Document 6RDkMmbxZ6E2NE6VoJjbVBG89

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M r - T c o p ^ ^ J lS L ^ j ^ L ^ tn ------- - dvNv,.n JL .+ m a ju o ^ ---- CiJ^ l/ i Acession #: 9 Title of Project: DMP Deer and Elk Study Status of Project: in progress Location of Sampling: Oregon and Washington Herbicide Use: forest Sample Media: deer and elk adipose tissue Range of Lim it o f D e t e c t i o n : RTP: 0.8 - 6 . 0 ppt WSU : 8 .0 - 34.0 ppt Average Limit o f D e t e c t io n : RTP: 2.7 ppt WSU: 14.8 ppt # of Samples C o l l e c t e d : deer: 6 elk : 9 # of Samples Analyzed: IS # of Samples Suspected P o s i t i v e : : deer: 3 o f 6, 50.0% e l k : 7 o f 9, 77.8% Comments: I t is important to note t h a t four samples were analyzed below 50% recovery o f the 37C1 TCDD standard and th e r e fo r e require r e - e x t r a c t i o n and r e - a n a l y s i s according to standard DMP procedures. These samples are # OR-D-1, WA-D-4, WA-D-8, and WA-E-7. N e v e r t h e l e s s , the data i n d i c a t e the presence of 2 , 3 , 7 , 8 TCDD residues in both deer and e l k . Also i t is important to note t h a t RTP reported the presence of other minor isomers in a d d i t i o n to the major 2 , 3 , 7 , 8 TCDD isomer (based on the DMP a n a l y t i c a l c r i t e r i a ) . The r e s u l t of t h i s study i n d i c a t e s t h a t a d d i t i o n a l sampling may be u s e fu l in providing more information about the e x t e n t ^ of dioxin contamination associated with the use of 2 ,4 ,5 -T and s i l v e x i n t h i s geographi c are a ..  m m * , -y ID Adipose Sample Description Toxicant Analysis Center Sample# ppt() Native TCDD Added to Sample deer deer /deer deer deer deer /deer Method B l a n k ^ elk elk elk deer elk elk elk elk Standard Solution^) elk ! n elk OR-D-5 W A-D -1 O R -D -1 Control W A-D -4 O R -D -6 W A-D -8 W A-E-2 O R -E -1 1 W A-E-6 Control O R -E -7 O R -E -9 WA-E-5 OR-E- 8 W A-E-4 W A-E-7 7 0 21 14 * Research Triangle Park T t1/2*1 Wright State University TCDD TCDD 37CL*d ) RTP Detected Detection % Recovery # (ppt)() Limit ( p p t ) ^ TCDD TCDD Detected Detection (ppt) te) Limit ( p p t ) ^ 37CL % Recovery 1 2 4 (e) 125 126 12? ND ND 127 9 128 1 2 9 (e) 130 ND rl $ T fi 131 1 3 2 (e) 133 1 3 4 (e) 1 3 5 (e) ND kf'':,l-f ND '5*1 ' 34 "4 136 137(e) 24 1 138 l 3 9 (e) 2i T'V ' 4 140 23 2 2 4 2 3 2 4 3 2 2 0 .8 6 3 3 3 1 2 1 4 1 (e) 142 '21 y ND 2 4 89 \ 50 34 78 36 84 31 50 67 71 100 50 50 100+ 57 87 100+ \ x (g) 2 3 <s) 4 NA 5 6 tg> 7 NA 8 9 10<*> 11 12 (g ) i3 (g) 14 15 76 16<*> 36 1 7 (g) 311 ND NP 14 9 21 21 34 13 14 i. ND ND 13 24 8 ND ,,fit- 68 - 41 I 29' 1 ND ND ND 17 8 10 .8 8 8 19 10 11 21 '11 ND 25 46 46 29 51 59 33 105 97 79 150 102 98 43 84 76 70 32 (ci) Method mk: extraction procedure using solvent only (b) Standard Solution: a standard solution containing 2,3#7,8-tetrachloro/dibenzo-p-dioxin (c) ppt: parts per trillion (d) corrected for recovery loss (e) two other isomers in addition to 2,3, 7,8-tetrachlor< in the major isomer^ were observed (f) WSU analyses reported here are the average of two runs (g) evidence of PCB contamination in sample X ND: Not Detected NA: Not analyzed by WSU due to limited amount of sample NB: Sample wt. for all samples was 5g.; 37CL TCDD addition to each sample is 5 nanograms (ng) except Control Sample RTP-135/WSU-10 which contained 1.33 ng. A UNITED STATES ENVIRONMENTAL PROTECTION AGENCY HEALTH EIT-F.CTS RESEARCH LABO RATO RY RESEARCH TRIANGLE PARK NORTH CAROLINA 2771 1 DATE: December 17, 1979 SUBJECT: Results of Capillary Column GC/HRMS Analyses Performed on Extracts of Deer and Elk Adipose Tissue for 2,3,7,8-TCDD Residues FROM: Robert Harless s&utZ'*'* HERL, ETD, ACB (MD-69) TO: Mike Dellarco, Dioxin Project Manager Special Pesticide Review Division (TS-791) Office of Toxic Substances 401 M Street, SW US EPA Washington, DC 20460 THRU: Dr. R. G. Lewis, Chief Analytical Chemistry Branch (MD-69) HERL, ETD Per your request,., these samples were not analyzed until the most high priority samples, "Vertac", etc. were analyzed. This shipment of sample extracts was received from the EPA Pesticide Monitoring Laboratory, Bay-St. Louis, Mississippi, 9-6-79, and was assigned the identification number ACB-108. The sample extracts were subjected to a previously described capillary column GC/HRMS multiple ion monitoring method of analysis for 2,3,7,8-TCDD residues utilizing a Varian 311A mass spectrometer interfaced with an SE-30 WCOT glass capillary column. The criteria utilized for confirmation of 2,3,7,8-TCDD were: 1. Capillary column GC/HRMS retention time of 2,3,7,8-TCDD. 2. Co-injeccion of sample fortified with^Cl-TCDD and 2,3,7,8TCDD standard. 