1 15 3 Dublin Core The Dublin Core metadata element set is common to all Omeka records, including items, files, and collections. For more information see, http://dublincore.org/documents/dces/. Title A name given to the resource Alvin L. Young Collection on Agent Orange Description An account of the resource The Alvin L. Young Collection on Agent Orange comprises 120 linear feet and spans the late 1800s to 2005; however, the bulk of the coverage is from the 1960s to the 1980s and there are many undated items. The collection was donated to Special Collections of the National Agricultural Library in 1985 by Dr. Alvin L. Young (1942- ). Dr. Young developed the collection as he conducted extensive research on the military defoliant Agent Orange. The collection is in good condition and includes letters, memoranda, books, reports, press releases, journal and newspaper clippings, field logs and notebooks, newsletters, maps, booklets and pamphlets, photographs, memorabilia, and audiotapes of an interview with Dr. Young. For more about this collection, <a href="/exhibits/speccoll/exhibits/show/alvin-l--young-collection-on-a">view the Agent Orange Exhibit.</a> Text A resource consisting primarily of words for reading. Examples include books, letters, dissertations, poems, newspapers, articles, archives of mailing lists. Note that facsimiles or images of texts are still of the genre Text. Box The box containing the original item. 094 Folder The folder containing the original item. 2396 Series The series number of the original item. Series IV Subseries IV Dublin Core The Dublin Core metadata element set is common to all Omeka records, including items, files, and collections. For more information see, http://dublincore.org/documents/dces/. Creator An entity primarily responsible for making the resource Hutter, R. M. Philippi Description An account of the resource Corporate Author: Regione Lombardia, Ufficio Speciale di Seveso Date A point or period of time associated with an event in the lifecycle of the resource 1984-09-01 Title A name given to the resource Typescript: Paper from September 1984 conference entitled, "Studies on Microbial Metabolism of TCDD under Laboratory Conditions" Subject The topic of the resource dioxin microbial populations herbicide toxicology Dublin Core The Dublin Core metadata element set is common to all Omeka records, including items, files, and collections. For more information see, http://dublincore.org/documents/dces/. Title A name given to the resource Alvin L. Young Collection on Agent Orange Description An account of the resource The Alvin L. Young Collection on Agent Orange comprises 120 linear feet and spans the late 1800s to 2005; however, the bulk of the coverage is from the 1960s to the 1980s and there are many undated items. The collection was donated to Special Collections of the National Agricultural Library in 1985 by Dr. Alvin L. Young (1942- ). Dr. Young developed the collection as he conducted extensive research on the military defoliant Agent Orange. The collection is in good condition and includes letters, memoranda, books, reports, press releases, journal and newspaper clippings, field logs and notebooks, newsletters, maps, booklets and pamphlets, photographs, memorabilia, and audiotapes of an interview with Dr. Young. For more about this collection, <a href="/exhibits/speccoll/exhibits/show/alvin-l--young-collection-on-a">view the Agent Orange Exhibit.</a> Text A resource consisting primarily of words for reading. Examples include books, letters, dissertations, poems, newspapers, articles, archives of mailing lists. Note that facsimiles or images of texts are still of the genre Text. Box The box containing the original item. 056 Folder The folder containing the original item. 1510 Series The series number of the original item. Series III Subseries II Dublin Core The Dublin Core metadata element set is common to all Omeka records, including items, files, and collections. For more information see, http://dublincore.org/documents/dces/. Creator An entity primarily responsible for making the resource Andersen, Kenneth J. Edith G. Leighty Mark T. Takahashi Source A related resource from which the described resource is derived Journal of Agricultural and Food Chemistry Date A point or period of time associated with an event in the lifecycle of the resource 1972 Title A name given to the resource Evaluation of Herbicides for Possible Mutagenic Properties Subject The topic of the resource herbicide testing chromosomal aberrations microbial populations ao_seriesIII https://www.nal.usda.gov/exhibits/speccoll/files/original/9346e6068481f593b699f8853bda813b.pdf 553b9c533e1e82c5b45c3ef79964b2ae PDF Text Text Item ID Number Author CorpOratB Author 00184 Patrick, Michael A. Environics and Human Factors Office, Air Force Armament Laboratory, Armament