Document 2qxZgqd4j3nXQjvxn7jwZLXG6

TRADE SECRET Study Title H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Laboratory Project ID: DuPont-6544 AR226-3195 DuPont-6544 AUTHOR: Carol Finlay, B.A. STUDY COMPLETED ON: November 26, 2001 PERFORMING LABORATORY: E.I. du Font de Nemours and Company Haskell Laboratory for Health and Environmental Sciences Elkton Road, P.O. Box 50 Newark, Delaware 19714-0050 &tf^ 1 CO^ ^ QO' ^ ,<c,' 0^ ^ f^ C^ Page 1 of 83 H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 CERTIFICATION We, the undersigned, declare that this report provides an accurate evaluation of data obtained from this study. Reviewed by: (ji^^j(/^-^^ /PanlHinderliter.Ph.D. ' Postdoctoral Fellow Z^-Mf - if< Date Reviewed by; D.A.B.T.Date (^JjbLi^xLC- ^^cUtU______ // Judith C. Stadter. Ph.D., '/ Director S^-/!/)[/-2.60 / Issued by Study Director:______/! ^i/l<!L lb'kJA^^_________ CarolrfnIay.B.A. ^ Staff Scientist ^UP -hvU ^'t-'Of Date ^^ ^^ ^^ 1- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 TABLE OF CONTENTS Page CERTIFICATION........:................................................................................................................! LIST OF TABLES........................................................................................................................^ LIST OF FIGURES.......................................................................................................................4 LIST OF APPENDICES ...............................................................................................................5 STUDY INFORMATION.............................................................................................................6 STUDY PERSONNEL...................................................................................................................8 SUMMARY...................................................................................................................................^ INTRODUCTION........................................................................................................................11 MATERIALS AND METHODS ................................................................................................11 A. Test Substance and Positive Controls................................................................................11 B. Test Species .......................................................................................................................11 C. Animal Husbandry .............................................................................................................11 D. Quarantine and Pretest.......................................................................................................12 E. Study Design........................................................:.............................................................13 F. Assignment to Groups and Study Start..............................................................................13 G. Dosing Material Preparation and Administration..............................................................13 H. Body Weights.....................................................................................................................14 I. Mortality and Clinical Observations.................................................................................. 14 J. Collection and Analysis of Blood, Livers, and Fat ............................................................14 K.. Treatment of Fluorine Data................................................................................................15 L. Statistical Methods.............................................................................................................15 RESULTS AND DISCUSSION..................................................................................................16 A. m-Life Toxicology.............................................................................................................16 B. Liver Weights.....................................................................................................................16 C. Fluorine Data.....................................................................................................................17 CONCLUSIONS..........................................................................................................................18 RECORDS AND SAMPLE STORAGE ....................................................................................19 TABLES........................................................................................................................................20 FIGURES......................................................................................................................................27 APPENDICES ..............................................................................................................................38 Sanitized. Do.. not contain TSCACBI Company .3- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 LIST OF TABLES Page 1. MEAN BODY WEIGHTS...................................................................................................................................21 2. MEAN BODY WEIGPTT GAINS........................................................................................................................22 3. MEAN BODY AND LIVER WEIGHTS............................................................................................................. 23 4. MEAN BLOOD FLUORINE LEVELS...............................................................................................................24 5. MEAN BLOOD FLUORINE CONCENTRATION NORMALIZED TO DOSE ...............................................24 6. MEAN LIVER FLUORINE LEVELS.................................................................................................................25 7. MEAN LIVER FLUORINE CONCENTRATION NORMALIZED TO DOSE ................................................. 25 8. MEAN FAT FLUORINE LEVELS .....................................................................................................................26 9. MEAN FAT FLUORINE CONCENTRATION NORMALIZED TO DOSE...................................................... 26 LIST OF FIGURES Page 1. MEAN BODY WEIGHTS...................................................................................................................................28 2. COMPARISON OF MEAN RELATIVE LIVER WEIGHTS: TEST SUBSTANCE AND NEGATIVE 29 CONTROL................................................................................................................................................;.......... 3. COMPARISON OF MEAN RELATIVE LIVER WEIGHTS: TEST SUBSTANCE AND POSITIVE CONTROLS.........................................................................................................................................................30 4. MICROMOLAR EQUIVALENTS IN RAT BLOOD .........................................................................................31 5. NORMALIZED H-24943 AND POSITIVE CONTROL BLOOD AUCINF/D RESULTING FROM A 10-DAY ORAL GAVAGE ..................................................................................................................................33 6. MEAN LIVER FLUORINE CONCENTRATION NORMALIZED TO DOSE .................................................34 7. COMPARISON OF RELATIVE LIVER WEIGHT AND MEAN LIVER FLUORINE CONCENTRATION FOR H-24943 AND NEGATIVE CONTROL.................................................................. 35 8. MEAN FAT FLUORINE CONCENTRATION NORMALIZED TO DOSE...................................................... 36 9. COMPARISON OF MEAN BLOOD, MEAN LIVER, AND MEAN FAT FLUORINE CONCENTRATION NORMALIZED TO DOSE...............................................................................................37 Company Sanitized. Does not contain TSCA CB8 4- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 LIST OF APPENDICES Page A. INDIVIDUAL BODY WEIGHTS.......................................................................................................................39 B. INDIVIDUAL CLINICAL OBSERVATIONS...................................................................................................^ C. TERMS AND CALCULATIONS FACTORS INFLUENCING INTERPRETATION OF KINETIC ANALYSIS..........................................................................................................................................................65 D. INDIVIDUAL FLUORINE LEVELS IN BLOOD.............................................................................................. 69 E. INDIVIDUAL FLUORINE LEVELS IN LIVER................................................................................................ 