3. Molecular ion chlorine isotope ratio (m/e' 320 and m/e 322). 4. Capillary column GC/HRMS simultaneous multiple ion monitor ing response (m/e 320, m/e 322, and m/e 328) for TCDD. 5. M/e 320 and m/e 322 MS response greater than 2.5 mm noise level. The capillary column GC/HRMS retention time of 2,3,7,8-TCDD was 15 min utes + 1 5 seconds. The MS mass resolution (8,500) was sufficient to resolve TCDD from contamination. The masses monitored during these analyses were: 1. m/e 318.9793 PFK reference 2. m/e 319.8965 TCDD 3. m/e 321.8935 TCDD 4. m/e 327.8847 37C1-TCDD Results The results are shown in Table 1. The recovery of 37 Cl-TCDD was below 50% for specific sample extracts. Therefore, the results should not be reported. These specific samples should be subjected to analytical clean-up and GC/HRMS analysis again to confirm the reported results. TCDD isomers (3) were detected in RTP-124, 129, 132, 134, 135, 137, 139, and 141. The isomer having exact GC/HRMS retention time as 2,3,7,8TCDD was the major isomer detected (high concentration). No problems were encountered in these analyses. Summary 2,3,7,8-TCDD and two (2) TCDD isomers were detected in deer and elk/ adipose tissue samples. CC: Dr. William Durham, Director Environmental Toxicology Division (MD-66) Dr. Nancy Wilson, Chief Chemical Characterization Section (MD-69) TABLE 1 ACB #108 ANALYTICAL RESULTS FOR 2,3,7,8-TCDD RESIDUES Sample ID RTP-124' RTP-125 RTP-126 RTP-127 RTP-128 RTP-129' RTP-130 RTP-131 RTP-132* RTP-133 RTP-134RTP-135 * RTP-136 RTP-137 RTP-138 RTP-139RTP-140 RTP-141 ' RTP-142 Sample Weight (g) 5 5 5 5 5 5 5 5 4 5 5 5 5 5 5 5 5 5 5 ND = not detected 37C1-TCDD Fortification Level (ng) 5 5 5 5 5 5 5 5 4 5 5 1.33 5 5 5 5 5 5 5 37C1-TCDD % Recovery 'y. 89 50 34 78 36 84 3150 67 71 100 50 50 100+ 57 87 100+ 76 36 TCDD Detection Limit (ppt)* 2 2 4 2 3 2 4 3 2 2 0.8 6 3 3 3 1 2 2 4 TCDD Detected (ppt) * *12 ; ND ND 9f ND 9j 71 V ND 9? 4 ND 54 j 34 | 5I 121 41 23 5 21 ^ ND ppt = parts per trillion * s corrected for recovery losses Comments Deer Adipose II II II II II II II II l l II Elk Adipose II II l II II l l II II II II II II ll II l l II II II II II '<` 7 ( Rationale and Study Design for Proposed TCDD Analysis of Region X Elk and Deer Adipose Tissue samples. 1. Reasons why such a study is needed: A. Because of so many variables, mother's milk TCDD levels alone do not provide much valuable information about TCDD in the environment. Utilizing human subjects presents great diversity in terms of diet, mobility in the environment, medication, and exposure history. Because of the nature of the TCDD molecule, it should be more concentrated in adipose tissue than elsewhere. Milk with its butterfat content would thus not be an unreasonable sample type, but adipose tissue would be much better. The difficulties of obtaining adipose biopsies from a selected population of humans are obvious. B. A TCDD monitoring study of carefully selected samples of elk and deer will provide the following advantages: Elk and deer live in discrete forest areas all of their lives. The herds are closely monitored by professional wildlife biologists. Exposure information from past spraying histories of the areas can be obtained. This localized nature of the herds enhances research design. Elk and deer are browsers and grazers, eating only vegetation from the forest areas they inhabit. Their diet thus reflects their environment (in terms of residues of any environmental contaminants) far more accurately than does a typical diet of an "exposed" human. Elk and deer are consumed by large numbers of people. Knowledge of possible contaminants in these animals is thus of publics health importance. ? In