Development and Test Center, Eglin AFB, Florida Toxicological and Recalcitrant Properties of a Proposed Propellant Ingredient, Triaminoguanidine Nitrate (TAGN). I. Microbiological Study Journal/Book Title Year 1976 November Wnr Number of Images Project No. 5066; Task No. 01 ; Work Unit No. 001 Friday, January 05, 2001 Page 184 of 194 �AFATL-TR-76-139 TOXtCOLOGICAL AND RECALCITRANT PROPERTIES OF A PROPOSED PROPELLANT INGREDIENT, TRIAMINOGUANIDINE NITRATE (TAGN) I. MICROBIOLOGICAL STUDY ENVIRONICS AND HUMAN FACTORS OFFICE NOVEMBER 1976 FINAL REPORT: APRIL - NOVEMBER 1976 Approved for public release; distribution unlimited AIR FORCE ARMAMENT LABORATORY AIR F O R C E S Y S T E M S COMMAND • UNITED S T A T E S AIR F O R C E EGLIN AIR F O R C E B A S E , F L O R I D A �UNCLASSIFIED SECURITY CLASS'FICATION OF THIS =>AG€ (When Rate Entered) READ INSTRUCTIONS BEFORE COMPLETING FORM 2. GOVT ACCESSION NO, 3. RECI^'FWT'S CATALOG NUMBER REPORT DOCUMENTATION PAGE 1, REPORT NUMDER AFATL-TR-76-139 4. TITLE (and Subtitle) 5. TVFE OP REPORT * PERIOD COVERED Final Report - April to November 1976 TOXICOLOGICAL AND RECALCITRANT PROPERTIES OF A PROPOSED PROPELLANT INGREDIENT, TRIAMINOGUANIDINE NITRATE (TAGN). I. MICROBIOLOGICAL STUDY 6. PERFORMING ORG. REPORT NUMBER 7. AUTHORfs) 8. CONTRACT OR GRANT NUMBERfa) Michael A. Patrick, Lt, USAF 10. PROGRAM ELEMENT. PROJECT, TASK APE* ft WORK UNIT NUMBERS 9. PERFORMING ORGANIZATION NAME AND ADDRESS Environics and Human Factors Office Air Force Armament Laboratory Eglin Air Force Base, Florida 32S42 Project No. 5066 Task No. 01 Work Unit No. 001 tl. CONTROLLING OFFICE NAME AND ADDRESS 12. REPORT DATE Air Force Armament Laboratory Armament Development and Test Center Eglin Air Force Base, Florida 32542 November 1976 14. MONITORING AGENCY NAME ft AODRESSfff different from Controlling Office) 15. SECURITY CLASS, (at thlt report) 13. NUMBER OF PAGES 72 UNCLASSIFIED 15«. OECLASSIFtC ATI ON/ DOWNGRADING SCHEDULE 16. DISTRIBUTION STATEMENT (of this Report) Approved for public release; distribution unlimited. 17. DISTRIBUTION STATEMENT (of th» abttroct entered in Block 20, If different tram Report) 18. S U P P L E M E N T A R Y NOTES Available in DDC. 9. K E Y WORDS (Continue on reverse side It necessary and identify by block number) Toxicological Properties Recalcitrant Properties Triaminoguanidine Nitrate Microbial Populations 20. ABSTRACT (Continue on reverse mid* If necessary end Identity by block number) The toxicological and recalcitrant properties of a proposed propellant ingredient, triaminoguanidine nitrate (TAGN), were investigated. Pure cultures of microorganisms isolated from Eglin Air Force Base, Florida, as well as cultures obtained from US Army Natick Laboratories, Natick, Massachusetts were exposed to TAGN and evaluated. During the course of this investigation, it was determined that microbial populations were not adversely affected by short-term exposure to TAGN. The following parameters were not significantly DD 1473 EDITION OF 1 NOV 65 IS OBSOLETE UNCLASSIFIED SECURITY CLASSIFICATION OF THIS PAGE (When Dmtu Entm-nS) �UNCLASSIFIED SCCURITY CLASSIFICATION OF THIS FAG£fiMj«n Da** Entered) (Item 20 concluded) altered by TAGN concentrations up to 50 ppm: ' growth rate, respiratory activity, and viability. At concentrations greater than 100 ppm, TAGN was bacteriostatic but not bacteriocidal. Of the two bacteria tested, Pseudoaonas aerugi.nosa and Escherichia coli, both were capable of removing (degrading) TAGN from aqueous solution. UNCLASSIFIED SECURITY CLASSIFICATION OF THIS PAGEfflfmn D*>* �PREFACE This technical report is the result of research conducted by the Air Force Armament Laboratory, Armament Development and Test Center, Eglin Air Force Base, Florida, from April 1976 to November 1976 under Air Force Exploratory Development Project 50660101. Reference to specific manufacturers or suppliers of scientific equipment used in this study is for the sole purpose of identification and does not constitute endorsement of these products by the United States Air Force. The assistance of Cadet Ron Alford, USAF Academy, in the bacteriocidal portion of this study is gratefully acknowledged. This report has been reviewed by the Information Office (01) and is releasable to the National Technical Information Service (NTIS). At NTIS, it will be available to the general public, including foreign nations. This technical report has been reviewed and is approved for publication. FOR THE COMMANDER: A. FARMER Chief, Environics and Human Factors Office i (The reverse of this page is blank) ��TABLE OF CONTENTS Section I II III IV Title Page 1 INTRODUCTION MATERIALS AND METHODS, . . . Cultures . . . . . Inhibitory Studies . . Bacteriocidal Effects Oxygen Uptake. . . . . . . . TAGN Degradation . . . . . . . . ..... ... RESULTS AND DISCUSSION CONCLUSIONS. . . . . . . . ill 3 3 3 3 3 4 5 . . 