Louisiana State University LSU Digital Commons LSU Historical Dissertations and Theses Graduate School 1993 Sulfadimethoxine Analysis in Channel Catfish: A Model Drug Residue Monitoring Program for Aquaculture. Calvin Charles Walker Louisiana State University and Agricultural & Mechanical College Follow this and additional works at: https://digitalcommons.lsu.edu/gradschool_disstheses Recommended Citation Walker, Calvin Charles, "Sulfadimethoxine Analysis in Channel Catfish: A Model Drug Residue Monitoring Program for Aquaculture." (1993). LSU Historical Dissertations and Theses. 5681. https://digitalcommons.lsu.edu/gradschool_disstheses/5681 This Dissertation 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 Historical Dissertations and Theses by an authorized administrator of LSU Digital Commons. For more information, please contact [email protected]. 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Ann Arbor, MI 48106 Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. SULFADIMETHOXINE ANALYSIS IN CHANNEL CATFISH: A MODEL DRUG RESIDUE MONITORING PROGRAM FOR AQUACULTURE A Dissertation 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 Doctor of Philosophy in The Interdepartmental Program in Veterinary Medical Sciences through the Department of Veterinary Physiology, Pharmacology and Toxicology by Calvin Charles Walker B.S., Mississippi State University, 1973 D.V.M., Auburn University, 1978 December, 1993 Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Dedicated to Rosanne, Phillip, and James ii Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. ACKNOWLEDGEMENTS I would like to express my appreciation to the graduate faculty members of my doctoral advisory committee for their consultations and guidance: Steven A. Barker, Ph.D., Chair Jay C. Means, Ph.D. Samuel P. Meyers, Ph.D. Steven S. Nicholson, D.V.M. David H. Swenson, Ph.D. Ronald L. Thune, Ph.D. I would also like to thank Dr. Gary L. Ter Haar and the Ethyl Corporation for providing financial support for the Ethyl Corporation Graduate Fellowship in Toxicology. Special gratitude is expressed to Dr. Steven Barker for freely sharing his knowledge of analytical toxicology and for providing the direction and support needed during this course of study. Finally, I am thankful for the support and sacrifices of my wife, Rosanne, and my children, Phillip and James, during this period of study. iii Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. TABLE OF CONTENTS DEDICATION........................................... ii ACKNOWLEDGEMENTS .................................. iii LIST OF T A B L E S .................................... vii LIST OF FIGURES.................................... ix ABSTRACT ............................................. xi CHAPTER 1 GENERAL INTRODUCTION ........................ 1 BACKGROUND .............................. 1 RESIDUE ANALYSIS ........................ 2 PURPOSE .................................. 9 REFERENCES ................................10 2 REVIEW OF LITERATURE ........................ 13 EXTRACTION AND ANALYSIS OF DRUGS IN AQUATIC S P E C I E S ........................ 13 INTRODUCTION ..............................13 DRUG RESIDUE PROBLEMS IN AQUACULTURE . 17 METHODS OF DRUG RESIDUE ANALYSIS IN AQUATIC SPECIES .......................... 21 MICROBIAL INHIBITION TESTS............. 28 SWAB TEST ON P R E M I S E S ...........30 CALF ANTIBIOTIC AND SULFA TEST . 31 LIVE ANIMAL SWAB T E S T ...........32 FAST ANTIBIOTIC SCREEN TEST .... 33 DELVOTEST P .....................34 BRILLIANT BLACK REDUCTION TEST . 