California, State agencies have interpreted the precautions on the 2,4,5-T label to apply to both deer (California does not have appreciable elk populations) and livestock. The state thus opposes 2,4,5-T spraying 2 weeks prior to and during the deer season. Elk and deer herds in Region X coastal forests often consist of sufficient quantities of older animals to allow for sampling them preferentially as indicators of any bioaccumulation in an "exposed" environment over a several year timeframe. As explained later, several of our elk samples are from animals in the 10 to 13 year age class. were labeled as to: species, date sampled, sex, age, site and date of kill, name, address and telephone of donating hunter, name, address and telephone of biologist making the collection, and miscellaneous comments by the sampling officer. They were then frozen and taken to a central site where they were then picked up by EPA Region X. Labels were made of paper and affixed to the bottles with strong twine. Colecting dates were as follow: Oregon deer, 11-5-77; Oregon elk, 12-3-77; Washington deer, 10-22 and 10-23-77; and Washington elk, 10-31-77 through 11-21-77. * At present, the samples are stored frozen at the EPA Region X Laboratory in Seattle. They are labelled and sealed since their collection, and can be shipped at once whenever a suitable analytical laboratory can be found and utilized. E. Sample data are as follow: Oregon elk totalled 15. All of these animals were taken from the Coos Bay to Rosenburg area. Specifically, all of the animals came from the Millacoma and Coos River drainages, stretching across predominantly Weyerhaeuser land eastward from the vicinity of Allegany. The elk herdjis monitored closely by game biologists, and has'been experiencing anexcess of reproductive vp r p b l e i ^ #>;;ineludlx^sSill!^fns"vand: teratisms during recent f .^.ygars#.J' Ages range from 2 to 13 years (mean: 7.5 years). Mean size of fat sample is 43.5 grams. All are female, because only cows are sacrificed and studied from this herd annually by the game biologists. Oregon deer totalled 10. All were taken from hunters at check stations in the Tillamook area. Age ranged fr om 1.5 to 5.0 years (mean: 2.8 years). Mean size of fat sample is 58.8 grams. Seven males and three females were sampled. Washington deer totalled 9. They were taken in the general area stretching from Aberdeen toward the Pacific Coast. Mean sample size was 22.3 grams. Ages ranged from 0.5 to 5 years (mean: 1.9 years). Seven males and two females were sampled. Region X begem this study in the Fall of 1977 as a corollary to the mother's milk sampling study for TCDD, for reasons already outlined at the beginning of this paper. We have informed OSPR of this effort since its inception emd have requested assistance numerous times in the analysis of the samples for TCDD, chiefly through Mr. Reising of OSPR. We have received no response whatsoever from OSPR. Wright State University Dnyton, Ohio 45431 / November 29, 1979 Mr. W. T. Hollaway U.S. Environmental Protection Agency Office of Toxic Substances Special Pesticide Review Div. (TS-791) Crystal Mall #2 1921 Jefferson Davis Highway, Rm. 728 Arlington, Virginia 22202 1rnr\ /p 1 4'.i I 1 j; 'i!i ! ! ! i1 UEC-8.&?* !j \ L Dear Mr. Hollaway: Attached is a table showing the results of analyses of the remaining samples in the batch of 17 which we have been analyzing. Results for the first nine samples were reported in our recent Quarterly Report. We are proceeding as rapidly as possible with the MS-30 modifications to permit isomer-specific TCDD determina tions, and will provide you with a status report on those develop ments in the near future. Sincerely, TOT/aam Attachment Thomas 0, Tiernan, Ph,D, Professor of Chemistry and Director of the Brehm Laboratory 7 TABLE 1 RESULTS C? GC-HIGH RESOLUTION MS ANALYSES OF SAMPLE EXTRACTS PROVIDED BY EPA FOR TETRACHLORODIBENZO-p-DIOXIN (TCDD) BY WRIGHT STATE UNIVERSITY EPA Sample No. WSU-10 WSU-11 WSU-12 WSU-13 WSU-14 WSU-15 WSU-16 WSU-17 Run 1 Native TCDD Detected (parts-per-trillion) m/e 320 m/e 322 Average Run 2 Native TCDD Detected (parts-per-trilHon) m/e 320 m/e 322 Average Average Native TCDD (ppt) 31 43 37 25 23 24 000 000 000 15 20 18 18 26 22 000 48 . 