7 �LIST OF FIGURES Figure 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 Title Page Standard Growth Curves of (A) Pseudoaonas aeruginosa QMB 1468 and (B) Bacillus megaterium QMB 1605 Exposed to TAGN . . 8 Standard Growth Curves of (A) Staphyj.ococcus aureus QMB 1458 and TO Bacillus cereus QMB 1597 Exposed to TAGN. . 10 Standard Growth Curves of (A) Escherichia. coli QMB 1557 and (B) SR 401 Exposed to TAGN , 12 Standard Growth Curves of (A) SR 403 and (B) SR 404 Exposed to TAGN , 14 Standard Growth Curves of (A) SR 405 and (B) SR 407 Exposed to TAGN 16 Standard Growth Curves of (A) SR 408 and (B) SR 409 Exposed to TAGN . 18 Standard Growth Curves of (A) C 1 and (B) C 4 Exposed to TAGN . . . . . . . . . . . . . . . . . . . . 20 Standard Growth Curves of (A) T 6 and (B) Arthrobacter sp. QMB 1631 Exposed to TAGN. 22 Standard Growth Curves of (A) Bacillus subtilis QMB 1611 and (B) Serratia marcescens QMB 1466 Exposed to TAGN 24 Standard Growth Curves of (A) SR 402 and (B) SR 406 Exposed to TAGN . . . . . . . . . . . . . . . . 26 Standard Growth Curves of (A) T 4 and (B) T 100 Exposed to TAGN ............ 28 Endogenous (A) and Exogenous (B) Oxygen Uptake by Pseudomonas aeruginpsa QMB 1468 Exposed to 500 ppm TAGN . 30 Endogenous (A) and Exogenous (B) Oxygen Uptake by Bacillus megaterium QMB 1605 Exposed to 500 ppm TAGN 32 Endogenous (A) and Exogenous (B) Oxygen Uptake by Staphylococcus aureus QMB 1458 Exposed to 500 ppm TAGN. . . . . . 34 Endogenous (A) and Exogenous (B) Oxygen Uptake by Serratia_ marcescens QMB 1466 Exposed to 500 ppm TAGN 36 Endogenous (A) and Exogenous (B) Oxygen Uptake by Escherichia coli QMB 1557 Exposed to 500 ppm TAGN . . . . . . . . 38 Endogenous (A) and Exogenous (B) Oxygen Uptake by Arthrobacter sp. QMB 1631 Exposed to 500 ppm TAGN . . . . . . . . 40 Endogenous (A) and Exogenous (B) Oxygen Uptake by Bacillus cereus QMB 1597 Exposed to 500 ppin TAGN 42 IV �LIST OF FIGURES (CONCLUDED) Figure 19 20 Title Page Endogenous (A) and Exogenous (B) Oxygen Uptake by SR 402 Exposed to 500 ppm TAGN 44 Endogenous (A) and Exogenous (B) Oxygen Uptake by SR 404 Exposed to 500 ppm TAGN 21 22 23 24 46 Endogenous (A) and Exogenous (B) Oxygen Uptake by SR 406 Exposed to 500 ppm TAGN . 48 Endogenous (A) and Exogenous (B) Oxygen Uptake by SR 407 Exposed to 500 ppm TAGN 50 Endogenous (A) and Exogenous (B) Oxygen Uptake by SR 408 Exposed to 500 ppm TAGN. 52 Endogenous (A) and Exogenous (B) Oxygen Uptake by SR 410 Exposed to 500 ppm TAGN 25 54 Endogenous (A) and Exogenous (B) Oxygen Uptake by C 4 Exposed to 500 ppm TAGN 26 27 56 Disappearance of TAGN from Cultures of (A) Pseudomonas aeruginosa QMB 1468 and (B) Escherichia coli QMB 1557 as a Function of Cell Density (O.D.) Thin-Layer Chromatogram Depicting the Disappearance of TAGN from the Cell-Free Supernatant of a Growing Culture of Pseudomonas aeruginosa as a Function of Time 58 . 60 LIST OF TABLES Table 1 2 Title Page Exposure of Bacterial Cultures Obtained from US Array Natick Laboratories to TAGN 61 Exposure of Bacterial Cultures Indigenous to Eglin AFB, Florida, to TAGN. . 63 v (The reverse of this page is blank) ��SECTION I INTRODUCTION An experimental propellant consisting of approximately 45 percent triaminoguanidine nitrate (TAGN), 19 percent nitrocellulose (NC), 30 percent cyclotetramethylene tetranitramine (HMX), 5 percent isodecyl pelargonate, and I percent resorcinol is being considered by the Air Force for use in gun ammunition employing high-density, armor-piercing penetrators. It is a matter of environmental policy to determine toxicity and evaluate methods for disposal of new propellant constituents before inventory acquisitions. Prior to this study, no information was available concerning the biodegradation or toxicity of the major component, TAGN. The objectives of this initial study were to investigate whether TAGN is degradable by microorganisms and to determine if TAGN adversely affects microbial populations indigenous to soil and water habitats where appreciable amounts of TAGN may accumulate during propellant testing and disposal. 1 (The reverse of this page is blank) ��SECTION II MATERIALS AND METHODS CULTURES Bacterial strains used in this study were obtained either from US Army Natick Laboratories, Natick, Massachusetts, or isolated from soil or water samples collected at Eglin Air Force Base, Florida. Original cultures were preserved under liquid nitrogen. Subcultures were maintained on Trypticase soy agar (TSA) at 4°C and were transferred monthly. INHIBITORY STUDIES Starter cultures were grown overnight in Trypticase soy broth (TSB) with agitation at 20°C and were diluted 1:10 in 15-mM phosphate buffer (pH 6.6) prior to use. Experiments were initiated by inoculating 0,1-aJt cells into S.Q-m£ filter-sterilized (0.45-um Millipore filters) TSB containing