76 F. INDIVIDUAL FLUORINE LEVELS IN FAT.................................................................................................... 80 Company Sanitized. Does not contain TSCA CBI -5- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats TEST SUBSTANCE: STUDY INFORMATION Substance Tested: Synonyms/Codes: I Haskell Number: 24943 J Composition: i Known Impurities: POSITIVE CONTROL: Substance Tested: Potassium perfluoroalkyi sulfonate Synonyms/Codes: H-24019 DuPont-6544 Haskell Number: 24019 Known Impurities: Unknown Company Sanitized. Doss not ecs-.tain S'SCA CBS -6- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 STUDY INFORMATION (Continued) POSITIVE CONTROL: Substance Tested: Octanoic acid, pentadecafluoro-, ammonium salt Synonyms/Codes. H-24020 C-8 Perfluorooctanoate, ammonium salt APFO Ammonium perfluorooctanoate Haskell Number: 24020 Composition: Known Impurities: Sponsor; E. I. du Pont de Nemours and Company Wilmington, Delaware 19898 U.S.A. Study Initiated/Completed: May 22, 2001 / (see report cover page) In-Life Initiated/Completed: May 22,2001 / August 23, 2001 ID Company Sanity. Does not contain TSCA CS1 7- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 STUDY PERSONNEL Study Director: Management: Primary Technician: Carol Finlay, B.A. Judith C. Stadler, Ph.D., D.A.B.T. James C. Mackay II Fluorine Data Analysis: Paul M. Hinderliter, Ph.D. Management: Matthew S. Bogdanffy, Ph.D., D.A.B.T. Toxicology Report Preparation: Wanda F. Dinbokowitz Laboratory Veterinarian: William Singleton, D.V.M., A.C.L.A.M ^Panysan; 'ooes-^^^ 8- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 SUMMARY The objective of this study was to evaluate the potential for H-24943, when administered by gavage, to be absorbed and to accumulate in a mammalian system. Two groups of 5 male Crl:CD(SD)IGSBR rats each were exposed to 1000 mg/kg/day of H-24943 for 10 consecutive days. Blood was collected from the orbital sinus of 5 rats (group I) approximately 2 hours after dosing on test days 1 and 5. Approximately 2 hours after the last dose, these rats were euthanized and blood, livers, and fat were collected. Blood was collected from the orbital sinus of the remaining 5 rats (group III) on test days 13, 24, and 52. These rats were euthanized on test day 94 and blood, livers, and fat were collected. Body weights and clinical signs were recorded on each day of dosing and then weekly during the recovery period. Additionally, a negative control, deionized water, and 2 positive controls, H-24019 (10 mg/kg/day) and H-24020 (20 mg/kg/day), were tested as described for H-24943. No deaths occurred. One rat dosed with the test substance exhibited black nasal discharge during the dosing period. Comparison of body weights was complicated by the fact that there was a difference in age on test day 1 between the rats dosed with the test substance and those dosed with the positive controls or negative control. This difference resulted in differences in mean body weights on test day 1. Accounting for the age difference at study start and the expected rate of body weight gain, the mean body weights and mean body weight gains of the rats dosed with H-24943 were comparable to the weights of the positive and negative control rats. The mean relative liver weights (liver/body weight) of rats dosed with the test substance, H-24943, were similar to the weights of the negative control rats on day 10 and day 94. The mean relative liver weight of rats dosed with the positive control H-24019 was 38% higher at day 10 than the negative control group. The mean relative liver weight of rats dosed with the positive control H-24020 was 88% higher on day 10 than the negative control group. By day 94, the mean relative liver weights of rats dosed with H-24019 and H-24020 were similar to the negative control group. A steady-state for fluorine levels in whole blood was not achieved during 10 consecutive days of dosing with 1000 mg/kg H-24943. An area under the curve (estimated to infinity) was calculated and normalized for fluorine content for the test substance and each positive control. The AUCINF/D for the fluorine component of the test substance, H-24943, was 1.62xl03 compared .to AUCINF/D values of5.22xl05 and 8.15xl04 for H-24019 and H-24020, respectively. The concentration of fluorine in the livers from rats dosed with the test substance, H-24943, was 47.7 pM equivalents on day 10 and 2.9 uM equivalents on day 94. On day 10, mean pM equivalent concentrations of fluorine in the livers from rats dosed with the positive control materials were approximately 100-fold (H-24019) and 18-fold (H-24020) greater than the fluorine concentration in livers from rats treated with the test substance. By day 94, the Company Sanitised-0Dooe-s. n_o.t contain TSCACBf -9- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats_____________ DuPont-6544 concentrations were approximately 450x and 6x the fluorine concentration in rats treated with H-24943. The fluorine concentration in the fat from rats dosed with the test substance was 41.3 uM equivalents on day 10. On day 94, the concentration was 6.6 uM equivalents. The fluorine concentration of the positive control H-24019 was approximately 5x higher than H-24943 on day 10. By day 94, the uM equivalent concentration of fluorine in the fat from rats dosed with the positive control H-24019 was approximately 2x higher than H-24943. The fluorine concentration of the positive control H-24020 was slightly higher on day 10 than the concentration in fat from rats dosed with the test substance. There was no detectable fluorine by day 94 in the fat from rats dosed with H-24020. The uM equivalents of fluorine in the liver and fat of animals dosed with the test substance were higher than levels in the blood. Under the conditions of this study, there was limited absorption and no retention of fluorine in the blood in rats dosed with H-24943. Administration of the test substance, H-24943, to male rats for 10 consecutive days resulted in some absorption and retention of fluorine in the liver; however, there was no evidence of increased relative liver weight, either at the end of dosing or following the recovery period. Fluorine levels in blood and liver in rats dosed with H-24943 were lower than the levels in rats dosed with the positive control materials, H-24019 and H-24020. Some retention of fluorine in the fat was evident. Fluorine levels in the fat from rats dosed with the test substance were lower than the levels in fat from rats dosed with the positive control H-24019 and were slightly lower on day 10 than the levels in fat from rats dosed with the positive control H-24020, but higher on day 94. Company Sanitized. Does not contain TSCA CBI 10- H-24943: Biopersistence Screening 10-DoseOral Gavage Study in Rats DuPont-6544 INTRODUCTION The objective of this study was to define the potential of H-24943 to be absorbed and to bioaccumulate in a mammalian system, as indicated by analytical determination of total fluorine in blood, liver, and fat. The test substance was compared to 2 positive controls that were materials previously shown to bioaccumulate in mammals. The daily dosage of 1000 mg/kg for the test substance was selected based on available toxicity data. The dosage of 1000 mg/kg was selected for the limit dosage for this project. The limit dosage of 1000 mg/kg was chosen for the study and was expected to produce less than a 10% difference in mean body weight over 10 days when compared to the negative control. MATERIALS AND METHODS A. Test Substance and Positive Controls The test substance, H-24943, was supplied by the sponsor as a cloudy yellow-amber liquid. The positive controls, H-24019 and H-24020, were supplied by the sponsor as white solids. The test substance and positive controls appeared to be stable under the conditions of the study. No evidence of instability, such as a change in color or physical state, was observed. B. Test Species Male Crl:CD(SD)IGSBR rats were received from Charles River Laboratories, Inc., Raleigh, North Carolina. The Crl:CD(SD)IGS BR rat was selected on the basis of extensive experience with this strain and its suitability with respect to longevity, hardiness, sensitivity, and low incidence of spontaneous diseases. C. Animal Husbandry 1. Housing Environment Rats were housed singly in stainless steel, wire-mesh cages suspended above cage boards. Animal rooms were maintained on an approximate 12-hour light/dark cycle (fluorescent light) and at a temperature of 23 1C and a relative humidity of 50 10%. 2. Feed and Water Tap water was provided adiibitum. All rats were fed PMI Nutrition International, Inc. Certified Rodent LabDiet 5002 chow. The feed is guaranteed by the manufacturer to meet specified nutritional requirements and to be free of specified contaminants. Company Sanitized. Does not contain TSCA CB1 H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats___________________________________DuPont-6544 3. Identification Each rat was assigned a unique identification number which was recorded on a card affixed to the cage. The last 3 digits of the number were tattooed on the tail of each rat. 4. Animal Health Monitoring Program As specified in the Haskell Laboratory animal health and environmental monitoring program, the following procedures are performed periodically to ensure that contaminant levels are below those that would be expected to impact the scientific integrity of the study: Water samples are analyzed for total bacterial counts, and the presence ofcoliforms, lead, and other contaminants. Feed samples are analyzed for total bacterial, spore and fungal counts. Samples from freshly washed cages and cage racks are analyzed to ensure adequate sanitation by the cagewashers. Certified animal feed is used, guaranteed by the manufacturer to meet specified nutritional requirements and not to exceed stated maximum concentrations of key contaminants, including specified heavy metals, aflatoxin, chlorinated hydrocarbons, and organophosphates. The presence of these contaminants below the maximum concentration stated by the manufacturer would not be expected to impact the integrity of the study. The animal health and environmental monitoring program is administered by the attending laboratory animal veterinarian. Evaluation of these data did not indicate any conditions that affected the validity of the study. D. Quarantine and Pretest Upon arrival at Haskell Laboratory, the rats were removed from shipping cartons and quarantined for 6 days. The rats were weighed 3 times during the pretest period and examined daily for any clinically apparent signs of disease