43 35 32 00 00 00 14 16 17 20 00 45 34 0 0 0 15 19 0 41 j 29 0 0 0 17 j 21 | 0 Average M.D.C. (PPt) Average % Recovery 8 150 8 102 8 98 19 43 10 84 11 76 11 70 25 32 Notes a., -- a. a. a. -- a. a. a. PCB contamination of sample 1s evident, b. This sample was supposedly spiked with only 1,33 ng total 37C1-TCDD, The Indicated recovery however Indicates a possible error 1n spiking. QUANTITATION OF PARTS-PER-TRILLION LEVELS OF 2,3,7,8-TETRACHL0R0DIBEHZ0-p-DI0XIN IN EPA FURNISHED EXTRACTS OF BIOLOGICAL MATERIALS EPA Cooperative Agreement No. CR806846-01 Submitted by Brehm Laboratory Wright State University Dayton, Ohio 45435 Thomas 0. Tiernan, Joseph G. Solch, Garrett VanNess, Michael L. Taylor, and Thomas Mazer Quarterly Report September 23, 1979 Submitted to William T. Hollaway Coordinator - Dioxin Implementation Plan U.S. Environmental Protection Agency Office of Toxic Substances Special Pesticide Review Division (TS-791) Crystal Mall #2 1921 Jefferson Davis Highway, Rm. 728 Arlington, Virginia 22202 I. INTRODUCTION Analyses of EPA-furnished extracts of deer and elk adipose for tetrachlorodibenzo-p-dioxin content were accomplished by Wright State University during the third quarter of the program being conducted under EPA Coopera tive Agreement No. CR806846-01. Results of these analyses are reported herein and other pertinent topics are discussed. . II. EXPERIMENTAL PROCEDURES The experimental procedures applied for the analyses reported herein are essentially the same as those described previously in the Quarterly Report of March 23, 1979. III. RESULTS AND CONCLUSIONS The results of analyses of nine extracts of deer and elk adipose are listed in Table 1. Since these extracts were based on five gram samples, whereas the Cooperative Agreement specified that ten gram samples would be extracted by EPA, it was not possible to achieve the specified detection sensitivity for TCDD. However, the minimum detectable TCDD levels in most of the samples are quite reasonable, being 13 parts-per-trillion (ppt) or less for five of the nine samples. In all cases where higher detection limits were imposed, the recoveries, based on the 37C1-TCDD Interrial standard, were extremely poor. It is noteworthy that recoveries were less than 50% for four of the nine samples, and were in the acceptable range for only three samples. This suggests that the EPA extraction and cleanup methodology is not really adequate for these types of samples, and that improvements are desirable. Three of the nine samples analyzed exhibited very large apparent polychlorinated biphenyl (PCB) Interferences, and the recoveries for these samples were the lowest observed 1n this set, as shov/n 1n Table 1. IV. PROJECT STATUS AND PLANS In order to be consistent with the Cooperative Agreement schedule, V/right State would have been required to analyze twenty-one (21) sample extracts during the present third quarter reporting period. As noted in the previous Quarterly Report submitted to EPA, Wright State analyzed nine (9) samples in excess of the requirements during the previous quarter, which reduces the normal sample analysis.load (30 samples/quarter) to twentyone (21), as just mentioned. However, EPA supplied V/right State with only seventeen (17) sample extracts during the present reporting period, and these were not received until September 20, 1979, only three days before the end of the third quarter of the Cooperative Agreement. In view of this late shipment of samples, it was not possible to complete analyses of all seventeen (17) extracts prior to the end of the reporting period. Analyses of nine of the seventeen were completed and are reported herein, as noted above. It must be emphasized that EPA has not provided Wright State with the numbers of samples specified in the Cooperative Agreement for the present quarter, and that the sample shipment which was made was not timely. It Is urged that attempts be made to rectify this situation in the coming quarter, and that the appropriate number of samples be shipped as soon as possible. Wright State should receive a minimum of thirty-four