TAGN at concentrations of 500, 100, 50, or 10 ppm. Control samples contained no TAGN. All experiments were performed in triplicate at 20°C with agitation on a gyratory shaker (120 rpm). Growth was monitored periodically by recording the optical density (O.D.) of each sample with a Bausch and Lomb Spectronic 20 Spectrophotometer at 520 nm.1 All sanples were corrected for O.D. discrepancies due to tube variations and TAGNinduced absorption. BACTERIOCIDAL EFFECTS Cultures were grown overnight in 50-mH TSB at 25°C with agitation and were harvested at late log or stationary phase by centrifugation at 6,000 rpm for 15 minutes in an IEC/B20 refrigerated centrifuge. Following resuspension in sterile phosphate buffer, the cells were diluted to give a final O.D. reading of 0.2 at 520 nm. Diluted samples (5.0 mi) were added in duplicate to 5.0-m£ phosphate buffer containing TAGN at final concentrations of 0, 500, and 2,000 ppm. Samples were shaken in sterile 15-m£ centrifuge tubes at 25°C for 1-hour or 5-hour time periods. Afterwards, the exposed cells were centrifuged at 6,000 rpm for 15 minutes to remove TAGN and were resuspended in equal amounts of phosphate buffer. Samples were subsequently diluted with buffer to a final titer of 3.0 x 10a 3.0 x 10s colony-forming units/mi (CFU/m£) and were spread plated in triplicate. Following incubation overnight at 34°C, the plates were counted with the aid of a Quebec Colony Counter. OXYGEN UPTAKE Cultures were grown at 25°C, harvested, and resuspended in phosphate buffer as previously described. Endogenous preparations contained 1.5-ntfc �cells, 1.5-ml phosphate buffer, and a final TAGN concentration of 500 ppm, Exogenous preparations contained 1.5-mi cells, 0.5-mH phosphate buffer, 1.0-m£ TSB, and a final TAGN concentration of 500 ppm. Exogenous samples were pre-incubated 30 minutes at 30°C prior to data collection. All control samples were identically prepared but contained no TAGN, Oxygen uptake measurements were performed with a YSI 5331 Oxygen Probe at 30°C with airsaturated solutions. TAGN DEGRADATION Twelve liter fermentors (Virtis Research Equipment, Gardiner, New York) containing 10 £ of a mineral salts medium consisting of 1,79 g/Ji KH2POi», 1.65 g/£ Na2HPO% • 7H20, 0.12 g/£ MgSQK, 0,03 g/£ CaCl2, and 5.0 g/£ glucose were inoculated with 25 m£ of an actively growing culture of either Pseudomonas aerujinosa or Escherichia coli.2 Immediately following inoculation, filter-sterilized TAGN was added to give a final concentration of 61 ppm for the P. aeruginosa culture and 75 ppm for the E_, coli culture. NaNOs (0.7 g/£) was added to each fermentor at a designated time following inoculation. Periodically, samples were aseptically withdrawn, centrifuged at 6,000 rpm for 15 minutes, and analyzed for TAGN by a modification of the ninhydrin assay.3 Freshly prepared ninhydrin reagent (3.0 m£3 was added to l.Q-mJl sample and heated for 5 minutes in a boiling water bath. The sample was cooled, and the resulting optical density was determined at 570 nm against a blank containing no TAGN. Culture densities were aeasured prior to centrifugation at 520 nm. In order to rule out the effects of other ninhydrin reacting substances, 50 u£ supernatant samples were spotted on silica gel GF thin-layer plates (Analtech, Inc., Pittsburgh, Pennsylvania) and were developed against a solvent system of methanol-water-methyl sulfoxide (40:30:30). Plates were sprayed with 0.2 percent ninhydrin in watersaturated butanol. �SECTION III RESULTS AND DISCUSSION Growth of the majority of the bacterial isolets examined was not adversely affected by TAGN concentrations up to 50 ppm. However, at 100 ppm and above, 16 of the 22 bacterial cultures tested were markedly inhibited (Figures 1 to 8). Since continued incubation of these cultures for up to seven days did not result in further growth initiations, it must be assumed that the inhibitory effects of TAGN were absolute and not merely the result of greatly extended lag periods. Of the 6 remaining bacterial cultures capable of growth in the presence of 100 ppm TAGN (Figures 9 to 11), all exhibited prolonged lag phases prior to logarithmic growth. Although the overall growth rates of these cultures were considerably retarded, the ultimate cell densities attained, when compared to controls, were not significantly affected by exposure to TAGN. At the present time, inadequate evidence is available to explain the inhibitory actions of TAGN on bacterial cultures. However, it is clear that the inhibitory effects were not due to TAGN-induced pH changes, since the addition of this substance had no pronounced influence on initial hydrogen ion concentration of the media. To determine whether TAGN was bacteriocidal, cells were suspended in buffered solutions of TAGN at concentrations of 500 or 2,000 ppm for up to 5 hours. These