Are Enzymes Accurate Indicators of Postmortem Interval?: a Biochemical Analysis Karly Laine Buras Louisiana State University and Agricultural and Mechanical College
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Louisiana State University LSU Digital Commons LSU Master's Theses Graduate School 2006 Are enzymes accurate indicators of postmortem interval?: a biochemical analysis Karly Laine Buras Louisiana State University and Agricultural and Mechanical College Follow this and additional works at: https://digitalcommons.lsu.edu/gradschool_theses Part of the Social and Behavioral Sciences Commons Recommended Citation Buras, Karly Laine, "Are enzymes accurate indicators of postmortem interval?: a biochemical analysis" (2006). LSU Master's Theses. 177. https://digitalcommons.lsu.edu/gradschool_theses/177 This Thesis is brought to you for free and open access by the Graduate School at LSU Digital Commons. It has been accepted for inclusion in LSU Master's Theses by an authorized graduate school editor of LSU Digital Commons. For more information, please contact [email protected]. ARE ENZYMES ACCURATE INDICATORS OF POSTMORTEM INTERVAL? A BIOCHEMICAL ANALYSIS A Thesis Submitted to the Graduate Faculty of the Louisiana State University and Agricultural and Mechanical College in partial fulfillment of the requirements for the degree of Master of Arts in The Department of Geography and Anthropology by Karly Laine Buras B.S., Louisiana State University, 2001 May 2006 ACKNOWLEDGEMENTS I would like to thank the members of my thesis committee for their continued encouragement and help throughout my program. My heartfelt thanks go out to Bob Tague, Mary Manhein, and Grover Waldrop for helping me along the way. I would especially like to thank Grover Waldrop for his contributions to my thesis expenses, as well as for the use of his lab for my research. I would not have been able to do any of this without his help. To my statistics professor, Dr. Lynn Lamotte, I owe my eternal gratitude for all of his help with my data and statistical procedures. I would also like to thank the people at the Louisiana State University School of Veterinary Medicine, who helped me with my initial research on the rats used in the study. Finally, I would like to thank my family and friends who have supported me along the way. I couldn’t have done this without each of them. ii TABLE OF CONTENTS Acknowledgements……………………………………………………………………….ii List of Tables……………………………………………………………………………..iv List of Figures………………………………………………………………………….….v Abstract………………………………………………………………………………….vii Introduction……………………………………………………………………………….1 Literature Review…………………………………………………………………………6 Materials and Methods…………………………………………………………………..16 Results……………………………………………………………………………………20 Discussion.……………………………………………………………………………….75 Works Cited……………………………………………………………………………...80 Vita……………………………………………………………………………………….83 iii LIST OF TABLES 4.1. PI Values of Knots in Figure 4.1…………………………………………………...21 4.2. PI Values of Knots in Figure 4.2…………………………………………………...23 4.3. PI Values of Knots in Figure 4.3…………………………………………………...23 4.4. PI Values of Knots in Figure 4.4…………………………………………………...25 4.5. PI Values of Knots in Figure 4.5…………………………………………………...27 4.6. PI Values of Knots in Figure 4.6…………………………………………………..27 4.7. PI Values of Knots in Figure 4.7…………………………………………………..29 4.8. PI Values of Knots in Figure 4.8…………………………………………………..29 4.9. PI Values of Knots in Figure 4.9…………………………………………………..31 4.10. PI Values of Knots in Figure 4.10………………………………………………..33 4.11. PI Values of Knots in Figure 4.11………………………………………………..33 4.12. PI Values of Knots in Figure 4.12………………………………………………..35 4.13. PI Values of Knots in Figure 4.13………………………………………………..35 4.14. PI Values of Knots in Figure 4.14………………………………………………..37 4.15. PI Values of Knots in Figure 4.15………………………………………………..39 4.16. PI Values of Knots in Figure 4.16………………………………………………..39 4.17. PI Values of Knots in Figure 4.17………………………………………………..41 4.18. PI Values of Knots in Figure 4.18………………………………………………..43 4.19. PI Values of Knots in Figure 4.19………………………………………………..43 4.20. PI Values of Knots in Figure 4.20………………………………………………..45 iv LIST OF FIGURES 4.1. LDH activity in outside liver samples.…………………………….………………22 4.2. LDH activity in cold room liver samples………..……………...…………………24 4.3. CK activity in outside liver samples……….………..……………………………..26 4.4. CK activity in cold room liver samples…………..………………………………...28 4.5. AST activity in outside liver samples………………………………………………30 4.6. AST activity in cold room liver samples…………….……….…………………….32 4.7. ADH activity in outside liver samples……………………………………………...34 4.8. ADH activity in cold room liver samples…………….……………….……………36 4.9. LDH activity in outside muscle samples…………..……………………………….38 4.10. LDH activity in cold room muscle samples…………….…………………….…...40 4.11. CK activity in outside muscle samples…………...