samples from EPA for analyses during the fourth quarter (prior to December 23, 1979) in order to satisfy the projected schedule. During this reporting period, authorization was received from EPA (letter of September 13, 1979 from W. T. Hollaway) to accomplish an Internal budget-transfer of funds for the Cooperative Agreement. Accord ingly, $8000 v/as transferred from the equipment line to the supply line of expenditures for the Agreement, as authorized. Also during this reporting period, Dr. T. 0. Tiernan attended the National Meeting of the American Chemical Society in Washington, D.C., where he participated in a Symposium on the Chemistry of Chlorinated Dibenzodioxins and Dibenzofurans. Dr. Tiernan also visited Mr. Hollaway and Mr. DeLarco, at their EPA offices during this trip, to discuss the status of the Cooperative Agreement and future plans. In the course of these discussions, Dr. Tiernan reviewed some recent data obtained by Wright State, relevant to the detection of TCDD in combustion samples. Preliminary data showing separation and determination of TCDD isomers using capillarycolumn gas chromatography in conjunction with low-resolution mass spectro metry were presented by Dr. Tiernan, and the detection of TCDD in fireplace soot by Wright State was noted. Dr. Tiernan further emphasized the desir ability of proceeding with the proposed modifications to our MS-30 Mass Spectrometer to incorporate capillary-column GC, in view of these findings and the reports by Dow Chemical of TCDD in combustion samples. EPA Sample No. TA&w. 1 RESULTS OF GC-HIGH RESOLUTION MS ANALYSES OF EPA-SUPPLIED SAMPLE EXTRACTS BY WRIGHT STATE UNIVERSITY Run 1 Native TCDD Detected (parts-per-trill1on) m/e 320 m/e 322 Average Run 2 Native TCDD Detected (parts-per-trillion) m/e 320 m/e 322 Average Average Nati ve TCDD (PPt) Average Minimum Detectable Concentration (PPt) Average at to Recover: WSU-la * 30 25 28 (deer adipose) 46 WSU-2 000 (deer adipose) 0 WSU-3a * 000 (deer adipose) 0 WSU-4 14 14 14 (deer adipose) 14 WSU-5 14 11 13 (deer adipose) 16 WSU-6a* 000 (deer adipose) 0 WSU-7 000 (deer adipose) 0 WSU-8 000 (elk adipose) 0 WSU-9 72 64 68 (elk adipose) 65 a. PCB contamination of samples 1s evident. 21 34 0 0 00 13 14 14 15 00 00 00 70 68 31 # 0 0 14 # 14 * 0 0 0 68 ? 21 21 34 13 13 24 8 8 10 46 46 29 51 59 33 105 97 79 u' DCf*A.TMCNT Or CHCMiSTBY T H E UNIVERSITY O F N E 3 R A S K A - U N C O L N LINCOLN. NEBRASKA 6 S 5 SS July 13 Dr. Hugh Black U.S. Forest Service Fisheries and Wildlife Management Staff 630 Sansoroe Street San Francisco, CA 94111 ' ' t -f jwo "rj f ' Dear Hugh: .' Enclosed is-a-copy of-the final-.report for analysis of TCDD in deer tissue and in other samples (mylar panel and leaf washings). In the report, I outline sample work-up (only briefly since Or. Dupuy did the work and he should report on this), sample custody, and the details of the GC/MS analysis. Some typical outputs are also given as well as a tabularreport of all the results. If there questions, errors, or sections which require amplification, please let me know and I will submit a revised report. In my judgments, all "positives" (see Table IV) are indeed "positive'.' You can see from the Figures 2-3 that the data for a sample at ca. 5 parts-pertrillion is rather unambiguous. Of course, there is always a possibility for contamination in the work-up or sample handling. f l have taken the liberty of sending Dr. Dupuy a copy of the report because he has served as an; important coworker in this project. Upon your approval of the report, I will ask my University Grants, and Contracts Office to send a bill to you for full payment. Although the contract arrange ments were made for sample work-up, we have done a number of additional analyses (particularly the mylar panel and leaf washings). In addition, each "positive" (and others) were validated by means of a full, second analysis monitoring m/z 320 and 322. From these data,, we could check the ratio of isotopic cluster signals and quantitate by a second, absolute method. Neither of these research activities were required by the contract. Therefore, I propose that they be considered as a full substitute for the workups. Thank you for the opportunity to assist you with this interesting project. I hope the results will be of value to you in understanding the possible bioaccumulation of dioxin in wildlife. Finally, I would appreciate the opportunity to study the sample code and comment on our performance on control samples. Sincerely'"yours, Michael L, Gross Professor THE UNIVERSITY OF NEQRASKA-UNCOLN THE UNIVERSITY OF NEBRASKA AT OMAHA THE UNIVERSITY O F NEBRASKA MEDICAL CENTER aig-q, f SUBJECT TO REVISION FINAL REPORT Ultratrace Analysis of Tetrachlorodibenzo.