concentrations were high enough to prevent cell proliferation in a suitable medium such as TSB, but as shown in Tables 1 and 2, viability of the cultures was unaffected. In every case the bacteria were capable of renewed growth following TAGN removal by centrifugation. Therefore, while TAGN was bacteriostatic under the specified conditions of this test, it was neither bacteriocidal nor significantly toxic to the microbial cultures examined . Oxygen uptake, a method of evaluating cellular oxidative capabilities, was investigated at a constant TAGN concentration of 500 ppm (Figures 12 to 25). In several instances oxygen uptake was markedly depressed by 500 ppm TAGN, as in the case of the common soil inhabitants Arthrobacter sp. (Figure 17) and Bacillus cereus (Figure 18). But the majority of the bacteria tested were not significantly influenced by exposure to TAGN. In most cases, the bacteria assimilated oxygen at nearly identical rates to those determined for the controls. A few isolets, such as StaphylocQccus^ aureus (Figure 14) and SR 406 (Figure 21), were even stimulated by exposure to TAGN. Both Pseudomonas aeruginosa and Escherichia coli were examined for their ability to degrade TAGN under batch fermenter conditions (Figure 26), In each case the bacteria significantly reduced the quantity of TAGN available in solution, persumably through degradation, although no methods were �available to determine active bioaccumulation or adsorption. Thin-layer chromatography of the resulting cell-free supernatants of the Pseudpmonas aeruginosa culture (Figure 27) failed to show the presence of any soluble TAGN byproducts, but did provide an additional method to verify the reduction of TAGN in aqueous solutions under these culture conditions. �SECTION IV CONCLUSIONS The results of this study indicate that, while TAGN was generally bacteriostatic at concentrations of 100 ppm and above, the bacteria tested were not otherwise adversely affected by concentrations as high as 2,000 ppm for contact periods up to 5 hours. Under the experimental conditions of this study, TAGN was neither bacteriocidal nor did it appreciably affect the respiratory activity of the cells. Following exposure and subsequent removal of TAGN from solution, all bacteria tested were capable of normal growth resumption. Moreover, some bacteria were capable of degrading or at least removing TAGN from solution, thereby effectively reducing the aqueous concentration of this compound as might result from testing and disposal of this proposed propellant. �1.6 1 (A) PseydoMonas aeruginosa 1,4 X 1.2 •M •H in (3 4> O 1.0 <D u o O.f <M 1/5 (fl c 0.6 (U Q en u p. o 0.4 0.2 10 Time (hours) 12 �1.6 -.1 (B) Bacillus megaterlum 1.4 , . X *J 1.2 •H t/> Q 2 1.0 SC O 0.8 - - X •M •H tfi g 0.6 Q Q •H 0.4 0.2 -- 8 10 12 14 16 18 20 Time (hours) Figure 1. Standard Growth Curves of (A) Pseudoiaonas aerugino sa QMB 1468 and (B) Bacillus megaterium QMB 1605 Exposed to TAGN �1.6 -r 2 (A) Staphylococcus aureus 1.4 - o 10 n Time (hours) 14 16 20 �l.fi-r 2 (B) Bacillus cereus 1,4- - X 1.2 • H tn c i.o- - O C. S to X g C.6 Q o • r4 10 12 14 16 18 Time (hours) Figure 2. Standard Growth Curves of (A) Staphylococcus aureus QMB 1458 and (B) Bacillus cereus QMB 1597 Exposed to TAGN �1.6 — 3 (A) Escherlchia coll 1.4- X 1.2. tn (3 O Q 1.0 - fl> U 3E C 0.8 - - CM LTS X +-> •H w C (U Q 0.6 - - tfl o •r-l *J 0.4 . . 0.2 . . Time (hours) �1.6 T 3 (B) SR 401 1.4 - - X1.2-- c Q -« 1.0 - i—i u 35 o o.e + LO X +J g 0-6 + Q ,0,4 - - 0.2 - - 50 ppm 100 ppm 500 ppm I Time (hours) Figure 3. Standard Growth Curves of (A) Escherichia coli QMB 1557 and (B) SR 401 Exposed to TAGN �4 (A) SR 403 1.4. . X 1.2. _ w C tt» o l.C o CJ s: O 0.8-- LD X 4-1 •—! £ 0.6. - ctt o .-4 a. 0.4 . _ o 0.2 - - Time (hours) 20 �1.6 -r Time (hours) Figure 4. Standard Growth Curves of (A) SR 403 and (B) SR 404 Exposed to TAGN �1.6 _ 1.4 - X 1.2 to 1.0 4» U s^/ as O 0.8 CM tn X 4-1 Pi 4> 0.6 Q O 0.2 -- 10 Time (hours) 12 14 16 18 20 �1,6-*5 (B) SR 407 1.4-- 1.2-0> o 1.0- 0) u O 0.8- CM LO W g 0.6' O n) U •H a, 0.4 O 0.2 -- 10 12 14 16 Time (hours) Figure S. Standard Growth Curves of (A) SR 405 and (B) SR 407 Exposed to TAGN 18 20 �1.6 10 Time (hours) 12 14 16 18 20 �<£> 10 Time (hours) 12 u Figure 6. Standard Growth Curves of (A) SR 408 and (B) SR 409 Exposed to TAGN 18 �7 (A) C 1 K) o Time (hours) �1.6 —. 8 10 12 14 U Time (hours) Figure 7. Standard Growth Curves of (A) C 1 and (B) C 4 Exposed to TAGN �N) K> 10 Time (hours) 12 14 16 18 �8 (B) Arthrobacter sp. 1.4. . X 1.2. . o> cs 2 i.o4- o 0.84- CN U^ •Jl g 0.6-JQ td o •H cL 0.4- . 0.2 - - 10 12 14 Time (hours) Figure 8. Standard Growth Curves of (A) T 6 and (B) Arthrobacter sp. QMB 1631 Exposed to TAGN 16 18 20 �1.6 — 9 (A) Bacillus subtilis 1.4. . X 1.2 •H c OJ a 1-0 tt» u 2 0 0.8 r-4 tn X g 0.6 o 03 O 0.2 20 Time fhours) �9 (B) Serratia marcescens a> Q •-< 1.0 - - u v_/ z O 0.8 r-j NJ cn tn g 0.6- - a 0.4-- 0.2- - 10 12 16 18 Time (hours) Figure 9. Standard Growth Curves of (A) Bacillus subtilis QMB 1611 and Serratia marcescens QMB 1466 Exposed to TAGN �1.6 -_. 