or injury. The rats were observed daily for mortality and signs of illness, injury, or abnormal behavior. On the bases of acceptable body weight gains and freedom from clinically apparent signs of disease or injury, the rats were released from quarantine by the designee of the laboratory animal veterinarian. ^ Company Sanitized. Does not contain TSCA CBj 12- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 E. Study Design Substance Negative Control Deionized water Positive Controls H-24019 H-24020 Test Substance H-24943 Vehicle Not applicable Acetone/Corn Oil Acetone/Corn Oil Not applicable Dosage (mg/kg) 0 10 20 1000 Number of Animals 10 10 10 10 F. Assignment to Groups and Study Start After the quarantine period, the rats were selected on the bases of adequate body weight gain and freedom from any clinical signs of disease or injury. The selected rats were arbitrarily assigned to each group. After assignment to groups, each rat was housed individually. The rats were 7 or 8 weeks of age at the time of dosing. Dosing began on test day 1. Rats that were not assigned to the study were released for other laboratory purposes, or were sacrificed by carbon dioxide asphyxiation and discarded without pathology evaluation. G. Dosing Material Preparation and Administration 1. Test Substance H-24943 was dosed as received. The amount of test substance each rat received was based on the body weight collected on each day of dosing and the test substance density of 1.06 g/mL. The test substance was stirred on a magnetic stir plate throughout the dosing procedure to maintain homogeneity. 2. Positive Controls It was necessary to dissolve H-24019 and H-24020 in acetone before suspending them in corn oil. The ratio of acetone to corn oil was 20:80. The amount each rat received was based on the body weight collected on each day of dosing and the suspension concentration. The rats were dosed at a volume of 1 mL/100 g of body weight. The dosing preparations were stirred on a magnetic stir plate throughout the dosing procedure to maintain homogeneity. 3. Negative Control Deionized water was chosen as the negative control. The rats were dosed at a volume of 1 mL/100 g of body weight. These rats were dosed in a separate room from the rats dosed with the test substance or positive controls. Company Sanitized. Does not contain TSCA CBi - 13- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 H. Body Weights All rats were weighed on each day of dosing and weekly during the recovery period. I. Mortality and Clinical Observations Cage-site examinations to detect moribund or dead rats and abnormal behavior and appearance among rats were conducted at least once daily throughout the study. At every weighing, each rat was individually handled and examined for abnormal behavior and appearance. J. Collection and Analysis of Blood, Livers, and Fat At time points selected for blood sampling other than sacrifice days, approximately 1 mL of blood was collected into EDTA tubes from the orbital sinus of each rat. Rats designated for sacrifice were euthanized by carbon dioxide anesthesia and exsanguination and blood, livers, and fat were collected according to the following schedules: Group I I I III III III III Dosing Days 1-10 1-10 1-10 1-10 1-10 1-10 1-10 Tissue Collected Blood Blood Blood, Liver, and Fat Blood Blood Blood Blood, Liver, and Fat Sampling Time Test day 1 (2 hours post dosing) Test day 5 (2 hours post dosing) Test day 10 at sacrifice (2 hours post dosing) Test day 13 Test day 24 Test day 52 Test day 94 As much blood as possible was collected into EDTA tubes at sacrifice. The livers and fat were weighed. The fat weights were used only for the calculation of fluorine levels. The liver weights were used for the calculation of fluorine levels and for the calculation of liver weight relative to total body weight. The blood from all rats was refrigerated and the livers were frozen. The livers and fat were appropriately packaged and shipped refrigerated to Jackson Laboratory, Deepwater, New Jersey where they were analyzed for total fluorine. The liver and fat samples remained frozen while shipped. The total fluorine content of the samples was determined by using a Wickbold torch combustion method, followed by analysis with a fluoride ion selective electrode. The samples were decomposed or volatilized in the presence of wet oxygen and swept through an oxy-hydrogen flame in a closed quartz apparatus. The combustion products were collected in an aqueous absorbing solution and analyzed with a fluoride ion selective electrode. Kinetic analysis of the data received (ppm F m each sample) was performed by Haskell Laboratory personnel for evaluation of fluorine biopersistence. ^'"""ys.,^ ^'^nr^, - 14- k. ) ./--"^ y-" H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats K. Treatment of Fluorine Data _______ DuPont-6544 Since the test substance and positive control materials had different toxicity profiles, it was not possible to administer a uniform mg/kg dose for all test materials. Properly conducted kinetic comparisons in situations with varied doses required the use of a normalized dose. The dosenormalization was conducted in umolar units to accommodate test material molecular weight differences and was adjusted to an arbitrary normalized dose of 0.1 mmoles/kg. H-24019 and H-24020 were used as positive controls and a dosing diluent was used as the negative control. The percent of fluorine and molecular weight of the test substance and positive controls were used as provided by the sponsor. The measured total fluorine values were used as received (ppm F) from Jackson Laboratory. For the Wickbold torch method, the background fluorine level is 0.2 ppm. This is the limit of detection (LOD) of this method and was subtracted from each sample. The limit of quantification (LOQ) for this method is 0.5 ppm, and any values listed as less than 0.5 ppm were excluded from further treatment. Since the dosages and molecular weight for positive controls and test material differed, the data were converted from a mass to a molar basis. In addition, the data were standardized to a single reference dose to allow comparison between compounds of differing fluorine content. The doses were first converted from a mg of test material basis to millimoles of fluorine. Raw fluoride ion data (ppm F) were then normalized to a 0.1 millimole dose of active component. Finally the molar dosage and normalized concentration were combined to yield the umolar (uM) equivalents of active component in the tissue. Detailed calculations can be found in Appendix C. The uM equivalents can be compared across compounds provided the considerations listed in Appendix C are observed. Noncompartmental analysis was conducted on fluorine data derived from rats dosed with H-24943 and the positive controls using WinNonlin Version 3.1 software (Pharsight Corp, Mountain View, CA). WinNonlin software provided a means of computing derived pharmacokinetic parameters from experimental data. All analysis was calculated using the uM equivalent in the data. The maximum observed concentration in blood was Cmax (uM equivalent). Biopersistence was assessed by quantifying terminal elimination blood half-life (T%, days). The points included in determination of the T% were selected manually and included only points after apparent log-linear elimination was achieved. Internal exposure was determined by calculating the blood areaunder-the-curve (AUC). AUC, which is simply the integral of blood concentration over time, is the most common means for expressing internal dose. With the calculated half-life, the AUC can be extrapolated to infinity (AUCINF) to reflect the elimination of the compound. AUCINF normalized to dose (AUCINF/D) can be used to compare the relative exposures of different compounds and dosages. L. Statistical Methods Descriptive statistics (e.g. mean, standard deviation) were used. Company Sanitised. Does not contain TSCA CB! - 15- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 RESULTS AND DISCUSSION A. In-Life Toxicology (Tables 1-2, Figure 1, Appendices A-B) No deaths occurred. Black nasal discharge was observed during the dosing period in a rat dosed with the test substance. Hair loss observed in 2 rats dosed with the test substance was considered spurious. A swollen mouth was observed during the dosing period in a negative control rat. One negative control rat exhibited black nasal discharge during the recovery period. Hair loss was observed in rats dosed with H-24019 and H-24020, and a wound was observed in a rat dosed with H-24020. Ocular discharge, dark eyes, comeal opacity, enophthalmus, and exophthalmus observed in several rats are considered to be a result of orbital sinus bleeding. Comparison of body weights was complicated by the fact that there was a difference in age on test day 1 between the rats dosed with the test substance and those dosed with the positive controls or negative control. This difference resulted in differences in mean body weights on test day 1. The rats dosed with the test substance, H-24943, were older and heavier in weight than the positive and negative control rats. Accounting for the age difference at study start and the expected rate of body weight gain, the mean body weights and mean body weight gains of the rats dosed with H-24943 were comparable to the positive and negative control. B. Liver Weights (Table 3, Figures 2, 3, and 7) 1. Test Substance The mean relative liver weights (liver/body weight) of rats dosed with the test substance, H-24943, were similar to the weights of the negative control rats on day 10 and day 94. 