-g_-dioxin in Samples of Deer Tissue.by Gas-Chromatography/ High Resolution-Mass Spectrometry (Contract No. 53-91S8-8-6133) to United States Department of Agriculture ; Forest Service (Region-5) 630 Sansome St. San Francisco, CA 94111 r Dr. Hugh B1 ack ' Project. Coordinator IItI II ! i I i i* Michael L. Gross Professor of Chemistry University of Nebraska Lincoln, NE 68588 July 10, 1979 It Introduction t This is a report of the analytical support work for a project to examine the possible accumulation of tetrachlorodibenzo--dioxin (TCDD) in the tissue of deer exposed to TCDO via spraying of 2,4,5,-T. The project was organized and conducted by the U.S. Forest Service-, Dr. Hugh Black, project coordinator. II. Sample Preparation and Custody , The various deer tissue samples and controls were digested in base, extracted with hexane, washed with sulfuric acid and submitted to small scale liquid chromatography ("acid-base" work-up) by Dr. Aubry Dupuy and his coworkers at. the Pesticide Monitoring Laboratory, USEPA, in Bay v St.. Louis, Mississippi. The extracts along with any controls were coded without regard to origin and shipped frozen in dry ice to our laboratory- Upon reception, the samples were stored in a freezer until they were analyzed- All arrived in perfect condition. Various-samples of"washings of & Mylar panels-and 5 leaf-samples were passed through a silica gel column and concentrated by Dr. Anthony S. Wong of California Analytical' Laboratories. These samples we r e . collected after the 2,4,5-T spraying at the deer confinement area. Sample Coding __________ ''Date Shipped' '' .... .... Date'Received FS-T through FS-14' Mylar 2,4,34,38,42,44 and 7672-1, 7635-2,7635-1, 7696-1, 7635-3 FS-15 through FS-27 FS-28 through FS-48 February 14, 1979 March 1,1979 March 26, 1979 May 9, 1979 February 15, 1979 March 2, 1979 March 27,1979 May 10, 1979 SU9? 9 I I I . Analysis of TCDD "At the time of analysis, each sample tube provided by Dr. Dupuy was opened, the total volume measured using a 100y syringe, and an % aliquot removed (usually between 20- arid 35% of the total sample) for GC/HRMS. The Mylar Panel and leaf washing were analyzed by simply removing an aliquot (the total volume Was measured by Dr. Wong and not by us) and injecting onto the GC/HRMS. ' 'EaclTJbT^the"rex:tracts--was ^analyzed for tetrachl o.rodibenzo--dioxin using gas chromatography/high resolution mass spectrometry (GC/HRMS). The GC was a Perkin Elmer Sigma II and the mass spectrometer was a Kratos MS-50.. The interface consisted.of a. single glass lined capillary tube followed by a capillary leak directly into the mass spectrometer The source region was pumped with a 1300 1/sec diffusion pump and, thus, no molecular separator was employed. Typical source pressures were about 1 x 10`5 torr at the ion gauge located on the 6" pump tube. The: gas chromatographic column (180 cm x 2mm ID. glass) contained 0.50% OV-17 + 0.40% Poly S-179 coated on 100% methyl silicone bonded to 80/100 mesh Chromosorb W-AW, and was operated with a helium flow rate of 15 ml/min.. The temperature was held at 250 C for 1.5 min and then increased linearly to 300C at a rate of 10C/min; the retention time of authentic TCDD was 3.3 min. The entire effluent from the gas chromatographic column was admitted to the mass spectrometer ion source.and ionized using 70eV electrons. The source temperature was 250C, the accelerating voltage 8kV, and the mass spectral resolution was 10,000 (10% valley definition). TCDD was quantitated by dual ion monitoring using peak matching. One channel was centered at m/z 327.8848 (^Cl^-TCDD, the