANTS. SO ALL THESE DIFFERENT THINGS NEED TO BE LOOKED AT USING SELECTIVE TESTING. I HAVE ECONOMICALLY MOTIVATED ADULTRINS ON HERE, UNFORTUNATELY THE DIETARY SUPPLEMENT IS A HUGE TARGET FOR THESE ADULTERATION AND AS JIM HARLEY SPOKE ABOUT, THERE'S A LOT OF ADULTERATION. SO I WILL GIVE MORE ON THE SAME TOPIC BECAUSE SADLY A LOT IS GOING ON THERE. I THREE UP A COUPLE OF STRUCTURES FROM ANALYTICAL POINT OF VIEW. ONE OF THE BASIC TOOLS WE LIKE TO USE IS HPLC WITH A UV DETECTOR. SO SO A LOT OF LABS HAVE THAT, TA WORKS REALLY WELL WHEN YOU HAVE COMPOUNDS LIKE THIS WHERE YOU HAVE ALTERNATING DOUBLE BONDS. THOSE -- THAT STRUCTURE ALLOWS THE CHEMICAL TO ABSORB UV OR VISIBLE LIGHT, IT GIVES A STRONG SIGNAL IN OUR CHROMATOGRAPHY. SOME OF THE OTHER COMPOUNDS LACK THAT ABSORPTION BONDING PATTERN. THESE ARE ONES WE HAVE TO BE LOOKING FOR DETECTION METHOD THAT MAYBE LESS SELECTIVE AND UNIVERSEAL. TO TRY TERMNOID COMPOUNDS, THOSE ALSO LACK STRONG ABOUTSOR BEARS. ABSORBER, THOSE DETECTORS ARE NOT AS SELECTIVE BUT WE NEED A THAT BECAUSE THIS LACKS SENSITIVITY ON A TYPICAL UV DETECTOR. SO WHETHER WE'RE DOING ANALYSIS WE NEED TO KNOW WHETHER WE'RE GOING TO VERIFY CLAIMS FOR US, WE'RE OFTEN LOOK AT THE LABEL, WE WANT TO LOOK AT THE LABEL CLAIM. IS IT CLAIM FOR A SINGLE CHEMICAL OR IS IT FOR A LARGE MULTI-CONSTITUENT CLASS OF CHEMICALS SO WE MAY HAVE TO USE A LESS SELECTIVE METHOD IF WE'RE TRYING TO CONFIRM A CLAIM FOR A LARGE CLASS. THESE ARE ECHINACEA LABELS THAT I PULLED OFF IS THE INTERNET ONE DIDN'T TELL ME ANYTHING. IT SAID AERIAL PARTS THEN GIVES A QUANTITY HERE. THIS IS ONE OF TOES THINGS THAT I'M SURPRISED A LOT OF PEOPLE ASK THE ANALYTICAL CHEMISTRY, WHY CAN'T YOU PROVE THERE'S 400-MILLIGRAM OF ECHINACEA IN THIS PRODUCT OF THIS SURVEY. REMEMBER THAT'S THE FIBER THE MOISTURE, THAT'S A LOT OF EXCIPIENTS AN -- AND OTHER THINGS, IT'S A QUALITATIVE STATEMENT, NOT A QUANTITATIVE STATEMENT. OTHER LABELS HERE IS AN EXAMPLE THEY SAID THE SPECIES AND THE PLANT PART BUT NOW IT'S PART OF A PROPRIETARY BLEND. NOT TELLING YOU ANYTHING ABOUT PROPORTIONS. THIS PANEL THEY GIVE A STANDARDIZED EXTRACT HERE SO THEY TELL YOU HOME POLYSACCHARIDES BUT THEY HAVE THROWN IN ANOTHER ECHINACEA INGREDIENT SO IF YOU WERE TRYING TO DO ANALYSIS OF THESE MARKER COMPOUNDS, THEY'RE GOING TO GET CONTRIBUTED TO BY THIS OTHER INGREDIENT SO FROM A FINISH PRODUCT POINT OF VIEW THIS IS HARD TO QUANTIFY. IT CERTAINLY SHOULDN'T BE BELOW THESE NUMBERS. THAT'S FOR SURE. SO KNOWING WHAT TECHNIQUE WE WANT TO USE DEPENDS ON IF WE DO A QUALITATIVE ANALYSIS, VERSUS A QUANTITATIVE. IF WE DO QUALITATIVE, WE WANT THE TECHNIQUE TO DIFFERENTIATE THE MATERIAL FROM INTERFERENCES AND DIFFERENT ADULTERANTS SO THE KEY IS SELECT TESTIFITY FACTOR. YOU ALSO NEED TO BE CONCERNED ABOUT SENSITIVITY TOO BECAUSE YOU NEED TO KNOW HOW MUCH MATERIAL IS IT GOING TO TAKE TO GIVE YOU A POSITIVE TEST. FOR QUANTITATIVE WE NOT ONLY NEED SELECTIVITY, SENSITIVITY BUT WE ALSO NEED ACCURACY AND REPRODUCIBILITY. SO THERE'S A WHOLE SLEW OF TOOLS WE CAN USE, WE WANT TO FIND THE RIGHT TOOL FOR THE JOB AT HAND. WITHOUT A REFERENCE STANDARD THOUGH WE'RE DEAD IN THE WATER. ESPECIALLY WITH TARGETED ANALYSIS. WE REALLY NEED TO FOCUS ON THESE ISOLATED PEER COMPOUNDS TO GIVE THE QUANTITATIVE ANALYSIS. BUT AS JIM TALKED ABOUT, KNOWING WHAT THE FULL PLANT PROFILE LOOKS LIKE IS ESSENTIAL AS WELL, WHETHER THAT'S THE PLANT MATERIAL OR SOME TYPICAL PROCESS MATERIAL LIKE EXTRACT THAT HAS BEEN AUTHENTICATED. WE NEED A MULTITUDE OF REFERENCE STANDARDS TO MAKE THE MOST OUT OF TARGETED -- SO GINKGO BILOBA IS A GREAT EXAMPLE BECAUSE IT GIVES -- ILLUSTRATES A LOT OF ISSUES THAT NEED TO BE ADDRESS WHEN LOOKING AT ANIMAL PRODUCTS AND HERBAL. THE FLAVINNOIDS, WE TALKED ABOUT OFTEN IT'S STANDARDIZED, MATERIAL IS 24% PIPELINE AND 50% TRITERMNOIDS. BUT THE FLAVINNOIDS THERE'S 20 BEFORE THE HYDROLYSIS, AND AFTER IT WORKS OUT TO BE THE THREE, THE CAMPEROL AND ISORAMPTIN. EARLY ON DOING TARGETED ANALYSIS WE FOCUS ABOUT THE RATIOS OF THOSE THREE FLAVINOIDS AND SAY THEY HAVE TO MATCH THE NATURAL RATIO. BUTLY SHOW YOU HOW THAT IS NOT ENOUGH ANY MORE. THE TRY TERPINOIDS, THERE IS MORE CHALLENGES BECAUSE THEY DON'T HAVE STRONG SENSITIVITY ON HPLC UV SYSTEM, SO WE HAVE TO USE OOH DETECTOR LIKE SCATTERING DETECTOR. THIS DETECTOR, IT CAN BE INFLUENCED BY INTERFERENCES, IT DOESN'T GIVE YOU THAT FINGERPRINT INFORMATION TO SAY YES I KNOW I'M LOOKING AT THE CHEMICAL I THINK I'M LOOKING AT. MASS SPEC IS A GOOD WAY OF DOING IT BUT A LOT OF LABS ARE STILL DON'T HAVE ACCESS SO IT'S NOT JUST THE GO TO TOOL FOR EVERYBODY. SO THE GINGKO NEEDS TO BE THE RIGHT PLANT, RIGHT PLANT PART, LEAVES MORE COMMON, BUT THERE ARE SOME SEEDS IN COMMERCE. AND TALK ABOUT THE PRESENCE OF THE GIN COLIC ACID. THERE'S AN EU LIMIT BUT IT'S NOT CONTROLLED IN THE U.S. MARKETPLACE. WHAT WE SEE IS GOOD PROCESSERS ARE DEALING WITH THIS AND GETTING RID OF IT. BUT LOW QUALITY MATERIAL THAT SHOWS UP, WITH THE GINGKO TOXIN, IF THERE'S CHANCE THE SEEDS GOT MIXED WITH THE LEAF, PEOPLE SHOULD CHECK THE GINGKO TOXIN. SO HERE IS THE BASIC CHROMATOGRAPH OF WHEN YOU TAKE GINKGO BILOBA MATERIALS YOU LOOK FOR THE FLAVINOIDS, HYDROLYZED IT, SO YOU STARTED OFF WITH SIGHINGER BASED MOIETIES, YOU CLICK THE SUGAR BASED HYDROLYSIS AND YOU GET THE PEAKS. THE PROBLEM IS COURSEETIN IS RELATIVELY CHEAP, READILY AVAILABLE, SO PEOPLE WOULD ORIGINALLY THEY WOULD THROW BUNCH OF THIS COURSEETEN IN AND YOU SEE A HIGH PEAK HERE AN LOWER PEAKS SO YOU CAN SAY IT'S ADULTERATED. THE CAMPEROL SYNTHETIC FORM IS ALSO RELATIVELY CHEAP. SO NOW THEY'RE ADDING BOTH OF THESE AT A RATIO THAT MATCHES WHAT WOULD NATURALLY OCCURRING, SO AGAIN, THEY'RE TRYING TO TRICK THIS TEST. IT SHOWS YOU THIS TEST IS REALLY INADEQUATE TO DIFFERENTIATE QUALITY MATERIAL FROM LOW QUALITY MATERIAL. THIS IS THIN LAYER CHROMATOGRAPHY TAKING SAMPLE AND NOT HYDROLYZING, STARTING OUT THE GATE. THE LEANS THE ROUGH LANES REFERENCE ONE, REFERENCE 2 AND THEN REFERENCE 1 IS RAN AGAIN, OR IN THESE AISLES. THESE ARE THE AREAS WHERE THE FLAVINOIDS THE GLYCOSIDES, THE TRITERINOIDS SHOW UP SO THIS IS AFTER HYDROLYSIS SO AS JIM HARNLY WAS TALKING ABOUT, A LOT OF MATERIAL, A LOT OF SAM P BILLS ARE ADULTERATED. ANYWHERE YOU SEE A SAMPLE WITH BREAKS GOING DOWN HERE THEY HAVE TAKEN AND ADDED THE AGLYCONE SO THIS IS NOT A NATURAL MATERIAL COMING OUT OF THE PLANT EXTRACT. HERE IS THE SAME SAMPLE, MOSTLY RUTIN, THIS IS A ROW THAT SHOW ROOTEN AND COURSEETIN. IT WILL CONVERT COURSEETEN. THIS IS SCARY IN TERMS OF SHOWING US THAT A LOT OF PRODUCTS WERE BAD BUT THERE WERE A COUPLE OF GOOD ONES IN THERE BUT MORE BAD THAN GOOD. THIS IS THE GIN COLIC A ACID AND AS WE SAID, THE POOR QUALITY EXTRACTS EXIST OUT THERE WHERE THERE WAS LARGE AMOUNTS OF GIN COLIC ACID WHEREAS HIGH QUALITY YOU DON'T SEE THOSE PEAKS. LOOKING AT ECHINACEA, FOCUSING ON DIFFERENT PLANT PARTS HAVING DIFFERENT CHEMISTRIES, WE ARE TYPICALLY RUNNING A FENOLICS PROGRAM FOR HE CAN NAY SHARKS WE USE THE AC SCALE UV TO LOOK AT SPECIES IDENTIFICATION AS WELL AS QUANTIFICATION FOR TOTAL FENOLICS. YOU CAN SEE THE GREEN LINE, IT HAS CLOROGENIC ACID, THE PURPOREA, THE PURPLE LINE HERE. DOESN'T HAVE THAT COMPOUND BUT IT HAS CAMPERIC ACID SO A WAY TO USE TO DIFFERENTIATE THE DIFFERENT SPECIES. IF WE WERE NOT DOING THE CHROMATOGRAPHY AND WE WERE JUST LOOKING AT THE UV SCAN, COMPOUNDS ARE HARD TO DIFFERENTIATE, THE CHEMO METRIC IS NECESSARY BECAUSE IF YOU LOOK BY EYE THEY HAVE A SIMILAR PATTERN. SO YOU HAVE TO HAVE COMPLIMENTARY METHODS THAT TELL YOU DIFFERENT THINGS. THE BLACK COHOSH IS A CHALLENGING ONE. THE MARKER COMPOUNDS AND HPLC ELSD IS COMMON FOR THE IDENTIFICATION AND DETERMINATION OF THESE TRITERPINE COMPOUNDS. THIS NON-SELECTIVE DETECTOR CAN GET US IN TROUBLE AND IF YOU DO HAVE RETENTION TIME SHIFT, THERE CAN BE MISIDENTIFICATION OF PEAKS ADDED SO YOUR QUANTITATION MIGHT BE SKEWED HIGH. THERE'S ONLY A COUPLE OF REFERENCE STANDARDS TOO, TO DETERMINE MAYBE TEN STAFF IN THIS MATERIAL SO HAVING NOT ENOUGH REFERENCE STANDARDS MAKES IT EVEN HARDER TO MAKE SURE THAT THE PEAKS YOU'RE SEEING MATCH UP. SO A AUTHENTICATION OF BLACK COHOSH IS PARTICULARLY CHALLENGING. COMP MEN TEAR METHODS ARE -- COMPLIMENTARY METHODS ARE ADVISED. THERE'S RECENTLY A PUBLICATION BY THE AMERICAN BOTANICAL COUNCIL WITH AHP AND P P P P P NC AND PR GROUP, THAT GIVES MORE DETAILS ABOUT ALL THE CHALLENGES THAT EXIST WITH VERIFYING THIS MATERIAL. SO IT'S A PRETTY GOOD PAPER. SEW WHAT QUESTIONS DO WE ASK CAN WHEN TEST SOMETHING WE HAVE TO FIGURE OUT THE RISKS. YOU GOT TO KNOW THE PRODUCT, YOU HAVE TO KNOW THE INGREDIENT WHERE IT WAS COLLECTED HOW IT WAS PRODUCED, WHAT PARTS OF THE PLANTS ARE USED. AND REALLY THE KEY AS ANALYTICAL CHEMIST IS KNOWING WHAT TEST METHODS DRIVING THE C OF A BECAUSE THERE'S PEOPLE THERE TRYING TO TRICK THESE TESTS DAY IN AND OUT AND THEY'RE DOING IT BECAUSE THERE'S AN ECONOMIC ADVANTAGE OF THAT BUT YOU NEED TO KNOW THE PAST HISTORY, HAVE THERE BEEN RECALLS TO IDENTIFICATION, ADVERSE EFFECTS FOR TOXICOLOGY CONCERNS. SO ALL IN ALL YOU HAVE TO KNOW WHAT QUESTIONS TO ASK TO MAKE SURE THE BEST SELECTED WILL PROVIDE THE RESULTS FOR CONFIRMING THE QUALITY OF THE HERBAL MATERIAL. SO THANK YOU. [APPLAUSE] >> OUR NEXT SPEAKER DR. STEVE FERGUSON IS A SCIENTIST AND MOLECULAR TOXICOLOGY IN GENOMICS GROUP AT NTP. HE'S AN EXPERT IN IN VITRO LIVER MODELS DRUG METABOLISM AND DRUG DRUG INTERACTION PATHWAY AND HIS TALK WILL DESCRIBE NTP'S EFFORT TO EVALUATE THE BOTANICAL SIMILARITY OF VARIOUS GINKGO BILOBA EXTRACTS IN SANDWICH CULTURES OF PRIMARY HUMAN HEPATOCYTES. >> THANK YOU. FOR THE OPPORTUNITY TO SPEAK TODAY. I'M GOING TO TRY TO HIGHLIGHT SOME OF NTP EFFORTS TO UNDERSTAND THE BIOLOGICAL RESPONSE SIMILARITY. WITH PHYSIOLOGICALLY RELEVANT IN VITRO LIVER MODELS WITH GINKGO BILOBA EXTRACT AND THEIR CONSTITUENTS. AND WE WANT TO USE THESE MODELS BECAUSE THEY'RE PREDICTIVE OF ME MILLIONS OF DRUG DRUG INTERACTIONS AND METABOLISM FEATURES THAT HAPPEN IN HUMANS. SO WE WILL APPLY THE MODELS THAT WAY. I HAVE NOTHING TO DECEMBER CLOSE AN STATEMENTS ARE MY OWN. SO SIGNATURE ADVANCES ARE MADE IN THE USE OF IN VITRO LIVER MODELS TO UNDERSTAND HUMAN RESPONSE TO XENOBIOTICS. IN PARTICULAR THESE ADVANCES IN THE LAST 10 TO 15 YEARS ARE MADE BECAUSE WE REALIZE THE IMPORTANCE OF INCORPORATING ORGANOTYPIC STRUCTURES PRESENT IN VIVO. WHEN WE DO THIS TO CELL COULD CANTURES THEY'RE ABLE TO RECONFIGURE IN A WAY THAT MIMICS MANY OF THE BIOLOGICAL PROCESSES THAT HAPPEN IN VIVO IN THE TISSUE. DATA GENERATED FROM THESE IN PARTICULAR WITH DRUG METABOLIZING ENZYMES I'LL HIGHLIGHT THE ENZYME ENDUCTION PATHWAYS CAN CAN BE QUANTITATIVELY TRANSLATED WITH PHYSIOLOGICALLY BASED PHARMACOKINETICS MODELS TO PREDICT THE BLOOD PLASMA RATIO CHANGES OR THE AREA UNDER THESE CURVES IF YOU HAVE INHIBITORS OR INDUCERS OF CYTOCHROME P 450 THAT MIGHT ALTER CO-ADMINISTERED DRUGS. CURRENTLY US FDA REGULATES THIS AS A DRUG SERIES OF GUIDANCE DOCUMENTS ON IT. AND NEGATIVE RESULT IN THESE STUDIES WOULD ELICIT A LABEL ON A DRUG AND NO FURTHER EVALUATIONS FOR DRUG DRUG INTERACTIONS FOR LIVER INDUCTION WOULD BE NECESSARY. A POSITIVE HOWEVER WOULD REQUIRE PHARMACEUTICAL COMPANY TO DO CLINICAL TRIALS TO ASSESS THAT. SO THESE MODELS HAVE A LITTLE MORE POWER THAN TYPICAL DIFFERENTIATED CANCER CELL MODELS. WE WANTED TO STUDY PRIMARY HEPATOCYTES AND LIVER ENZYME INDUCTION BECAUSE GINKGO BILOBA EXTRACTS WITH WERE FOUND IN HUMANS TO INCREASE THE CLEARANCE OF CO-ADMINISTERED DRUGS FOR THIS PARTICULAR EXAMPLE SHOWS A SIGNIFICANT DIFFERENCE IN THE MIDAZOLAM AREA UNDER THE CURVE SO BECAUSE THESE GINKGO BILOBA EXTRACTS ARE LINKED TO LIVER ENZYME INDUCTIONS WE WANTED TO USE THESE MODELS TO BE ABLE TO UNDERSTAND. HOWEVER SOME LITERATURE IS INCONSISTENT AND MAYBE RELATED TO DOSING AND CHANGES OF CONSTITUENT LEVELS ACROSS LOTS EVALUATED. SO THE GOAL WERE TO EMPLOY THE PHYSIOLOGICALLY RELEVANT MODELS TO EVALUATE SIMILARITY WITH 29 GINKGO BILOBA EXTRACTS. LOOK AT VARIOUS OUTCOMES TO ASSESS SIMILARITY. WHAT IS THE BIOLOGICAL RESPONSE SIMILARITY AND WHAT DOES THAT TELL ABOUT CHEMICAL SIMILAR RARITY? WE EVALUATED 12 RECIPIENTS TO SEE IF WE CAN LINK EFFECTS WITH THE GINKGO BILOBA EXTRACTS. THE THIRD GOAL WAS TO SEE IF ACROSS SENTINEL GENES FOR THE FIVE HEPATIC RECEPTOR PATHWAYS, THREE WHICH ARE REGULATED BY FDA IN A DRUG DRUG INTERACTION GUIDANCE, CAN WE OBSERVE ACTIVATION OF GENE EXPRESSION THAT WOULD SUGGEST THAT WE HAVE ACTIVATED RECEPTOR PATHWAYS AND TELL US SOMETHING MECHANISTICALLY HOW THE GINGKO EXTRACTS AND THEIR CONSTITUENTS MIGHT BE ACTIVATING PATHWAYS. FINALLY, TO ARCHIVE RNA AND SPENT CULTURE MEDIA FOR NEAR TERM TRANSCRIPTOMICS AND METABALOMICS STUDY EMERGING FROM THE NEW PLATFORMS AT NTP. SO STUDY DESIGN WAS SIMPLE WE USE 96 WELL CULTURE OF SANDS WITCH CULTURE OF PRIMARY HUMAN HEPATOCYTES SO A LAYER OF COLLAGEN UNDERNEATH LAYER OF HEPATOCYTES THAT ARE CONFLUENT AND LAYER ON TOP IS MATRIGEL SO CREATES A SANDWICH TYPE STRUCTURE TO GIVE THE CELLS MORE DIMENSIONALITY. CULTURED THIS THIS ENVIRONMENT WILL POLARIZE AND BEGIN TO FORM STRUCTURES SUCH AS BIOCANICOLA, THESE ARE DERIVED FROM A FEMALE PATIENT, 47 YEARS OF AGE, CAUCASIAN. FOR THE END POINTS, WE WANT TO LOOK AT CELL HEALTH SO WE HAVE ACYTE ZOOM TO ALLOWS PHOTO MICROGRAPHS FROM WELL IN THE CELL CULTURE PLATE SO WE CAN DOCUMENT MORPHOLOGY THROUGH THE STUDY. A ATP DEPLETION WAS USED AS THE CYTOTOXICITY CONTROL AND WE MONITORED REACTIVE OXYGEN SPECIESES WITH A NEW ASSAY FOR -- THAT MONITORS HYDROGEN PEROXIDE LEVELS WITH LUMINESCENCE ASSAY. FOR LIVER ENZYME ENDUCTION, WE LOOK AT FIVE TARGET GENES NORMALIZED TO THREE ENDOGENOUS CONTROL GENES AND THE FIVE GENES ARE ZONE HERE, THREE CYTOCHROME P 450 AND THESE ARE PROTOTYPICAL ENZYMES THAT ARE ASSOCIATED WITH HUMAN DRUG DRUG INTERACTIONS. WE ALSO HAVE AN E FLUX TRANSPORTER OF THE EXPORT PUMP GENE ABC 11 AND CHOLESTEROL SYNTHESIS GENE AND THESE ARE REFLECTIVE SENTINEL GENES, THESE KEY HEPATIC RECEPTOR PATHWAYS SO WE SAW AS A SMALL TARGETED FIRST STEP THIS WOULD BE A GOOD APPROACH. WE ALSO ASSESS LIVER ENATIC ACTIVITY, WE DON'T HAVE TIME TO TALK ABOUT THIS TODAY