2. Positive Controls The mean relative liver weight of rats dosed with one of the positive controls, H-24019, was 32% higher at day 10 than the liver weight of rats dosed with the test substance, H-24943. By day 94, the weights were similar. The mean relative liver weight of rats dosed with the other positive control, H-24020, was 80% higher at day 10 than the liver weight of rats dosed with H-24943. By day 94, the weights were similar. The mean relative liver weight of rats dosed with H-24019 was 38% higher at day 10 than the negative control group. By day 94, the mean relative liver weight of rats dosed with H-24019 was similar to the negative control group. The mean relative liver weight of rats dosed with Company Sanitized. Does not contain TSCA CB! - 16. H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats __ ________ H-24020 was 88% higher on day 10 than the negative control group. were similar. DuPont-6544 __ By day 94, the weights Therefore, liver weights of rats dosed with the test substance, H-24943, were not affected. C. Fluorine Data (Tables 4-9, Figures 4-9, Appendices C-F) 1. Factors Influencing Interpretation of Analysis The data used in the kinetic analysis were derived from a limited screen, and therefore several caveats and considerations are important. A couple of considerations of particular importance are (1) a single dosage was used and kinetics may or may not be linear, (2) the kinetics apply only to blood, (3) steady-state may not have been achieved, and (4) the sample size is low and may impact calculation of the terminal half-life. A more complete list of considerations is shown in Appendix C. 2. Positive Controls The positive controls were H-24019 and H-24020. The H-24019 and H-24020 normalized uM equivalents in rat blood continued to rise throughout the dosing period and may not have reached steady-state (Figures 4A and 4B). The Cmax for H-24019 was 541.45 50.37 uM equivalents (Mean SD) with a terminal half-life of 42.2 days. The Cmax for H-24020 was 1043.08 54.57 uM equivalents (Mean SD) witK a terminal half-life of 15.1 days. For each of the positive controls, blood was sampled at seven time points throughout the study, with only four of them occurring post-dose. The small sample size and analytical variability should be taken into account when using the derived terminal half-life for comparative purposes. The total internal exposure resulting from a normalized dose was described by AUCINF/D and was the basis for comparison between positive controls and the test substance. The AUCINF/D for the fluorine component was 5.22x105 for H-24019 and 8.15xl04 for H-24020. The concentrations of fluorine in the livers on day 10 from rats dosed with the positive control materials were 4802.15 and 866.67 uM equivalents for H-24019 and H-24020, respectively. By day 94, the concentrations were 1292.92 and 17.10 uM equivalents. The concentrations of fluorine in the fat on day 10 from rats dosed with the positive control materials were 194.46 and 57.25 uM equivalents for H-24019 and H-24020, respectively. By day 94, the concentration of fluorine in the fat from rats dosed with H-24019 was 15.38 uM . equivalents. There was no detectable fluorine by day 94 in the fat from rats dosed with H-24020. 3. Test Substance The H-24943 normalized pM equivalents in rat blood rose rapidly and did not reach steady-state (Figure 4C). The Cmax for H-24943 was 4.65 1.49 uM equivalents (Mean SD) with a terminal half-life of 19.7 days. Blood was sampled at seven time points throughout the study, with only four of them occurring post-dose. The small sample size and analytical variability t}^ Companv Sanifaed - 17- n^ Q^ntgin T3CA Cfll H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 should be taken into account when using the derived terminal half-life for comparative purposes. Since the day 24 and 94 values were below the limit of quantification, only two points were used in the half-life calculation. The total internal exposure resulting from a normalized dose was described by AUCINF/D and was the basis for comparison between H-24943 and the positive controls. The AUCINF/D for the fluorine component of H-24943 was 1.62 xl03 as compared to AUCINF/D values of5.22xl05 and 8.15xl04 for H-24019 and H-24020, respectively. Levels of total fluorine in livers from rats dosed with the test substance, H-24943, were lower than the levels in livers from rats dosed with the positive control materials. The total fluorine concentration in the liver from rats dosed with H-24943 was 47.7 uM equivalents at day 10 and 2.9 pM equivalents at day 94. For the positive control H-24019, the liver concentrations were approximately lOOx higher (day 10) and approximately 450x higher (end of study) than H-24943. For H-24020, the liver concentrations were approximately 18x higher (end of dosing) and 6x higher (end of study) than H-24943. Levels of total fluorine in fat from rats dosed with the test substance were lower than the levels in fat from rats dosed with the positive control H-24019. Fluorine levels in the fat from rats dosed with the test substance were slightly lower on day 10 than the levels in fat from rats dosed with the positive control H-24020 but higher on day 94. The fluorine concentration in the fat from rats dosed with the test substance was 41.3 uM equivalents on day 10 and 6.6 uM equivalents on day 94. The fluorine concentration in the fat from rats dosed with the positive control H-24019 was approximately 5x higher than H-24943 at day 10 and 2x higher on day 94. The fluorine concentration on day 10 in the fat from rats dosed with the test substance was slightly lower than the concentration in fat from rats dosed with the other positive control, H-24020, By day 94, there was no detectable fluorine the fat from rats dosed with H-24020. The uM equivalents of fluorine in the liver and fat of animals dosed with the test substance were higher than levels in the blood. CONCLUSIONS Rats dosed for 10 consecutive days with 1000 mg/kg H-24943 exhibited no mortality, body weight effects, or liver weight effects. One rat dosed with the test substance exhibited a clinical sign. A steady-state for fluorine in the blood was not achieved during the 10-day dosing period with the test substance. Dose-adjusted areas under the curve (AUCINF/D) for positive controls, H-24019 and H-24020, were approximately 320x and 50x the AUCINF/D for the test substance. Under the conditions of this study, there was limited absorption and no retention of fluorine in the blood following dosing with H-24943. Administration of the test substance, H-24943, to male rats for 10 consecutive days resulted in some absorption and retention of fluorine in the liver; however, there was no evidence of increased relative liver weight, either at the end of dosing or following the recovery period. Fluorine levels in blood and liver in rats dosed with . H-24943 were lower than the levels in rats dosed with the positive control materials, H-24019 tj and H-24020. Some retention of fluorine in the fat was evident. Fluorine levels in the fat from - Company Sanitized. Does not contain TSCA CBI - 18- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 rats dosed with the test substance were lower than the levels in fat from rats dosed with the positive control H-24019 and were slightly lower on day 10 than the levels in fat from rats dosed with the positive control H-24020, but higher on day 94. RECORDS AND SAMPLE STORAGE All original records will be retained at Haskell Laboratory, E. I. du Font de Nemours and Company, Newark, Delaware or at Iron Mountain Records Management, 200 Todds Lane, Wilmington, Delaware. Company Sanitized. Does not contain TSCA CB! | 19- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 TABLES Company Sanitized. Does not contain TSCA CBi .20- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats TABLE 1 MEAN BODY WEIGHTS (g) Test Days 1 2 3 4 5 6 7 8 9 10 17 24 31 38 45 52 59 66 67 73 80 87 94 Negative Control Deionized Water 189.6 195.5 204.6 211.7 222.1 225.0 235.2 242.3 248.3 257.0 293.3 325.5 359.3 380.9 407.4 420.5 436.4 - 452.8 472.6 489.6 497.7 511.7 Positive Controls H-24019 H-24020 184.4 184.1 189.2 187.8 199.5 197.8 206.4 204.8 216.2 212.5 222.0 216.0 229.4 233.7 223.6 226.9 240.4 246.5 234.2 243.0 290.1 313.3 297.4 338.6 348.4 381.9 370.8 404.9 403.2 434.4 422.8 460.6 439.6 483.1 - - 455.6 480.1 496.5 512.2 524.6 502.7 525.6 542.1 546.2 570.5 Indicates the animals were not weighed. DuPont-6544 Test Substance H-24943 254.5 259.6 269.6 276.1 285.4 293.1 299.3 302.5 310.1 316.5 363.8 392.1 420.3 442.6 469.4 495.9 504.2 521.0 - 528.1 546.8 566.7 580.4 Company S a n r t i i e d . Does "ot contain TSCAC85 21 - H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats TABLE 2 MEAN BODY WEIGHT GAINS (g) Test Days 1-5 5-10 1-10 Negative Control Deionized Water 32.5 34.9 67.4 Dosing Positive Controls H-24019 H-24020 31.8 28.4 30.3 30.5 62.1 58.9 DuPont-6544 Test Substance H-24943 30.9 31.1 62.0 Test Days 10-17 17-24 24-52 52-94 10-94 Negative Control Deionized Water 36.3 32.2 95.0 91.2 254.7 Recovery Positive Controls H-24019 H-24020 43.6 54.4 23.2 41.2 109.5 122.0 101.8 109.9 278.1 327.5 Test Substance H-24943 47.3 28.3 103.8 84.5 263.9 compa^^^18 not contain ^ACBI 22- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 Test Days 10 94 TABLES MEAN BODY AND LIVER WEIGHTS (g) DEIONIZED WATER (NEGATIVE CONTROL) Absolute Body Weight Liver Weight 258.8 10.814 511.7 17.868 Mean Relative Liver Weight (Liver/Body Weight) 0.042 0.035 Test Days 10 94 H-24019 (POSITIVE CONTROL) Absolute Body Weight____Liver Weight 243.9 14.205 524.6 19.296 Mean Relative Liver Weight (Liver/Body Weight) 0.058 0.037 Test Days 10 94 H-24020 (POSITIVE CONTROL) Absolute Body Weight____Liver Weight 243.1 19.174 