internal standard) and the other at m/z_ 321.8936 (. (the most abundant molecular Ion of TCDO having natural isotopic elemental abundances). The complete peak profiles were acquired by scanning at a frequency of 2Hz, corresponding in each case to a mass range of 300 ppm (0.096 amu). The output was accumulated using a Nicolet Model 1170 signal averager and the resulting signals were submitted to a 3-point smoothing routine prior to output to an X-Y recorder. The concentration of TCDD was calculated-using the ratio of the intensities (maximum peak heights) at m/^ 327.8848 and 321.8936; if no signal was observed, the detection limit was set at 2.5 times the noise amplitude (a 2.5:1 S/N criterion). The calculation is made possible by obtaining the slope of a calibration plot based on various standard mixtures of TCDO and TCDO-37Cl^. These standard mixtures were interspersed between unknowns and-were analyzed in the same manner. The-value of the slope was 1.5-_+ 0.1. Positives were validated by analyzing a second aliquot, this time monitoring u 319.8966 (TC00-35C14) and 321.8936 (TCDD-35C1?37C1). The theoretical ratio l for the intensities of these two ions is Q..77 We anticipate the uncertainty in this measurement is +; 10% relative. Certainly, those "positives" giving a 0 .^ 7 * 7 ratio less than.0.60 should not be considered as validated, and, therefore, they are judged to be "negatives" . The validation also presented the opportunity to quantitate the amount of TCDD in each "positive" sample. This was done using absolute mass spectrometer response factors determined by injection of known amounts of TCDO (native) and measurement of peak heights at m/z 321.8936. Because most recoveries are not 100%, the concentration of TCDD determined in this absolute manner will be too low, and it has been corrected by dividing the concentration by the recovery expressed as a fraction. The agreement between the concentration determined by this method and by the ratio method is usually quite good. '' ' ' When the validation studies were initiated, the results using the internal standard method had not yet been worked up carefully. Consequently, some SLSLdOf samples which turned out to be "negative" by the internal standard method had been subjected to the validation study. They were "negative" by this method (see FS-5, 10,15, 22, 26, 32, 44, and 48), also. The concentration of TCDD in the Mylar and leaf washing was determined only by this method as no" standard TCDD-37Cl^ had been added to the solution. Thus the internal standard method was not applicable. The results are reported in Tables I, Il`, and III. Each Table represents data taken during separate analysis periods beginning on the dates specified below. Table Table I Table II Table III Samples FS-1 through FS-27 FSt 28 through FS-48 Mylar and Leaf Washings Date April 20, 1979 May 17, 1979 April 20, 1979 Typical data representing a standard mixture, an unknown "positive", validation of that "positive", and a typical "negative" are shown in Figures 1-4 respec tively. ' IV. Conclusions: . A.- Precision and Accuracy: No detailed study of precision and accuracy below levels of 10 parts-per-trillion (ppt) has been made to date. Consequently, it is difficult to estimate these parameters. How ever, errors and precision on the order of 100% (relative) would not be surprising in view of the difficulty in working at these low levels. It should be pointed Out that our work at levels greater than 9 parts-per-trillion has been validated by an EPA study conducted in 1977. Reports of that study have been made public and may be obtained from the Dioxin Project manager of EPA. Dr. Ralph Ross of USDA has Figure Legends Figure 1: Typical output for calibration mixture of 64 picograms (pg) of native 2,3,7,8-TCDD and 1100 pg of 2,3,7,8-TC00-37Cl* (Internal standard). The gain x30/x3" refers to the .gain of the low and high mass ions respectively, and "at 2K" pertains to the attenuator on the signal averager. Note `that the slope of the calibration plot based on this single analysis is 1.49. "RT" is retention time in minutes. Figure 2: A typical output for an unknown (FS-23) giving a positive result. m/z .321.8936 is on the right. Figure 3: Validation study for FS-23 monitoring m/z 319.8966 and m/z 321.8936. The observed intensity ratio is 0.72 (theory=0.77) Figure 4: A typical output for an unknowh (FS-38) giving a "not detected" (nd) for TC00. Note that the detection limit is taken by multiplying the observed noise level at 321.8936 by a factor of 2.5 (see output on left). > ^2^03 Figure 1 A 100*4- a Figure 2 \S l iD Nature of 'Samp]e TABLE I ANALYSIS OF TCDD BY GLC HIGH RESOLUTION MASS SPECTROMETRY Cone. Detection Limit Percent Recovery Cone. (i.