20 Time (hours) �1.6 10 (B) SR 406 1.4 - - X 1.2 - . <D Q 1.0 - 0) u 3E O 0.8 fM LA tn C 0.6 o Q 0.2. . 16 Time (hours) Figure 10. Standard Growth Curves of (A) SR 402 and (B) SR 406 Exposed to TAGH IB 20 �1.6 ro 00 8 W Time (hours) 12 14 16 18 �1.6 -r 11 (B) T 100 Time (hours) Figure 11. Standard Growth Curves of (A) T 4 and (B) T 100 Exposed to TAGN �100 T 12 (A) PseudoiBonas aeruginosa (Endogenous) ( N O fti O CO 80- - o (U o TAGN (U O. Relative Rates: .Control (100 %) .500 ppm TAGN (86"r) 70- - 60 i I 10 Time (min") 12 14 16 18 �100 T 12 (B) Pseudoroonas aeruginosa (Exogenous) ^ Control \ D . 40 J_ Relative Rates -Control (100?) 500 ppm TAGN 20 10 14 16 Time (min) Figure 12. Endogenous (A) and Exogenous (B) Oxygen Uptake by Pseudomonas aeruginosa (^ffi 1468 Exposed to 500 ppm TAGN 18 20 �100 T 13 (A) Bacillus rnegaterium (Endogenous) Control 70 - - Relative Rates: V .Control (1002) .500 ppm TAGN (lOOt) I 60 10 Time (min) 12 14 I 16 18 20 �13 (B) Bacillus megateriurn (Exogenous) Relative Rates: Control (IOCS) 500 ppm TAGN (114%) 20 8 10 12 14 16 Time (rain) Figure 13. Endogenous (A) and Exogenous (B) Oxygen Uptake by Bacillus megaterium QMB 1605 Exposed to 500 ppm TAGN 18 �100 - . 14 (A) Staphilococcus aureus (Endogenous) 70 Relative Rates: .Control (100%) ,500 ppm TAGN (142%) 60 10 Time (min) 12 14 16 18 20 �100 T 14 (B) Staphylococcus aureus {Exogenous) Control 90 . _ o 3 r—i O 80. . D U SH 0> \ 500 ppm \ TAGM V \ \ N a. 70 . _ Relative Rates: .Control (100$) ,500 ppm TAGN (200%) f 60 S 10 12 a 14 16 Time (min) Figure 14. Endogenous (A) and Exogenous (B) Oxygen Uptake by Staphylococcus aureus QMB 1458 Exposed to 500 ppm TAGN 18 20 �100 . 15 (A) Serratla marcescens (Endogenous) 90 - - 500 ppm TARN <M o CD *—I o 80 - - c d> u o> ^ a. 70 . _ Relative Rates: .Control (100°*) ,500 ppm TAGN (112%) 60 10 Time 12 14 +— 16 f 18 20 �100 15 (B) Serratla inarcescens (Exogenous) 90 - - CM O ppm TAGN JO 3 80 - - c 4) o H 4) Q. 70 - - Relative Rates: .Control (100%) — -.-.—500 ppm TAGN (93%) 60 10 iz 14 Time (min) Figure 15. Endogenous (A) and Exogenous (B) Oxygen Uptake by Serratia inarcescens QMB 1466 Exposed to 500 ppm TAGN IB �100 -r- 16 (A) EscheH.chla colj (Endogenous) 90 - - . Control ( M O 0 ) 500 ppm TAGN •s r-t i—i O in 80 - CM 00 4) u 70 . . Relative Rates: .Control (100X) .500 ppm TASN (103%) 60 10 Time (min) 12 14 16 is 20 �100 16 (B) Escherichia coll (Exogenous) to OJ ( X 40 4- Relative Rates: .Control (100%) .500 ppm TAGN (118S) 20 10 12 14 16 Time (min) Figure 16, Endogenous (A) and Exogenous (B) Oxygen Uptake by Escherichia cpli QMB 1557 Exposed to 500 ppm TAGN 18 20 �100 .*17 (A) Arthrotuctar sp. (Endofenous) 500 ppir, ' v. TAGN CM O Control 3 i—i O 80. *-> 4> U f-i 70 Relative Rates: .Control (100%) ,500 ppm TA6N (56%) 60 10 Time (min) 12 14 16 18 20 �100 TT 17 (B) Arthrobacter sp. (Exogenous) 80 4- (M O 4> ! >. 500 ppm v TAGN V—* . 0 O CO 60-4- -(-I Q> CJ ^H 0) a. 404Relative Rates: .Control (1005:) .500 ppm TAGN (90?.) 20 10 Time (rain") Figure 17. Endogenous ( ) and Exogenous (B) Oxygen Uptake by A Arthrobacter sp. QMB 1631 Exposed to 500 ppm TAGN �100 T 18 (A) BacjVTus cereus (Endogenous) O C <£> O (H (U a, Relative Rates: .Control (10OT) _ _ _ _ _ _ 500 ppm TA6N (103'* 80 10 Time (min) 12 14 16 18 �100-t- 18 (B) Bacillus cereus (Exogenous) 500 ppm TAGN 90- - Control O 0> rH £3 •—t O CO 80- . c <u u CD ^ D, 70. - Relative Rates: Control (1005) 500 ppm TAGN (69°",) 60 10 14 16 Time (rain) Figure 18. Endogenous (A) and Exogenous (B) Oxygen Uptake by Bacillus cereus QMB 1597 Exposed to 500 ppm TAGN 18 20 �100 19 (A) SR 402 (Endogenous) 500 ppm TAGN 90- - Control o 0> i-H • § t-H O !) / 80- - <o o f-l 0) .. 0 70. _ Relative Rates: .Control (100%) ,500 ppm TAGN (72%) 60 10 Time (min) 1.2 14 16 18 20 �100 __- 19 (B) SR 402 (Exogenous) 90- 500 ppm TAGN (M o 3 i—t O 80 _ _ O> o !-. 03 O. 70 _ . Relative Rates: .Control (100%) 500 ppm TAGN (99*) 60 10 12 14 16 Time (min) Figure 19, Endogenous (A) and Exogenous (B) Oxygen Uptake by SR 402 Exposed to 500 ppm TAGN 18 20 �100 _ 20 (A) SR 404 (Endogenous) 90- - O tu r-1 , 0 3 c 0 U 500 ppm TAGN Control 70- . Re1atl¥e Rates: , Control (100*) 500 ppm TAGN (84%) 60 10 Time (rain) 12 34 16 18 20 �100 _ 20 (B) SR 404 (Exogenous) 80- - CM o , 0 <—I O CO 60- - c <o u ft Relative Rates: 40- - .Control (100?) 500 ppm TAGN (1125) 20 H \ 8 10 12 16 Time (min) Figure 20. Endogenous (A) and Exogenous (B) Oxygen Uptake by SR 404 Exposed to 500 ppm TAGN 18 20 �100, 21 (A) SR 406 (Endogenous) 4:1. oo 70. . Relative Rates: Control (100%) 500 ppm TASN (140%) 60 10 Time (min) 14 16 18 20 �100--. 