570.5 19.590 Mean Relative Liver Weight (Liver/Body Weight) 0.079 0.034 Test Days 10 94 H-24943 (TEST SUBSTANCE) Absolute Body Weight____Liver Weight 306.3 13.394 580.4 19.512 Mean Relative Liver Weight (Liver/Body Weight) 0.044 0.034 ^ ^ - ^CACBI .23- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 TABLE 4 MEAN BLOOD FLUORINE LEVELS Test Days 1 5 10 13 24 52 94 Nega,tive Control Deio nized Water (ppm) 0.603 c I.IO3 c 0.53 c c Positive (controls H-24019 H-24020 (ppm) (ppm) 2.60 (0.1)" 9.40 (2.4) 31.32 (1.1) 74.92 (7.0) 68.00 (3.5) 61.76 (5.5) 53.98 (1.2) 29.52 (4.9) 39.62 (3.4) 11.18 (2.9) 23.56 (2.1) 12.60 (1.2) 2.26 (1.1) 0.85'1 (0.1) Test Substance H-24943 (ppm) 1.1 (0.6) 2.2 (0.4) 2.4 (0.7) 1.8 (0.1) c 0.7'1 (O.I) c a One of 5 values. Four of the values were below the limit of quantification (LOQ) or non-detectable. b Standard deviation is in parentheses. c All values were below the LOQ or non-detectable. d Mean of 2 of the 5 values. Three of the values were below the LOQ. TABLE 5 . MEAN BLOOD FLUORINE CONCENTRATION NORMALIZED TO DOSE Positive Controls Test Substance Test H-24019 H-24020 H-24943 Days____jiM F Equivalents___pM F Equivalents_____uM F Equivalents 1 36.92 (I.I)3 66.67 (17.5) 2.0 (1.3) 5 478.77 (17.6) 541.45 (50.4) 4.3 (0.8) 10 1043.08 (54.6) 446.09 (39.7) 4.6 (1.5) 13 827.38 (19.0) 212.46 (35.6) 24 606.46 (53.1) 79.57 (20.8) 52 359.38 (32.7) 14.93 (7.7) 94 190.77 (19.2) 4.711' (0.5) 3.4 (0.2) c 1.01' (0.2) c a Standard deviation is in parentheses. b Mean of 2 of the 5 values. Three of the values were below the limit of quantification (LOQ). c All values were below the LOQ. -\ Compos ^^^'nrsc.cs/ -24- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 TABLE 6 MEAN LIVER FLUORINE LEVELS Test Negative Control Deionized Water Positive Controls H-24019 H-24020 Test Substance H-24943 D1a0ys____0_.9(0pp(0m.2))3_____3_1_2.(3p4p(m19)._7)_____11_9(.p80pm(3.)5_) _______(p22p.4m()3.4) 94_____0.78 (0.0) 84.24 (7.4)_____2.56 (1.3)_______1.6 (0.2) a Standard deviation is in parentheses. TABLE 7 MEAN LIVER FLUORINE CONCENTRATION NORMALIZED TO DOSE Positive Controls Test Substance Test H-24019 H-24020 H-24943 Days____uM Equivalents_____)-iM Equivalents 10 4802.15 (303.8)3 866.67 (25.5) uM Equivalents 47.7 (7.2) _94______1292.92 (114.1)______17.10 (9.8)_________2.9 (0.4) a Standard deviation is in parentheses. ^^San,^"^^^con^TSS .25- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 TABLE 8 MEAN FAT FLUORINE LEVELS Negative Control Positive Controls Test Substance Deionized Test Water H-24019 H-24020 H-24943 T O a 1 2 . 8 4 (1.1)19.4 Days_____(ppm)________(ppm)________(ppm) (Ppm) (I.?)"8.10 (5.3) (0.9) 94_______^__________1.20 (0.3)_______^___________3.3 a All values were below the limit of quantification (LOO) or non-detectable. b Standard deviation is in parentheses. TABLE 9 MEAN FAT FLUORINE CONCENTRATION NORMALIZED TO DOSE Positive Controls Test Substance Test H-24019 H-24020 H-24943 Days_____]iM Equivalents_____uM Equivalents_____uM Equivalents 10 194.46 (26.8)3 57.25 (7.8) 41.3 (11.3) 94_______15.38 (4.1)__________^___________6.6 (1.9) a Standard deviation is in parentheses. b All values were non-detectable. Company Sanitized. Does nontf cont. ai. n TSCA CBf 26- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 FIGURES --as'-' Company Sanitized. Does not contain TSCACB1 27- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats FIGURE 1 MEAN BODY WEIGHTS (g) -28- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats FIGURE 2 COMPARISON OF MEAN RELATIVE LIVER WEIGHTS: TEST SUBSTANCE AND NEGATIVE CONTROL a. S N (0 0. 0 0 (C U) o 3 --} 01 0 > 0 -BB- Test Days -29- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats FIGURE 3 COMPARISON OF MEAN RELATIVE LIVER WEIGHTS: TEST SUBSTANCE AND POSITIVE CONTROLS -3 N (0 Q. 0 0 <o w 3 0 t5a'. -V\) 0 > -e- Test Days -30- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 FIGURE 4 MICROMOLAR EQUIVALENTS IN RAT BLOOD A. Normalized Rat Blood H-24019 pH Equivalents Resulting from a 10-Day Oral Gavage 1200 Micromolar (pM) equivalents of H-24019 (positive control) in rat blood resulting from a 10-day oral gavage exposure. Values are means and error bars are standard deviation. R^ y Micromolar (uM) equivalents ofH-24020 (positive control) in rat blood resulting from a 10-day oral gavage exposure. Values are means and error bars are standard deviation. Company Sanitized. Doas not contain TSCA CBi -31 - H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 C. Normalized Rat Blood H-24943 uM Equivalents Resulting From a 10-Day Oral Gavage Limit of Quantification / 10 20 30 40 50 60 70 80 90 100 Time(days) Micromolar (uM) equivalents of H-24943 (test substance) in rat blood resulting from a 10-day oral gavage exposure. Values are means and error bars are standard deviation. Company Sanitized. Does no? contain TSCA CB! 32- ^ H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats 600000 FIGURE 5 NORMALIZED H-24943 AND POSITIVE CONTROL BLOOD AUCINF/D RESULTING FROM A 10-DAY ORAL GAVAGE 500000 400000 300000 200000 100000 BAUCINF/D H-24020 8.15E+04 H-249 1.62E 33- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats FIGURE 6 MEAN LIVER FLUORINE CONCENTRATION NORMALIZED TO DOSE Day 10 .34. Day 94 ^u H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats FIGURE 7 COMPARISON OF RELATIVE LIVER WEIGHT AND MEAN LIVER FLUORINE CONCENTRA FOR H-24943 AND NEGATIVE CONTROL Day 10 Day 94 Day 10 Da D Deionized Water (Negative Control) B H-24943 (Test Substance) -35- ^ H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats FIGURE 8 MEAN FAT FLUORINE CONCENTRATION NORMALIZED TO DOSE CT" -d U. 36- -^ H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats FIGURE 9 COMPARISON OF MEAN BLOOD, MEAN LIVER, AND MEAN FAT FLUORINE CONCENTRATION NORMALIZED TO DOSE 6000 5000-r " 4000 3000 3 0" Id '5. 2000 Blood / Liver / Fat Blood / Liver / Fat -37- Blood / Liver H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 APPENDICES Company Sanitized. Does not contain TSCA CBI 38- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 APPENDIX A Individual Body Weights Company Sanitized. Does not contain TSCA CBJ 39- H-24943; Biopersistence Screening 10-DoseOral Gavage Study in Rats INDIVIDUAL BOD^ WEIGHTS ABBREVIATIONS: FD SD - found dead sacrificed by design EXPLANATORY NOTES DuPont-6544 c^mysa"?M^''o".a,,,T8C,CO, 40 H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats ANIMAL NUMBER 646932 646933 646934 646935 646936 Day 1 197.1 191.9 182.6 203.0 196.8 Day 2 202 .1 195.4 188.9 208.9 201.4 DEIONIZED WATER (NEGATIVE CONTROL) INDIVIDUAL BODY WEIGHTS (g) OF MALE RATS GROUP I Day 3 TEST DAY Day 4 Day 5 Day 6 212 .9 205 .0 195 .4 218 .0 208 .6 221.5 213 .2 207.0 228.1 204.0 233 .6 226 .5 213 .9 239 .7 216 .7 234.1 219.3 215.9 243 .0 218.6 ANIMAL NUMBER 646932 646933 646934 646935 646936 Day 10 264.7 254.4 248.6 278.0 248.1 SD test day 10 SD test day 10 SD test day 10 SD test day 10 SD test day 10 TEST DAY . Day 7 242.4 232 .9 224.2 257.2 229.9 -41 - H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DEIONIZED WATER (NEGATIVE CONTROL) INDIVIDUAL BODY WEIGHTS (g) OF MALE RATS GROUP III ANIMAL TE ST DAY NUMBER Day 1 Day 2 Day 3 Day 4 Day 5 Day 6 Day 7 D 646937 184. 5 186.8 199.0 205.9 213. 6 221. 0 229 .6 23 646938 163. 7 173.8 180.9 189.3 200. 6 204. 1 215 .9 22 646939 193. 3 204.2 214.6 221.1 234. 4 240. 7 248 .9 2 646940 191. 5 197.1 203.8 209.8 218. 5 222. 1 229 .3 2 646941 191. 6 195.9 207.9 216.8 223. 1 231. 2 241 .3 2 ANIMAL TEST DAY NUMBER Day 10 Day 17 Day 24 Day 31 Day 38 Day 45 Day 52 Da 646937 249.8 282.6 322.3 366.1 395.0 425.0 443.9 4 646938 237.2 270.4 296.5 325.8 342.9 367.5 373.7 3 646939 277.9 329.7 372.8 415.2 443.2 475.2 484.1 5 646940 244.5 271.5 291.3 317.9 332.8 356.3 367.0 3 646941 266.3 312.4 344.8 371.6 390.7 412.8 433.6 4 0 j V 1 ANIMAL NUMBER Day 73 Day 80 Day 87 TEST DAY Day 94 (0 a ss 646937 5' 646938 (0 0. 646939 S S 646940 646941 W 504.3 417.4 540.1 407.3 493.7 520.5 431.0 568.9 424.0 503.5 533.7 432.1 578.1 430.8 513.8 545.7 450.6 586.9 449.4 525.8 SD test day 94 SD test day 94 SD test day 94 SD test day 94 SD test day 94 5 (7 0 (3J, T 5" i-tji 0 > <3 -42- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats H-24019 (POSITIVE CONTROL) INDIVIDUAL BODY WEIGHTS (g) OF MALE RATS GROUP I ANIMAL TEST DAY NUMBER Day 1 Day 2 Day 3 Day 4 Day 5 Day 6 Day 7 D 646910 646911 646912 646913 646914 184 .3 184 .4 180 .5 186 .4 185 .5 185.3 187.1 185.9 189.7 189.6 197.3 195.7 195.9 198.1 198.8 204 .8 202 .0 202 .5 205 .7 203 .8 212 .7 211 .7 211 .9 221 .8 215 .4 216 .3 217 .2 217 .7 225 .0 222 .7 226.4 225.2 227.2 228.6 226.7 ANIMAL NUMBER 646910 646911 646912 646913 646914 Day 10 241 .1 236 .6 241 .9 257 .0 243 .0 SD test day 10 SD test day 10 SD test day 10 SD test day 10 SD test day 10 TEST DAY -43- ^y H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats H-24019 (POSITIVE CONTROL) INDIVIDUAL BODY WEIGHTS (g) OF MALE RATS GROUP III ANIMAL TE ST DAY NUMBER Day 1 Day 2 Day 3 Day 4 Day 5 Day 6 Day 7 D 646915 182 .7 188 .3 200 .1 206 .3 211 .1 220. 6 225.8 2 646916 185 .0 192 .7 205 .5 211 . 6 220 .0 226. 9 239.5 2 646917 187 .8 194 .3 205 .8 210 .4 221 .8 227. 5 233.9 2 646918 175 .5 181 .9 189 .8 197 .6 207 .0 212. 4 220.2 2 646919 192 .0 196 .9 207 .9 218 .9 228 .3 234. 0 240.8 2 ANIMAL TE ST DAY NUMBER Day 10 Day 17 Day 24 Day 31 Day 38 Day 45 Day 52 Da 646915 240 .4 283 .4 302.9 326 .7 337 .9 371.1 399.7 4 646916 257 .0 310 .0 334.4 383 .2 411 .9 450.3 469.8 4 646917 250 .2 280 .3 308.2 339 .7 362 .8 387. 9 409.0 4 646918 237 .1 270 .6 292.7 323 .2 350 .8 379.4 396.7 4 646919 260 .2 306 .1 328.5 369 .2 390 .4 427.3 439.0 4 ANIMAL NUMBER 646915 646916 646917 646918 646919 Day 73 461 .2 544 .2 457 .5 448 .1 489 .5 Day 80 475 .0 565 .5 464 .7 472 .5 504 .8 Day 87 498 .1 578 .5 479 .5 480 .8 524 .2 TEST DAY Day 94 513. 