BUT THEY'RE ANALYZED AND WE FROZE SAMPLES FOR TRANSCRIPTOMICS AND METABALOMICS. SO IT WAS BROKE TON TWO STAGES, ONE WAS ARRANGED FINDING WHERE WE KID DUPLICATE CONCENTRATIONS, 11 CONCENTRATIONS, IN ORDER THE FIND OUT SEEING OVERT CYTOTOXICITY BECAUSE WE CAN'T ISOLATE RNA AND DO THE GENE EXPRESSION ANALYSIS. SO ARE FROM THAT WE WENT TO STAGE 2 AND DID A TRIPLICATE DOSING OF EACH COMPOUND OR EACH EXTRACT. AT SEVEN CONCENTRATIONS WHERE FOR THE MOST PART WE HAD ELIMINATED ANY ISSUE OF CYTOTOXICITY. SO THE FIRST ASSAY I WANT TO TALK ABOUT IN TERMS OF RESULTS WAS THE ROSS GLOW ASSAY, WE FOUND ALL 29 LOTS PRODUCED STATISTICALLY SIGNIFICANT VEHICLE CONTROL CHANGES IN ROSS AND MOST OF THESE WERE DOSE RESPONSES OF VARYING DEGREES OF BEAUTY. BUT IN THIS CASE WHAT WE ALSO FOUND IS THE GBEs THEMSELVES CONTAIN SUBSTANTIAL LEVELS OF HYDROGEN PEROXIDE SO THAT IS TELLING US SOMETHING BUT WHEN WE PUT THOSE ON THE CELLS WE SAW THOSE GO DOWN DRAMATICALLY. SO ONE PRESUME CATALASE IS FUNCTIONING IN THE CELLS AS YOU MIGHT EXPECT. DOWN HERE BELOW YOU CAN SEE HOW THE LOT SEPARATED SO I HAVE HIGHLIGHTED GVEAA WHICH WAS THE NTP REFERENCE LOT EVALUATED. YOU CAN SEE DISTANCE WHERE IS TBEU HAD MORE RAW SIGNAL IN TERMS OF AREA UNDER THE CURVE. THE GBEH A HAD THE LEAST RAW SIGNAL ACROSS THAT. YOU CAN SEE IT FELL NEAR LOWER END OF THAT RANGE. WE OBSERVED CONSTITUENTS GINGKO TOXIN AND COURSEETIN WERE THE TWO INDIVIDUALS EXAMINED BUT PRODUCE AD SIGNIFICANT RAW SIGNAL SO WE'RE STILL FIGURING HOW TO USE THE DATA BUT WE THINK THEY'RE TELLING US SOMETHING ABOUT SIMILARITY ACROSS THE LOTS. CELL TITER GLOW ASSAYS WITH WERE RUN TO EVALUATE CYTOTOXICITY AND ALL 29 LOTS ALSO WITH CONCENTRATIONS EXAMINED PRODUCE DECREASES IN ATP CONTENT. THAT WERE CONCENTRATION RELATED. THESE ARE REPRESENTATIVE CURVES. AND WE DID OBSERVE VARYING POTENCIES. ACROSS THESE, OOZE ONE MIGHT EXPECT. CONSTITUENTS DID NOT REDUCE ARCTP CONCENTRATIONS EXAMINE. SO IT WAS REALLY MOST CONCENTRATION ADJUSTMENTS HAD TO BE DONE WITH THE GINKO LOT EXA TRACT. IF U YOU DO A HIERARCHICAL CLUSTERING ON CYTOTOXICITY ASSAY DATA YOU SEE A GROUPING FALL OUT FROM THAT. SOME MADE SENSE WHEN WE FIRST LOOK AT THINGS, AC CLUSTERING TOGETHER. WE SAW GBW IN A LEAGUE OF ITS OWN. THIS IS THE CURVE FOR GBEW. JIM WAS SUGGESTING THAT WAS A SMALL CHANGE BASINAL CHANGE CHEMICALLY MAY MANIFEST TO LARGE CHAIN IF DRIVING THE RESPONSE. MORE ANALYSIS IF NEEDED BUT EXCITED TO GET RESOLUTION EVEN JUST FOR THE SIMPLE CYTOTOXICITY ASSAY. THIS IS A CONSTELLATION PLOT GIVING A DEPICTION HOW DIFFERENT GBEW WAS AND GROUPINGS OF THE OTHER LOTS. YOU RANK RELATIVE AREAS UNDER THE CURVE CAN, IN THIS CASE RECIPROCAL AC UCGBW WAS BY FAR THE MOST EFFICACIOUS AND POTENT AND THE GBEs FROM NTP FELL IN THE MIDDLE RANGE. AND GENERALLY WE SAW THESE THAT JIM HIGHLIGHTED AS ROOTIT TYPE LOSS, WE'RE FALLENING A GROUPING TOGETHER MULTIPLE ASSAY END POINTS. IN GENERAL WE DID SEE A DISTRIBUTION OF EFFICACIES AND WE USE THE DATA TO PICK THE DOSE LEVEL FOR INDUCTIONS STUDY. THIS IS JUST A GRAPHICAL REPRESENTATION OF THE DIFFERENT LEVELS THAT WE INITIALLY EVALUATED. YOU CAN SEE HOW DRAMATICICALLY WE HAD TO SHIFT GBEW TO AVOID THE CYTOTOXICITY THAT WAS QUITE POTENT. AND SO ALL THESE WERE ADJUSTED TO DO INDUCTIONS STUDY. SO TYPICALLY WHEN WE RUN THESE STUDIES WE ALSO RUN CLINICAL INDUCERS LIKE RA TAMP SIN AND PHENO BARBITOL TO SEE IF IT'S REFLECTIVE OF TYPICAL HEPATOCYTES AND WE SAW TYPICAL RESPONSES ACROSS THE PATHWAYS AND SO WE WERE REASONABLY CONFIDENT GOING FORWARD WITH THE LOOKING A AT THE DATA. SO FOR THE GBEs, QUESTION GENERALLY SAW INCREASED SIP 3, 4 AND SIP 2B 6 INDUCTIONS COMPARABLE TO CLINICAL INDUCERS RAFAMPACIN AND PHENO BARBITOL. WE ALSO SAW SUPPRESSION OF SIP 3A 4 WITH GBE THAT GENERALLY WERE FALLING INTO THAT VERY DISSIMILAR CATEGORY. FOR THE GBE TYPE OR SIMILARS, WE GENERALLY SAW ARCPC 11 AND HMB SUPPRESSION. LIKE THESE EXAMPLES YOU SEE HERE. AND WE OBSERVED THAT WITH ALMOST ALL THE LOTS. A GOOD PROPOSITIONER OF THEM. BUT WE DID SEE MODEST INCREASES WITH A FEW OUTLIARS THAT MAYBE EXPLAINED BY JIM GROUPINGS AND SOME UNIQUE INDIVIDUAL COMPONENTS. GINGKO RIDE WAS A CONSTITUENT THAT ALSO SUPPRESSED ABCB 11. FINALLY SIP 1A 2, THERE'S REPORTS ABOUT THE INDUCIBILITY. IF YOU RUN A STUDY ON GBE TO SAY IS THIS LIKELY TO PRODUCE CLINICALLY RELEVANT INDUCTIONS OF SIP 1A 2? NO. THE EFFICACY HERE RELATIVELY MARGINAL COMPARED TO IN THIS CASE 300 FOLD INDUCIBILITY OF THIS PARTICULAR DO MORE PREP. SO EFFICACY DOESN'T FALL TO A CATEGORY TRANSLATING TO A CLINICAL CHANGE. IT'S THIS GENE HAPPENS TO BE SENSITIVE IN MANY CASES, INTERESTING IT DID NOT INDUCE SIP 1A 2. WHEN WE TRIED TO PUT THE DATA TOGETHER TO LOOK AT EVERY DATA POINT DOSE LEVEL AT ONCE, SCOTT AURBACH ON OUR TEAM WAS NICE ENOUGH TO GENERATE THE HEAT MAP TO ANALYZE THE DATA. THERE WERE GROUPINGS THAT STARTED TO MAKE SENSE. LIKE A A AB AND AC CLUSTERED IN A SIMILAR AREA BUT WE ALSO SAW SOME THINGS THAT REPORT OF -- LIKE WE WERE AGGREGATING THE DATA AND MAYBE NOT SEEING SOME OF THE MORE MINUTE FEATURES. GLENN MADE THE POINT LOOKING AT AN INDIVIDUAL END POINT MIC BETTER THAN AGGREGATING EVERYTHING TOGETHER. SO WE MAY HAVE TO DO A LITTLE BIT OF BOTH OF THOSE DEPEND TONG QUESTION. ONE OF THE WAYS TO PULL OFF THE DATA TOGETHER IS DOING A MULTI-VARIANT ANALYSIS SO USE JUMPTIN TO DO THIS WITH CHEMICAL DISTANCES. JUDGE TALKED ABOUT THAT. IMAGINE HIS NUMBERS TRANSLATED WITH SCOTT AURBACH DOING CORRELATION ANALYSIS TO COME UP WITH CHEMICAL DISTANCES FROM THE NTP LOT. IN GENERAL THE NONE OF THE ENZYMES IN THIS OR NONE OF THE GENE TRANSCRIPTS IN THE ANALYSIS ROSE TO A STRONG CORRELATION WITH ANY CHEMICAL DISTANCES. BUT A NON-PARAMETRIC CORRELATION INCLUDING THE SPEARMAN'S ROW CORRELATION DID SHOW SIGNIFICANT RELATIONSHIPS, IF WE DON'T HAVE ENOUGH LOTS AT THIS POINT COULD BE THAT THE RELATIONSHIPS IN TERMS OF TRYING TO SIMPLIFY THE DATA INTO A DISTANCE MAY NOT EXPLAIN THE FULL COMPLEXITY OF THE DATA AND TYPE ANALYSIS. WE HAD THE LIMITATION FOR THE DATA WITH WE WERE ANALYZING SIX MOST DISTINCT LOTS DID NOT HAVE DISTANCE VALUES SO DIFFICULT FOR DISCRIMINATING LOTS TO NOT INCLUDE THOSE IN THE ANALYSIS. HOWEVER HEALTHCARE WHEN YOU LOOK AT THE DATA QUALITATIVELY, LOOK AT THE CURVES, THIS IS HOW I INITIALLY LOOK AT THE DATA, WE NOTICE LOTS LOOKING LIKE GBE IN TERMS OF SIMILARITY, TENDED TO CALL SIP 3, 4 INDUCTION, NOT UNEXPECTED WHILE THE LEAST SIMILAR LOTS TENNED TO SUPPRESS SIP 3 AND 4. WE SAW THE SAME TRANSFER ABCB 11 SUPPRESSION WHERE THEY TENDED TO INDUCE. SO THERE'S LIKELY DIAGNOSTIC PROPERTIES HERE WE HAVEN'T TEASED OUT AT THIS POINT IN THE ANALYSIS. WE DID HIERARCHICAL CLUSTERING ON TWO METRICS, ONE WAS THE FOLD CHANGE E MAX VALUES, Q1 THAT INCLUDES E MAX WITH A POTENCY FACTOR. YOU CAN SEE SEVERAL LOTS INCLUDED SHOWING GBEW AND V BECAME MORE SIMILAR TO THE NTP LOT WHEN WE INCLUDED THE POTENCY SO THAT'S ANOTHER FACTOR TO CONSIDER WHEN DOING THIS ANALYSIS. SUMMARIZING THE GROUPINGS. IN GENERAL WE FOUND AA AB AND AC GROUPED TOGETHER WITH NYZ AND E. THEN ABCFGH AND M SEEM TO BE CLEARLY DISTINCT FROM THE NTP LOT. WE WANT TO LOOK AT CONSTITUENTS. SO GINGKO LIED A AND B WERE THE MOST EFFICACIOUS SIP 3 AND 4 INDUCERS YOU CAN SEE IN GRAY AND PINK. THEN NTP IS IN THE BLACK AREA. SO LOOKED LIKE THEY WERE SIMILAR, IN FACT WHEN WE NORMALIZE THE CONCENTRATIONS OF OF THE GBEAA 2 IS RELATIVE LEVELS OF GINGKO LIED A YOU CAN SEE IT LINED UP WITH THE POTENCY SO CONSISTENT WITH THE RESPONSE. SO WE SEE C AND J PRODUCEING A LESS POTENT RESPONSE AND BELOW LESSER -- LESS POTENT EFFICACIOUS RESPONSE. COLLECTIVELY WE SAW THIS TYPE OF GROUPING WITH EVERYTHING. WHEN WE NORMALIZE THE DATA WE FOUND WE TIGHTENED UP THE POTENCIES OBSERVED WHEN WE NORMALIZE TO GINGKO LIED A AND B CONCENTRATIONS. BUT IF WE NORMALIZE SUM OF ALL ALL OF IT, IT BROADENED SO SUGGESTIVE THAT MAYBE ALL THE TERPENE LACTONES TOGETHER ARE NOT EXPLAINING THE VARIATION WE HAVE SEEN WITH GLENN CO-LIDE A AND B, DRIVEN BY THE RELATIVE QUANTITIES. LOOKING AT A CORRELATION ANALYSIS WITH SIP 3, 4 GINGKOLIDE A, THE TOP CONSENT TRYINGS THAT WE LOOKED AT, WE DID SEE A RELATIONSHIP, IT'S NOT PRETTY RELATIONSHIP. BUT IT CERTAINLY IS A RELATIONSHIP THAT HAS A MODEST R SQUARED OF .4 BUT THERE SEEMS TOM TRENDS HERE SO WE'LL CONTINUE TO DIG INTO THIS FURTHER BUT WE PROBABLY NEED MORE LOTS TO REALLY DO THIS EFFECTIVELY. WE SAW SIMILAR RELATIONSHIPS WITH ABCB 11 SYNTHASE 2. A AT THE LOCAL ALSO PRODUCED THE SAME CURVE THAT WE SEE WITH GINGKOLIDE A. FINALLY I WANT TO GO INTO THE POWER OF THE INDUCTIONS THAT WE SAW. WHEN WE SEE RESPONSES IN VITRO AND SAY THAT WAS LIKELY TO HAPPEN IN THE CLINIC. IF TOWER TAKE AT THE DOSE TESTED. SO LOOKING AT THE DATA IN TERMS OF PERCENT OF POSITIVE CONTROL, WHICH IS FDA GUIDANCE ON THIS WORK IN 2006. WHAT WE FIND IS QUITE A FEW OF THE LOTS ACTUALLY PRODUCE MORE THAN 40% POSITIVE CONTROL WHICH IS IN 2006 TO 2011 WAS THE GUIDANCE FOR HOW YOU MAKE THIS DECISION TO DECIDE IF YOU RUN A CLINICAL TRIAL. ASSUMING YOU HAD RELEVANT DOSE LEVELS. SO WE CERTAINLY HAVE THAT FOR THE DATA AND GINGKOLIDE ARC AND B ROSE TO THE LEVEL. GINGKOLIDE C ALSO DID AND JAY DID NOT SO WE HAVE THE RANKINGS AND IT CERTAINLY SAYS SOMETHING ABOUT THE LOTS AND SIMILARITY. BUT FURTHER IV IB IS NEEDED AND WE PLAN TO DO IT SOON TO CALCULATE THE PROJECTED AUC CHANGES WITH THE 2012 GUIDANCE RELEASED. IN GENERAL GINGKOLIDES WERE EFFICACIOUS INDUCERS OF SIP 3 AND 4. FINALLY, THE BIG QUESTION THAT CAME TO ME DOING ANALYSIS, ARE THE GINGKOLIDE CONCENTRATIONS WE'RE TESTING IN THE LOT ITSELF RELEVANT TO HUMAN EXPOSURES. I FOUND THIS PAPER, THIS 2010 PAPER THAT DID A SINGLE DOSE OF 120-MILLIGRAMS OF EGB 761. THAT TRANSLATED TO A PEAK BLOOD CONCENTRATIONS OF 41 NANOMOLAR. THE SO WHEN YOU LOOK AT OUR IN VITRO GINGKO LIED A LEVELS IN NTP LOT, WE SAW A 370 NANOMOLAR CONCENTRATION AND DOSE RESPONSE WELL COVERED NANOMOLAR, RIGHT IN THE MIDDLE OF A RANGE. CONCENTRATIONS WE EXAMINE SEEMED TO BE RELEVANT AND IT MAY EXPLAIN WHY IF WE'RE NEAR THE EDGE OF CAUSING SUFFICIENT EFFICACY WHILE WE SEE STUDIES THAT PRODUCE EFFECTS CLINICALLY AND SOME THAT SAY THERE AREN'T, YOU MAY BE NEAR THE BOUNDARY. SO TO SUMMARIZE WE'RE CONTINUING TO EVALUATE CHEMICAL SIMILARITY BUT IN THE DATA ANALYSIS WE HAVE HAD A CHANCE TO DO THUS FAR WE WORRY ABLE TO MAKE CLASSIFICATIONS THAT DO HIGHLIGHT THINGS THAT MADE SENSE BUT ALSO MAYBE OPEN THE QUESTION WHETHER SOME OF THESE SMALL CHANGES SEEING IN THE CHROMATOGRAMS AND CHEMISTRY WERE HAVING BIGGER BIOLOGICAL RESPONSES THAN WE THOUGHT SO THAT MAY END UP USEFUL WAY TO CLASSIFY THINGS GOING FORWARD. DBE MOST SIMILAR TO THE NTP LOT INTENDED CAR PXR AND TARGET GENES EFFICACY AND SUPPRESS ARCBC 11, SYNTHASE 2 WITH A MARGINAL SIP 1A 2 INDUCTION OF WE ALSO NOTICE NON-SIMILAR ARE ONES THAT MAYBE ROOTEN OR SOME OTHER NON-TBE LIKE RESPONSES, THIS CALLED BIOMODAL INDUCTIONS AT LOW HIGHER POTENCY AND THAT WOULD FOLLOWED BY SUPPRESSION OF SIP 2B 6, HALLMARK ACROSS ALL. THE LOT RANKED NEAR MEDIAN OR MAIN EFFICACY FOR MOST OF THE SEVEN END POINTS, SO IT LOOKS TO BE AN AVERAGE LOT IF YOU LOOK ACROSS THE LOTS WE WERE ABLE TO PROCURE. IS THE CONSTITUENTS ALL INDUCED 2B 6 AND ARE CONSISTENT BECAUSE OF RELATIVE PROPORTION OF SIP 3A 42B 6 INDUCTIONS TO BEP CAR ACTIVATORS BASED ON DATA WE HAVE SEEN WHEREAS -- HAD THE OPPOSITE TREND A LARGER MORE EFFICACIOUS INDUCTIONS OF SIP 2B 6 THAN THREE OF FOUR SO THESE DATA SUGGEST THAT MAYBE IT'S A CONTACT ACTIVATOR. THEN FINALLY, 19 OF 29 DID PRODUCE RESPONSES AND WE'LL GO FORWARD WITH CALCULATING AUC CHANGES FOR EACH OF THESE LOTS AS WELL AS CONTINUING TO UNRAVEL THE SIMILARITY DIFFERENCES. WITH THAT, I WILL THANK YOU AND AS FOR YOUR ACKNOWLEDGMENT OF THE COLLABORATORS AND PARTICULARLY SHRENA PAUL AND JULIE WHO DID THE LAB WORK, SCOTT AURBACH WHO HELPED WITH THE DATA ANALYSIS, AND CYNTHIA RIDER WHO WAS THE PILOT OF THE PROJECT AND THEN THE CHEMISTRY FOLKS FOR ALL THEIR SUPPORT PARTICULARLY HELPING GUIDE THE CHEMISTRY SIDE OF THIS, AND MIKE DEVITO AND WENDY BOYD, KEY PLAYERS IN THE PROGRESSION OF THE PROJECT. THANK YOU FOR YOUR TIME. [APPLAUSE] >> I WILL TAKE A PREROGATIVE HERE SINCE THE CAFETERIA DOWNSTAIRS IS OPEN UNTIL 3 O'CLOCK, SOME OF YOU MAY HAVE BEEN RUSHED FOR LUNCH SO LET' TAKE OUR AFTERNOON BREAK NOW. PLEASE TRY TO HOLD IT TO 15 MINUTES BECAUSE THROUGH NO FAULT OF OUR OWN, THE SPEAKERS ARE GREAT STAYING ON TIME. WE'RE STILL BEHIND. SO IF YOU CAN ALL RETURN AT APPROXIMATELY 10 UNTIL, THAT WOULD BE GREAT, APPRECIATE IT. GOING TO GO AHEAD AND START AGAIN. GOOD TO SEE SO MANY NICE CONVERSATIONS GOING ON BUT WE'RE GOING TO HAVE TO RESUME OUR TALK. SO OUR NEXT SPEAKER IS DR. STEPHANIE SMITH-ROE, GENERAL TECH TOXICOLOGIST AND DNA EXPERT IN TOXICOLOGY GROUP AT NTP. SHE'S EXPLORING BRASH COHOSH AND OTHER SAMPLES USING IN VITRO SYSTEM. THIS AFTERNOON SHE'LL PRESENT OUR HER FINDINGS ON THE COMPARATIVE GENO TOXIS OF SEVERAL COHOSH SAMPLES. STEPHANIE. >> GOOD AFTERNOON. I WOULD LIKE TO BEGIN BY THANKING CYNTHIA FOR THE OPPORTUNITY TO TALK ABOUT THE GENETICKING TO DOG GROUP COHOSH SAMPLES. IN ADDITION TO GINGKO, BLACK COHOSH EXTRACT IS A BOTANICAL OF CONSIDERABLE INTEREST TO THE NTP. IT WAS NOMINATED TO THE PROGRAM BY THE NCI AND NIEHS, THIS IS THE PLANT AND THE EXTRACT, MADE FROM THE ROOT. AND THERE ARE SEVERAL REASONS TO CONDUCT A TOXICOLOGICAL ASSESSMENT OF BLACK COHOSH EXTRACT. FOREMOST, EAT A POPULAR BOTANICAL PRODUCT SIMILAR TO GINKGO BILOBA, TOP TEN SALES SO WIDESPREAD EXPOSURE. THAT'S A FEW ON THE MARKET, THE EXTRACT SOLD AS GROUP POWDER AND ALSO FOUND AS OTHER BOTANICALS AS WELL. IT'S MARKETED TO RELIEVE SYMPTOMS OF MENOPAUSE AND VIEWED AS ALTERNATIVE TO HORMONE REPLACEMENT THERAPY BUT ALSO MARKETED TO WOMEN OF CHILD BEARING AGE TO REGULATE THE MENSTRUAL CYCLE AND ALLEVIATE SYMPTOMS OF PM AND USED TO INDUCE LABOR. THERE ARE HAVE BEEN A FEW REPORTS OF HEPATOTOXICITY BUT OTHERWISE VERY LITTLE IS KNOWN ABOUT THE SAFETY OF BLACK COHOSH EXTRACT. SO THE FIRST ORDER OF BUSINESS AT NTP IS TEST ARTICLE AND THE NTP TOOK A PRACTICAL APPROACH BY ASKING WHAT IS THE TOP SELLING PRODUCT ON THE MARKET AND WHAT WOULD BE