……………………………….42 4.12. CK activity in cold room muscle samples..…..…………………………………...44 4.13. AST activity in outside muscle samples…………...…………………….………..46 4.14. AST activity in cold room muscle samples……………………………………….47 4.15. LDH activity in outside heart samples…………………………………………….48 4.16. LDH activity in cold room heart samples………………………………………....49 4.17. CK activity in outside heart samples…………...…………………………………50 4.18. CK activity in cold room heart samples…………………………………………...51 4.19. AST activity in outside heart samples…………...………………………………..52 4.20. AST activity in cold room heart samples…………….…………………………....53 v 4.21. Lactate Dehydrogenase (Outside Liver Samples)……….……………………….55 4.22. Lactate Dehydrogenase (Cold Liver Samples)….…………..……………………56 4.23. Creatine Kinase (Outside Liver Samples)……………..…..……………………..57 4.24. Creatine Kinase (Cold Liver Samples)….………….………..…………………...58 4.25. Aspartate Aminotransferase (Outside Liver Samples)….…….………………….59 4.26. Aspartate Aminotransferase (Cold Liver Samples)….………..………………….60 4.27. Alcohol Dehydrogenase (Outside Liver Samples)……………..………………...61 4.28. Alcohol Dehydrogenase (Cold Liver Samples)…………………..………………62 4.29. Lactate Dehydrogenase (Outside Muscle Samples)……….………..……………63 4.30. Lactate Dehydrogenase (Cold Muscle Samples)……….………………………...64 4.31. Creatine Kinase (Outside Muscle Samples)……………..……………………….65 4.32. Creatine Kinase (Cold Muscle Samples)……………..…………………………..66 4.33. Aspartate Aminotransferase (Outside Muscle Samples)….……………………...67 4.34. Aspartate Aminotransferase (Cold Muscle Samples)….…………………………68 4.35. Lactate Dehydrogenase (Outside Heart Samples)……….……………………….69 4.36. Lactate Dehydrogenase (Cold Heart Samples)……….…………………………..70 4.37. Creatine Kinase (Outside Heart Samples)……………..…………………………71 4.38. Creatine Kinase (Cold Heart Samples)……………...……………………………72 4.39. Aspartate Aminotransferase (Outside Heart Samples)…..……………………….73 4.40. Aspartate Aminotransferase (Cold Heart Samples)…..…………………………..74 vi ABSTRACT There are numerous ways to estimate postmortem interval (PMI), or time since death, including body temperature, rigor mortis, insect activity, and decomposition. Individually, many of these indicators are prone to inaccuracy due to the influence of the external environment upon them. This study proposed that in addition to or in conjunction with these and other indicators, certain enzymes could be used to accurately determine PMI, namely aspartate aminotransferase (AST), alcohol dehydrogenase (ADH), creatine kinase (CK), and lactate dehydrogenase (LDH). In this project, 18 rats were studied postmortem to determine how ethanol consumption and different environments affect decomposition and enzyme activity. Three groups of six rats each were studied at three different times. Two rats in each group of six received Treatment 1 (0 mg/dL ethanol), two received Treatment 2 (75 mg/dL ethanol), and two received Treatment 3 (200 mg/dL ethanol). One hour after treatments were given, the rats were euthanized in a CO 2 chamber. After euthanasia one rat from each treatment pair was put into a cold room (approximately 40 ° F) and the other was put outside. Liver tissue, cardiac muscle, and skeletal muscle samples were taken periodically and frozen at -80 ° C. Samples were homogenized and centrifuged at 12x g for one hour and put back into -80 ° C until analysis on the spectrophotometer. After this analysis, the data were compiled using the SAS/IML version 9.1 program. Graphs for each enzyme and tissue type were made, illustrating the positions of all data points in relation to the prediction interval deduced from the statistical computations. Visually and statistically, the graphs vii did not seem to exhibit any discernible pattern with regard to enzyme velocity. Alcohol did not appear to affect the enzyme activity. This study did not find a pattern for using enzyme velocity to determine PMI. Further research projects could attempt to study other variables not considered in this project. Studies using larger animals, different environments, and different taphonomic conditions would all be projects that could enhance this subfield of forensics. viii INTRODUCTION Postmortem interval (PMI), or time since death, is a matter of crucial importance in any death investigation, but especially those involving homicide. In the investigation of deaths from any cause, the evaluation of changes in the body requires careful consideration of all variables one may encounter. One must record pertinent variables such as ambient temperature, relative humidity, covering or clothing of the body, general structure and supporting structure of the body, presence and activity of scavengers, wind currents, and other influencing factors. Also considered must be additional items, including events which occurred prior to death: existence of injury; relationship of injury and death; interval between injury and death; cause, mode,