3 590. 9 495. 7 500. 3 522. 9 SD test day 94 SD test day 94 SD test day 94 SD test day 94 SD test day 94 -44- ^ H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats ANIMAL NUMBER 646921 646922 646923 646924 646925 Day 1 204.5 178.3 185.9 188.7 172.2 Day 2 204.9 180.9 185.1 194.6 176.0 H-24020 (POSITIVE CONTROL) INDIVIDUAL BODY WEIGHTS (g) OF MALE RATS GROUP I Day 3 TEST DAY Day 4 Day 5 Day 6 215.7 188.9 196.0 209.5 183.9 222.2 197.4 202.3 212.6 193.4 233.3 197.7 203.5 230.4 202.7 238.8 202.4 204.9 225.9 202.7 ANIMAL NUMBER 646921 646922 646923 646924 646925 Day 10 262.6 218.6 240.6 259.1 234.8 SD test day 10 SD test day 10 SD test day 10 SD test day 10 SD test day 10 0 i i r ^ (ii a ? Ul 0 n* 0 a" ^ ^P. ,, ,,,- ... .- 0 ,.,,.,,-., TEST DAY -45-., .. . Day 7 244.3 202.4 219.1 236.2 215.1 H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats ANIMAL NUMBER 646926 646927 646928 646929 646930 Day 1 182 .0 172 .2 190 .4 178 .6 187 .9 Day 2 187 .8 178 .8 195 .3 182 .5 191 .6 H-24020 (POSITIVE CONTROL) INDIVIDUAL BODY WEIGHTS (g) OF MALE RATS Day 3 . GROUP III TEST DAY Day 4 Day 5 Day 6 197 .0 191 .1 204 .5 190 .7 201 .0 203.6 195.1 212 .3 198.3 210.4 206.0 206.6 222.1 203.5 219.6 213.2 213.2 230.1 204.2 224.4 Day 7 222 .9 218 .2 236 .1 214 .6 226 .7 ANIMAL TEST DAY NUMBER Day 10 Day 17 Day 24 Day 31 Day 38 Day 45 Day 52 D 646926 646927 646928 646929 646930 0 1-0 ANIMAL NUMBER (A te 3 646926 <K0T 646927 p. 646928 0 646929 0 646930 <B b) 3 5, r> 0 3 nT 3' V) 0 > 0 V 250 .8 243 .3 254 .1 225 .5 240 .3 Day 73 535 .3 551 .2 540 .3 502 .6 498 .5 297 .2 306 .2 310 .6 280 .9 292 .3 Day 80 551 .9 574 .5 557 .0 509 .6 517 .6 349 .3 355 .1 349 .4 323 .2 316 .2 Day 87 564 .4 541 .3 568 .3 526 .7 530 .3 383.8 404.0 392.8 376.0 352.7 412.1 424.8 422.4 396.8 368.4 432.1 463 .2 443.5 426.4 406.8 TESTDAY Day 94 580.6 602.5 581.9 544.0 543.4 SD test day 94 SD test day 94 SD test day 94 SD test day 94 SD test day 94 -46- 467 .8 484 .0 464 .8 450 .2 436 .0 H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats H-24943 (TEST SUBSTANCE) INDIVIDUAL BODY WEIGHTS (g) OF MALE RATS GROUP I ANIMAL TEST DAY NUMBER Day 1 Day 2 Day 3 Day 4 Day 5 ' Day 6 Day 7 D 648927 249.1 252.6 265.4 272.2 277.6 289.1 295.1 3 648928 253.4 254.5 265.6 273.1 282.3 285.1 294.7 3 648929 261.0 262.0 271.0 275.8 284.7 287.7 285.3 3 648930 240.7 246.9 259.8 264.3 276.0 277.4 277.3 2 648931 255.0 258.8 274.2 274.8 287.5 288.3 301.5 3 ANIMAL NUMBER 648927 648928 648929 648930 648931 0 0 ^ 0) to e> 1 0 0 (6 (A g 0 0 s 5" 5 S S Day 10 316.6 317.4 304.3 275.0 318.4 SD test day 10 SD test day 10 SD test day 10 SD test day 10 SD test day 10 TEST DAY -47- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats H-24943 (TEST SUBSTANCE) INDIVIDUAL BODY WEIGHTS (g) OF MALE RATS GROUP III ANIMAL TE;3T DAY NUMBER Day 1 Day 2 Day 3 Day 4 Day 5 Day 6 Day 7 D 648932 246. 2 255. 3 265.8 271.1 277. 3 297. 2 293.5 29 648933 254. 7 266. 3 270.4 278.5 287. 4 295. 0 307.8 30 648934 264. 1 269. 9 280.1 288.3 299. 8 308. 5 317.9 32 648935 259. 0 263. 1 270.4 281.4 285. 9 297. 1 309.2 30 648936 261. 7 266. 4 273.7 281.2 295. 0 305. 6 310.9 31 ANIMAL TEST DAY NUMBER Day 10 Day 17 Day 24 Day 31 Day 38 Day 45 Day 52 Da 648932 311.2 352.8 387.8 410.4 431.4 453.0 477.5 47 648933 326.6 368.3 391.7 428.1 450.4 472.2 499.7 51 648934 335.2 378.7 413.7 451.6 479.1 517.0 546.8 55 648935 325.7 351.1 377.7 402.4 425.0 448.8 481.6 48 648936 334.9 367.9 383.6 408.8 427.3 456.2 474.1 48 ANIMAL NUMBER 648932 648933 648934 648935 648936 Day 73 515 .0 541 .9 563 .4 515 .7 504 . 6 Day 80 535. 3 563. 5 585. 8 535. 1 514. 1 Day 87 556 .4 586 .7 605 .0 555 .8 529 .4 TEST DAY Day 94 576. 9 600. 3 609. 0 572. 9 543. 1 -48. H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 APPENDIX B Individual Clinical Observations Company Sanitized. Does not contain TSCA CBI 49- ^.sl..^ H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DEIONIZED WATER (NEGATIVE CONTROL) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS GROUP I Animal 646932 646933 0 I 646&4 < W a S N ra p. a o ro V) 3 0 Observation General observation. No Abnormality Detected Eye Observations, Exophthalmus, Left Eye Observations, Bled via Orbital for Clin Path, Left Discharge, Eye left. Black Sacrificed by design General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Dark, Left Discharge, Eye left. Black Swollen Observations, Mouth Sacrificed by design . General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Discharge, Eye left. Black Sacrificed by design Days 1 2-9 1,5 10 10 1-5 1,5 6-10 6-7 10 10 1-5,10 1,5 6-9 10 3 Ul 0 f 0 CO -50- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Animal 646935 646936 DEIONIZED WATER (NEGATIVE CONTROL) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS GROUP I (Continued) Observation . Days General observation. No Abnormality Detected 1-10 Eye Observations, Bled via Orbital for Clin Path, Left 1,5 Sacrificed by design 10 General observation. No Abnormality Detected 1-10 Eye Observations, Bled via Orbital for Clin Path, Left 1,5 Sacrificed by design 10 ? I tu .3 0) S 0. I 0 3' ^ g 0 CD -51 H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Animal 646937 646938 I 0 CO 3.646939 3" o a3 } ) i DEIONIZED WATER (NEGATIVE CONTROL) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS Observation GROUP III Days General observation. No Abnormality Detected 1-10,17-94 Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Sacrificed by design 13 24,52 94 General observation. No Abnormality Detected 1-10,17-45 Eye Observations, Enophthalmus, Right Eye Observations, Exophthalmus, Right 80-94 52-73 Eye Observations, Bled via Orbital for Clin Path, Bilateral Eye Observations, Bled via Orbital for Clin Path, Right Eye Observations, Corneal Opacity, Right Discharge, Eye right. Black Sacrificed by design 13,52 24 80-94 59 94 General observation, No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Eye Observations, Bled via Orbital for Clin Path, Sacrificed by design Left Right 1-10,17-94 13 24,52 94 -52- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Animal 646940 646941 DEIONIZED WATER (NEGATIVE CONTROL) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS Observation GROUP III (Continued) Days General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Discharge, Nose, Black Sacrificed by design General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Bilateral Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for din-Path, Right Sacrificed by design 1-10,17-80,94 13 24,52 87 94 1-10,17-94 52 13 24 94 -53 H-24943: Biopersistence Screening 10-DoseOral Gavage Study in Rats Animal 646910 646911 646912 646913 646914 H-24019 (POSITIVE CONTROL) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS GROUP I Observation Days General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Sacrificed by design General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations. Dark, Left , Sacrificed by design General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Sacrificed by design General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Sacrificed by design General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Sacrificed by design 1-10 1,5 10 1,10 1.5 2-9 10 1-10 1 5 10 1-10 1,5 10 1-10 1,5 10 -54- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Animal 646915 646916 646917 H-24019 (POSITIVE CONTROL) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS Observation GROUP III Days General observation. No Abnormality Detected 1,17-38,73 Eye Observations, Bled via Orbital Discharge, Eye right. Red Hair Loss, Forelimb, Right for din Path, Left 13,24,52 2-10 80 Hair Loss, Forepaw, Right Sacrificed by design General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Bilateral Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Sacrificed by design . 45-67,87-94 94 1-10,17-94 24 13 52 94 General observation, No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Sacrificed by design 1-10,17-94 13,24 52 94 -55- x^y H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Animal 646918 646919 H-24019 (POSITIVE CONTROL) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS Observation GROUP III (Continued) . Days General observation. No Abnormality Detected 1-10,17-94 Eye Observations, Bled via Orbital Eye Observations, Bled via Orbital Sacrificed by design for Clin Path, for Clin Path, Left Right 13,24 52 94 General observation. No Abnormality Detected 1-10,17-94 Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Sacrificed by design 13,24 52 94 -56- V H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats H-24020 (POSITIVE CONTROL) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS GROUP I Animal 646921 646922 646923 Observation General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Dark, Left Sacrificed by design General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Sacrificed by design General observation. No Abnormality Detected Eye Observations, Exophthalmus, Left Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Eye Observations, Dark, Left Sacrificed by design Days 1,5-10 1,5 2-4 10 1-10 1 5 '10 1,7-10 4-6 1 5 2-6 10 -57. H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Animal 646924 646925 H-24020 (POSITIVE CONTROL) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS GROUP I (Continued) Observation Days General observation. No Abnormality Detected 1-10 Eye Observations, Bled via Orbital for Clin Path, Left 1 Eye Observations, Bled via Orbital for Clin Path, Right 5 Sacrificed by design 10 General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Sacrificed by design 1-10 1 5 10 -58- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Animal 646926 646927 6ffl5928 O ^ r H-24020(POSITIVE CONTROL) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS Observation GROUP III Days General observation, No Abnormality Detected 1-10,17-94 Eye Observations, Bled via Orbital for Clin Path, Bilateral Eye Observations, Bled via Orbital for Clin Path, Left Sacrificed by design 52 13,24 94 General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Bilateral Eye Observations, Bled via Orbital for Clin Path, Left Sacrificed by design 1-10,17-94 52 13,24 94 General observation, No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Sacrificed by design Left 1-10,17-94 13,24,52 94 -59- ^ H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Animal 646929 646930 H-24020(POSITIVE CONTROL) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS Observation GROUP III (Continued) Days General observation, No Abnormality Detected 1-10,17-31 Eye Observations, Bled via Orbital for Clin Path, Left 13,24 Eye Observations, Bled via Orbital for Clin Path, Right 52 Hair Loss, Forelimb, Bilateral Hair Loss, Neck, Left 52-73 73 Hair Loss, Neck, Ventral Wound, Superficial, Face Sacrificed by design 38-67,80-94 59 94 General observation. No Abnormality Detected . 