REPRESENTATIVE OF EXPOE SHIRE. THE ANSWER WAS -- EXSURE. -- EXPOSURE. STANDARDIZED BLACK COHOSH EXTRACT PRODUCED IN GERMANY SINCE '50s, THEY COMPARED A CHROMATOGRAPHIC PROFILE OF WITH BLACK COHOSH STANDARD MATERIAL AND THE ANALYSIS ARE FAIRLY SIMILAR. THE NEXT TASK WAS TO FIND A LOT OF BLACK COHOSH FROM SUPPLY THAT SHOW AD PROFILE THAT INDICATED IT WAS SIMILAR TO THE REFERENCE MATERIAL AND ALSO TO REMEFEMEN T PRODUCT ON THE MARKET AND LUCKILY THERE WAS MATERIAL THAT MET THE CRITERIA. THE NTP TESTED THE MATERIAL IN A VARIETY OF SHORT TERM AND 90 DAY ASSAYS. SINCE BLACK COHOSH EXTRACT IS USED TO ALLEVIATE SYMPTOMS OF GYNECOLOGICAL AILMENTS THE NTP TESTED WHETHER IT HAS ESTROGENIC EFFECTS. OUR SAMPLE BLACK COHOSH EXTRACT WHICH IS SIMILAR TO PRODUCT ON THE MARKET WAS NEGATIVE IN THE UTERO TROPIC ASSAY, IT DID NOT INCREASE UTERINE WEIGHT AND DIDN'T ATTENUATE EFFECTS OF CO-ADMINISTRATION WITH ESTRADIOL. ALSO BLACK COHOSH EXTRACT HAD LITTLE ESTRA CYCLE IN RODENTS AND THESE FINDINGS ARE FAIRLY SIMILAR TO WHAT'S BEEN REPORTED IN THE LITERATURE SO OTHERS HAVE REPORTED THAT BLACK COHOSH EXTRACT IS NEGATIVE IN UTERO TROPIC ASSAYS IN RODENTS AND ALSO BLACK COHOSH EXTRACT IS NEGATIVE IN VITRO TESTS DESIGNED TO DETECT ACTIVITY. THIS IS A LITTLE AD ODDS FOR ALLEVIATING SYMPTOM OF GYNECOLOGICAL AILMENTS BUT IT IS -- IT MUST BE DOINGEN SOMETHING CONTAINING SAIL SILLIC ACID AND DOPAMINERGIC COMPOUNDS WHICH TALKS ACT WHY CONSUMERS HAVE PEN FROM TAKING THIS BOTANICAL. A FEW CASES OF HEPATOTOXICITY IN HUMANS BUT THERE WAS LITTLE EVIDENCE OF LIVER TOXICITY IN RODENTS, LIVER WEIGHTS INCREASE BY 15% INMYCIN IN RATS AT THE HIGHER DOSES THAT WERE USED IN THE STUDY. AT LEAST WITH THE RATS THERE ARE A FEW RATS THAT SHOWED A VERY MILD TO MINIMAL TO MILD LIVER NECROSIS AT TOP DOSE. SO SO FAR, ON THE TOXICOLOGICAL EFFECT OF BLACK COHOSH EXTRACT ARE NOT PARTICULARLY DRAMATIC. BUT ALSO NOMINATED TO THE NTP THEY GO TO GENE TOX TESTING AND TURNS OUT, BLACK COHOSH EXTRACT WAS POSITIVE IN THE GENOTOXICITY IN THE MICRONUCLEUS ASSAY. SO IN MICE THERE WERE SIGNIFICANT DOSE DEPENDENT INCREASES IN MICRONUCLEAR RETICK RECITES. AND IN RATS THERE WAS INCREASES IN MICRONUCLEAR RETHE TICK LOW SITES AT TESTIMONY -- RETICULOSITES AT TOP DOSE. BECAUSE OF THE FINDING IT'S GENO TOXIC AND BECAUSE OF WIDESPREAD EXPOSURE, IT'S BEING EVALUATED IN THE TWO YEAR RODENT CAN CANCER BIOASSAY. MICRONUCLEI WERE VALE WAITEDDED TWO INTERIM TIME POINTS BUILT INTO THIS ASSAY AT 90 DAY AND ONE YEAR AND BOTH TIME POINTS SIGNIFICANT DOSE DEPENDENT INCREASES MICROKNEW CLEAR RETICULOSITES AND ERYTHROCYTES WHICH REPLICATED OUR INITIAL FINDING HERE. THERE WAS ANOTHER INTERESTING FINDING IN THE STUDY, MICE AND RATS DEVELOPED A NON-REGENERATIVE MACROCYTIC ANEMIA WHICH WAS MORE PRONOUNCED IN MICE COMPARED TO RATS AS WELL AS ALSO SEEN WITH THE MICRONIGH COLLIE EYE. SO TOGETHER, THESE RESULTS SHOW THAT THE BONE MARROW COMPARTMENT IS THE TARGET FOR BLACK COHOSH INDUCED TOXICITY. WHAT'S PARTICULARLY INTERESTING ABOUT THIS ANEMIA, IS IN HUMANS DEVELOPMENT OF A SIMILAR EMIA IS DUE ALMOST EXCLUSIVELY TO DRUGS THAT INTERFERE WITH THE FOLATE METABOLISM PATHWAY AND TURNS OUT, MICRONUCLEI CAN BE CAUSED BY THE DISRUPTION OF THE THE METABOLISM PATHWAY SO TAKEN TOGETHER THIS IS AN INDICATION OF WHAT A POSSIBLE MECHANISM OF ACTION MIGHT BE FOR THE GENO TOXIC EFFECTS OF BLACK COHOSH EXTRACT. AND IN ORDER TO FOLLOW THAT UP WE NEED IN VITRO APPROACHES. IN VITRO APPROACHES CAN ALLOW US TO ASK OTHER QUESTIONS AS WELL. AND TO FORMALIZE THOSE QUESTIONS AND SET UP A FRAMEWORK FOR THE TALK, WE HAVE ASKED WHETHER THE GENO TOXIC EFFECTS OF NTP ARTICLE CAN BE REPLICATED IN VITRO. WE LOOKED WHETHER GENOTOXICITY IS A CHARACTERISTIC THAT IS COMMON IN COHOSH SAMPLE AND WE HAVE ASKED WHETHER THE GENO TOXICS COMPONENT OR COMPONENTS OF COHOSH COULD BE IDENTIFIED. AS YOU KNOW IN THIS ROOM THAT'S A REALLY TOUGH QUESTION. SO IN ORDER TO THE ADJUST THE -- ADDRESS THE FIRST QUESTION WE EXPOSED HUMAN LYMPHOBLASTOID CELLS WHICH ARE A WORK HORSE CELL LINE IN GENE TOX WORLD TO THE SAMPLE BLACK COHOSH EXTRACT GENO TOXIC IN ANIMALS. AND FOR THIS IN VITRO ASSAY, WE TESTED MULTIPLE CONCENTRATIONS UP TO ASSAY DEFINED CYTOTOXICITY, ABOUT 50%. DOSE RESPONSE AND INDIVIDUAL DOSE POINT ARE CONSIDERED AND RATHER THAN YES OR NO ANSWER FOR SUBSTANCES CONSIDERED TO BE GENO TOXIC. AND THESE DATA MET RHODE ISLAND TIERIA FOR A POSITIVE CALL -- MET OUR CRITERIA. AND SINCE TESTING THESE UNITS ARE IN MICROGRAMS PER ML. BUT THE -- USING THE MOLECULAR WEIGHT OF THE COUPLE OF COMPONENTS THE IN BLACK COHOSH EXTRACT 100 MICROGRAMS IN THIS ASSAY, TURNS OUT WE'RE TESTING ABOUT THE 100 TO 200 MICROMOLAR INCH. SO HAVING PERFORMED THIS PROOF OF PRINCIPLE EXPERIMENT, WE CAN THEN ADDRESS THE SECOND QUESTION, GENOTOXICITY AND CHARACTERISTIC THAT IS COMMON AMONG SAMPLES AN INSTEAD OF TESTING THE COHOSH THE NTP HAD ON HAND, THIS TABLE SHOWS COHOSH EXTRACT IN MICRONUCLEUS ASSAY WITH HUMAN TK 6 CELLS, THESE ARE EXTRACTS, FIRST IS BLACK COHOSH EXTRACT STANDARD REFERENCE MATERIAL AND THE GRAY ROW BELOW IT WAS COHOSH GENO TOXIC IN ANIMALS. THE FIRST COLUMN INDICATES WHETHER THE COHOSH SAMPLE HAS A CHEMICAL SIGNATURE, MORE CONSISTENT WITH BLACK COHOSH CHINESE COHOSH, OR IF IT HAS A CHEMICAL SIGNATURE THAT LOOKS LIKE COHOSHs ARE PRESENT, BOTH OF THOSE DISEASE. I LIKE TO POINT OUT THAT IN SOME CASES, WE HAVE MORE THAN ONE LOT FROM A SUPPLIER. AND ALSO SOMETHING THAT'S INTERESTING IS THAT WHAT WE'VE ACQUIRE THESE AT DIFFERENT TIME POINTS, THESE ARE EARLY 2000 AND THE COHOSH WE HAVE ACQUIRED RECENTLY ALL APPEAR MIXTURE OF COHOSH. SO WHO IS GENO TOXIC? THE SISTER LET OF THE SAMPLE GENO TOXIC IN ANIMALS WAS ALSO GENO TOXICSE AND TWO OTHER SUPPLIES WERE GENO TOXIC IN ASSAYS UP TO THE TESTED DOSE RANGE. AND I WANT TO POINT OUT THAT FOR SOME OF THESE SAMPLES WE DIDN'T QUITE REACH THE LIMIT FOR CYTOTOXICITY. SO THE ONES THAT ARE LISTED AS NO IN THAT COLUMN, NEED TO BE TESTED AT HIGHER CONCENTRATION TO CONFIRM WHETHER THEY ARE OR NOT GENO TOXIC IN OUR ASSAY. ALSO GENE TOX DATA IS USUALLY DOWN TO YES OR NO ANSWER. IN THE ATTEMPT TO APPLY DIFFERENTIATION TO RESULTS SCOTT OAR BATCH AND THE BIOMOLECULAR SCREENING BRANCH DID SOME CALCULATIONS TO DETERMINE BENCHMARKS FOR SOME INFORMATION ON POTENCY. AS YOU CAN SEE THERE ARE SOME DIFFERENCES OF POTENCY BUT NOT PARTICULARLY DRAMATIC. WE'RE NOT ORDER OF MAGNITUDE CONSISTENT POTENCY. ALSO THE BLACK COHOSH EXTRACT TESTED IN ANIMALS, ALONG WITH SISTER LOT WAS THE THOSE ARE THE MOST POTENT SAMPLES WE LOOKED AT. SOMETHING ELSE YOU MAY HAVE NOTICED IS THE BLACK COHOSH EXTRACT STANDARD REFERENCE MATERIAL ITSELF IS NOT GENO TOXIC, AT LEAST NOT IN OUR TESTED DOSE RANGE SO WOULD BE INTERESTING TO PERHAPS USE THIS LOT AND MAYBE OTHERS TO TRY TO DETERMINE WHAT THE GENO TOXIC COMPONENTS OF BLACK COHOSH MIGHT BE. WE MADE A GREAT FOR RAY SEEING WHETHER THE ROOTS OF DIFFERENT SPECIES OF COHOSH HAS GENO TOXIC ACTIVITY. WE TESTED THE POWDER GROUP MATERIAL CHINESE RED AND YELLOW STANDARD REFERENCE MATERIALS AND AMONG THESE POWDERED SAMPLES, THE CHINESE COHOSH MET OUR CRITERIA FOR POSITIVE CALL. AND THESE COHOSH NEED TO BE TESTED AT HIGHER CONCENTRATIONS. WE ALSO LIKE A BLACK COHOSH POWDER GROUP SAMPLE AND TEST AS WELL FOR FOR COMPARISON, IT'S OF INTEREST TO CONSIDER MAKING EXTRACTS FOR THESE POWDERED ROOT MATERIALS TO SEE WHETHER THE PROCESS OF EXTRACTS COULD POSSIBLY INFLUENCE THE GENO TOXICITY OF THE SAMPLE. WITH REGARDS TO THE LAST QUESTION, WE HAVE SEEN SOME DIFFERENCES IN GENOTOXICITY AMONG OUR SAMPLES, I WILL IT SEEMED WORTH IT TO GIVE IT A TRY, TO SEE IF ANY KNOWN CHEMICAL MARKERS IN BLACK COHOSH EXTRACT CORRELATED WITH GENOTOXICITY. AND THUS FAR NEARLY 100 CONSTITUENTS HAVE BEEN REPORTED, IN THE LITERATURE TO BE PART OF BLACK COHOSH EXTRACT, ACCORDING TO INFORMATION GATHERED BY THE NATURAL PRODUCTS DATABASE. THESE 13 CHEMICALS HAVE BEEN USED AS MARKERS BY THE NTP TO HELP DISTINGUISH COHOSH SAMPLE AND TO EVALUATE THEIR STABILITY OVER TIME. IN PARTICULAR I WANT TO POINT OUT THAT THE INFUSION APPEARS TO BE A RELIABLE IDENTIFIER, THE CHINESE COHOSH. AN APPROACH TO MAKE SIMPLE OBSERVATIONS REGARDING CHEMICAL SIGNATURE AND GENOTOXICITY. AT THE NTP CREATED THE -- CHEMICAL MARKERS THAT THE NTP HAS BEEN USING TO DIFFERENTIATE COHOSHES. THE BRIGHTER THE COLOR, THE GREATER THE AREA PERCENT OF THE PEAK FROM THE CHROMATOGRAM FOR EACH CHEMICAL. ON THE LEFT AXIS YOU CAN SEE THAT THERE ARE TWO MAJOR CLADES. AND THE TOP CLADE CORRESPONDS TO COHOSH SAMPLES BLACK COHOSH TYPE SIGNATURE. BOTTOM CORRESPONDS TO COHOSH IDENTIFIED AS MIXTURE OF BLACK AND CHINESE COHOSHES. THIS IS THE COHOSH SAMPLE THAT WAS GENO TOXIC IN ANIMALS. HEARSAY THE SISTER LOT,S THIS THE BLACK COHOSH STANDARD REFERENCE MATERIAL THAT ALSO INCLUDED THREE DIFFERENT LOTS OF REMIFEMIN, ALL THESE DELONG TO THAT SIGNATURE CLADE. WITH REGARDS TO GENOTOXICITY, THE BAR RIGHT HERE ON THE LEFT SHOWS THE YES, NO RESULT FOR THE GENO TOX TEST, WITH A DOSE RANGE, HERE IS THE SAMPLE THAT WAS GENO TOXIC IN VIVO. HERE ARE SAMPLE CONSIDERED TO BE GENO TOXIC IN YOU ARE ASSAY. SO YOU CAN SEE THEY DON'T NECESSARILY CORRELATE WITH ANY ONE PARTICULAR CLADE AND THEY DON'T ALL CLUSTER TOGETHER. WE ALSO HAVE INFORMATION ABOUT POTENCY, THOSE BENCHMARK DOSES WE LOOKED AT THOSE TO SEE IF THERE'S REFINEMENT HERE. BUT AGAIN, THE MOST POTENT SAMPLES COULD BE FOUND IN EITHER CLADE, FOUND ACROSS THESE CHEMICAL SIGNATURES. ALSO NO SINGLE CHEMICAL CORRELATED WITH GENOTOXICITY AND WE LOOKED AT THIS A NUMBER OF WAYS, WE LOOK AT THE AREA PERCENT FOR THE PEAK FROM THE CHROMATOGRAM, WE ALSO LOOK AT LATE PERCENT OF CHEMICALS. E LOOK AT GENE TOX RESULTS IN A COUPLE OF WAYS AND NOTHING REALLY SHOOK OUT OF THE DATA. AND MOST SURPRISING IS NOT EVEN ACID CORRELATED WELL WITH ANY OF THE GENE TOX RESULTS. AND THE NTP HAS DONE STUDY SHOWING THAT ACID INDUCES MICRONUCLEI IRRITANTS. SO TAKEN TOGETHER WITH REGARD TO THIS QUESTION, GENOTOXICITY IN OUR TESTED DOSE RANGE CAVEAT CORRELATE WITH SIGNATURE MARKERS OF CHEMICALS AN THUS FAR WE'RE NOT SEEING ANY STRONG RELATIONSHIP BETWEEN WHAT WE KNOW IS IN BLACK COHOSH SAMPLES GENE TOX DATA, AGAIN WITH ANOTHER CAVEAT THAT THE GENE TOX DATA TENDS NOT TO HAVE A LOT OF DIFFERENTIATION. WE DO HAVE MORE BIOLOGICAL INFORMATION ON THE WAY. STEVE FERGUSON AS IN IS PROCESS OF APPLYING THE SAME APPROACH HE USED FOR GINGKO AILNAL SEIZE TO COHOSH SAMPLES. USING A MODEL PREDICT GIVE OF HUMAN METABOLISM, THESECLETURES OF PRIMARY HUMAN HEPATOCYTES, LOOKING AT QUITE A FEW BIOLOGICAL END POINTS WITH HIS APPROACH. THESE ADDITIONAL END POINTS ADD DIMENSIONALITY TO ANALYSES WITH THE COHOSH SAMPLE. SO IN SUMMARY BONE MARROW IS TARGET OF BLACK COHOSH EXTRACT INDUCED TOXICITY AND IT DEMONSTRATED BIOLOGICAL EFFECT OF PLAQUE COHOSH EXTRACT IS GENOTOXICITY A SAFETY RISK. HOPEFULLY WE CAN COMPARE COHOSH SAMPLES TO PERHAPS ZERO IN ON THE GENO TOXIC COMPONENTS OF COHOSHES AND ALSO INVESTIGATING THE POSSIBILITY THAT A MECHANISM OF ACTION FOR PLAQUE COHOSH EXTRACT INDUCED GENOTOXICITY COULD BE DISRUPTION OF THE METABOLISM PATHWAY. ALSO POINT OUT BLACK COHOSH EXTRACT IS MARKETED TO WOMEN OF CHILD BEARING AGE AND DISRUPTING FULLY MEMBER TAB LIMB COULD BE AN ISSUE WITH NEURAL TUBE DEFECTS. ALSO KNOWN MARKER P COMPOUNDS DO NOT APPEAR TO CORRELATE WELL WITH GENOTOXICITY AND UNTARGETED ANALYSIS MAY PROVIDE WITH GREATER INFORMATION. ADDITIONAL WORK IS UNDERWAY. AND IN ORDER TO CORRELATE CHEMICAL COMPOSITION WITH BIOLOGY. WITH THAT, I WOULD LIKE TO THANK MY COLLEAGUES AT THAT TIME NATIONAL TOXICOLOGY PROGRAM AND NIEHS. I WOULD ALSO LIKE TO ACKNOWLEDGE CONTRACTOR INTEGRATED LABORATORY SYSTEMS FOR THE TESTING OF IN CYTOMICRONUCLEUS TOOK PLACE. THANK YOU VERY MUCH. [APPLAUSE] >> OUR NEXT SPEAKER THIS AFTERNOON IS DR. CHRIS GENNINGS, SHE'S A RESEARCH PROFESSOR OF PREVENTIVE MEDICINE AND RESEARCH PROFESSOR OF POPULATION HEALTH SCIENCE AND POLICY AT THE MOUNT SINAI HOSPITAL. DR. GENNINGS WILL PRESENT A CASE STUDY OF GINKGO BILOBA EXTRACT TO ILLUSTRATE THE DETERMINATION OF SUFFICIENT SIMILARITY OF RELATED BOTANICALS. >> GOOD AFTERNOON, I HAVE TO BEGIN BY SAYING I APPRECIATE BEING HERE, IT'S INTERESTING TO HAVE THIS CASE STUDY. SO NOW WE CAN CALL CYNTHIA RIDER THE QUEEN OF MIXTURES AND GENIUS QUEEN OF MIXTURE FOR ORGANIZING THIS. IT'S FABULOUS TO SEE THIS IN DIFFERENT ANGLES. AS YOU WILL SEE A LOT OF WHAT HAS BEEN TALKED ABOUT ALREADY IS VERY DIFFERENT THAN WHAT I'M GOING TO PRESENT. I'M A BIOSTATISTICIAN AND THAT'S MY DISCLAIMER LIKE MY ROOMMATE IN COLLEGE SHE ALWAYS SAID I'M JUST A MUSIC MAJOR, I CAN'T DO THIS. I'M NOT A MUSIC MAJOR. ANYWAY. WHEN TALKING ABOUT RISK ASSESSMENT MIXTURES AS WHOLE MIXTURES THAT'S THE PREFERRED AROACH RELATIVE TO COMPONENT BASED MIXTURES. TOXICOLOGISTS BACK IN THE DAY IN THE '80s AND '90s WORK WAS FOCUSED IN THAT DIRECTION. BUT I GET MORE INTEREST FOCUSED ON THIS IDEA OF WHOLE MIXTURE APPROACH AND THE INTERESTING THING TO ME, THE COMMUNITY NEEDS TO DECIDE IF THAT'S THE DIRECTION WE GO, WE NEED TO FILL MANY THE DATA GAPS THERE. THERE'S EFFORTS UNDERWAY TO DO THAT. WHOLE MIXTURE STRATEGIES. SO IN ORDER TO TALK ABOUT WHOLE MIXTURES, WE CAN'T OBVIOUSLY EVALUATE POSSIBLE MIXTURES SO WE NEED TO HAVE THIS CONCEPT OF SUFFICIENT SIMILARITY. SO THAT WE CAN HAVE REFERENCE MIXTURES SIMILAR MIXTURES EVALUATED IN SIMILAR WAY IN TERMS OF RISK ASSESSMENT. SO THE CONCEPT THAT I'M GOING TO SHOW YOU IS BASED ON WORK I DID IN RICHMOND AT VIRGINIA COMMONWEALTH UNIVERSITY WORKING WITH Ph.D. STUDENTS THERE WHO WON A TRAINING GRANT FROM NIEHS WORKING WITH ME ON THIS CONCEPT OF SUFFICIENT SIMILARITY AND THE IDEA OF SUFFICIENT SIMILARITY BUT IN TERMS OF DOSE RESPONSE SIVENESS SO NOT CHEMICAL CONSIDERATIONS BUT DOSE RESPONSIVENESS. SO LET ME EXPLAIN THE CONCEPTS IN A TWO DIMENSIONAL SCHEMATIC RECOGNIZING THAT THE TWO DIMENSIONS CAN EASILY GENERALIZE TO HIGHER DIMENSIONS. SO IMAGINE SOME -- YOU LOOK AT ACT SEIZE CHEMICAL ONE AND TWO, CHEMICAL ONE AND TWO WOULD BE COMPONENTS IN A PICTURE, TWO COMPONENTS IN SUCH A HIXTURE. AND THE DOTS, THE LITTLE CIRCLES MIGHT REPRESENT WHAT I'M GOING TO CALL BENCHMARK, LOW DOSES OF THOSE CHEMICALS AND YOU CAN RECOGNIZE THAT IF YOU DRAW -- HERE I HAVE GOT GROUPS WHERE WE SAY MAYBE THERE'S ONE MIXTURE THAT'S RELATED TO LOW LEVEL CHEMICAL TWO AND HIGH LEVEL 1, AND LOW LEVEL CHECK CAM 1 AND HIGH OF CHEMICAL 2. THAT WOULD THEN EACH MIXTURE THERE'S VARIABILITY ABOUT WHAT YOU ACTUALLY HAVE IN THE MEEKTURE SO IN THIS CASE DIFFERENT BLOCKS OF GINGKO, IT'S NOT THE SAME, THIS VARIABILITY IN THAT SO THAT'S THAT CONCEPT THERE. SO TALKING DOSE RESPONSIVENESS OR THE DOSE -- SIMILARITY IN THEMES OF DOSE RESPONSIVENESS WHAT I'M IMAGINING IS THAT ALONG A GIVEN WAY, THIS BOTTOM ONE IF I CAN GET IT STRAIGHT, WOULD BE REPRESENTED BY A FIXED RATIO OF CHEMICAL 1 AND 2 TOGETHER. SO YOU CAN GO OUT ALONG THE RAY, STILL HAVE THE SAME RATIO BUT YOU'RE INCREASING IN TOTAL DOSE. YOU'RE INCREASING THE TOTAL AMOUNT BUT THE TICKETED RATIO IS THE SAME. SIMILAR FOR THE OTHERS SO WHAT WE WANT TO DO IS TO FIGURE OUT, OF THESE RAYS WHICH ONE OF THOSE MIXTURES ARE SIMILAR. SO LET'S SELECT A REFERENCE DOSE AND THINK ABOUT IT AS BEING THE REPRESENTATIVE COME POINT OR MIXTURE THAT WE WANT TO STUDY. BUT WHAT WE WANT IS TO HAVE METRIC TO DESCRIBE DIFFERENCE BETWEEN THAT REFERENCE DOSE AND OTHER DOSES. OR OTHER MIXTURES I SHOULD SAY. SO THE WAY TO THINK ABOUT THAT IS IN THE SENSE OF THE DISTANCE BETWEEN SOME PLACE ON EACH OF THOSE TOTAL DOSE RAYS. AND SO WE WERE KICKING AROUND IDEAS AND WE SAID LET'S GET SOMETHING DOWN IN THE LOW DOSE RANGE. LET'S CHOOSE THE A BENCHMARK DOSE, IT COULD BE ABOUT ANYTHING YOU CHOSE BUT FOR CONCEPTUALLY IT MAKES SENSE TO THINK ABOUT DISTANCE BETWEEN ONE MIXTURE AND ANOTHER BASED ON A LOW WHAT WE MIGHT CONSIDER ENVIRONMENTALLY RELEVANT RANGE OF DOSE RESPONSE CURVE. SO IMAGINE IF YOU HAVE A REFERENCE MIXTURE WHICH IS IN WHAT COLOR DO YOU THINK THAT IS? ORANGE, YELLOW, GOLD, SO THERE COULD BE A DOSE HERE. WE HAVE ANOTHER ONE HERE SO I WILL CALL MIXTURE 1 AND 2 CANDIDATE PICTURES. 