21 (B) SR 406 (Exogenous) 90 - - o 0> I—I 3 O r—I 80-- -ft10 0) o (H 0) a. Control 70 Relative Rates: ..Control (100*) — _ — _. 500 ppm TAGN (87% 60 8 Figure 21, ^ 10 Time (rain) 12 14 16 Endogenous (A) and Exogenous (B) Oxygen Uptake by SR 406 Exposed to 500 ppm TAGN 18 �100 T 22 (ft,) SR 407 (Endogenous) 90- - CM O £L, V 500 ppm TAGN S Control 70. Relative Rates: .Control (100*) .500 ppm TAGN (91%) 60 I I 10 Time (min) 12 14 16 18 20 �1CW T 22 (B) SR 407 (Exogenous) 80 . . r-j O 3 i-H O 60 - - O !~i 0> a. 500 ppm\ TAGN \ 40 - - Relative Rates: Control (100%) 500 ppm TAGN (134%) 20 10 14 16 Time (min) Figure 22. Endogenous (A) and Exogenous (B) Oxygen Uptake by SR 407 Exposed to 500 ppm TAGN 18 20 �100-_ 23 (A) SR 408 (Endogenous) in r-o 70 Relative Rates: .Control (lOOt) 500 ppm TASN (107%) 60 10 Time (min) 12 14 16 18 20 �90-_23 (B) SR 408 (Exogenous) 70. - CN o O en 50- - 500 ppra TAGN o UJ F-t <u Cu 30. Relative Rates: .Control (100%) 500 ppm TAGN (77%) 8 10 12 14 16 Time (min) Figure 23, Endogenous (A) and Exogenous (B) Oxygen Uptake by SR 408 Exposed to 500 ppm TAGN 18 �100 T 24 (A) SR 410 (Endogenous) 90 . > o w TAGN GJ F-H •§ o I—I t/3 80 . _ Control C CO u f-l 70 _ . Relative Rates: .Control (10021) .500 ppm TAGN (102%) 60 10 Time (min) 12 16 18 �100 T 24 (B) SR 410 (Exogenous) CM O 0) f—t O 60 •M C 0) u 40 - - Relative Rates: .Control (1001) ,500 ppm TAGN (77° 20 8 10 12 14 16 Time (roin) Figure 24. Endogenous (A) and Exogenous (B) Oxygen Uptake by SR410 Exposed to 500 ppm TAGN 18 20 �100 25 (A) C 4 (Endogenous) 90 CM O 4) O to 80 c t-t O 01 500 ppm TAGN 80 __ 0) u \ N 0) o. 70 __ — Relative Rates: .Control (100S) 500 ppm TAGN (92%) 60 8 10 12 16 Time (min) Figure 25. Endogenous (A) and Exogenous (B) Oxygen Uptake by C 4 Exposed to 500 ppm TAGN �—.0,16 80 __ 26 (A) Pseudowonas aeruglnosa 70. . & O, ui z 'J DO Culture Density (O.D.; pp« TAGN 80 100 Time (hour) 120 140 160 180 �0.16 26 (B) Escherich^ia coil * en 20 40 60 80 100 1?0 140 160 Time (hour) Figure 26. Disappearance of TAGN from Cultures of (A) Pseudomonas aeruginosa QMS 1468 and Escherichia coli QMB 1557 as a Function of Cell Density ( . . 0D) �Solvent Front R f 0.11 Origin 0 hr 48 hr Mobile Phase: 72 hr 96 hr 120 hr 140 hr Standard TAGn Methane1 - Water - Dimethyl Sulfoxide (40:30:30) Figure 27. Thin-Layer Chroaatogram Depicting the Disappearance of TAGN from the Cell-Free Supernatant of a Growing Culture of Pseudomonas aeruginosa as a Function of Time 60 �TABLE 1. EXPOSURE OF BACTERIAL CULTURES OBTAINED FROM US ARMY NATICK LABORATORIES TO TAGN (N is the number of replicate samples; a is the observed significance level) Sample Incubation (hr) TAGN (ppm) N Standard Deviation Mean a 1 0 6 2.2xlO? 0.6xl07 .. . 1 500 5 l.SxlO7 0.9x10 0.114 1 2000 6 1.4x10 5 Pseudomonas aeruginosa 0 5 6.3x10 7 7 7 0.4x10 7 1.1x10 7 5 500 6 3.8x10 5 2000 4 3.2x10 1 0 6 11x10 7 1.0x10 7 7 0.2x10 6 Bacillus megaterium 6 1 500 6 5.8xl0 2.7xl0 1 2000 6 6 6.7x10 1.2x10 5 0 6 2.0x10 6 — 0.004 0.002 6 2.0x10 6 0.011 6 — 0.003 0.002 6 0.7x10 5 5 l.OxlO 0.3xl0 5 Bacillus cereus 500 — 0.007 2000 6 2.8xl06 0.6xl06 0.022 1 0 6 2.1x10 0.6x10 1 500 6 2.6xl06 2.0xl06 1 2000 6 2.9x10 5 0 6 2.7x10 5 500 6 1.2x10 5 2000 6 2.6x10 61 6 6 6 6 6 6 2.0x10 5 6 6 0.147 0.103 6 0.7x10 6 0.5x10 1.4x10 6 0.002 >0.2 �TABLE 1. EXPOSURE OF BACTERIAL CULTURES OBTAINED FROM US ARMY NATICK LABORATORIES TO TAGN (CONCLUDED) /v (N is the number of replicate samples; a is the observed significance level) Sample Incubation (hr) TAGN (ppm) N Mean Standard Deviation A- a 6 7,3xl07 I.SxlO7 500 6 7.9xlO? I.SxlO7 0.13 2000 6 6.6xlO? 1.9xl07 0.128 5 0 6 7.7xi07 O.SxlO7 5 500 6 S.lxlO7 2.1xl07 0.169 5 2000 6 7.9xlO? 1.2xl07 0.182 1 0 6 7.0xlO? 0.4xlO? 1 500 6 8.2xl07 3.3xlO? 0.109 1 2000 6 llxlO7 2,0xl07 0.002 5 0 6 8.2xlO? 2.0xl07 5 500 6 7.2xlO? 1.4xl07 5 Eseherichia coli 0 1 Serratia marcescens 1 1 Staphylococcus aureus 2000 6 8.2xl07 I.SxlO7 1 0 6 l,3xlO? 0.7xlO? 1 500 6 1.6xlO? 0.9xl07 0.138 1 2000 6 i.SxlO7 O.SxlO7 0.002 5 0 3 6,9xl07 3.1xl07 5 500 6 2.8xlO? I.SxlO7 0.042 5 2000 6 2,lxlO? 0.7xlO? 0.031 62 0.097 �TABLE 2. EXPOSURE OF BACTERIAL CULTURES INDIGENOUS TO EGLIN AFB, FLORIDA, TO TAGN A (N is the number of replicate samples; a is the observed significance level) N Mean Standard Deviation 0 6 l.SxlO7 0.2xlO? 1 500 5 l.SxlO7 0.3xl07 1 2000 6 l.SxlO7 5.6xlO? 5 0 4 2.8xl07 6.3xlO? 7 7 Sample Incubation (hr) SR 409 1 TAGN (ppm) 5 6 l.SxlO O.lxlO 5 SR 404 500 2000 6 2.3xl07 O.SxlO7 1 0 5 O.SxlO6 0.4x10 6 f\ a ___ 0.033 >0.2 — 0.194 >0.2 1 500 6 l.SxlO O.lxlO — 0.013 1 2000 5 l.lxlO6 0.2xl06 0.100 5 0 4 7.0xl06 0.6xl06 6 6 6 — <0. 