^) * Cone. ^322^ ''320/1322 -1 Deer Muscle nd -2 If nd ;-2 U nd -3*1 I-4*1 5-5 tf ifffj- 3.3: It Y-f ] .6 nd S-6 . 5-7 n14 nd 7.4 S-8 ti 3.4 'S-9*2 . hd :S-T0*2 it nd rS-ll n 4.5 FS-12 it nd FS-13 u nd FS-14 M .* . n<* Iv/r,d, L-^ FS-15*2 deer adipose* 2-0 FS-T6 it nd FS-17*2 n l. 9 FS-18 FS-19 it 12 N fr y.n>Tw f 2 i I1I - 1 .2 .& 1 1 *9 T ;8 1 .1 .5 .7 .2 .6 .4 2 .9 1 2 2 50 ' 75 105 85 125 85 130 .95 95 . 95 * 80 , 85 ' 75 85 100 45 65 70 - 75 135 -` 4.1 3.0 < .1 '5.6 ` 1.9 1.2 4.7 <2 1.8 6.1 3,0 4.8 3.0 -- 5.9 2.0 1.3 5.5 * 2.6 '8.3 3.0 _ .60 .64 .79 .69 .48 .77 ** - ,54 ^ 0.t .57 ,83 .67 Q2_-SLLrOvP^ 7O ' TABLE II .. ' ANALYSIS OF TCDO BY GLC HIGH RESOLUTION MASS SPECTROMETRY *- ID NatS-uarmepl,eof C~one. Detection Limit Percent Recovery Cone.. (i322) Cone. (1322^ 1320/l FS-28 Deer Liver nd FS-29 46 14 FS-30 tl ^nd FS-31 FS-32 It n z -5 nd FS-33 n nd FS-34 it 2.1 , FS-35 . it nd TS-36 u * Wfti* 4.5 7S-37 II nd FS-38 tt nd FS-39 It nd CO TS-40 If TS-41 It - FS-42 tl 3J FS-43 Bone Marrow FSt44*4 II * FS-45 M FS-46 M FS-47 M FS-48 If nd 2 3,9 nd nd nd 2 .8 1 1 2 3 1 2 2 .5 2 2 2 1 3 5 2 3 7 7 3 40 75 - 70 75 75 35 65 55 55 30 75 70 65 80 130 .65 60 80 70 70 70 1 .1 <1 .1.3 2.9 1 9.2 -6.4 <1 2.8 <3 ' --- - -- 1 .5 .80 --- - 2.0* .i ,76 - 5.3 .75 -- - -- 14 .82 - 6,4 . -. .69 - - 3.5 .71 _* - ** TABLE I I I ANALYSIS OF TCDD BY GLC HIGH RESOLUTION-MASS SPECTROMETRY ID Mylar 2 Mylar 4 Mylar 34 Mylar 38 Mylar 42 Mylar 44 7635-1 ' 7635-2 7635-2 7635-3 7672-1 7696-T. Cone. 110 pg/yi 61 pg/yi, <1.4 pg/y2. 18 pg/ yi 9.4 pg/y - yy pg/yJt, .7 pg/yZ <.3 jog/viJl <..4 pg/yZ < 2 pg/ yZ <*T pg/y <-3 pg/yZ' 1320/l .80 .76 -- .81 .70 .84 .52 -- -- -- Table IV f Summary of Positives || ID No. in Deer Tissue V Sample Type * FS-3 FS-4 FS-7 FS-8 FS-H FS-18 FS-I9 FS-20 FS-23 FS-24' FS-27 FS-31 FS-34 FS-36 FS-40 FS-42 FS-45 .5 Deer Muscle it n 'M. II <F FvV in' 6 1* It ir It *- 4-' /f Deer Adipose rz*. {- tV d. it> n HM . ' It" r'r-i f-L* is n - It" H r-jTr F>' 3 Deer Liver * ;* 1 I* F <1 W n *t nBone Marrow r-rf-' K'1 ^ r -f-V \r e <? F i^-or r-< -S - ^ <U.C\ ^*'v-- ^ ' * .ro ' Nature, of -Sample Cone. r etection Limit Percent Recovery Cone./ ^322^ f S-20 Deer adipose 2.9 :S-21*2 n *^1 is-* 3 -S-22*3 FS-23 FS-24 FS-25 FS-26 FS-27 " 1.6 ; lbff*~4.9 If- 2.2 II nd II (02.3 ^ * 3.5 1 3 1 1 T 1 1 .7 ;85 3,0 95 5.4 50 , <1 60 4.5 55 'fc- 4.6 65 65 <2 75 3.1 rLOs. f * Cone. 1320/322 ^322^ 3.5: 5.7 -- 7.7 8.4- - 4-.1 % ,75 .57 .62 .72 .68 - .73 c } j 1 ! .* * iI 1 R A o 'l r rO- Une<Dn . . Fv <C.VO.. C\~ U ,/ / MCMS/y.'lL >21(Ul RUN NO.' T/Cl$ SAMPLE F 5 'f GAIN KcoA J AT______ f K VOL. RATIO 17J 97J_ R.T. 2>-f ` SWEEPS Y / ------------------- ---------- \I \ tnotes: 1-J22 corrected from less than 100% recovery to 100% recovery. 1. Ratio distorted due to large PC8 enhancement at m/e 322 decreasing the ratio i320/1322. ' 2. The ratios of 0.60 or 1ess are suggestive of residuals in the initial standard and should be considered negative. 3. S/N poor on validation, however 132(/1322 is extractable from the data. 4. S/N poor on validation, and peak heights not measurable. Should be considered as negative. 5. Isotope ratio not in accord with a positive. ' V J 1 I \ \ t3- n studied this report. -' B. Samples giving "Positive11 Results: The validity of our assignment of "positives" can be judged by examing the control samples. At this writing., these are unknown to us. The following Table IV is a summary of samples judged definitely to be positive based, on the internal standard method and the validation work. All other samples should be considered "negative" at the reported detection limit. I o /y