1 IS CANDIDATE MIXTURE AND WE MIGHT DESCRIBE IT AND SAY IT'S BENCHMARK DOSE IS HERE. SO WHAT WE'RE DOING IS FINDING THE DISTANCE BETWEEN BENCH MARK DOSE OF REFERENCE MIXTURE TO ANY CANDIDATE MIXTURE SO WE DEFINE THAT DISTANCE EUCLIDIAN DISTANCE, WE MIGHT WEIGHT BASED ON POTENCY SO SOMETHING LIKE THAT BUT SOME MEASURE OF DISTANCE WE WOULD DEFINE. MIXTURE ONE IS CLOSER IN TERMS OF THE BENCHMARK DOSE TO THE REFERENCE MIXTURE COMPARED TO MIXTURE TWO, THAT'S A FURTHER DISTANCE THERE. AND WHAT WE NEED TO DO THEN IS THINK ABOUT HOW FAR IS CLOSE ENOUGH AND HOW FAR IS TOO FAR. BECAUSE WE LOOK TO SHOW SUFFICIENT SIMILARITIES WE WILL SWITCH AROUND AND FIND CONCEPT OF SIMILARITY SO WE'LL USE EQUIVALENCE TESTING USED IN DRUG TRIAL FOR GENERIC DRUGS AND THINGS LIKE THAT. SCHEMATICALLY HERE IF THERE'S A MIXTURE WITH A BENCHMARK DOSE WITHIN THIS RADIUS WHERE WE SAY THAT IS -- THAT CIRCLE IS OUR SIMILARITY REGION WE WOULD SAY THAT'S THEN ENOUGH FOR US TO SAY THAT MIXTURE IS SIMILAR TO OUR REFERENCE MIXTURE. LIKE YOU CAN'T SEPTEMBER A NULL HYPOTHESIS, IF YOU HAVE STATISTICS YOU KNOW WHAT I MEAN, IN THIS CASE IF WE FAIL TO FIND SIMILARITY, WE ARE NOT ABLE TO SAY MIXTURE TWO IS DIFFERENT, WE CAN SAY IT'S NOT SIMILAR. SO WE HAVE TO BE A LITTLE CAREFUL ABOUT THAT. WEIGHT A MINUTE, YOU'RE TALKING ABOUT HAVING DOSE RESPONSE CURVE ON EVERY SINGLE ONE OF THESE MIXTURES. THAT'S NOT GOING TO WORK. THAT'S A BAD THING. THIS IS THE DATA SCENARIO. THIS IS THE SCHEMATIC WHERE WE'RE TRYING TO SET THE STRUCTURE WHAT WE'RE GOING TO DO BUT WE DO IT ALSO IN A DATA POOR CASE SO I GOT AHEAD OF MYSELF BUT THIS IS WHAT I SAID, THIS DISTANCE BETWEEN THE TWO BENCHMARK DOSES. SO BACK TO THE DATA, HOLD THAT THOUGHT. IN ORDER TO DO E CAN QUESTION I HAVE LENS TESTING THEN, WHAT WE WOULD DO IS FIGURE OUT WHAT WE MEAN BY SIMILARITY, I WILL SHOW YOU TWO WAYS OF THAT TODAY. ONE THAT THIS CASE STUDY IS HELPED US DETERMINE WORKING WITH CYNTHIA WE CAME UP WITH A SECOND APPROACH WHAT WE MEAN BY SIMILARITY OR DEFINING THAT. BUT ONCE YOU KNOW WHAT IS SIMILAR ENOUGH, WE WOULD DETERMINE THAT IN TERMS OF THIS LETTER R THE RADIUS OF THAT SPHERE IN HIGHER DIMENSION SO THE TEST FOR SIMILARITY IS EQUIVALENCE TESTING WHERE WE SAY P IF THAT DISTANCE BETWEEN BENCHMARK DOSE OF REFERENCE AND OF CANDIDATE MIXTURE, IF LESS THAN THE RADIUS INSIDE THE CIRCUMSTANCE IS L WE HAVE EVIDENCE OF SIMILARITY. SO WE NEED TO HAVE ENOUGH EXPERIMENTAL DESIGN SO THAT WE HAVE GOOD ABILITY TO DO THAT. AGAIN, SO OUR NULL HYPOTHESIS IS THAT DISTANCE IS TOO FAR OUT AND WE LOOK FOR EVIDENCE IT'S CLOSE ENOUGH SO WE REJECT, SO WE HAVE NINE STATISTICAL RIGOROUS PROPERTIES IN THIS SORT OF FRAMEWORK. THE WAY WE DO THAT IS ESSENTIALLY COME UP WITH A CONFIDENCE LIMIT AROUND THE DISTANCE BETWEEN EACH CANDIDATE MIXTURE AND THE REFERENCE PICTURE AN COMPARE UPPER 95% CONFIDENCE LIMIT, IF LESS THAN R WE SAY THAT'S CLOSE ENOUGH, THAT'S WHY I SAY IT'S GOT NICE STATISTICAL PROPERTY BECAUSE THE DESIGN HAS TO BE SUCH THAT THAT CONFIDENCE LIMIT IS NARROW. YOU CAN'T DO THAT W TWO ANIMALS YOU HAVE A RIGOROUS ENOUGH DESIGN SO THAT THE VARIABILITY IS SUCH THAT THAT IS A SMALL CONFIDENCE INTERVAL. LIKE I SAID BEFORE, WE'RE NOT GOING TO EXPERIMENTALLY EVALUATE ALL MIXTURES, WE SWITCHED TO THE CONCEPT OF SUFFICIENT SIMILARITY. SO WHAT I HAVE DONE IN THIS SCHEMATIC IS IDENTIFY THAT SUPPOSE WE HAVE DOSE RESPONSE INFORMATION ON A REFERENCE MIXTURE, WE HAVE AN ESTIMATE FOR THE BENCHMARK DOSE SAY OF THAT MIXTURE AND WE HAVE SOME CONCEPT OF WHAT WE MEAN BY SIMILAR SO WE HAVE GOT THE RADIUS OF SIMILARITY REGION. BUT I HAVE CHANGED THESE NOW TO DASH LINES MEANING WE MIGHT KNOW WHAT THE MIXTURE IS, WE MIGHT BE ABLE TO CHARACTERIZE THESE DIFFERENT CANDIDATE MIXTURES BUT WE DON'T HAVE DOSE RESPONSE DATA ON THAT. SO ARE WE STUCK OR CAN WE WIGGLE OUR WAY OUT? WE CAN MAKE SIMPLIFYING ASSUMPTIONS SO ANY TIME YOU HAVE A DATA POOR SCENARIO YOU HAVE TO MAKE ASSUMPTIONS. IN THIS CASE THE ASSUMPTIONS ARE AS FOLLOWS, ALLOW A LITTLE BIT OF ALGEBRA HERE, MATRIX ALGEBRA. IF YOU THINK ABOUT A BENCHMARK DOSE IN K DIMENSION, K CHEMICALS IN OUR MIXTURE, AND SUPPOSE THE A HERE IS VECTOR OF PROPORTIONS SO HOW THOSE CHEMICALS WOULD THE PROPORTION OF EACH COMPONENT IN THAT MIXTURE. IN TERMS OF TOTAL DOSE, T SUBR IS THE TOTAL OF THOSE BENCHMARK DOSE SO MULTIPLY THE BENCHMARK DOSE IS 1 HUNKER ON THAT MIXTURE, AND YOU KNOW PROPORTION OF EACH COMPONENT IN THE MIXTURE, YOU CAN FIGURE OUT WHAT THE DOSE IS OF EACH COMPONENT IN THAT BENCHMARK DOSE SO GOING FROM A HIGHER DIMENSIONAL VECTOR HERE TO A SCALE HERE. SO ALONG THIS REFERENCE MIXTURE THERE IS A SINGLE VALUE, IF YOU GIVE ME THAT MIXING RATIO THERE ARE'S A SINGLE VALUE FOR BENCHMARK DOSE, THAT'S WHAT I MEAN BY CAPITAL T. WHAT WE'RE GOING -- THE ASSUMPTION THAT WE HAVE TO MAKE OR DO MAKE, IS IF MIXTURES ARE SIMILAR ENOUGH TO THE REFERENCE MIXTURE, THE TOTAL DOSE FOR BENCHMARK DOSE ISN'T GOINGTOR THAT DIFFERENT. SO IF YOU HAVE A MIXTURE HERE THAT HAS A BENCHMARK DOSE OF 100 THE BENCHMARK DOSE ISN'T TWO, IT'S PROBABLY 105 OR 98 OR 93, SOMETHING LIKE THAT BUT IN THAT RANGE. SO P IF YOU'RE CLOSE ENOUGH, THAT NUMBER IS GOING TO BE CLOSE. SO THAT'S WHAT WE'RE SAYING HERE, THE TOTAL DOSE MIXTURE FOR CANDIDATE MIXTURES WOULD BE SIMILAR. SO IN OUR CASE WE'RE GOING TO SUBSTITUTE WHAT'S UNKNOWN HERE, THE T SUBJ WHAT'S KNOWN FROM THE REFERENCE MIXTURE T SUBR, THAT GETS US DOWN THE ROAD, THAT WE'RE GOING TO USE TO FIGURE OUT WHAT THE BENCHMARK DOSE ESTIMATE IS FOR ANY MIXTURE THAT WE WANT, WE WILL KNOW PROPORTION OF THE COMPONENTS IN THAT MIXTURE WE WILL USE THE REFERENCE OF TOTAL CAN DOSE, THAT T SUBR AND GET THE DATA J. WHY DO WE NEED THAT? BECAUSE WE WANT TO BE ABLE TO WE WANT TO DEFINE DISTANCE. AND IN ORDER TO DO THAT I NEED TO HAVE THE HIGHER DIMENSIONAL VALUE FOR EACH OF THE DIFFERENT MIXTURES. TO DETERMINE THAT MIXTURE, I WILL SHOW YOU MORE OFNA IN A MINUTE. WE CAN ACTUALLY WEIGHT THINGS SO WE HAVE SOME CHEMICALS, SMALL CHANGE MAYBE MORE IMPORTANT THAN SAME AMOUNT OF CHANGE IN OTHER CHEMICALS AND DETERMINING BY POTENCY SO WHEN WE HAVE THAT WE LIKE TO WEIGHT THEM BUT IN THE CASE TODAY WE WEREN'T ABLE TO DO THAT. SO WHAT WE'RE GOING TO HAVE IS DISTANCE BETWEEN EACH OF THE REFERENCE BENCHMARK DOSE, AND ADJUSTED DOTES FOR EACH CANDIDATE MIXTURES. BUT NOW TALK ABOUT WHAT WE MEAN BY THE -- HOW WE DETERMINE R. HOW DO WE DETERMINE WHAT WE MEAN BY SIMILAR ENOUGH. GLENN WAS ON THE TEAM WORKING WITH US, ORIGINALLY WE SAID HOW TO DETERMINE R, MAYBE DO THAT BASED ON DOSE RESPONSE INFORMATION. SO IF YOU IMAGINE HERE, SUPPOSE WE HAD GINGKO MIXTURE HERE AND I PLOTTED IT SAY ON A MIXTURE OF DOSE PLUS ONE LOG SCALE THERE. WE HAVE RELATIVE LIVER WEIGHT ON THE Y AXIS, SO ON ORANGE COLOR SUPPOSE WE SAY THIS IS OUR BENCHMARK REFERENCE LEVEL AND THIS THE BENCHMARK DOSE CORRESPONDING TO THAT AND WE SAY IF THE RESPONSE IS INCREASES NO MORE THAN WHAT'S UP TO THAT GREEN LINE, THAT'S NOT ENOUGH OF A CHANGE IN THE RESPONSE THAT WE WOULD BE WORRIED ABOUT. SUPPOSE THAT'S WHAT WE WERE WILLING TO ASSUME. WE CAN FIGURE OUT WHAT THE ED WHATEVER IS FOR THAT, AND COME OVER AND GET THIS DONE SO THAT DISTANCE, THAT DIFFERENCE BETWEEN THOSE TWO PLACES COULD BE WHAT WE CONSIDER R TO BE. WE TRIED TO DO THAT IN AN EXAMPLE HERE, AND IT ENDED UP NOT WORKING WELL. SO WE BACKED UP AND CAME UP WITH ANOTHER STRATEGY, WHICH I THINK IS INTERESTING. THAT'S THIS. SO THIS IS THE GINGKO DATA THAT WE HAVE, I WILL SHOW YOU MORE ABOUT THAT IN JUST A MINUTE. BUT THERE -- I THINK IT'S LABEL AD DIFFERENT WAY ON SOME OF THESE TALKS BUT THIS IS THE REFERENCE MIXTURE THAT WAS ACTUALLY EXPERIMENTALLY EVALUATED IN THESE FIVE DAY TOXICITY STUDIES, SO THAT'S THE BLACK LINE. THEN THERE ARE TWO OTHER LINES HERE, THE GREEN AND THE GOLD, THE N AND THE T, ALL THREE ARE DOSE RESPONSIVE IN TERMS OF THE CHANGES IN RELATIVE LIVER WEIGHT. IN COMPARISON TO THAT BLUE DOWN HERE, THERE'S NO ACTIVITY AT ALL. SO I MIGHT CONSIDER THESE BEING SAY A NEGATIVE CONTROL IS THESE TWO DOWN HERE AND A POSITIVE CONTROL MEANING THAT IT IS ACTIVE JUST LIKE THE REFERENCE MIXTURE IS, WOULD BE UP HERE. ONE WAY WE CAN THINK ABOUT WHAT WE MEAN BY SIMILAR IS IF WE CAN DEFINE A DISTANCE BASED ON WAY I HAVE DESCRIBED TO YOU FROM EACH MIXTURES OF INTEREST AND WE SAY THE DISTANCE IS NO MORE THAN WHATEVER THE FURTHEST DISTANCE IS FROM THE REFERENCE DOSE TO ANY OF THE POSITIVE CONTROLS SO WE ALREADY KNOW SOMETHING ABOUT THE OTHER MIXTURES SO THAT'S A LITTLE EXTRA INFORMATION THAN ORIGINALLY THOUGHT WE WOULD HAVE WHEN WE STARTED TO DO THIS WORK. SO WE CAN USE THAT BY SAYING AS LONG AS IT'S NO MORE DIFFERENT THAN THAT, THEN THAT'S STILL DOSE RESPONSIVE IN A SIMILAR ENOUGH WAY, THAT WOULD BE CONSIDERED SUFFICIENTLY SIMILAR. BY COMPARISON THESE ARE GUYS THAT WOULD SAY WE THINK THAT'S TOO FAR SO THAT MIGHT BE TOO FAR OF A BOUNDARY FOR OUR REGION. BUT THAT'S GOING TO LEAVE US IN BETWEEN HERE, WHERE IT'S THE NO MAN'S LAND WE DON'T KNOW. SO HERE IS THE INVESTIGATING CASE STUDY. LIKE YOU HAVE ALREADY SAID WE HAVE THE GINGKO BELOB ASH EXTRACT. WE HAVE IN DIFFERENT LOTS, THAT WE'RE EVALUATED AND WE HAVE DIFFERENT -- ACROSS THE INDUSTRY. SO WE HAVE THE REFERENCE MIXTURE AND WE HAVE 22 DIFFERENT SUPPLIERS OF OTHER MIXTURES THAT WE WOULD BE INTERESTED IN, THAT'S WHAT I CALL THE CANDIDATE MIXTURE TO SEE IF THEY'RE SUFFICIENTLY SIMILAR. WE LOOK AT IN VIVO HEPATOTOXICITY IN RATS BASED ON LIVER WEIGHT. SO ONE STEP THAT TOOK A LITTLE BIT OF TIME WITH THIS IDEA OF HOW DO YOU MAKE THIS IN TERMS -- HOW DO YOU MAKE GINGKO COMPONENTS IN TERMS OF PERCENTAGE OF THE MIXTURE. AND THAT -- I DIDN'T DO THIS CALCULATION, CYNTHIA I SENT IT TO CYNTHIA, SHE -- OKAY. SOMEONE FIGURED THAT OUT. I THINK THAT WAS LIKE A OWEMAN'S TASK, THAT WASN'T NECESSARILY AN EASY THING TO DO. BUT FOR THIS METHOD TO WORK I THINK I DON'T -- I COULDN'T FIGURE HOW TO DO WITHOUT THE PERCENTAGE, I WAS THINKING MAYBE ANOTHER METRIC WE CAN USE BUT IT SEEMS BASED ON THAT PERCENT SO THAT WAS DONE PERCENTAGE OF THE WEIGHT, WE HAD THIS ONE MIXTURE, THE REFERENCE MIXTURE, THE GBE 1, AND THE POSITIVE AND NEGATIVE CONTROL AND WE HAVE THESE TWO DIFFERENT VERSIONS, SO I WILL DO TWO ANALYSES, WHEN PERCENTAGES ARE BASED ON HYDROLYZED GINGKO MIXTURE AND WHEN NON-HYDROLYZED. SO LET ME SHOW YOU WHAT YOU'RE LOOKING AT HERE. THERE WERE OTHER COMPONENTS HERE OF THE MIXTURE BUT THEY WERE VERY SMALL RELATIVE TO THESE IN TERMS OF PERCENTAGE. SO THESE ARE THE ONLY SETS THAT I HAVE PLOTTED HERE. SO THIS IS THE LACTONE AND FLAVANOL GLYCONE SO THE RED LINE CAN IS CONNECTING THE PROPORTION, THIS IS THE PERCENT OF THE MIXTURE MASS FOR HYDROLYZED MIXTURE, THIS IS THE REFERENCE MIXTURE THAT GBE. WHAT YOU SEE THE LITTLE OTHER LITTLE BLUE LINES ARE FOR EACH OF THE OTHER MIXTURES, OTHER 22 MIXTURES SO YOU SEE THERE ARE SOME OF THESE GUYS WAY BACK HERE THAT DON'T LOOK LIKE THAT AND SOME THAT ARE HIGH UP HERE. BUT THAT'S THE KIND OF VARIABILITY, THAT'S A NICE PICTORIAL TO SEE THE VARIABILITY ACROSS THE DIFFERENT 23 MIXTURES. HERE IS THE SAME THING EXCEPT FOR THE NON-HYDROLYZED MIXTURES SO THERE'S SOME WHERE PERCENTAGES ARE VERY DIFFERENT AND OTHERS WHERE THEY CLUMP UP TOGETHER AND THERE'S SOME OUTLIERS. KIND OF INTERESTING THAT WAY. THIS IS FIGURE THAT I'M THREING TO DEMONSTRATE WHAT DO WE MEAN BY NEGATIVE AND POSITIVE CONTROLS? SO AGAIN, THIS IS NOW FOR THE HYDROLYZED MIXTURE BUT THE RED LINE AGAIN IS THE SAME REFERENCE MIXTURE, THE GREENS ARE WHAT I HAVE DETERMINED BEFORE AS POSITIVE CONTROL. SO YOU GET A SENSE OF -- REMEMBER THAT ORIGINAL FIGURE WHERE THE DOSE -- THEY WERE SIMILAR NO TERMS OF DOSE RESPONSE RELATIONSHIP AND HERE YOU'RE SEEING WHAT VARIABILITY THERE IS IN TERMS OF MIXING PROPORTION FOR SUBSET OF MIXTURES. IT'S NOT ALL COMPONENTS BUT IT'S THE ONES HERE THAT WERE IN THE SAME RANGE THAT MADE THE PLOT TOGETHER. SO THE GREEN TEND TO LOOK MORE SIMILAR TO THE RED, BELOW THE LINE WASN'T THAT SIMILAR. ACROSS THOSE, MAYBE THAT'S NOT AS IMPORTANT. BUT INTERESTING THE NEGATIVE CONTROLS BEFORE LOW HERE BUT ONE SHOT UP HERE FOR CAMPFEROL SO THE NEGATIVE CONTROLS WERE DIFFERENT IN TERMS OF THAT. HERE IS THE OTHER FIGURE. IN THIS CASE NEGATIVE CONTROL IS MORE POSITIVE CONTROL FOR