1-10,17-94 Eye Observations, Bled via Orbital Eye Observations, Bled via Orbital Sacrificed by design for Clin Path, for Clin Path, Left Right 13,24 52 94 -60- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats H-24943 (TEST SUBSTANCE) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS GROUP I Animal 648927 648928 648929 ) I Observation ' General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Sacrificed by design General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Sacrificed by design General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Discharge, Eye left. Black Sacrificed by design Days 1-10 5 1 10 1-10 5 1 10 1-5,7-10 5 1 5 10 -61 H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Animal 648930 648931 H-24943 (TEST SUBSTANCE) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS GROUP I (Continued) Observation Days General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left 1-4,7-10 5 Eye Observations, Bled via Orbital for Clin Path, Right 1 Discharge, Nose, Black 5-6 Sacrificed by design 10 General observation. No Abnormality Detected 1-10 Eye Observations, Bled via Orbital for Clin Path, Left 5 Eye Observations, Bled via Orbital for Clin Path, Right 1 Sacrificed by design 10 -62- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Animal 648932 648933 648934 H-24943 (TEST SUBSTANCE) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS Observation GROUP III Days General observation, No Abnormality Detected 1-10,17-94 Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right 13 24,52 Sacrificed by design 94 General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Sacrificed by design 1-10,17-94 13 24,52 94 General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Eye Observations, Corneal Opacity, Right Sacrificed by design 1-10,17-52,94 13 24, 52 59-87 94 -63- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Animal 648935 648936 H-24943 (TEST SUBSTANCE) INDIVIDUAL CLINICAL OBSERVATIONS IN MALE RATS Observation GROUP III (Continued) Days General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Hair Loss, Porepaw, Bilateral Sacrificed by design 1-10,17-24,66-94 13 24,52 31-59 94 General observation. No Abnormality Detected Eye Observations, Bled via Orbital for Clin Path, Bilateral Eye Observations, Bled via Orbital for Clin Path, Left Eye Observations, Bled via Orbital for Clin Path, Right Hair Loss, Abdomen, Medial 1-10,17 52 '13 24 24-45 Hair Loss, Abdomen, Right Sacrificed by design 52-94 94 -64- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 APPENDIX C Terms and Calculations Factors Influencing Interpretation of Kinetic Analysis 65- Company SamteeA. Uoas ^ico^ini 'ISGA.C81 H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 Terms: Active % Active Mol Wt Active Formulation Dose % F in Active Mol Wt F TERMS AND CALCULATIONS Fluorine containing compound The % of formulation that is made up of fluorine containing components The molecular weight of the fluorine containing components (g/mole) The mg of formulation given per kg of animal body weight The % fluorine in the fluorine containing components of the formulation (weight basis) The molecular weight of fluorine g/mol Compound Calculations: Dose Active (mg/kg) The mg of fluorine containing compound administered per kg of animal body weight. =(% active/I 00) x Formulation Dose Dose Active (mmole/kg) The mmole of fluorine containing compound administered per kg of animal body weight = dose [mg/kg] / Mol Wt Active [mg/mmol] Dose F (mg/kg) The mg Fluorine administered per kg of animal body weight = (% F in active/I 00) x Dose Active [mg/kg] Dose F (mmol/kg) The mmole of fluorine administered per kg of animal body weight = Dose F [mg/kg] / Mol Wt F [mg/mmol] Molar Ratio (Active/F) The moles of fluorine containing compound per mole of fluorine = Dose Active [mmol/kg] / Dose F [mmol/kg] Formulation Dose Normalization Factor The formulation dose that would be required to administer the amount of active needed to achieve the normalized dose = (Normalized dose of Active [mmol/kg] / Dose Active [mmol/kg]) x Formulation Dose Company Sanitised. Does not contain TSCA CBI -66- H-24943: Biopersistence Screening 10-DoseOralGavage Study in Rats DuPont-6544 TERMS AND CALCULATIONS Individual Animal Measurement: ppmF The ppm fluoride measured Individual Animal Calculations: ppm F minus Bkg 0.2 ppm The ppm fluoride measured minus the background fluoride measured in control animal. In this case the value was established at 0.2 ppm. ppmF normalized to 0.1 mmol/kg Dose The ppm fluoride minus background that would be expected if the active dose was 0.1 mmol/kg instead of the actual active dose. This assumes linearity between administered dose and blood fluorine levels, but is needed because different doses of active were used in the study. = (0.1 [mmol/kg] / Active dose [mmol/kg]) x (ppm F in blood minus background) umolar equivalents of active The umolar [umol/L] concentration of fluorine containing compound based on the ppm fluorine, normalized to 0.1 mmol/kg active dose. This assumes that all fluorine is derived from the fluorine-containing component in the formulation. Note: 1 ppm - 1 mg/L = (Normalized ppm [mg/L] fluorine / Mol Wt F [mg/mmol]) x molar ratio active/F [mmol active/mmol F] x 1000 umoVmmol Company sanded. Does "ot contain TSCACBI 67- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 FACTORS INFLUENCING INTERPRETATION OF KINETIC ANALYSIS Considerations: - The data used for kinetic analysis was from a limited screen and extrapolation should be done cautiously. - Sample size is low - Analytical data used without validation - Steady-state not reached - Terminal phase may not be reached - Some compounds are mixtures offluorinated compounds - Different active and formulation doses were used - Different vehicles were used to deliver formulations - Each compound may have very different potency for producing toxicity Assumptions: (May or may not be justified in all cases) - Fluorine concentrations are linear with respect to dose - Analytical method is appropriate for all types of compounds - Elimination kinetics can be determined based on total fluorine rather than on concentrations of individual components - Background Fluorine is 0.2 ppm - % F data is the % Fluorine of the active (Fluorine containing component(s) in the formulation) - Molecular weight is the molecular weight of the active component in the formulation Company Sanm.ed.Does not contain TSCACBf -68- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 APPENDIX D Individual Fluorine Levels in Blood Company Sanitized. Does not contain i SCA CBI 69- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Data for H-240119 DuPont-6544 Given: MoPWt. Active (g/imole): 497 Formulation Dose (mg/kg): 10 %Ac tive (F Containing) in Fcirmulation: 100 % F in Active: 65 Mol Wt. F (g/mc)!): 19 Calculatied Values: Dose Active (mg/k{?): Dose Active (mmol e/kg): Dose F (mmol/kg): 10 0.020 0.342 Molar Ratio (Ad;ive/F): Dose F (mg/kg): 0.059 6.5 Rat Number Test Day Sample ppm F in Blood Group I 646910 1 2.6 646911 1 2.7 646912 1 2.5 646913 1 2.6 646914 1 2.6 ppm F in Blood Minus Bkg 0.2 ppm 2.4 2.5 2.3 2.4 2.4 ppm F in Blood Normalized to 0.1 mmoles/kg Dose 11.93 12.43 11.43 11.93 11.93 umolar Equivalents of Active in Blood 36.92 38.46 35.38 36.92 36.92 Group I 646910 5 646911 5 646912 5 646913 5 646914 5 30.1 30.3 31.3 32.2 32.7 29.9 30.1 31.1 32.0 32.5 148.60 149.60 154.57 159.04 161.53 460.00 463.08 478.46 492.31 500.00 Group I 646910 10 71.5 646911 10 70.5 646912 10 66.9 646913 10 62.5 646914 10 68.6 71.3 70.3 66.7 62.3 68.4 354.36 349.39 331.50 309.63 339.95 1096.92 1081.54 1026.15 958.46 1052.31 ) Company San^taeti. Does not contain TSCA CB8 70- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Rat Number Test Day Sample ppmF in Blood Group III 646915 13 646916 13 646917 13 646918 13 646919 13 ^ 55.0 55.3 52.2 53.8 53.6 ppm F in Blood Minus Bkg 0.2 ppm 54.8 55.1 52.0 53.6 53.4 ppm F in B lood Normalize!dto 0.1 romoles>/kg Dose 272.36 273.85 258.44 266.39 265.40 DuPont-6544 umolar Equivalents of Active in Blood 843.08 847.69 800.00 824.62 821.54 Group III 646915 24 646916 24 646917 24 646918 24 646919 24 Group III 646915 52 646916 52 646917 52 646918 52 646919 52 38.9 43.8 37.6 35.4 42.4 23.2 20.3 25.9 24.9 23.5 38.7 43.6 37.4 35.2 42.2 23.0 20.1 25.7 24.7 23.3 192.34 216.69 185.88 174.94 209.73 114.31 99.90 127.73 122.76 115.80 595.38 670.77 575.38 541.54 649.23 353.85 309.23 395.38 380.00 358.46 Group