00025 5 6 3.9xl0 l.lxlO 5 SR 406 500 2000 4 J.OxlO6 2.4xl06 1 0 6 5.6xl07 l.OxlO7 _ — 1 500 6 5.7xlO? 0.9xlO? >0.20 1 2000 6 S.OxlO7 0.6xlO? 5 0 5 8.7xl07 l.OxlO7 7 7 — 0.001 5 500 5 8.4xl0 l.SxlO — 0.191 5 2000 6 7.6xl07 1.9xlO? 0.076 63 �TABLE 2. EXPOSURE OF BACTERIAL CULTURES INDIGENOUS TO EGLIN AFB, FLORIDA, TO TAGN (CONTINUED) (N is the number of replicate samples; a is the observed significance level) Sample Incubation (hr) SR 402 1 TAGN (pprn) Standard Deviation a 0 6 4.3xl07 l.OxlO7 — «.— 500 6 4.8xl07 2.0xl07 0.154 1 2000 6 5.8xl07 l.SxlO7 0.024 5 0 6 5.9xl07 0.9xlO? --- 5 500 6 lOxlO7 l.OxlO7 <0. 00025 5 2000 6 S.SxlO7 0.9x10 0.002 1 0 6 4.8x10 1.9xlO? 1 500 6 4.1xl07 O.SxlO7 0.114 I 2000 6 7.4xl07 l.lxlO7 0.009 S 0 6 3.7xlQ7 l.SxlO7 -__ 5 500 6 5.2xl07 2.1xl07 0.061 S SR 408 Mean 1 SR 407 N 2000 6 2.9X107 O.SxlO7 0,092 1 0 6 2.4xl07 0.6xlO? 1 500 6 1.9xlO? 0.2xl07 0.028 1 2000 6 S.lxlO7 0.6xlO? 0.029 5 0 6 3.2xl07 0.7xlO? 7 7 A. _ __ 5 500 6 2.8xl0 0.4xl0 — 0.083 5 2000 6 3.0xl07 O.SxlO7 0.150 64 �TABLE 2. EXPOSURE OF BACTERIAL CULTURES INDIGENOUS TO EGLIN AFB, FLORIDA, TO TAGN (CONCLUDED) (N is the number of replicate samples; a is the observed significance level) Sample Incubation (hr) SR 405 1 TAGN (ppm) Standard Deviation N Mean 0 5 1.0x10 1 500 6 1 2000 6 5 0 6 5 500 6 1.2xlO? 7 1.0x10 7 1.6x10 7 1.0x10 0.4xlO? 7 0.2x10 7 0.3x10 7 0.4x10 5 2000 6 0.3xlO? 0.2xl07 7 0.1x10 65 (The reverse of this page is blank) /•>• a 7 — 0.080 — — 0.009 <0. 00025 ��REFERENCES 1. Oster, G., and A.W. Pollister, (eds.). Physical Techniques in Biological Research, New York: Academic Press, 1955, Vol I, pp. 51-76. 2. Norris, J.R., and D.W. Ribbons, (eds.), Methods in Microbiology, New York: Academic Press, 1969, Vol I, pp. 473-504. 3. Housewright, R.D., and C.B. Thorne, "Synthesis of Glutamic Acid and Glutamyl Polypeptide by Bacillus anthracis; I. Formation of Glutamic Acid by Transamination," Journal of Bacteriology, 1955, 60:89. 67 (The reverse of this page is blank) ��INITIAL DISTRIBUTION DDC 12 AUL (AUL/LSE-70-239) 1 ASD/ENFEA USAF (AF/SAMI) 1 1 Ogden ALC/MMWM AFIS/INTA Veg Con Div (SAREA-CL-V) DDR§E (Tech Lib) 2 1 1 1 USAFA/DFCBS 1 AFLC (DS) Deseret Test Cen (Tech Lib} 1 1 AFLC/MMNO SAAMA/SFOT 1 1 NWC (Tech Lib) NWL (Tech Lib) USDA/Pesticide Coordinator USDA/Agr Env Qual Inst AFSC/SDW DDR§E (Env § Life Sci) Edgewood Arsenal (SAMUEA-SA) AFSC/DEV AEDC/DEE Edgewood Arsenal (SAREA-TS-L) 1 1 1 1 1 1 1 1 1 1 Edgewood Arsenal (SAREA-CL-V) 1 CINCPAC(JSAl) USAF Env Health Lab NASA Miss Test Facility NWC Env Eng AMD (RD) 1 1 1 1 1 USA Natick Lab 1 AMRL/THE AFCEC/EQ AMRL/THT Eglin AFB: ADTC/DEN ADTC/SGPE TAWC/TRADOCLO 1 1 1 1 1 1 AFATL/DL 1 AFATL/DLOSL 9 AFATL/DLV 10 ADTC/CSV 1 69 (The reverse of this page is blank) �� Dublin Core The Dublin Core metadata element set is common to all Omeka records, including items, files, and collections. For more information see, http://dublincore.org/documents/dces/. Title A name given to the resource Alvin L. Young Collection on Agent Orange Description An account of the resource The Alvin L. Young Collection on Agent Orange comprises 120 linear feet and spans the late 1800s to 2005; however, the bulk of the coverage is from the 1960s to the 1980s and there are many undated items. The collection was donated to Special Collections of the National Agricultural Library in 1985 by Dr. Alvin L. Young (1942- ). Dr. Young developed the collection as he conducted extensive research on the military defoliant Agent Orange. The collection is in good condition and includes letters, memoranda, books, reports, press releases, journal and newspaper clippings, field logs and notebooks, newsletters, maps, booklets and pamphlets, photographs, memorabilia, and audiotapes of an interview with Dr. Young. For more about this collection, <a href="/exhibits/speccoll/exhibits/show/alvin-l--young-collection-on-a">view the Agent Orange Exhibit.</a> Text A resource consisting primarily of words for reading. Examples include books, letters, dissertations, poems, newspapers, articles, archives of mailing lists. Note that facsimiles or images of texts are still of the genre Text. Box The box containing the original item. 017 Folder The folder containing the original item. 0184 Series The series number of the original item. Series II Dublin Core The Dublin Core metadata element set is common to all Omeka records, including items, files, and collections. For more information see, http://dublincore.org/documents/dces/. Creator An entity primarily responsible for making the resource Patrick, Michael A. Description An account of the resource Corporate Author: Environics and Human Factors Office, Air Force Armament Laboratory, Armament Development and Test Center, Eglin AFB, Florida Date A point or period of time associated with an event in the lifecycle of the resource 1976-11-01 Title A name given to the resource Toxicological and Recalcitrant Properties of a Proposed Propellant Ingredient, Triaminoguanidine Nitrate (TAGN). I. Microbiological Study Subject The topic of the resource microbial populations biodegradation