THE NON-HYDROLYZED. SO I THINK I WORK ON THESE FIGURES TO TRY TO SHOW YOU SCHEMATICALLY OR GRAPHICALLY WHAT WE'RE TALKING ABOUT IN TERMS OF THE DIFFERENCE MIXTURES BUT AGAIN LIMITATION HERE IS THAT THESE -- IT'S ONLY A SUBSET OF THE MIXTURES. HERE WE GO WITH DOSE RESPONSE STUDY. WE FIT A NON-LINEAR LOGISTIC MODEL TO THE DATA AND SO WHAT YOU SEE HERE IS IN TERMS OF LOG DOSE, YOU CAN SEE IT DOES FIT WELL. SO HERE IS JUST THE PARAMETERS THAT WE GOT FROM THAT AND WE HAVE A BENCHMARK DOSE ESTIMATE HERE AND THAT'S WHAT WE FIND BETWEEN BENCHMARK DOSE FOR IN VIVO STUDY BETWEEN THE 22 CANDIDATES. SO YOU PROBABLY CAN'T SEE THE NUMBERS BUT WHAT WE HAVE HERE IS THEN A RANKING OF THE DIFFERENT MIXTURES, THE DISTANCE THAT WECAL CAN QUEUELATED SO HERE IS THE REFERENCE MIXTURE, THE DISTANCE THAT WE CALCULATED BETWEEN REFERENCE MIXTURE AN EACH OF THE OTHERS AND I THINK IT SHOULD BE RANK ORDERED ACCORDING TOE THAT DISTANCE. AND THEN HERE IS THE UPPER CONFIDENCE LIMIT. P IF WE HAD A VALUE FOR R WE WOULD SIMPLY BE ABLE TO SAY, IF VALUE OF R FOR THREE, ALL THE CONFIDENCE INTERVALS THAT WERE LESS THAN THREE WHICH PUT YOU RIGHT HERE WOULD BE SIMILAR SO THE WORK HAS TO BE DUB TO GIRL HOW CAN WE FIGURE OUT WHAT SIMILAR MEANS SO I SHOWED TWO DIFFERENT WAYS BUT USING POSITIVE ANDEN NEGATIVE CONTROLS, THE GREEN IS FURTHEST THAT -- OF THE TWO POSITIVE CONTROLS WHICH I THINK ARE ONES HIGHLIGHTED HERE, THAT'S THE FURTHEST AWAY, SO YOU CAN SEE AN R VALUE SAY LESS THAN 2.22 OR 2.23, THAT'S HOW THE -- NOW REGION OF SIMILARITY SO ANY MIXTURE YOU GOT THAT WAS THAT CLOSE WE WOULD SAY WOULD BE SUFFICIENTLY SIMILAR SO THAT'S WE'RE REJECTING FOR THAT. WHAT WE SAY IS TOO FAR BUT AGAIN OPPOSITE OF OUR -- THE WAY WE SET THIS UP BUT JUST DEMONSTRATION IT IS THE CASE NEGATIVE CONTROLS DID HAVE THE FURTHEST DISTANCE. AND THEIR WAY DOWN HERE, THE YELLOW GUYS HERE ARE BETWEEN THAT WE WOULDN'T BE ABLE TO SAY ANYTHING ABOUT. BUT SEPTEMBER THAT THEY ARE FURTHER AWAY IN TERMS OF THE DISTANCE METRIC THAN THE POSITIVE CONTROLS THAT WE CHOSE TO LOOK AT. YOU SEE WE GET QUITE A BIT DIFFERENT RESULT WHEN WE HAVE THE NON-HYDROLYZED MIXTURES BUT STILL THE CONCEPT IS THE SAME OF SUFFICIENT SIMILARITY WITH GREEN IN BETWEEN PLACE AND THEN THE RED WOULD BE WHERE THERE WAS NEGATIVE CONTROL, THEY WEREN'T EVEN DOSE RESPONSES. SO AGAIN IF WE HAVE SOME POTENCY ESTIMATES THAT MAYBE WOULD HAVE IMPROVED WHAT WE HAVE DONE HERE, THOSE WERE NOT AVAILABLE, BUT WE SAW 68% OR 15 OF THE 22 WE WOULD CLAIM SUFFICIENTLY SIMILAR. FOR HYDROLYZED, IT'S 59%. SO CONCLUSION, I DO THINK THE STRATEGY IS AN OBJECTIVE APPROACH, THAT'S TRANSPARENT, REPRODUCIBLE AND ALL THAT. WHAT'S THE DISADVANTAGE THIS POINT IS THAT WE STILL ARE WORKING ON WHAT THE WAY TO ANY GUSH OUT WHAT SIMILARITY MEANS. JUST LIKE WHEN THEY STARTED DOING EQUIVALENCE TESTING, SOMEBODY HAD TO DECIDE WHAT WE MEAN BY A DRUG BEING GENERIC BEING E QUESTION I HAVE AND NOT BIOEQUIVALENT TO A MARKETED DRUG. SO THAT CONCEPT THAT THE COMMUNITY CAN START THINKING ABOUT WHAT THAT MEANS, THEN THAT WOULD HELP INFORM. BECAUSE THAT'S IN ORDER FOR THIS TO BE REALLY A RIGOROUS TEST WE HAVE TO HAVE THAT NAILED DOWN AND DECIDED BEFORE WE START LOOKING AT DISTANCE MEASURES AND THINGS LIKE THAT. SO I WANT TO THANK ALL THE PEOPLE THAT HELPED, THERE ARE PEOPLE THAT I DIDN'T WORK SO CLOSELY WITH DURING THIS PROCESS BUT I CONSIDER CYNTHIA CO-AUTHOR ON THIS THOUGH SHE'S NOT ON THIS SLIDE BUT WE HAVE THE ORIGINAL FOLKS WHO WERE WORKING THIS WHOLE CONCEPT OF SUFFICIENT SIMILARITY BACK IN THE DAY. APPRECIATE YOUR TIME AND EFFORT, LISTENING TO ME AND I'LL BE HAPPY TO TAKE QUESTIONS. THANKS. [APPLAUSE] >> WE'LL SAVE THE QUESTIONS TO THE END FOR THE PANEL DISCUSSION. OUR LAST SPEAKER IN THE AFTERNOON SESSION IS DR. SCOTT AURBACH, A TOXICOLOGIST AND GROUP LEADER FOR TOXICOINFOR MA TICKS AT NF AND HAS EXPERTISE IN 'NALSIS AND INTERPRETATION OF MULTI-VARIANT DATA SETS. SCOTT WILL DISCUSS INFERRING TOXICOLOGICAL SIMILARITY WITH MULTI-DIMENSIONAL RELATIONSHIP ANALYSIS. >> I THINK WEAL MAKE UP TIME BECAUSE I DRANK THIS ENTIRE BOTTLE. THAT TITLE WAS GIVEN BEFORE WE HAD DATA. SO WANT TO MAKE IT SOUND GOOD. SO SIMPLE ISSUE. YOU GUYS ALREADY GET THIS BUT WE'LL GO THROUGH IT AGAIN. SO IMAGINE YOU HAVE PERFORM AD COMPREHENSIVE TOXICOLOGICAL TEST ON ONE MECHANICALLY CYNTHIA TALKED ABOUT, GB, IT COSTS YOU ABOUT $4 MILLION IN SIX YEARS. SO LIKELY YOU'RE RUNNING ANOTHER ONE OF THESE STUDIES. SO YOU NEED TO FIGURE OUT HOW THE REGULATORY CAN DO IT SO THE REGULATORY AGENCY WANTS TO USE YOUR DATA TO MAKE A MARKET WIDE DECISION ON THE BOTANICAL YOU TEST SOD FIRST QUESTION OF COURSE TO ASK IS HOW DO YOU KNOW THE LOT IS YOU TESTED IS TOX COLOGICALLY EQUIVALENT ON THE MARKET. WE WILL ASSUME AT THIS POINT IT'S GB THAT'S THE TESTING WE HAVE BEEN WORKING THROUGH SO KEEP THAT QUESTION IN MIND AS WE GO THROUGH THESE DIFFERENT ITERATIONS AND NAIL SEIZE THAT PRESENT TO YOU. REVIEW OF THE NTP GB STUDIES, DOSE RANGE BETWEEN 62.5 AND THOUSAND MILLIGRAMS PER KILOGRAM AND THIS IS MALE RAT RESULTS THERE WERE OTHERS OBVIOUSLY IN THE FEMALE AND MICE. AFFECTS OF LIVER AND THYROID IN THOSE, LIVER BECAUSE THAT'S WHAT THYROID, NOSE AND KIDNEY IN THE LIVER YOU SAW HEPATOCYTE BUT YOU ALSO SAW BIOHYPERPLASIA, ORTHO NOTTIC NEOPLASIA AND NECROSIS. THERE'S NEOPLASTIC EFFECTS IN LIVER. SO YOU HAD 26 LOTS, THIS NUMBER KEEPS MOVING BECAUSE WE HAVE THE COMPREHENSIVE DATA SET 26 IS THE NUMBER WE HAVE TARGETED UNTARGETED ANALYSIS. CHEMICAL FINGERPRINT FIRST SO LOOK AT THIS IMAGE, MANY ARE DOING THE THING THAT WE'RE GOING DO THROUGHOUT THE ENTIRE TALK COMPUTATIONALLY, YOU'RE LOOKING AT PATTERNS, THE SAME WITH CHROMATOGRAM TURNED ON ITS SIDE O SO THE YELLOW, WHEN THE PEAK IS HIGHER AND BLACK ONE IS LOWER. DOWN AT THE BOTTOM THERE, LOT ONE IS WHAT WE CALL THE NTP LOT. FOR POINT OF REFERENCE. SO AS YOU CAN SEE, THIS GROUP HERE ALREADY HAD HIGH DEGREE OF CONSISTENCY. SO EVERYTHING IS FOR OUR PURPOSES OF THIS STUDY, IT'S SCALED AGAINST 1F AND OR 1, PARDON ME, FROZEN FORM OF 1 WE HAD AVAILABLE. WE DID FROM JAMES HARNLY YOU NEED TO GET USED TO THE LINE PLOTS, THERE'S A LOT HERE, JUST TO ORIENT YOU. SO EVERYTHING IS SCALED AGAINST ONE. AND ONE IS SHOWN HERE, ONE IS SHOWN HERE, SO WE HAD DIFFERENT ANALYSES OF ALL THESE DIFFERENT LOTS. THERE'S 26 LOTS SHOWN HERE, YOU COUNT UP ALL THE LETTERS IT'S 26. SO WE HAD A TARGETED HYDROLYZED ANALYSIS TARGETED HYDROLYZED UNTARGETED, UNHYDROLYZED AND THESE ARE JIM WENT THROUGH NUMBER OF THESE AND CYNTHIA HIGHLIGHTED THEM TOO. I ASKED WHICH ONE OF THE PEOPLE TYPICALLY USE FOR ANALYSIS SHE DONE HAVE A GOOD ANSWER SO WHAT WE DID, THESE SCALE AGAINST ONE, PREPARING TO DO RELATIVE SIMILARITY TO ONE, YOU PUT THEM TOGETHER. AND WHAT YOU GET FROM THAT IS AVERAGE CORRELATION IN THIS PLOT, WHAT YOU'RE GOING TO BE ORIENTED TO, THROUGHOUT THIS TALK. SO THESE PLOTS, THIS IS THE RELATIVE SIMILARITY SO MORE SIMILAR OVER HERE, ONE, LESS SIMILAR OVER HERE. ONLY SWITCHES ONCE. TRY TO DO RELATIVE SIMILARITY, OKAY WHAT'S TOX COLOGICALLY SIMILAR. MAYBE DRAW IT HERE. GOING TO BE REALLY CONFIDENT, MAYBE DRAW HERE AND SAY THESE ARE ALL SIMILAR ONES BUT YOU DON'T HAVE A GOOD ANSWER. SO LITTLE BIOLOGY. ANY MORE DATA? BIOLOGICAL FINGERPRINT SO WHAT DO WE CHOOSE TO DO? WE HAVE BEEN EXPLORING THIS IDEA OF DOSE RESPONSE TOXICOGENOMIC STUDY, THIS IDEA WAS PIONEERED AT THE ROCKY THOMAS AND FACT AND FIGURES TAKEN FROM ONE OF THE PAIN EFFORTS AND THE REASON WE WANT TO DO THIS IS TWO FOLD. WHEN YOU DO A TRANSCRIPTOME ANALYSIS YOU LOOK AT 20,000 DATA POINTS. SO YOU GET A NICE FINGERPRINT OF BIOLOGY, YOU CAN USE THAT BASED UPON PATTERNS. WHAT YOU CAN ALSO DO, RUSTY HAS SHOWN THIS, YOU CAN TAKE AND DO SHORT TERM STUDIES AND CALCULATE A PATHWAY LEVEL BENCHMARK DOSE SO A PEAK 3 PATHWAY OR VARIOUS SIGNALING XENOBIOTIC METABOLISM THAT TEND TO MOVE TOGETHER AND WORK TOGETHER. YOU COMPARE OR TWO YEAR STUDY, IT APPEARS THE -- APICAL BMD SO WE CAN USE THE QUANTITATIVELY TOO, WE CAN DO QUICK AND CHEAP AND GET NUMBERS THAT ARE USEFUL. SO WE'RE PURSUING THIS FOR TWO REASONS. WHAT DID WE SELECT? BACK TO OUR LINE DIAGRAM, IT'S NOT THE SAME COLORS BUT THE DISTRIBUTION RELATIVE TO ONE OF ALL THE LOTS WE HAD TO CHOOSE FROM. AND YOU CAN SEE HERE THE YELLOW IS ONE WE PICKED SO OBVIOUSLY ONE THAT'S A REFERENCE AND THEN WE PICK T AND WE PICKED NP AND G. THIS IS THE LINEAR VERSION OF THE PLOTS YOU SAW EARLIER FROM CINDY. SO WHAT WAS OUR STUDY? WE CHOSE WE TYPICALLY DO HARLEM SPRAY, THIS IS NTPS MOVEED THE STRAIN IT TESTED WITH, WE USE THE FISHER THAT IS WHAT THE BIOASSAY WAS DONE IN. WE DO AN ORAL WRAP LAVAGE WIDE DOSE RANGE, FIVE ANIMALS PER DOSE GROUP, FIVE REPEAT DOSES AND 24 HOURS LESS DOSE AN COLLECT LIVER AND WE ALSO COLLECTED CLINICAL OBSERVATIONS BODY AND ORGAN WEIGHT AND CLINICAL PATHOLOGY WE CAN RELATE TO. SO THIS IS SOME OF YOU PROBABLY MUCH MORE COMFORTABLE WITH THAT DATA THAN THE GENE EXPRESSION DATA SO IT'S NICE LENDING TO. ONE FIRST THING WEAL LOOK AT IS SUS -- THE SIZE OF DOTS ARE EQUIVALENT TO THE EFFECT AFFECT SIZE, THAT MEANS THEY'RE SIGNIFICANT IF PINK. SO WHAT YOU SEE IS ONE T AND N ARE ALL -- WEAL A LITTLE BIT SMALLER EFFECTS AND WE SAW MECHANICALLY. E CAN'T FIGURE THIS OUT, DR. BROOK AND I HAVE BEEN GOING BACK AND FORTH WHY THIS OCCURRED IT GOES UP, IT WENT DOWN AND WENT COUNT CONSISTENTLY SO IT'S AN ON VAGUER THAT'S CONSISTENT SO WE USED -- AN OBSERVATION THAT'S CONSISTENT. YOU SEE HERE IS ONE AN T SIGNIFICANT DECREASE AND ALSO DECREASE BUT NOT SIGNIFICANT IN P AND G, NOTHING SO NOTICE HOW IT'S TRACKING, NICELY WITH THE SIMILAR LAYERTY TO CHEMICALLY TO ONE. -- SIMILARITY. SO YOU WANT TO BE CONSERVATIVE DRAW THE LINE THERE, SAY THESE ARE SIMILAR, THOSE ARE NOT. IF YOU WANT TO BE MORE LIBERAL THESE ARE SIMILAR, FOR THOSE INTERESTED IN DOING THIS, WE HAVE TAKEN A BORROWED RUSTY THOMAS'S OFFER, BORROWED THE EPA SOFTWARE AND BUILT A SOFTWARE PACKAGE THAT ALLOWS YOU TO DO MASSIVE SCALE DOSE RESPONSE MODELING ON THESE HIGH DIMENSIONAL DATA SETS, IF YOU'RE INTERESTED IT'S CALLED BMD EXPRESS. IT'S A NICE SOFTWARE TOOL SO IN THE SHORT TERM STUDIES WE MEASURED RATS AND TRANSCRIPT IN THE ORGANS WE TAKE AND FILTER FOR EXPRESSED GENES, IT'S A VOLCANO PLOT FILTERED BY P VALUE. THEN FIT STATISTICAL MODELS, SO INDIVIDUAL GENES AND FROM THERE WE GOT A BENCHMARK DOSE VALUE. AND THERE BAG OF GENES THAT HAS DOSE VALUES AND SORT THEM TO THE PATHWAYS AND YOU SAY THIS PATHWAY IS FULL. LET'S REPORT FROM THAT PATHWAY. TYPICALLY WHAT YOU DO IS TAKE THE MEAN BND SO FIVE GENES IN THAT PATHWAY AND CHECK THE MEAN FOR THOSE FIVE GENES EACH ONE HAS DOSE RESPONSE OR MEDIAN. WE HAVE GONE BACK AND FORTH WHAT TO DO IN THIS CASE I TOOK MEDIAN. SO WHAT YOU HAVE IS -- IF YOU HAVE PATHWAYS RELATED TO HIGH PER TROPHY, THERE'S 30 GENES IN THE HIGH PER TROPHY PATHWAY, THAT HAS VALUES YOU REPORT THE MEDIAN GENE. THAT WOULD BE YOUR HYPERTROPHY BMD. SO QUICK ORIENTATION OF AFFECT SIZE, EFFECT SIZE AND AMPLITUDE OF THE RESPONSE FOR ALL THESE PLOTS WAS SMALL WITH THE EXCEPTION OF ONE. LOOKING AT THE EXPRESSED GENES LOOK AT 200 AND IT DROPS FROM THERE, THESE ARE IN THE RANGE OF 50 DOWN TO BASICALLY ALMOST NOTHING. THEN THE BOY LOGICAL PATHWAY POTENCY RELATED TO THE BMD SO TELL ME WHAT THE MOST SENSITIVE PATHWAY IS THAT'S CONSIDERED ACTIVE AND THE LARGER THE DOT, THE MORE POTENT IT WAS, LOWER THE BMD, SO ONE WAS POTENT FOLLOWED BY TNN AND IT DROPS OFF SIGNIFICANTLY FROM THERE. ONE OTHER THING WE QUALITATIVELY LIKE TO DO, IT'S NICE TO LOOK AT JUST THESE FIVE BUT WHAT IF I CAN PUT THEM IN BIGGER CONTEXT A BIGGER SPACE AND YOU CAN SAY WHILE THEY LOOK DIFFERENT HERE YOU PUT THEM IN BIGGER SAYS SPACE SIMILAR ANYWAY. SO THEY FIND EACH OTHER IN BIGGER SPACE RELATIVE TO THE BIGGER CONTEXT, THE SAME. SO WHAT WE DID, YOU CAN IMAGINE THIS IS THE PCA PRODUCT, THIS WAS EASIER TO UNDERSTAND. AS WE TOOK BASICALLY THE MAXIMUM FULL CHANGE SET OF GENES FROM EACH ONE OF THESE TREATMENTS AND IMMERSED THEM TO THE DATA SET DRUG MATRIX AVAILABLE AT NTP. IN THIS IT WAS 150 COMPOUNDS GENE EXPRESSION LIVER FROM A MAXIMUM TOLLLATED DOSE SO YOU GET -- TOLERATED DOSE. YOU GET A STRING OF INFORMATION, A BUNCH OF GENES HE CAN PRESENTATIONED THAT ENDS OF BEING A PATTERN. WHEN WE IMMERSE THEM IN THERE AND CHOLESTEROL THE CLUSTER THE DATA YOU HOPE THE ONES THAT ARE TOX COLOGICALLY SIMILAR, THEY SHOULD GROUP TOGETHER. AND TURNS OUT, MY LUCK PNG ONES HERE ON THE FAR RIGHT WHICH ARE CHEMICALLY -- OFF ARE ON THEIR OWN AND N 1 AND T ALL TIGHTLY CLUSTERED WHICH SPEAKS TO THE FACT THAT EVEN IN THIS LARGE CONTEXTUALIZED SPACE THESE TRACK WITH EACH OTHER. ONE OTHER NICE THING WE HAVE FROM DRUG MATRIX IS WHEN YOU DO ENOUGH CHEMICALS YOU HAVE PATHOLOGICAL MANIFESTATIONS WITH GENE EXPRESSION. THESE HAD HYPERPLASIA, THESE DID NOT. GIVEN GENE EXPRESSION, I HAVE IDENTIFIER GENES THAT DIFFERENTIATE BETWEEN ANIMAL THAT HAS BILL LAYER HYPERPLASIA AND ONE THAT DOES NOT. WE HAVE A NUMBER OF MODELS, THIS HAPPENS TO BE AN EXAMPLE, YOU HAVE THREE DIMENSIONS HERE, AND THE EXPRESSION OF GENE 1, GENE 2 AND GENE 3, LABELED DIFFERENTLY LIVERS SO YOU HAVE THIS MODEL THAT YOU CAN NOW HAVE GENES THAT INFORM ON MANIFESTATION OF-PHENOTYPE. SO WE WENT THROUGH AND WE SCORED TOOK HE IS SIGNATURES SUBSET OF GENES IN THE DATA SETS AND SCORED WHETHER OR NOT THESE FOUR WERE ACTIVE. SCORE IS COMBINATION OF PERCENT OF GENES IN SIGNATURE THAT WAS ACTIVE COMBINED WITH IMMEDIATE I CAN'T BE BENCHMARK DOSE SO COMPOSITE SCORE IS THERE. SO ONE HITS, THIS IS THE CHEMICAL FINGERPRINT SIMILARITY, YOU SEE T HAS STRONGEST ON HEPATIC ACCUMULATION SO THESE ARE WHAT WE SAW IN THE STUDY. BILL AREA HYPERPLASIA, THE ONE WE SAW IN THE TWO YEAR STUDY YOU CAN SEE THESE ARE HITS, ALBEIT WEAK HITS FOR TNN AND ONE 1 HASSEST EFFECT ON THIS SET OF GENES WITH TNN WITH SMALLER EFFECTS AND PNG NOT SIGNIFICANT. FINALLY HEMATOSITE HYPERTROPHY SCORE, INCREASED LIVER WEIGHT IS THE IDEAL SITUATION. STATE ATTORNEY'S OFFICE A STRONG SCORE FOR 1 AND T OR STRONG HIT, WEAKER FOR N AND NO HITS FOR P NG SO WHAT YOU SEE IS SIGNATURE SCORES ARE TRACKING CHEMICAL FINGERPRINT SIMILAR IS LAYERTY ASSUMING YOU'RE REFERENCING ONE AS YOUR POINT OF