35 BIOAUTOGRAPHY .................... 35 COLORIMETRIC METHODS ..................36 RECEPTOR BASED ASSAYS ................ 37 BACTERIAL CELL RECEPTOR ASSAY . 38 RADIO-IMMUNOASSAY...............41 ENZYME IMMUNOASSAY...............42 CHROMATOGRAPHIC ANALYSIS ............ 51 THIN LAYER CHROMATOGRAPHY ........ 51 CLASSICAL THIN LAYER CHROMATOGRAPHY............ 5 1 HIGH PERFORMANCE THIN LAYER CHROMATOGRAPHY ................ 55 TLC-BIOAUTOGRAPHY ............ 57 HIGH PERFORMANCE LIQUID CHROMATOGRAPHY .................... 58 GAS CHROMATOGRAPHY...............67 iv Reproduced with permission of the copyright owner. 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MASS SPECTROMETRY......................69 OTHER RESIDUE DETECTION METHODS .... 71 METHODS OF ISOLATION ......................72 EVOLVING METHODS OF RESIDUE ANALYSIS .. 79 SOLID PHASE EXTRACTION ................80 IMMUNOAFFINITY ........................81 SUPERCRITICAL FLUID EXTRACTION .... 85 MATRIX SOLID PHASE DISPERSION ........ 88 MSPD APPLIED TO AQUATIC RESOURCES .. 90 DISCUSSION ............................... 91 REFERENCES ............................... 94 3 EXTRACTION AND ENZYME IMMUNOASSAY OF SULFADIMETHOXINE RESIDUES IN CHANNEL CATFISH (Ictalurus punctatus) MUSCLE .............. 121 INTRODUCTION .......................... 121 EXPERIMENTAL .......................... 125 REAGENTS AND EXPENDABLE MATERIALS . 125 SAMPLES ........................ 126 EXTRACTION PROCEDURE .............. 127 ELISA PROCEDURES .................. 128 RESULT DETERMINATION .............. 131 RESULTS AND DISCUSSION ................ 133 PRECISION...................... 133 TEST PERFORMANCE .................. 140 REFERENCES ............................ 155 4 EXTRACTION AND LIQUID CHROMATOGRAPHIC ANALYSIS OF SULFADIMETHOXINE AND 4-N-ACETYLSULFADIMETHOXINE RESIDUES IN CHANNEL CATFISH (Ictalurus punctatus) MUSCLE AND PLASMA ........................ 160 INTRODUCTION .......................... 160 EXPERIMENTAL .......................... 163 REAGENTS AND EXPENDABLE MATERIALS . 163 EXTRACTION PROCEDURE .............. 166 LIQUID CHROMATOGRAPHIC ANALYSIS . 169 DATA AN A L Y S I S ................... 170 RESULTS AND DISCUSSION ................ 173 CHROMATOGRAPHIC SYSTEM ............ 173 LINEARITY OF RESPONSE .............. 174 LIMITS OF DETECTION AND QUANTITATION 184 INTRA-ASSAY VARIABILITY......... 184 INTER-ASSAY VARIABILITY......... 184 RECOVERY OF METHOD ................ 186 REFERENCES ............................ 195 V Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 5 PLASMA/MUSCLE RATIOS OF SULFADIMETHOXINE RESIDUES IN CHANNEL CATFISH (Ictalurus punctatus) 199 INTRODUCTION .......................... 199 EXPERIMENTAL .......................... 204 REAGENTS AND EXPENDABLE MATERIALS . 204 EXPERIMENTAL ANIMALS, DRUG ADMINISTRATION, AND SAMPLING.... 206 EXTRACTION PROCEDURE .............. 207 LIQUID CHROMATOGRAPHIC ANALYSIS .. 208 DATA A N A L Y S I S ................... 209 RESULTS AND DISCUSSION ................ 210 REFERENCES ............................ 225 6 SUMMARY AND CONCLUSIONS .................. 229 FIELD ANALYSIS OF RESIDUES ............ 230 LABORATORY ANALYSIS OF RESIDUES ........ 233 CONCLUSIONS......................... 236 APPENDIXES APPENDIX A: OPTICAL DENSITY VALUES OBTAINED WITH THE IDS SULFADIMETHOXINE ONE-STEP ELISA ........ 239 APPENDIX B: OPTICAL DENSITY VALUES OBTAINED WITH THE SIGNAL SULFAMETHAZINEEL ISA................. 242 VITA ............................................. 245 vi Reproduced with permission of the copyright owner. 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