III 646915 94 13.3 13.1 646916 94 11.7 11.5 646917 94 12.9 12.7 646918 94 11.0 10.8 646919 94 14.1 13.9 65.11 57.16 63.12 53.68 69.08 201.54 176.92 195.38 166.15 213.85 -fat.. Company Sanitbed. Does not conlafc TSCA CBI H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Data for H-240 20 DuPont-6544 Given: MoPkVt. Active (g/:mole): 426 Formulation Dose (mg/kg): 20 %Active (F Containing) in Fo:rmulation: 100 % F in Active: 69 Mol Wt. F (g/mc)!): 19 Calculatiid Values: Dose Active (mg/kg): Dose Active (mmole/kg): Dose F (mmol/kg): 20 0.047 0.726 Molar Ratio (Ac<tive/F): Dose F (mg/kg): 0.065 13.8 Rat Number Test Day Sample ppmF in Blood Group I 646921 1 10.6 646922 1 10.7 646923 1 9.2 646924 1 5.3 646925 1 11.2 ppm F in Blood Minus Bkg 0.2 ppm 10.4 10.5 9.0 5.1 11.0 ppm F in Blood Normalized to 0.1 mmoles/kg Dose 22.15 22.37 19.17 10.86 23.43 p molar Equivalents of Active in Blood 75.36 76.09 65.22 36.96 79.71 Group I 646921 5 78.7 646922 5 83.2 646923 5 77.0 646924 5 66.1 646925 5 69.6 78.5 83.0 76.8 65.9 69.4 167.21 176.79 163.58 140.37 147.82 568.84 601.45 556.52 477.54 502.90 Group I 646921 10 63.0 646922 10 69.1 646923 10 60.3 646924 10 62.5 646925 10 53.9 62.8 68.9 60.1 62.3 53.7 133.76 146.76 128.01 132.70 114.38 455.07 499.28 435.51 451.45 389.13 (; Company SanSjBws, SB tsorttsSftJSC^CBH 72- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Rat Number Test Day Sample ppmF in Blood Group III 646926 13 646927 13 646928 13 646929 13 646930 13 34.8 30.0 23.7 33.7 25.4 ppm F in Blood Minus Bkg 0.2 ppm 34.6 29.8 23.5 33.5 25.2 ppm F in BHood Normalize;dto 0.1 mmole s/kg Dose 73.70 63.47 50.06 71.36 53.68 DuPont-6544 umolar Equivalents of Active in Blood 250.72 215.94 170.29 242.75 182.61 Group III 646926 24 11.9 11.7 646927 24 10.4 10.2 646928 24 8.1 7.9 646929 24 15.7 15.5 646930 24 9.8 9.6 24.92 21.73 16.83 33.02 20.45 84.78 73.91 57.25 112.32 69.57 Group III 646926 52 2.5 2.3 646927 52 1.7 1.5 646928 52 1.3 1.1 646929 52 4.0 3.8 646930 52 1.8 1.6 4.90 3.20 2.34 8.09 3.41 16.67 10.87 7.97 27.54 11.59 Group III 646926 94 0.8 0.6 646927 94 0.5 * 646928 94 0.5 * 646929 94 0.9 0.7 646930 94 0.5 * Below LOQ (Limit of Quantification) 1.28 * * 1.49 * 4.35 * * 5.07 * Coatpa^atisa^ftaaflsontefe^afee@i .73- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Data for H-24943 DuPont-6544 Given: MolWt. Active (g/imole): 1500C0 % F in Active: 15.5 Formul ation Dose (mg/kg): 1000 Mol Wt. F (g/mc)!): 19 % Acti-ve (F Centalning) in Foirmulation: 30 CalculatedI Values: Dose A.ctive (mg/lq5): Dose A.ctive (mmol e/kg): DoseF (mmol/kg): 300 Molar Ratio (Ac1tive/F): 0 0 Dose F (mg/kg): 46.5 2.4 Rat Number Test Day Sample ppmF in Blood Group I 648927 1 0.9 648928 1 0.9 648929 1 0.9 648930 1 0.7 648931 1 2.2 ppm F in Blood Minus Bkg 0.2 ppm 0.7 0.7 0.7 0.5 . 2.0 ppm F in Blood Normalized to 0.1 mmoles/kg Dose 35.0 35.0 35.0 25.0 100.0 umolar Equivalents of Active in Blood 1.5 1.5 1.5 1.1 4.3 Group I 648927 5 2.8 2.6 130.0 5.6 648928 5 2.0 1.8 90.0 3.9 648929 5 2.4 2.2 110.0 4.7 648930 5 1.9 1.7 85.0 3.7 648931 5 1.9 1.7 85.0 3.7 Group I 648927 10 2.1 1.9 95.0 4.1 648928 10 1.8 1.6 80.0 3.4 648929 10 1.9 1.7 85.0 3.7 648930 10 2.5 2.3 115.0 4.9 648931 10 3.5 3.3 165.0 7.1 Ceimpany Sanitized. Does not contain TSC&, CBI -74- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Rat Number Test Day Sample ppmF in Blood Group III 648932 13 1.8 648933 13 1.9 648934 13 1.8 648935 13 1.8 648936 13 1.7 Group III 648932 24 0.5 648933 24 0.5 648934 24 0.5 648935 24 0.5 648936 24 0.5 Group III 648932 52 0.5 648933 5.2 0.7 648934 52 0.5 648935 52 0.5 648936 52 0.6 Group III 648932 94 0.5 648933 94 0.5 648934 94 0.5 648935 94 0.5 648936 94 0.5 ppm F in Blood Minus Bkg 0.2 ppm 1.6 1.7 1.6 1.6 1.5 0.5 0.4 ppm F in Bl()od Normalized to 0.1 mmoles/ kg Dose 80.0 85.0 80.0 80.0 75.0 25.0 20.0 DuPont-6544 u molar Equivalents of Active in Blood 3.4 3.7 3.4 3.4 3.2 1.1 0.9 Company Sanilszea. Oaesnoe contain TSCACBS 75- H-24943: Biopersistence Screening 10-Dose Oral Gavage^tudy in Rats DuPont-6544 APPENDIX E Individual Fluorine Levels in Liver Company Saa'tized. Does nol corrtain TSCA C8S 76- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Data for H-240 19 DuPont-6544 Given: Mol Wt. Active (g/:mole): 497 Formulation Dose (mg/kg): 10 % Active (F Containing) in Fcrmulation: 100 % F in Acth/e: 65 Mol Wt. F Gg/mol): 19 Calculated Values: Dose Active (mg/k{5): Dose Active (mmole/kg): Dose F (mmol/kg): 10 0.020 0.342 Molar Ratio (Active/F): Dose F (mg/kg): 0.059 6.5 Rat Number Group I 646910 646911 646912 646913 646914 Test Day Sample ppmF in Liver 10 320.9 10 334.0 10 318.6 10 281.6 10 306.6 ppm F in Liver Minus Bkg 0.2 ppm 320.7 333.8 318.4 281.4 306.4 Group III 646915 94 646916 94 646917 94 646918 94 646919 94 85.6 75.9 87.5 78.0 94.2 85.4 75.7 87.3 77.8 94.0 ppm F in Liver Normalized to 0.1 mmoles/kg Dose 1593.88 1658.99 1582.45 1398.56 1522.81 424.44 376.23 433.88 386.67 467.18 p molar Equivalents of Active in Liver 4933.85 5135.38 4898.46 4329.23 4713.85 1313.85 1164.62 1343.08 1196.92 1446.15 C&mpafty Sanitised. Does not ccntain TSCA CBS ^^w , 77- H-24943: Biopersistence Screening 10-DoseOralGavage Study in Rats Data for H-240;20 DuPont-6544 Given: MoPNt. Active (g/imole): 426 Formulation Dose (mg/kg): 20 %Ac tive (F Containing) in Fcirmulation: 100 % F in Active: 69 Mol Wt. F (g/mc)!): 19 Calculatiid Values: Dose Active (mg/k|?) Dose Active (mmol e/kg): Dose F (mmol/kg): 20 0.047 0.726 Molar Ratio (Ac1;ive/F): Dose F (mg/kg): 0.065 13.8 Rat Number Test Day Sample ppmF in Liver Group I 646921 10 118.3 646922 10 122.7 646923 10 114.3 646924 10 121.6 646925 10 122.1 ppm F in Liver Minus Bkg 0.2 ppm 118.1 122.5 114.1 121.4 121.9 ppm F in Liver Normalized to 0.1 mmoles/kg Dose 251.55 260.93 243.03 258.58 259.65 umolar Equivalents of Active in Liver 855.80 887.68 826.81 879.71 883.33 Group II][ 646926 94 1.8 1.6 646927 94 1.2 1.0 646928 94 2.2 2.0 646929 94 4.7 4.5 646930 94 2.9 2.7 3.41 2.13 4.26 9.59 5.75 11.59 7.25 14.49 32.61 19.57 Company Sanitized, o^g "ot contain TSCACBI 78- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Data for H- 24943 DuPont-6544 Given: MolWt:. Active (g/mole): 150i000 % F in Active: 15.5 Formuliition Dose (mg/kg): 100'0 Mol W t.F(g/rnol): 19 % Acti\'e (F Containing) in Foirmulation: 30 Calculated Values: Dose A>;tive (mg/k g)-. Dose A(;tive (mmo le/kg): Dose F l[mmol/kg): 300 Molar 1?.atio (Aictive/F): 0 0 DoseF (mg/kg;): 46.5 2.4 Rat Number Test Day Sample ppmF in Liver Group I 648927 10 648928 10 648929 10 648930 10 648931 10 23.5 21.0 21.5 27.4 18.4 ppmF in Liver Minus Bkg 0.2 ppm 23.3 20.8 21.3 27.2 ,18.2 Group III 648932 94 1.5 1.3 648933 94 1.8 1.6 648934 94 1.3 1.1 648935 94 1.6 1.4 648936 94 1.6 1.4 ppm F in I iver Normalized to 0.1 mmoles/kg Dose 1165.0 1040.0 1065.0 1360.0 910.0 65.0 80.0 55.0 70.0 70.0 umolar Equivalents of Active in Liver 50.1 44.7 45.8 58.5 39.1 2.8 3.4 2.4 3.0 3.0 -79- ooes "of contain TSCAC^ H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 APPENDIX F Individual Fluorine Levels in Fat ainTSCACBI 80- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Data for H-240 19 DuPont-6544 Given: Mol Wt. Active (g/mole): 497 Formulation Dose (mg/kg): 10 % Active (F Containing) in Fcrmulation: 100 % F in Acth/e: 65 Mol Wt. F (,g/mol): 19 Calculated Values: Dose Active (mg/kg): Dose Active (mmole/kg): Dose F (mmoVkg): 10 0.020 0.342 Molar Ratio (Active/F): Dose F (mg/kg): 0.059 6.5 Rat Number Test Day Sample ppm F in Fat Group I 646910 10 14.4 646911 10 14.9 646912 10 10.9 646913 10 11.6 646914 10 12.4 ppm F in Fat Minus Bkg 0.2 ppm 14.2 14.7 10.7 11.4 12.2 Group III 646915 94 1.3 1.1 646916 94 1.1 0.9 646917 94 1.3 1.1 646918 94 0.8 0.6 646919 94 1.5 1.3 ppm F in Fat Normalized to 0.1 mmoles/kg Dose 70.57 73.06 53.18 56.66 60.63 umolair Equivale nts of Active in Fat 218.46 226.15 164.62 175.38 187.69 5.47 4.47 5.47 2.98 6.46 16.92 13.85 16.92 9.23 20.00 Company SanHteeA Does not contain TSCA CBi 81 - H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats DuPont-6544 Data for H-240:'.0 Given: MolWt. Active (g/:mole): 426 Formul ation Dose (mg/kg): 20 % Actrve (F Contai.ning) in Fc)rmulation: 100 % F in Active: 69 Mol Wt. F (g/mo I): 19 Calculated1Values: Dose A -ctive (mg/kjS): Dose A .ctive (mmol.e/kg): DoseF (mmol/kg): 20 0.047 0.726 Molar Ratio (Active/F): Dose F (mg/kg): 0.065 13.8 Rat Number Test Day Sample ppmF in Fat Group I 646921 10 6.2 646922 10 8.5 646923 10 8.7 646924 10 8.8 646925 10 8.3 ppm F in Fat Minus Bkg 0.2 ppm 6.0 8.3 8.5 8.6 8.1 Group III 646926 94 ND ND 646927 94 ND ND 646928 94 ND ND 646929 94 ND ND 646930 94 ND ND ND Non-detectable. ppm F in Fat Normalized to 0.1 mmoles/kg Dose 12.78 17.68 18.11 18.32 17.25 ND ND ND ND ND u molar Equivalents of Active in Fat 43.48 60.14 61.59 62.32 58.70 ND ND ND ND ND ^nipanysanifaed.ooes "ot contain TSCACBI 82- H-24943: Biopersistence Screening 10-Dose Oral Gavage Study in Rats Data for H-24!)43 DuPont-6544 Given: MoUVt. Active (g/imole): 15000 0 % F in Active: 15.5 Formiillation Dose (mg/kg): 1000 Mol Wt. F (g/mo I): 19 %Ac tive (F Contai ning) in Foirmulation: 30 Calculatiid Values: Dose Active (mg/k^D-Dose Active (mmol e/kg): Dose F (mmol/kg): 300 Molar Ratio (Act ive/F): 0 0 Dose F (mg/kg): 46.5 2.4 Rat Number Test Day Sample ppmF in Fat Group I 648927 10 16.1 648928 10 18.6 648929 10 27.9 648930 10 20.1 648931 10 14.3 ppm F in Fat Minus Bkg 0.2 ppm 15.9 18.4 27.7 19.9 14.1 Group II [ 648932 94 2.6 2.4 648933 94 3.7 3.5 648934 94 3.1 2.9 648935 94 2.4 2.2 648936 94 4.6 4.4 ppm F in Fat Normalized to 0.1 mmoles/kg Dose 795.0 920.0 1385.0 995.0 705.0 120.0 175.0 145.0 110.0 220.0 umolar Equivalents of Active in Fat 34.2 39.6 59.6 42.8 30.3 5.2 7.5 6.2 4.7 9.5 ^""'^'"^'"^-^TSCACB, 83-