REFERENCE. ONE THING THAT RELATES TO, THIS IS ME GOING THROUGH THE DIFFERENT ITERATIONS TRYING TO ILLUSTRATE WHAT THE RELATIONSHIPS ARE. ONE THING WE CAN DO IS TAKE THE STRING OF INFORMATION WE GET, THE GENES THAT ARE -- THE FULL CHANGE VALUE FOR EACH OF THESE LOTS AND SAY OKAY, WHAT'S THE SIMILARITY BASED UPON THOSE VALUES BECAUSED UPON THE CHEMICAL FINGERPRINT. GENE EXPRESSION SIMILARITY IS HERE. AND CHEMICAL FINGERPRINT SIMILARITY IS HERE. THIS TRIQUETRAK WHAT IS YOU SAW -- TRACKS IN THE DENDROGRAM BUT YOU SEE CHEMICAL FINGERPRINT SIMILARITY TRACKS WELL WITH GENE EXPRESSION SIMILARITY, THIS SOMEWHAT -- NOT PERFECT I ONLY HAVE FIVE LOTS. YOU SEE SOME DEGREE OF SIMILARITY. IN THE TRACKING PROCESS. THE NEXT QUESTION, HOW DO YOU INTEGRATE THESE THINGS AND COME UP WITH PREDICTIONS ON WHAT THE LOTS YOU DIDN'T TEST, WHAT WOULD THEY DO? SO THIS IS VERY SIMILAR TO LAST TALK, MUCH LESS SOPHISTICATED. MUCH LESS SOPHISTICATED. WE'RE STILL WORKING HOW TO DO THIS. BASICALLY WE WENT THROUGH AND JUST LOOKED FOR RELATIONSHIPS BETWEEN PHENOTYPES MEASURED WITH THE FIVE LOTS AND WHAT THE LEVELS OF THE UNTARGETED QUANTITIES OF THE CONSTITUENTS WERE. AND WHAT YOU SEE HERE, NOT SURPRISINGLY WE KNOW THIS QUITE WELL NOW, THE GIN GOIZED AND PER TEEN LACTONES HAD STRONGEST CORRELATION WITH THE PHENOTYPES WHETHER LIVER WEIGHT, TRADITIONAL END POINTS, THESE ARE THE SIGNATURES, THIS IS JUST QUANTIFICATION OF COMPOSITE BIOLOGICAL ACTIVITY WE LOOK AT. AND THE MINIMUM PATHWAY BMD SO THE MORE FOR MEAN LACTONE THE LOWER -- TERPENE LACTONE THE MORE BMD WAS. WE RANKED THEM OUT AND WE CREATED AND WENT THROUGH AND CREATED SIMPLE MODELS BASED UPON WHAT COMPONENT WAS MOST STRONGLY CORRELATED WE CREATED MODELS AND MADE PRE-KICKS ON ONES WE DIDN'T HAVE DATA ON. SO WHAT YOU SEE HERE IS LIVER WEIGHT PERCENT INCREASE, CHEMICAL SIMILARITY TO ONE HERE SO REMEMBER THIS IS LIKE A LINE PLOT SO MORE SIMILAR TO ONE IS OVER HERE. AND THEN PREDICTED IN BLUE AND MEASURED IS IN PINK. SO MEASURED ONES SHOWN HERE AND WHAT WE PREDICTED BASED UPON THE REGRESSION THAT WE DID HAVE W CONSTITUENT THAT CORRELATED STRONGLY LIVER WEIGHT IS SHOWN HERE. SO AS YOU CAN SEE WITH THE EXCEPTION OF V P AND G ALMOST ALL PREDICTED TO INCREASE LIVER WEIGHT WITHIN THE DOSE RANGE WE USE. SO WE HAVE TO GO BACK AND VERIFY THAT, I DON'T THINK WE'LL DO 26 ANIMAL STUDIES BUT TRY A COUPLE MAYBE TO SEE IF THAT WORKS. SAME SCENARIO WITH CHOLESTEROL. ALMOST ALL PREDICTED TO INCREASE CHOLESTEROL. V P AND G, NOT SO MUCH. ALPHA PHOSPHATASE, THIS IS THE DECREASE, THIS IS -- THEY'RE ALL TRACKING EACH OTHER PARTLY BECAUSE OF TERPENE LACTONES DRIVING ALMOST ALL THE BIOLOGY. LIPID ACCUMULATION, THIS ONE IS DIFFERENT. YOU SEE -- WHILE YOU SEE A POSITIVE SCORE OVER HERE FOR LIPID ACCUMULATION, YOU ALSO -- YOU'RE WHAT IT'S SAYING MORE CHEMICAL SIMILAR YOU ARE TO THE NTP LOT, THE MORE LIKELY YOU ARE TO HAVE SHOW LIPID ACCUMULATION SCORE. SAME WITH BILL AREA HYPERPLASIA. SO PRETTY MUCH ALL ARE SIMPLE REGREG BASED UPON WHAT THE DATA WE HAVE BILAYER. SO WE'RE PRETTY REASONABLY CONFIDENT THAT MOST OF THESE LOTS WOULD PROBABLY INCREASE LIVER WEIGHT, PROBABLY ACTIVATE THE LIPID ACCUMULATION SIGNATURE. AND WITH THE EXCEPTION OF COURSE OF PNG AND MAYBE -- WE'RE GOING TO BEAT IT TO DEATH BIT SO JUST MOVE PAST THIS. FINALLY WENT THROUGH AND FORMULATE AD MODEL TO PREDICT THE MINIMUM BENCHMARK DOSE, THESE ARE SIMPLE MODELS, IT APPEARS THAT ONE IS THE MOST -- ONE WOULD BE THE MOST POTENT, AND MUCH WEAKER, MUCH WEAKER MUCH WEAKER. IN CONCLUSION THE GINGKO SHOW SIMILAR RANK RELATIONSHIPS TO THE NTP TEST IN CHEMICAL FINGERPRINTS AND BIOLOGICAL SPACE AS FAR AS WE CAN TELL. THE TERPENE LACTONES FOR EXAMPLE GINGKOLIDES ARE BIOLOGICAL AND TOXICOLOGICAL EFFECTS OF GINGKO. ON THE PER TEEN LACTONE CONCENTRATIONS ARE PREDICTIVE OF BIOLOGICAL TO POTENCY. WITH THE EXCEPTION OF LOSS TFG IF OTHER LOTS PREDICTED TO HAVE SIMILAR BIOLOGICAL OR TOXICOLOGICAL EFFECTS IN THE LIVER TO THE NTP TEST LOT. LOTS DO OR WILL CARRY -- VARY BY POTENCY BASED ON CONCENTRATION OF TERPENE LACTONE. AND FINALLY, QUICK ACKNOWLEDGMENTS, AT THE NTP, RAY COLLINS AND -- DID CHEMISTRY WORK RELATED TO THE LOTS AND FORMULATIONS OF THE STUDIES, STEVE FERGUSON HELPED ANALYSIS, CYNTHIA LED THE PROJECT, MOLLY RAN STUDIES AT MATTEL. AND THEN JIM -- JAMES HARNLY WHO PROVIDED NORMALIZED DATA, THAT WE USE TO PARTICIPATE IN THE EARLIER HEAT MAP. THEN FOLKS FROM -- JASON PHILLIPS WHO HELPED FORMULATE SOFTWARE AND ALLOWED BMD ANALYSIS AND MATTEL AND MRI. THANK YOU. [APPLAUSE] >> SO THANKS TO OUR SPEAKERS THIS AFTERNOON, THAT WAS REALLY GREAT. IF I CAN HAVE THE PANEL MEMBERS COME UP, SO WE DEVELOPED THESE CASE STUDIES, THEN INVITED PEOPLE NOT FROM THE NTP OR WHO HAVEN'T BEEN INTIMATELY INVOLVED IN THE CASE STUDIES TO JOIN PANEL AND TALK ABOUT HOW -- WHAT THEY THINK OF THE APPROACH THAT WE HAVE TAKEN. SOCCERRY WHO KNOWS ABOUT TARGETED TESTING AND JIM WHO DID THE UNTARGETED BUT HAS WORKED WITH DIFFERENT EXAMPLES, TAKE A SEAT. A COUPLE OF PANEL iS THAT YOU HAVEN'T HEARD FROM JIM MACGREGOR WHO HAS A LOT OF EXPERIENCE, IN GENE TOX WORK AND LOOKING AT IN VITRO DATA HOW IT RELATES HOW EFFORT IN BOTANICALS BOTANICALS FROM THE CHEMICAL DONE SIMILARITY EXAMPLES WITH OTHERS IN BOTANICALS. SO WE KIND OF WANTED TO GET SOME FEEDBACK ON THE SESSION AND THE QUESTIONS WE PUT UP AT THE BEGINNING ABOUT DATA REQUIREMENTS, PROS AND CONS OF DIFFERENT APPROACHES THAT WERE PRESENTED, BUT SINCE WE DIDN'T HAVE TIME DURING THE SESSION TO ASK QUESTIONS YOU ALL ARE ALSO WELCOME TO ASK QUESTIONS OF BOTH PANELISTS AND OF THE SPEAKERS IF YOU HAVE FOLLOW-UP QUESTIONS OR CLARIFICATION QUESTION. TO EVERYBODY WHO HAS MICS AVAILABLE AT THEIR SEAT, THE POST DOCS NATASHA CATLAND GEORGIA ROBERTS AND KELLY SHEPKOUSKI WILL MONITOR EMAIL FOR QUESTIONS FROM THE WEBCAST. SO WITH THAT, WE CAN OPEN THE SESSIONS. I DON'T KNOW IF YOU WANT TO MAKE INERTIAL FEEDBACK COMMENT -- INITIAL FEEDBACK COMMENT FROM EACH PANELIST IS A GOOD WAY TO START. >> NOT SURE WHAT YOU'RE LOOKING FOR HERE, CYNTHIA BUT WE USE UNTARGETED ANALYSIS. I WAS STRUCK WHEN SCOTT AND GLENN GAVE THEIR DATA, WHETHER SIZE OF MARKER COMPOUNDS YOU CONSIDER SOME OF THE NON-MARKER COMPOUNDS BECAUSE THE QUESTION WAS ASKED JOANNE MCGUIRE, THE MARKER COMPOUNDS, EFFICACIOUS ONES, SOMETIMES, SOMETIMES NOT, I THINK IT WOULD BE INTERESTING TO REPEAT SOME OF THE DATA OR SOME OF THE STUDIES LOOKING AT ALL THE DATA. >> SO JUST TO RESPOND TO THAT QUICKLY, I GUESS ONE OF THE QUESTIONS IS, WERE YOU CONVINCED BY THE CASE STUDY THAT THE TERPENE LACTONES FOR EXAMPLE WERE THE BIOLOGICALLY ACTIVE GROUP AND IN THAT CASE MAYBE YOU WOULDN'T HAVE TO LOOK BEYOND THEOSOPHER ACTIVES BUT IN THE CASE OF BLACK COHOSH WHERE THERE WASN'T ASSOCIATION BETWEEN THE CHEMICAL PROFILE AND THE BIOLOGY WE WERE LOOKING AT, THAT WOULD BE A PERFECT EXAMPLE OF WHEN WE WOULD NEED LOOK FOR ANOTHER ACTIVE. >> I WAS INTERESTED WITH SCOTT'S STUDY OF LOTS P AND G, WHAT WERE THEY, AND WHERE WAS EGB 761 IN THE BATCHES TESTED? BECAUSE THIS IS, DO I HAVE JIM WHAT YOU'RE ASKING IS LIKE HOW DOES THIS RELATE TO KNOWN GOLD STANDARD. >> YES. THAT'S A GREAT QUESTION. I CAN ANSWER THAT. SO AND ACTUALLY A LOT OF THE LOTS, SO A, B, C M AND H WERE EVEN IN SCOTTS ANALYSIS BECAUSE THOSE WERE THE STRAIGHT LINES WE SAW IN THE CHROMATOGRAM SO THOSE WERE REMOVED AND WE DIDN'T HAVE TARGETED DATA SO THEY WERE IN JIMS BUT NOT SCOTTS BECAUSE HE ONLY USED LOTS THAT HAD TARGETED DATA AVAILABLE. SO B WAS GINGKO LICK ACID, IT HAD NO OTHER PEAKS IN THE OTHER AREAS. THE UV XY AND Z WERE ALL EITHER THENIST STANDARDS OR THE EGB 761, AMONG CLOSEST TO THE NTP LOT. SO THOSE -- THAT GROUP AS JIM SHOWED WERE THE ONES THAT DIDN'T HAVE AGLYCONES IN THE UNHYDROLYZED SECTION. SO THAT'S KIND OF THOSE FELL OUT CLOSEST TO THE NTP LOT THOUGH BECAUSE IT HAS THE HIGHER TERPENE LACTONE LEVEL, IT WAS ON THE EDGE. >> I WAS STRUCK OBVIOUSLY THERE'S A STRONG CORRELATION AS YOU SAID BETWEEN THE TERPENES AND THE RESPONSE. I SAW U AND V WERE DIFFERENT. I KNOW FROM THE CHEMICAL ANALYSIS, THEY'RE IDENTICAL. I'M STRUCK WHY THERE WAS A DIFFERENCE IN THE TOXICOLOGICAL RESPONSE. >> WE HAVE AN ANSWER FOR THAT. >> GOOD. >> U AND V, SO THEY WERE -- BECAUSE OF THE NORMALIZATION WHEN WE DID THE UNTARGETED, IT BROUGHT THE PEAKS UP. BUT THE ACTUAL MEASURE CONCENTRATIONS IN B WERE VERY, VERY LOW BECAUSE OF THE KINDS STANDARD IT WAS LIKE A TABLET CONTAINING GINGKO SO THE CONCENTRATION OF GINGKO WAS LOW COMPARED TO THE U. >> CONGRATULATIONS. WONDERFUL SESSION TODAY I LEARNED A LOT. WHEN THE SCIENTISTS GET THEIR HEADS TOGETHER THEY CAN SOLVE THE PROBLEM, IT'S REALLY COMPLICATED. STORY THAT WE HEARD FROM CLINICAL PROFILING AND STATISTICAL DATA AND IN VITRO AND HOW TO DEAL WITH THE SITUATION. SINCE I'M PART OF THE ASPECT, COMMENT ON IT. I'M LOOKING AT WONDERFUL SCIENCE AND HOW MUCH EFFORT GOES INTO THESE THINGS THAT WE DO, WHAT ARE THE ELEMENTS TO REAL PUBLIC CONSUMER SAFETY. WE ARE TALKING ABOUT WHAT IS SIMILAR IN THE FRONT AND I APOLOGIZE HURT SOMEBODY'S FEELING BUT THROWING OUT OUT FOR DISCUSSION PURPOSE OUT THERE, WE'RE TALKING WHAT IS SIMILAR. BUT THE LAST PART OF THE FORMULATION, BECAUSE I SAID ON THE VARIATION BELEND TO END POINT OF EFFICACY TOXICITY. SO YOU END UP WITH REALLY COMPLEX AND END OF THE DAY PROBABLY NOT WITH ELEMENT TO THE CONSUMER, THE SAFETY POINT OF IT. THESE PRODUCTS WE TALK ABOUT A (INDISCERNIBLE), MORE THAN 22 YEARS NOT REALLY USE IT HERE AND CLINICAL TRIAL ISN'T PERFORMED IN THAT ONE. SO TO ME LOOK ACT THE MARKET KNOWING WHAT'S GOING ON IS LOT MORE TO SEE THE PURE COMPONENTS THE TOXICOLOGISTS CAN TEST RIGHT THERE. FOR EXAMPLE, EGCG WITH CAFFEINE, LOT OF SINGLE ACTUALS CAUSING MORE PROBLEMS. THAN GINGKO. I'M NOT SAYING THEY NOTER CAUSING PROBLEM BUT JUST TO LOOK AT. SO LOOKING AT THAT POINT OF VIEW I THINK THERE'S AN UPPER BOOST HUB AND SAFETY WHICH IS ALREADY PUBLISHED VOLUME TO CLASSIFY HUB AND (INAUDIBLE) CAN TALK ABOUT IT MORE AND I THINK HE'S HERE, CLASSIFY BASED ON SAFETY. SO TAKING SCIENTIFIC IS WONDERFUL BUT THIS IS REALLY SHOW THE POWER OF TOOLS IN THE STATISTIC KNOWLEDGE BUT SHOULD WE LET INDUSTRY COME TO ONE STANDARD AND USB WHAT GINKGO BILOBA LOOKS LIKE. THAT'S WHAT IT'S SUPPOSED TO BE ABOUT BUT OTHER THAN RADIATION. THIS IS A THING WE HAVE TESTED, YOU CHECK WITH AURBACH ASSOCIATE, THESE ARE SAFE, THEY ARE NOT SAFE, CREATE DATA AND KEEPING THE BOOK AND YOU CAN GO WHOLE NINE YARDS TO DO ALL THE STUDIES. THAT'S MORE RESULTS OR YENNED TO ME AND MORE SAFETY CONCERN. BECAUSE ALL GINGKO STUDIES, CANNOT TAKE IT OVER. SO END OF THE DAY, PLUS THIS PRODUCT WHENEVER YOU SEND A LETTER, THEY CHANGE THE LEVEL. SO HOW DO YOU COMBINE THESE SO I'M JUST THINKING FROM PRAGMATIC POINT OF VIEW, SCIENTIFICALLY, IT WAS WONDERFUL, I ENJOYED ALL THE TALK, THE WAY YOU CAME UP WITH MODEL THAT'S HOW IT SHOULD GO BUT MY OPINION LOT OF RESOURCES, LOT OF SCIENCE, FOR SOMETHING MORE COMPLEX AND WE WILL NOT GET AN ANSWER WHICH CAN BE STANDARD ACCORDING TO REGULATING BODY. MAYBE IT'S OFF THE CUFF GARMENTS BUT THIS IS SOMETHING WE HAVE TO PUT A LOT OF ENERGY RESOURCES ARE LIMITED, HUNDREDS OF PLANTS. IN HAWAII I HAD (INAUDIBLE). THIS IS MORE PURE CHEMICAL THAN PEOPLE ARE DOING CAUSING THE PROBLEM. GINGKO BLACK COHOSH, AND MORE PEOPLE WILL USE IT AND I'M SURE WE WILL SEE MORE PROBLEM BECAUSE THEY'RE SUPPOSED TO BE MEDICINE, MEDICINE. AND WE ARE USING SUPPLEMENT EVERY DAY SO IF YOU SEE A PROBLEM, I DON'T KNOW WHY PEOPLE ARE SURPRISED BUT THE THING IS, THIS EVEN DEFINING THE TEST, IS SUCH A COMPLEX ISSUE AND THEN END OF THE DAY SPENDING A LOT OF RESOURCES, AND I THINK THIS -- THERE IS TELLING 50% OF THE PEOPLE WHAT THE THERE IN THE MARKET THAT WE CAN TEST. I THINK THAT IS WONDERFUL. >> I CERTAINLY SHARE SOME OF THE SAME CONCERNS. I THINK THE APPROACHES WE DISCUSS PRINCIPALLY ARE VERY IMPORTANT? THE TYPE OF APPROACH THAT YOU HAVE TO TAKE TO ADDRESS THE PROBLEM BUT THE PROBLEM IS THAT THE SECONDARY METABOLITES IN PLANTS ARE SO COMPLEX. WE HAVE ALREADY SEEN EXAMPLES IN THE PREGNANT. SOMETIMES YOU HAVE A CORRELATION AND YOU HAVE A CLASS OF CHEMICAL AND YOU GET A GOOD CORRELATION AND THEN YOU HAVE THE FOR EXAMPLE IT WILL EXHAUST EXAMPLE WHERE THE CHEMICAL CORRELATION WITH THE BIOLOGIC CORRELATION FOR THE TWO BIOLOGICAL END POINTS ARE ALL DIFFERENT. BIOLOGIC END POINT DIDN'T CORRELATE WITH EACH OTHER. SO THERE ARE DIFFERENT CHEMICAL REACTIVITY. I THINK PART OF THE' ONE THING THAT WORRIES ME A LOT THAT'S NOT TALKED ABOUT, P MUCH, RESOURCES REQUIRED, I SPENT MUCH CAREER WORKING IN DRUG DEVELOPMENT, AND SO FOR A DRUG, IF YOU HAVE AN IMPURITY THAT'S DEPENDING ON THE DOSE THAT'S USED BETWEEN POINT 15 AND POINT 05, PERCENT OF THE DRUG EXPOSURE, YOU HAVE TO QUALIFY THAT BY DOING BIOLOGICAL TESTING, ET CETERA. THEN YOU HAVE THE SUPPLEMENT LAW THAT ALLOWS YOU TO HAVE THESE VERY COMPLEX MIXTURES THAT IF YOU'RE VERY LARGE AMOUNTS OF SECONDARY METABOLITES, IF YOU THINK ABOUT WHAT THOSE ARE, THEY'RE BASICALLY BAY LOGICALLY ACTIVE PLANT DEFENSIVE CHEMICALS OR CHEMICALS THAT HAVE BEEN EVOLVED IN THE PLANT TO MANIPULATE LIVING ORGANISMS LIKE INSECTS. THERE'S MORE THAN 100,000 SECONDARY METABOLITES IDENTIFIED IN PLANTS. THESE ARE INVOLVED IN PLANTS UNIQUE TO ACTIVE IN. '05 IN THE CASE OF THINGS SUPPLEMENTS ON THE FDA TO PROVE HARM IN CONTRAST COST IS ESTIMATED TO BE ABOUT $2 BILLION NOW TO DO ALL THE SAFETY TO PRESENT PERFORM THE STUDIES IMPROVE SAFETY. ALL THIS HAPPEN BECAUSE IT'S ENORMOUS WORK TO LOOK AT ALL THE DIFFERENT END POINTS TO LOOK AT MIXTURES TO DO THOSE CORRELATIONS AND THEN EXTRAPOLATE VERY EXPENSIVE AND BURDEN IS REALLY NOT ON INDUSTRY TO DO THAT. >> I GUESS -- I'M HEARING THAT THE TAKE HOME MESSAGE WAS THAT THE PROBLEM IS TOO COMPLICATED BUT I FELT THE OPPOSITE OF THAT IN THAT I WOULD SURPRISE -- I HAVE SURPRISED THREE OF FIVE LOTS CHEMICALLY DIVERGENT LOTS HAD ALMOST THE SAME EXACT BIOLOGICAL PROFILE. AND THAT THIS IS AN ANIMAL MODEL WITH ITS VARIOUS METABOLIZING CAPABILITIES AND THEY HAD DIFFERENT RATIOS AND DIFFERENT CONCENTRATIONS BUT IT WAS PRETTY CLEAR THEY HAD THE SAME BIOLOGICAL EFFECT AND THIS IS OKAY TO SAY THESE ARE GINGKO, THESE ARE NOT, AND IN TERMS OF THE GOALS, I MEAN I FEEL LIKE IT WILL HELP US IN THE FUTURE TO HAVE THESE -- THIS WHOLE DIFFERENT COMPARISON ACROSS FROM IN VIVO TO IN VITRO TO TAKE A SUBSET AND SAY THIS MAY BE HELPFUL IN TEST ARTICLE SELECTION IN THE FUTURE FOR DIFFERENT COMPLEX MIXTURES WHICH IS SOMETHING WE FACE ALL THE TIME. I'M STRUCK BY HOW THEIR TAKE HOME MESSAGE WAS DIFFERENT FROM MINE. DID YOU WANT TO -- >> GRATE FOR THE GINGKO EXAMPLE BUT IT WASN'T GREAT FOR THE CO-HASH EXAMPLE. IT ISN'T CLEAR AT ALL WHAT THE RESPONSIBLE COMPONENTS FOR THE BIOLOGICAL ACTIVITY ARE IN THERE, WASN'T CLEAR IN THE EXHAUST EXAMPLE EITHER. SO I THINK AS YOU GO THROUGH THE MANY PRODUCTS, CERTAINLY YOU WILL FIND SOME EXAMPLES WHERE THERE'S AN ACTIVE CLASS OF CHEMICALS. AND IT WILL CORRELATE WITH THE CHEMICAL COMPOSITION. THEY'RE GOING TO BE OTHER EXAMPLES, I'M CERTAIN WHERE THERE ARE SMALL CHEMICAL COMPONENTS THAT HAVE A DISPROPORTIONATE BIOLOGICAL ACTIVITY AND THEY WILL HAVE SPECIFIC ACTIVITIES. AND THE TOXIC ACTIVITY MIGHT NOT BE THE SAME CHEMICAL THAT'S THE EFFICACIOUS ACTIVITY. VERY COMPLEX SCENARIO. >> THAT BRINGS US TO WE WILL BE DISCUSSING TOMORROW THE ACTIVE CONSTITUENT IDENTIFICATION. WHICH IS ABSOLUTELY KEY TOWARDS APPLYING ANY OF THESE APPROACHES. GLENN DID YOU HAVE SOMETHING YOU WANTED TO -- >> COUPLE OF THINGS. FIRST JUST GENERAL IMPRESSION, I WANT TO GIVE KERRI AN OPPORTUNITY AS WELL. I THINK REALLY IMPRESSIVE STUDY, IT'S A GREAT STUDY, THIS IS OBVIOUSLY A LOT OF REALLY IN DEPTH THINKING AND HARD WORK THAT HAVE GONE INTO THIS. YOU GUYS HAVE DONE A NICE JOB WITH THE PROGRAM CONVEYING EVERYTHING THAT YOU DID IN A VERY DISTILLABLE ONE DAY FORMAT. SO COUPLE OF THINGS. FIRST ONE, IS HOW DID YOU ACTUALLY SELECT THE TOXICITY TEST? THAT YOU CHOSE? WHAT WAS THE PREMISE THAT SORT OF SAID OKAY WE'LL DO THE FOLLOWING. LIKE TO UNDERSTAND THAT. IT COULD HAVE EVOLVED. AND AGAIN I THINK THAT'S TOTALLY FINE AS WELL. THE SECOND IDEA TO THINK ABOUT, I GUESS I WOULD LIKE TO EXPLORE THAT WITH THE GROUP, AND THE AUDIENCE AS WELL, IS THIS IDEA YOU HAVE DONE A GREAT JOB OF DEVELOPING A LIBRARY OF RESPONSES TO A SET OF GINNING CO-S, GINGKO EXTRACTS. HOW TAKE THAT LIBRARY OF RESPONSES AND MAKE AVAILABLE SO IF INDUSTRY WANTED TO TEST THEIR PRODUCT AND SAY WHERE DOES THEIRS FIT IN TO THIS MIX, HOW DO WE TAKE THAT INFORMATION AND MAKE IT AVAILABLE TO THEM OR OTHER GROUPS, TO DO THE SAME THING. BECAUSE YOU HAVE A REAL LIBRARY OF RESPONSES THAT PEOPLE CAN SET THEIR OWN EXTRACTS AGAINST, FOR GINGKO OR OTHER TYPES OF THINGS. ANYWAY, THOSE ARE THE TWO INITIAL THINGS TO TOSS OUT THERE. I'LL YIELD TO CARRY. >> I FOUND THIS VERY INTERESTING BECAUSE I LOVE WHEN LIKE ALL DIFFERENT ANGLES ARE COMING TOGETHER TO TELL A BIGGER STORY. FROM MY PERSPECTIVE, WE'RE LOOKING AT A LOT OF FINISHED PRODUCTS. AND SEEING THAT A LOT OF THESE PRODUCTS ARE NOT WHAT THEY CLAIM TO BE. ON THE LABEL. SO THE IDEA YOU CAN USE A MULTIPLE WAY TO FURTHER ENSURE THAT YOU HAVE THE MATERIALS THAT ARE ENOUGH SIMILARITY THAT THEY CAN BE CALLED BLACK COHOSH OR CAN BE CALLED THE GINKGO BILOBA, SO CRITICAL BEFORE YOU TAKE THAT NEXT STEP TO DO THE TOXICOLOGY STUDIES OR DO EFFICACY STUDIES, BECAUSE OTHERWISE YOU HAVE THESE OUTLIER DATA POINTS THAT MAYBE EARLY ON YOU COULD HAVE FIGURED OUT THESE DON'T BELONG IN OUR STUDY. THOSE THE GIN COLIC ACIDS. WE ALREADY KNEW THERE WAS TOXICITY. SO YOU WOULDN'T HAVE WANTED THAT TO BE IN THERE. >> SO I THINK WHAT I SAID IS THE WORK YOU PEOPLE HAVE PRESENTED IS SO TREMENDOUS AND WHAT IT TAKES TO DO APPROACHES IN THINKING IS TREMENDOUS. BUT WHAT I WAS THINKING THAT THERE ARE SO MANY PRODUCTS OUTSIDE IN THE MARKET WHICH REALLY DO NOT HAVE ANYTHING LIKE THE T SHORT PRODUCT, THAT DON'T HAVE GINKGO BILOBA IN IT. IT'S BETTER TO WORK WITH THE INDUSTRY GROUP OR SOMEBODY WHO CAN SAY 50% OF THE PRODUCTS SOLD IN THE U.S. MARKET IS FROM THIS COMPANY, THIS IS WHAT THEY SELL. THAT WILL MAKE -- YOU CAN DO LOT MORE SCIENCE ON ONE PRODUCT AND GET ALL THE DATA AND YOU CAN MAKE MORE PROGRESS THAN INCLUDING ALL THE JUNK WE HAD, DID A LOT OF JUNK, DID NOT COLLECT ALL OF IT. YOU CAN REALLY THERE ARE ONLY A FEW PRODUCTS THAT I WILL SAY THAT YOU CAN RELY ON ALL THE TIME, RELIABLE PRODUCT, THE REST -- THE MORE YOU BUY THE MORE JUNK -- STATISTICALLY YOU CAN REALLY MAKE 20%. LIKE GINGKO, 20% IS THE REAL GINGKO 80% IS SUBSTITUTE DERIVED. THAT'S ALREADY PUBLISHED. SO YOU CAN ADD THE VARIABLES, IT ADDS A LOT OF DATA, A LOT OF ENERGY TO YOUR WORK SO WHAT I WAS SAYING THAT IF WE HAD A GROUP TO BRING PEOPLE TOGETHER, YOU SAY THIS IS REALLY THE MAJOR BRAND LIKE 24 GINGKO, EVERYBODY KNOWS IT T STANDARD IS CLINICAL TRIAL ALREADY PERFORMED THAT ONE YOU CLEAR IN YOUR DATABASE. THAT WILL BE LOT MORE GOOD DATABASE, WHERE YOU BUY THE PRODUCTS FROM, WE KNOW THERE'S A LOT OF JUNK. CREATING THIS DATA ON THIS PRODUCT PROUDLY WE CAN -- YOUR APPROACH, NEEDS TO BE DONE. SO MANY VARIABLES UP THERE WE ARE NOT LOOKING AT PLAN VARIATION. WE'RE LOOKING FOR THE PROCESS, THE PRODUCT OF GETTING THE MARKET AND ON TOP OF IT NOW PEOPLE ARE TALKING ABOUT DNA AUTHENTICATIONFUL IT'S GOING TO GET MORE COMPLICATED WE HAVE NOT TALKED IN 20 YEARS ABOUT GINGKO, THAT IT MEANS WHAT IT CAN DOES. WE'RE STILL TRYING TO FIND OUT (INDISCERNIBLE) IN GINGKO. FROM THE TOXICOLOGICAL END POINT AND THIS WORK WHAT IS THE RELATIONSHIP SO KEEP THE SAME APPROACH BUT LITTLE MORE THINKING HOW TO MIGHT BEMIZE THE SAM IT WILL -- MINIMIZE THE SAMPLE IF YOU CAN -- TO TELL THEM WHAT IS THE RIGHT PRODUCT BUT I WANTED TO BRING IT TO YOUR ATTENTION THAT THIS IS -- YOU CAN DO A LOT MORE IN USEFUL DATABASE. >> (INDISCERNIBLE) I WILL BE TALKING TOMORROW. PROFESSOR UNIVERSITY OF NORTH CAROLINA GREENS BORROW. I'M DEALING WITH A LOT OF SAME QUESTIONS YOU GUYS HAVE BEEN TALKED ABOUT TODAY AND I THINK -- I WANTED TO RESPOND A LITTLE BIT TO DR. MACGREGOR'S COMMENT ABOUT WHETHER IT'S WORTH DOING THIS WHEN PROBLEMS ARE COMPLEX. FOR ALL OF US WHEN TRAINED IN VERY REDUCTIONIST SCIENCE TO TRY TO TACKLE L THESE COMPLEX MIXTURES HARD, REQUIRE BRAIN UP GRACIOUS START BEING ABLE TO THINK IN LARGE DATA SETS AND WORK IN SOME OF THE UNTARGETED WAYS JIM LARNLY WAS TALKING ABOUT IN HIS DATA, IT DOES CHALLENGE THE PARADIGM DRUG DISCOVERY PARADIGM, THAT IS YOU HAVE TO KNOW ABSOLUTELY EVERYTHING THAT'S IN THE PRODUCT, EVERYTHING TESTED AND EVALUATED FOR EFFICACY AS COMPARED TO HAVING PRODUCTS WHERE WE'RE ACCEPTING THAT THERE'S STUFF IN HERE WE DON'T KNOW WHAT IT IS. AND YET THE TECHNOLOGY IS CHANGING SO RAPIDLY, SOMEBODY WORKING IN THE FIELD FOR 15 YEARS THERE'S THINGS WE CAN DO NOW, THE ROBUSTNESS OF THE DATA WE CAN GET IS TOTALLY DIFFERENT SO WE ARE STARTING TO TACKLE DIFFICULT COMPLEX QUESTIONS AN CERTAINLY NOT ABLE TO DO THIS DEPTH OF RIGOR THAT YOU HAVE DONE WITH GINGKO IN EVERYTHING SINGLE PRODUCT OUT THERE. BUT IT'S VERY IMPORTANT WE HAVE THESE DISCUSSIONS LIKE TODAY ABOUT WHAT'S THE BEST WAY TO DO THIS. AND CASE STUDY MULTIPLE EACH L ATTACK FROM DIFFERENT ANGLES THE PERFECT WAY TO DO THAT. SO MAYBE MOVE TOWARDS DISCUSSION OF HOW DO WE THEN -- WHAT ARE THE BEST PRACTICES FOR ACTUALLY CHARACTERIZING BOTANICALS THAT ARE GOING TO BE SOLD TO THE PUBLIC BUT I THINK THIS IS A REALLY IMPORTANT STEP FOR US TO DO THAT. I DO THINK IT'S TRACTABLE. >> COULD I CLARIFY MY COMMENT BECAUSE I CERTAINLY DIDN'T SAY IT WASN'T WORTH DOING. IMINO MEANS, THE APPROACHES ARE APPROPRIATE. WHAT I AM SAYING, IT'S VERY COMPLEX AND IF YOU'RE GOING TO USE THESE APPROACHES, YOU CAN'T FIND A SINGLE SAMPLE GET A CORRELATION AND MOVE AHEAD. THE NATURE OF SECONDARY METABOLITES IS INDUCED BY STRESS OR ATTACKS OR FUN GUY, THEY'RE BIOLOGICALLY ACTIVE SO YOU HAVE TO START ASSESSING CHEMICAL COMPOSITION SPECTRUM TO GIVE YOU MULTIPLE SAMPLES. THEN YOU HAVE TO THE CHEMICAL MAY NOT BE THE ONE AFFECTING ANOTHER SO IT'S COMPLEX AND THE THING THAT WORRIES ME IS HOW THE LAWS ARE WRITTEN A. THE LAWS UNLIKE DRUG LAWS PLACING BURDEN OF PROOF ON SPONSOR A DRUG PLACES THE BURDEN OF PROOF ON REGULATORY AGENCY TO TAKE ANY ACTION TO PROVE HARM WITH DIETARY SUPPLEMENT COMPANIES ABLE TO MARKET SIMPLY BY PUTTING A LITTLE DISCLAIMER MAKING A STRUCTURE FUNCTION CLAIM OPPOSED TO DISEASE CLAIM. WHICH THEN FREES YOU FROM DOING THOSE TYPES OF STUDIES THAT ARE BEING DONE AROUND THE TABLE. THAT NEED TO BE DONE SO SAYING WE NEED A BETTER WAY OF GETTING THIS MASSIVE DATA HITS MUCH MORE COMPLICATED THAN THE DEVELOPMENT OF CONVENTIONAL SINGLE PURE DRUGS. >> QUESTIONS. >> WHAT PEOPLE ARE CALLING SUPPLEMENTS BY THE WAY ARE BEING SOLD AS DRUGS. PEOPLE SAKE SAW PALMETTOS AND MAINTAINING PROSTATE HEALTH THEY TAKE IT TO TAKE PROSTATE DISEASE, IT'S A LABEL SAID THAT THERE'S $2 BILLION OF RESEARCH BUT BECAUSE IT'S STRUCTURE FUNCTION CLAIM THEY'RE NOT IN IT FOR ANYTHING. >> I HAVE A COMMENT. FIRST I ELECT THE BEGINNING STATEMENT THAT WE ARE WORKING ON DEFINING WHAT SIMILARITY MEANS. THE OTHER COMMENTS, I DON'T THINK -- ONE OF THE CONCLUSIONS AND ONE OF THE PRESENTATION THAT (INDISCERNIBLE) LOOKING AT THOSE STANDARDS I BELIEVE A FEW OF THEM MEET SOME STANDARDS AN MAJORITY DON'T IMMEDIATE MEAT ANY STANDARD. THE ISSUE I BELIEVE WE WERE MISLED BY THE LOOKING AT -- BY LOOKING A AT ONE CLASS OF COMPOUNDS, LOOKING AT SPECIFIC COMPOUNDS, LOOK AT GINGKO, AND WE SAY THAT GINGKO EXTRACT CONTAINS 24% FLAVINNOIDS WITHOUT CONSIDERING THE FACT THAT WHAT'S IN GENERAL FLIGHT SIDES OF (INDISCERNIBLE) FOCUS ON THE -- SO THE ISSUE WE LOOK AT WHEN CONSIDERING THIS INFORMATION, WE NEED TO LOOK AT THE TOTALITY OF INFORMATION, WE NEED TO GET PEOPLE WHO ARE -- HAVE THIS EXPERTISE INVOLVED SO THAT WE START WITH GOOD MATERIAL SO IN THIS REGARD I ALSO AGREE WITH (INDISCERNIBLE). THANK YOU. >> I THINK INDUSTRY NEEDS TO CHANGE THE STANDARDS AND IF YOU WANT TO LOOK AT AGLYCONES TO LOOK AT YOUR 24% THAT'S NOT GOING TO WORK AS YOU POINTED OUT, IT'S EASILY FOOLED. SO INDUSTRY CAN CHANGE IF THEY LOOK AT THE UNHYDROLYZED THEY GET A TOTALLY DIFFERENT PATTERN AND DIFFERENT SET OF RESULTS. GOING BACK TO THE COMPLEXITY OF THE SECONDARY METABOLITES, THERE'S A LOT OF THEM OUT THERE. AND JOB SECURITY FOR CHEMIST. BUT IF YOU DO SOMETHING LIKE FLOW INJECTION MASS SPEC, YOU LITERALLY GET HUNDREDS OF THOUSANDS OF MASSES AND IF YOU DO HIGH RESOLUTION MASS SPEC YOU MAY GET TENS OF THOUGHTS AND CERTAINLY IF YOU DO NMR YOU GET CLOSE TO 30,000 DIFFERENT POINTS. SO YOU'RE MUCH -- YOU'RE PRETTY MUCH COVERING A LOT OF SAMPLES, THE BIGGEST OR COMPONENTS, THE BEST GAP WILL BE BASED ON THE SOLVENT YOU USE, IF IT'S METHANOL WATER YOU CAN CHANGE IT TO 100% METHANOL OR WATER, YOU GET A DIFFERENT DISTRIBUTION BUT THE POINT IS FLOW INJECTION MASS SPEC TAKES THREE MINUTES TO RUN SO THE SAMPLES ARE ANALYZED VERY, VERY QUICKLY. YOU PROBABLY WILL FIND THE BIOLOGICAL OR TOXICOLOGICAL ASSAYS ARE YOUR LIMITING FACTOR. WE HAVE DONE THIS BEFORE WHERE WE TAKE A COMPONENTLIKE MAYBE TOXICITY MEASUREMENTS YOU RANK YOUR SPECTRA ACCORDING THE LARGEST RESPONSE TOX COLOGICALLY AND LOWEST. AND THEN YOU DO SOMETHING LIKE YOU DIVIDE THEM INTO VARIOUS TOP FIVE DIFFERENT CATEGORIES OR DO IT INDIVIDUALLY IF YOU WANT TO DO REGRESSION BUT YOU CAN NICELY ESTABLISH THEN FOR EACH MASS, WHICH ONES CORRELATE WITH THE TOXICOLOGICAL RESPONSE. AND THE ANALYSIS IN THAT WAY IS REALLY VERY QUICK. SO THE BIGGEST PROBLEM IS WHAT ARE YOU GOING TO USE FOR YOUR REFERENCE MATERIALS. COLLECTING AUTHENTIC MATERIALS IS THE BIGGEST EXPENSE. IF YOU'RE MANUFACTURING YOU HAVE SAMPLES THAT ARE GOOD OR BAD OVER THE YEARS AND YOU CAN EASILY ESTABLISH THOSE A LAB LIKE MYSELF, WE HAVE TO GO BUY THOSE OR GET SOMEONE TO DONATE THEM SO WHAT IS AUTHENTIC, WHAT DO YOU WANT TO USE AS YOUR REFERENCE IS PROBABLY YOUR INITIAL BIGGEST DECISION. PURCHASING THEM MAYBE THE OBSTACLE YOU RUN INTO. >> SCOTT DID YOU HAVE A QUESTION? >> JUST A COMMENT. I THINK WE THINK ABOUT THE SCALE OF BIOLOGY DIFFERENT -- AT LEAST I DO, THERE'S CAPABILITIES THAT ARE BEING DEVELOPED AT NIEHS AND ALSO BEING DEVELOPED AT THE BROAD INSTITUTE THAT YOU CAN DO THOUSANDS OF GENES AND GENERATE PATTERNS FROM A VARIETY OF DIFFERENT CELL TYPES FOR DOLLARS. IT Es NOT SIGNIFICANT, I CAN GO TO GNC NOW PULL EVERYTHING OFF THE SHELF PROBABLY RUN A SCREEN AND DOSE RESPONSE AND TEN DIFFERENT CELL TYPES FOR MAYBE $100,000. GENERATING A RESOURCE LIKE THAT, HAVING A DATABASE, AVAILABLE AS POINTED OUT, NOT AN UNREASONABLE THING ON BIOLOGIC SIDE, CHEMISTRY ISN'T HIGH THROUGH PUT IN THIS POINT BUT IT'S GOING TO HAPPEN IN THE NEXT FIVE YEARS IT WILL BE AVAILABLE MATTER OF HAVING GOOD PRACTICE CELL CULTURE. OBVIOUSLY YOU CAN'T DO ANIMAL STUDY ON THESE THINGS. THAT'S NOT POSSIBLE. HE CAN DO A GOOD JOB LOOKING AT BIOLOGICAL ACTIVITY AND DIVERSE I HAVE TO EFFECT ACROSS A NUMBER OF DIFFERENT CELLULAR SYSTEMS. IN HIGH DIMENSION QUICKLY. I DO AGREE THE BIGGEST PROBLEM IS CONSULTING NON-CLINICAL DRUG DEVELOPMENT SO I'M USED TO LOOKING AT THE FLIP SIDE OR INDUSTRY DOES PAY FOR EVERYTHING. BUT PRIOR TO MY CONSULTING WORK, I WORKED IN PETROLEUM INDUSTRY. I DID SEE WORKING WITH VERY COMPLEX MIXTURES VERY -- A LOT OF US, MOST UNKNOWNS. IN YOUR MIXTURE. WHAT DR. KHAN WAS SAYING, IN TRYING TO GO TO MANUFACTURER, WHEN YOU'RE TRYING TO SELECT TEST MATERIAL A GOOD PLACE TO START WOULD BE WITH MANUFACTURERS ON WHAT THEY'RE SELLING WHAT THE HIGH SELLERS AND THOSE NUMBERS ARE NOT GOING TO BE -- NOT A PROBLEM OF BUYING A SAMPLE THE PROBLEM OF FINDING OUT WHAT PEOPLE ARE TAKING YOU MIGHT GET COMPANY ANNUAL REPORTS, THAT WOULD BE DATA MINING. INSTEAD OF TO USE AN OLD METAPHOR, OR ANALOGY, MOVING UNDER THE LAMP POST WHERE WE HAVE DATA, GO OUT THERE, FIND WHAT PEOPLE ARE REALLY TAKING, IN LARGE QUANTITIES AND GINGKO FALLS IN THAT CATEGORY. I ABSOLUTELY AGREE IN PICTURE TOXICOLOGY DECIDEING WHAT TO TEST THAT'S 90% ISSUE, HOW TO TEST IT AND YOU'RE ON THE DOWNHILL PART OF THE SLOPE. WORKING WITH THE MANUFACTURER AND WE DID HAVE ONE SPEAKER THIS MORNING WITH TRADE ASSOCIATION AND YOU'LL FIND COMPANIES ARE GOING TO BE GETTING MORE AND MORE COOPERATIVE IN THIS AREA. AND AGAIN, IF THEY'RE NOT, LOOK AT THE SALES FIGURES. >> PAUL HOWARD CURRENTLY WITH FOOD AND DRUG ADMINISTRATION. I PREDICTED TOX 21 WOULDN'T WORK, IT SURE HAS. WHEN YOU APPROACH MED A YEAR AGO, THIS ISN'T GOING TO WORK SO TAKE THAT INTO CONSIDERATION OF MY COMMENTS, SOMETIMES I'M NOT QUITE PREDICTIVE. REALLY TWO QUESTIONS. ONE, WHAT IS THE END GAME? IF THE END GAME IS TO DETERMINE IF A PRODUCT IS OUTSIDE THE NORM OF WHAT YOU EXPECT, I THINK THAT THIS EFFORT IS TREMENDOUS IN THAT YOU WILL BE ABLE TO ADVISE INDUSTRY TO SAY HERE ARE THE PARAMETERS TO USE TO CONVINCE PEOPLE YOU HAVE AN APPROPRIATE PRODUCT ON THE MARKET. IF THAT'S THE END GAME I THINK IT'S GREAT. IF THE END GAME IS TO DRIVE AN UNDERSTANDING WHAT'S GOING ON IN HUMANS I'M CONCERNED BECAUSE CRAIG HOSPITAL IN THIS TALK THIS MORNING, EVERY CLINICAL TRIAL THESE THINGS HAS FAILED. SO ALL YOUR PREDICTIVETY IS GOING AGAINST A FAILED EFFICACY STUDY. IS TA THE END GAME OR IS IT MORE FOR PROVIDING INDUSTRY WITH TOOLS TO KNOW IF THEY HAVE SOMETHING THAT MEETS NORM. I WILL GUARANTEE THEY KNOW WHEN THEY'RE DILUTING THEIR MATERIAL WITH MALTOSE OR ANYTHING ELSE THEY KNOW WHITHER DOING AN TRYING TO GET AWAY WITH IT SO WHAT'S THE END GAME, MAYBE THAT'S A QUESTION FOR THE PANEL. >> A LOT OF TIMES, THE MANUFACTURER THE RESPECTABLE MANUFACTURER MAY NOT KNOW, BECAUSE THE SUPPLIERS OF THAT MATERIAL ARE DOING THE DELUSION, DOING THE ADULTERATION AND SO I DO AGREE THAT WE NEED TO WORK COOPERATIVELY WITH RESPECTABLE MANUFACTURERS BUT SUPPLIERS HAVE RESPONSIBILITY, REGULATORY WISE THEY DON'T -- THEY'RE NOT OBLIGATED IN THAT WAY. THAT'S CONCERNING. THE OTHER COMMENT TO MAKE IS SOMEONE ELSE, ANALYTICAL CHEMISTRY INVOLVED OTHER TEST METHODS EVOLVED WE'RE ABLE IN A SHORT AMOUNT OF TIME TO GET A LOT OF DATA. YEARS AGO WE DID TAKE MORE REDUCTIONIST ATTITUDE TO ANALYZE FOR THESE FEW THINGS SO LET'S FOCUS ON THAT. THAT OPENED THE GATE FOR ALL THESE PEOPLE THAT ARE NOW CHEATING TO MAKE AN ECONOMICALLY INFERIOR PRODUCT AND IT'S CASCADING. SO TO OPEN UP AND USE NON-SELECTIVE NON-TARGETED ANALYSIS, THAT ARE SEEING EVERYTHING IN THE PLAN, YOU THEN START TO WEED OUT THE BAD MATERIAL. AND THEN YOU CAN TAKE ADVANTAGE OF THE SYNERGIES IN THAT NATURAL MATERIAL THAT MAKE THE WHOLE PLANT BENEFICIAL OR TRADITIONAL USE OF THAT PLANT BENEFICIAL. I THINK COMING TOGETHER FROM A LOT OF ANGLES AND USING MORE DATA, WILL HELP US ANSWER THESE QUESTIONS. >> REAL QUICK BACK TO PAUL, I WOULD SAY YOU UNDERSTAND WHAT NTP DOES, WE SELECTED TEST ARTICLE AND WE TESTED IT AND THEN PROVIDE INFORMATION. I THINK THIS EFFORT WAS AN ATTEMPT TO PROVIDE DON TEXT TO THE INFORMATION. SO IF WE TEST ONE THING AND WE REPORT RESULTS WE'RE ALWAYS OPEN TO THE CRITICISM THAT THAT THING IS NOT LIKE ANYTHING ELSE. YOU TESTED SOMETHING VERY SPECIFIC. THIS IS A WAY OF ACTUALLY QUANTIFYING IS IT ONE SPECIFIC THING OR DOES IT RELATE TO THIS LEVEL OF THINGS. OUTSIDE OF IT. SO I THINK THIS IS AN ATTEMPT TO REALLY SYSTEMATICALLY LOOK AT THAT QUESTION SO IN THE FUTURE NEXT TIME WE TEST SOMETHING WHICH WE WILL TEST MANY OTHER COMPLEX MIXTURES, WE ARE TRYING TO DEVELOP THESE APPROACHES TO SAY OKAY HERE IS HOW WE MOVE THROUGH THAT QUESTION, HOW IT RELATES TO OTHER THINGS. >> CAN I WORK ON -- (INDISCERNIBLE) ONE THING WE SHOULD UNDERSTAND THE WHOLE THING IS EVOLVING. LIKE SAINT JOHNS WORK, PEOPLE TRIED TO REMOVE IT SO THE PRODUCT AS PEOPLE ARE USING MORE CLINICAL TRIAL AND SAFETY STUDY IS BEING DONE, THESE PEOPLE HAVE NOTHING TO LOOSE FORMULATION (INDISCERNIBLE), THEY CHANGE LABEL, CHANGE THE TIME SO THE PRODUCT WE HAVE IN THE MARKET MAY NOT HAVE A PRODUCT FIVE YEARS OR MAYBE TOMORROW YOU NEVER NO KNOW. SO THE MARKET IS FLOODED WITH ALL THESE THINGS RIGHT NOW, THE THING WE'RE TALKING ABOUT GINGKO AND BLACK COHOSH (INDISCERNIBLE) IN THE GINGKO PRODUCT BEFORE THEY HAVE BEEN DEVELOPED BASED ON SCIENCE SO WHILE WE ARE LOOKING, THAT ONE MORE POINT IF WE STICK TO THE FINAL STANDARDS, SOME STANDARDS THAT WILL REMAIN THE SAME, THIS IS SAINT JOHN'S WART, THIS IS GINGKO, WE HAVE THE LIFE SPAN OF THIS THING A LOT LONGER THAN IF YOU LOOK IN THE MARKET PRODUCT BECAUSE MONOGRAPH THAT CHANGE OVER THE LIFETIME. (INDISCERNIBLE) (INDISCERNIBLE) TOMORROW. >> YES. HI, DR. PRETTY ANN HOFFMAN FORMERLY OF FDA, NOW HETEROGENEITY CONSULTING COMPANY, WE DEAL WITH FOODS, MOSTLY DRUGS, THERE ARE TWO ON THE U.S. MARKET, ONE IS WITH THE GREEN TEA EXTRACT, TOPICAL, AND THE OTHER IS ACTUALLY A PERUVIAN LATEX PRODUCT. FOR HIV -- HIV ASSOCIATED GONORRHEA. THESE BOTH WENT THROUGH THE DRUG PROCESS, THEY DID TAKE SIGNIFICANTLY LESS MONEY THAN THE $2 BILLION. $100 MILLION I WOULD BE SURPRISED. SO BASICALLY THEY WERE COST EFFECTIVE TO GO THROUGH. THEY BOTH GO THROUGH THE FULL TOX COLOGICAL STUDIES, THEY WENT THROUGH EVERYTHING AND SOLD AS PRESCRIPTION DRUGS BUT MY QUESTION IS REALLY, THE PROCESS IS PRODUCT, PRODUCT IS FROM PROCESS, MANY DIFFERENT SCENARIOS, WHAT ARE YOU LOOKING AT HERE, LOOKING TO TRY TO REMOVE PRODUCTS FROM THE U.S. MARKET? THAT DON'T MEET STANDARD? IS THAT THE GOAL OR IS THE GOAL I COULD SEE YOU DEVELOPING A REFERENCE STANDARD THE FDA COULD USE TO SAY YOU DON'T HAVE THIS PRODUCT. BUT WHEN YOU MOVE AWAY FROM SAY GINGKO, IT'S NOT JUST GINGKO, IT'S THE PART, THE PLANT WRIT'S GROWN, HOW IT'S GROWN AND PROCESSED. SO THERE'S RAW MATERIAL THAT YOU CAN LOOK AT AND TEST AND PAY AND LOOK AT THE INTERESTING TOXIC EFFECTS OF THAT PARTICULAR PARTICULAR PLANT -- BUT WHEN YOU GET THE ACTUAL FINISH PRODUCT IT WAS PROBABLY NEGATIVE BECAUSE IT WAS STANDARDIZED HIGH -- AND NOT HYPERFLORIN AND FLAVINNOIDS, THE OTHER THINGS CONTRIBUTED TO THE IN DIFFERENT TYPES OF EXTRACTS THAT WERE ON THE MARKET. AND THE NIH DID HAVE -- I'M OLD SO I KNOW, THE NIH DID HAVE A PROBLEM IN NOT CHOOSING THE MANUFACTURER WHO GOT THE PRIOR POSITIVE RESULT. THEY WERE TAKING DIFFERENT PRODUCTS, THEY -- DIFFERENT PROCESSES TO TEST SO THEY DIDN'T GET THE SAME BIOLOGICALS WHICH THOSE WERE DRUG TRIALS, THEY WERE NOT FOOD TRIALS. SO MY BIG QUESTION IS, WHAT IS THE PURPOSE, I CAN SEE A PURPOSE IF TRYING TO SAY GINGKO LEAVES CAUSE CERTAIN TOXIC EFFECTS. THEY SHOULDN'T BE USED BEYOND LEVEL IN PERHAPS OLD PRODUCTS. JUST THROWING THAT OUT. BUT TO TRY TO MAKE A STATEMENT FOR ALL GINGKO AND ALL MANUFACTURERS THERE ARE LOTS OF PROPRIETARY PROCESSES. >> I THINK THE GOAL SHOULD BE TO DEFINE STANDARDS THAT ASSURE WHEN A PRODUCT IS SOLD IT DOES WHAT EAT CLAIMED TO DO. ON THE BENEFIT SIDE AND DOES NOT PERFORM AND DOES NOT HAVE ADVERSE EFFECTS. >> BUT THERE'S NO BENEFIT FOR FOODS. THERE'S SAFETY BUT IT ISN'T A RISK BENEFIT, YOU DON'T HAVE -- THE LAW REQUIRES THE FDA TO LOOK AT RISK VERSUS BENEFIT VERSUS RISK FOR DRUGS. THAT DOESN'T EXIST FOR FOODS. >> THERE'S TWO SAFETY CLAIMS FOR FOODS. THERE'S STRUCTURE FUNCTION CLAIMS AN SAFETY CLAIM CLAIMS. A STRUCTURE FUNCTION CLAIM IS CLAIMING A CERTAIN EFFECT IS OCCURRING AS A RESULT OF USING THIS PRODUCT. AND MY OPINION IS THAT REGULATORY TAN CARDS SHOULD ASSURE THOSE ARE ACCURATE. >> THAT'S FOR THE FEDERAL TRADE COMMISSION TO DECIDE. THEY'RE THE ONES WHO HAVE TO LOOK AT THE STUDIES SEQUENTIALLY, NOT FDA. >> FTC IS ADVERTISING, FDA IS A LABEL ADULTERATION. >> MIKE DEVITO NTP. SO I'M GOING TO TRY TO REITERATE WHAT SHOT SHOWED IN THE FIRST SLIDE. WE WANTED TO TEST SAFETY CHRONIC TOXICITY CARCINOGENICITY IN THESE. WE WANT TO UNDERSTAND THEIR SAFETY. WE TESTED A PARTICULAR LOT. WE CHOOSE -- WE'RE ONLY GOING TO DEAL WITH ONE. THE REAL QUESTION IS, HOW DOES THIS LOT REPRESENT WHAT'S ON THE MARKET? AND WHEN WE PRESENTED IT THERE WAS A CHEMISTRY ARGUMENT OUR LOT WAS NOT IDENTICAL SO WHAT'S ON THE MARKET. SO WE THOUGHT WE WOULD TRY DOING SHORT TERM BIOLOGICAL SCREENS TO SUPPLEMENT CHEMISTRY DATA. AND THE REAL QUESTION WE'RE TRYING TO ASK IS WAS THAT HELPFUL? CAN WE COME IN AND SAY THESE ACT A LOT BIOLOGICALLY. SHOULD THE DECISION OF WHETHER THERE'S SIMILAR OR NOT BE DRIVEN SOLELY BY MINOR CHEMICAL CHANGES IN THE COMPETITION OR SHOULD AFTER -- CAN A BIOLOGICAL DATABASE OF SHORT TERM ASSAYS HELP US IN UNDERSTANDING WHETHER THINGS ARE SIMILAR OR NOT? AND I THINK WE'RE TRYING TO LOOK FOR SO MANY EXCEPTIONS TO THIS, SIMPLE QUESTION, WE HAVE LOST SITE OF THAT IS WHERE WE WERE TRYING TO GO, NOT TRYING TO TAKE THING OFFICE THE MARKET, WE'RE NOT TRYING TO TALK ABOUT EFFICACY OR WHETHER THEY'RE SAFE. WE WERE DOING SAFE, THAT'S RIGHT. MY BAD. BUT THAT SORT OF THE CONTEXT OF WHERE THIS IS COMING FROM. HOW DO WE ASSESS THE SAFETY OF THESE PRODUCTS GIVEN THE FACT THAT A DRY SEASON LOOKS BETTER THAN WET SEASON. >> SO THE ANSWER IS YES. >> I SAY THAT AND I WILL THROW THIS OUT THERE, I THINK THE BUY LOGICAL TEST, IF YOU CAN LINK THEM THROUGH AOP CONCEPT OR SOME OTHER CONCEPT, IF YOU CAN LINK THOSE PERTURBATIONS YOU MEASURE IN VIVO AND VITRO, WHATEVER IT'S GOING TO BE TO AN APICAL OUTCOME, WITH A PLAUSIBLE HYPOTHESIS, I THINK THAT'S A LOT STRONGER ARGUMENT FROM A SUFFICIENT SIMILARITY PERSPECTIVE. THEN THE IDEA WOULD BE TO TAKE THAT AOP AND TURN IT OVER AND SAY CAN WE GO BACK. AND LINK TO THE CHEMISTRY. OR LINK IT TO SOME SORT OF OTHER MOLECULAR INITIATING EVENT AT A CELLULAR LEVEL, THEN YOU'RE VERY FIRM GROUND BECAUSE YOU CAN GO ALL THE WAY BACK THROUGH THAT CHAIN OF EVENTS. AND SAY YES. MAKES THIS FIT TOGETHER IN ONE PIECE. IF THEY CAN'T YOU'RE LEFT WITH WHAT DAY YOU BASE THIS ASSESSMENT ON. IF YOU BASE ON THE BIOLOGY DATA YOU NEAR A STRONGER POSITION. >> I SAW LOTS OF HANDS BUT DIDN'T -- >> JUST RESPOND I THINK THE ANSWER IS YES. BUT YOU ALSO SAID WHEN THEY DID STUDIES PEOPLE SAID THIS IS NOT WHAT WE USE. MY POINT IS, YOU CAN GO DO ALL THE SCIENCE YOU WANT, THEY WILL STILL SAY THE SAME THING. DOESN'T MATTER HOW MANY STUDIES TO DO. BUT IF YOU GET THEM INITIALLY IN YOUR PROCESS BY SITTING WITH THEM, THEY'RE DOING ANY SIGNS BUT THIS IS OUR PRESENTED MODEL THE ONE SAMPLE YOU SELECTED IS THE ONE THAT 10% OF THE PEOPLE USE, YOU ARE GOOD TO GO. THAT'S THE ONLY THING YOU NEED. SCIENCE WISE YOU'RE GOING TO DO IS SCIENCE BUT BELIEF ME WE ARE DEALING WITH NSF, WE ARE DEALING, DOESN'T MATTER YOUR SCIENCE SAY, >> (INDISCERNIBLE) CONSULTING I THINK THE EXERCISE TRYING TO DEFINE SIMILARITY AND LOOK AT CHEMICAL ASPECT AND BIOLOGICAL ASPECT IS EXTREMELY IMPORTANT. I THINK WE HAVE TO GO FORWARD WITH THIS. THE ISSUE IS THAT EVERYBODY OUT THERE, THE AMERICAN CONSUMER AND MOST OF THE PRACTITIONERS ASSUME IF THEY HAVE THE COMMON NAME FOR A MATERIAL, THEN IT'S EQUIVALENT. SO THEY'LL ASSUME THAT THEY WILL SAY YOU MIGHT TRY TAKING SAW PALMETTO OR GINGKO. BUT THERE ARE NO INSTRUCTIONS BEYOND THAT. SO THE CONSUMER GOES OUT AND BUYS SOMETHING FROM COSTCO OR FROM A HEALTH FOOD STORE AND THEY MAY GET A REALLY GOOD PRODUCT OR THEY MAY GET SOMETHING WITH NO GINGKO AT ALL. SO I -- HOW DO WE ADDRESS THIS? I WORK WITH SOME MANUFACTURERS THAT PRODUCE PRODUCT. AND A LOT OF TIMES THEY WILL SAY WE WANT TO INCLUDE A GINGKO EXTRACT IN THIS FORMULA. SO THE CONTRACT MANUFACTURER, THIS IS HOW IN MOST PRODUCTS ARE NOT SO PROPRIETARY LIKE IN EUROPE, THEY HAVE A CONTRACT MANUFACTURER. THEY COME UP WITH AN INGREDIENT OR THEY PROPOSE AN INGREDIENT LIKE RIGHT NOW GINSNG IF YOU ASK FOR THAT YOU WILL GET GINSEN LEAF, I HAVE TO TURN IT DOWN TWICE AND THE MANUFACTURER SAYS TO ME, MARILYN YOU'RE SO PICKY CAN, THIS IS EXPENSIVE GO THROUGH THE ROOT. SO THEY'RE LIKE GET IT OF CONSULTANT AND WE'LL GO ON AND MAKE THE PRODUCT. RIGHT? YOU LAUGH BUT THIS IS IS REALITY BUT THE OTHER THING IS LIKE WITH GINGKO, GINGKO IS UNUSUAL BECAUSE WE HAVE A GOLD STANDARD. WE HAVE ETP 761. FOR MOST BOTANICALS WE DON'T HAVE A GOLD STANDARD OR A BARE ASPIRIN YOU CAN COMPARE -- YOU CAN DO GENETIC STUDIES, AND COMPARE AND FIND WHAT IS THE GOLD STANDARD. SO WE'RE SEARCHING AROUND LIKE SOMETIMES WE KNOW WE MIGHT TAKE REMIFEMIN BUT WHEN WE GET TO BOTANICALS RESEARCHED IN EUROPE, WE DON'T HAVE A GOLD STANDARD. >> I BROUGHT UP THE IDEA OF LIBRARY EARLIER AND I THINK IT GOES TO THE POINT YOU'RE MAKING, AS IF I'M WHOEVER IN THE GOVERNMENT THIS IS A NEW PRODUCT. LET'S DO A COUPLE OF SIMPLE SHORT TERM ASSAYS SAME CHEMISTS SO MAYBE BOTH BUT THE IDEA IS YOU SORT OF HAVE THIS LIBRARY YOU SET UP AND YOU SAY WELL, THIS MAYBE A BRAND NEW PROCESS, IT MAYBE VERY, VERY DIFFERENT BUT BY GOSH, THE TEST IS THE SAME. SO LOW AND BEHOLD IT TESTS DIFFERENTLY BUT YOU HAVE THIS LIBRARY SET UP ALREADY THAT YOU CAN COMPARE THIS NEW MIXTURE TO. THAT YOU HAVE ALREADY DEVELOPED. THAT'S WHY THINK THAT'S A REALLY IMPORTANT CONCEPT TO COME OUT OF THE WORK THAT YOU GUYS ARE DOING. >> SO GLENN TALKED ABOUT THE LIBRARY AND BEING ABLE TO COMPARE. I'M CHIEF NTP LABS, A LOT OF WORK WAS DONE IN OUR LABS. THIS DATA WILL BE AVAILABLE ON THE WEB IN RAW FORMAT. RIGHT AROUND THE TIME WE PUBLISH IT. SO PEOPLE WILL BE ABLE TO ACCESS THE DATABASE TO COMPARE THEIR PRODUCTS TO OURS AND SIMILAR ASSAYS AND OTHERS. >> ONE COMMENT. ACCORDING TO SCOTT'S DATA T CLOSER THE COMPOSITION WAS TO SAMPLE NUMBER ONE, OR 1F, THE NIEH STANDARD, THE MORE TOX RESPONSES THE HIGHER THE TOX RESPONSES, SO THE MANUFACTURERS WHO ARE ACTUALLY DILUTING SAMPLES BE VARIOUS CAMPEROL ARE ACTUALLY MAKING THE PRODUCT SAFER FOR US. >> THAT'S A GOOD POINT. >> JAMES STOLE MY THUNDER ON THAT, BUT ONE THING I DIDN'T SEE AND HAVEN'T SEEN -- AND IF TO HAVE BEEN THE ICBS MEETING MANY TIMES I ALWAYS BRING THIS UP BUT ONE THING I DIDN'T SEE HERE IN OUR TALKS TODAY ARE IS THAT WHAT A LOT OF THESE IN VITRO SYSTEMS SEE IS NOT -- I MEAN THESE IN VITRO SYSTEMS ARE NOT SEEING THESE PURIFIED CHEMICALS. THEY'RE SEEING METABOLITES OF PHYTOCHEMICALS. ONCE THROUGH THE GUT THINGS CHANGE. IT'S A DIFFERENT ANIMAL AT THAT POINT. SO IT WOULD BE REALLY NICE TO SEE SOME CONCENTRATIONS IN METABOLITES IN FOR AT THAT TIME BLOOD AND ALSO IN THE TISSUES THEMSELVES. SO THAT'S SOMETHING WE CONSTANTLY TRY TO LINK BIOLOGIC EFFECTS OR TOX COLOGIC EFFECTS TO THE INDIVIDUAL PHYTOCHEMICALS. I WOULD SAY 99 TIMES OUT OF 100 IT'S A METABOLITE OF THAT OR BIOTRANSFORMED ASPECT SO I WANTED TO THROW THAT OUT THERE FOR WHAT IT WAS WORTH. >> GREAT POINT. ALSO WE HAVE BEEN TO OUR DAY 2 DISCUSSION OF ADME WHICH WE'RE LOOKING FORWARD TO. SO WE'RE ABOUT 15 MINUTES BEHIND SCHEDULE BUT IT'S BEEN A GREAT DISCUSSION. I ENCOURAGE Y'ALL TO CONTINUE DISCUSSING THIS TOPIC. THANK YOU SO MUCH FOR YOUR ATTENTION TODAY. [APPLAUSE]