替硝唑在鸡体内的药物动力学和残留研究
摘要
本文采用HPLC法为定量手段，研究替硝唑在三黄鸡体内的单剂量(30mg／Kg. b. w)静注和口服的药物动力学过程以及多剂量给药后组织中的残留。静注和口服给药后，分别在0.08～24h和0.25～24h定时从翼静脉采血，分离血浆，—20℃避光保存。血浆中药物用二氯甲烷：甲醇(98：2v/v)提取，有机层于50～55℃水浴下氮气吹干，残渣以三蒸水溶解，用反相高效液相色谱法(RP-HPLC)测定。检测采用Spectrasystem HPLC系统，YWG-C_(18)柱(250×4.6mm，10μm)，以甲醇：20％乙腈：水(25：25：50 v/v)为流动相，流速1.0ml／min，检测波长318nm。替硝唑对照品的最低检测限为0.0625μg／ml，浓度在0.125～64.0μg／ml时与峰面积呈良好线性关系，线性方程为y=72.57x-22261.65(r=0.9998)。替硝唑水溶液浓度在0.125、4.0、64.0μg／ml时，日内、日间变异系数分别为4.27％和6.21％，1.47％和2.24％，0.08％和1.03％。血浆标准曲线线性范围为0.2～51.2μg／ml，线性方程为y=38.34x-8167.38(r=0.9989)。血浆中替硝唑浓度在0.2、1.6、51.2μg／ml时，日内、日间变异系数分别为3.26％和7.58％，1.28％和5.72％，0.20％和2.31％，平均回收率大于90％。鸡单剂量静注和口服给药后的血药浓度—时间数据及药物动力学参数用PKANALYST软件包处理。静注给药后的药—时过程符合二室开放模型，其主要药动学参数为：分布半衰期(t_(1/2α))为0.279h，消除半衰期(t_(1/2β))4.705h，中央室表观分布容积(Vc)0.857L／kg，稳态表观分布容积(V_(dss))1.293L／kg，体清除率(Cl_b)0.206L／h．kg，药时曲线下面积(AUC)153.280mg．h／L。口服给药的药—时数据符合一级吸收一室模型，其主要动力学参数为：一级吸收速度常数(K_a)1.915h~(-1)，一级消除速度常数(K)0.099h~(-1)，表观分布容积(V)2.600L／kg，药时曲线下面积(AUC)123.446mg．h/L，生物利用度(F)80.536％，达峰时间(t_(max))2.5h，峰浓度(C_(max))10.720μg／ml。根据鸡单剂量药物代谢动力学参数设计多剂量给药方案，给药间隔(τ)为24h，其先导剂量为34.484 mg／Kg．b．w，维持剂量为30 mg／Kg．b．W，稳态平均血药浓度可至4.275μg／ml。多剂量(30mg／Kg．b．w，每日一次，连续5天)口服替硝哗后检测其在组织中的残留。于末次给药后12、24、48、
替硝哇在鸡体内的药物动力学和残留研究
72h颈静脉放血致死，取肝脏、肌肉、肾脏、脂肪，一20℃避光保存。匀
浆后用提取液二氯甲烷:甲醇(98:Zv/v)提取，50~55℃水浴下氮气吹干
后检测。测定时，室温下解冻，匀浆、提取、HPLC分析。替硝哇在鸡组
织中的最低检测限为0.025 pg/g，组织中替硝哇浓度在0.05、0.5、8
pg/g时，日内、日l’ed变异系数分别(5%和(l既(n二3)，回收率在72.0一
96.08%之间，其平均回收率为85.044%。肌肉中替硝哇含量最高，12h时
为1.1一4士0.045 pg/g，脂肪其次为0.552士0.0一8 pg/g，肾脏再其次为
0.262士0.008 pg/g，肝脏中最低，为0.160士0.006“g/g;24h后，仅肌
肉中可检测出，为0.237士0.008协g/g，其它组织均检测不出。
结果表明，反相高效液相色谱法适用于替硝哇在鸡体内的药物动力学
和残留的研究。替硝哇在鸡体内吸收快，分布广，半衰期较短，生物利用
度高。依照给药方案投药，稳态平均血药浓度高，可达到有效治疗浓度.
组织中残留量从低至高依次为肝<肾<脂肪<肌肉，休药期建议为5天.
The studies on the Pharmacokinetics of Tinidazole (30mg/kg.b.w.) in 20 chickens after single dose of intravenous and oral administration, and on the residues of Tinidazole (30mg/kg.b.w. oral administration, five days) in tissues are reported in this paper. Plasma samples were collected from brachial vein at every different interval after administration. Tinidazole in the plasma was extracted by dichloromethane: methanol(98:2 v/v) ,and evaporated to dryness at 50-5 5 water bath. The contents were determined by reversed-phase high performance liquid chromatography. Tinidazole was separated on Spectrasystem HPLC instrument using YWG-C 18 column (250 4.6mm, 10 u m),the mobile phase consisted of a degassed mixture methandol:20% acetonitrile: water (25:25:50 v/v), the range of 0.125~64.0 g/ml (r=0.9998). The plasma calibration curve was linear in the range of 0.2~51.2 g/ml (r=0.9989) ,the average recovery was >90%,the inter-day coefficient of variance was <5%(n=3),and intra-day coefficient of variance was<10%.T
he single dose intravenous and oral administrations concentration-time data were analyzed by pharmacokinetic compartment analysis soft (PKANALYST). The procession after intravenous was fit the two-compartment open model, and its main pharmacokinetic parameters were followed :t1/2, 0.279h;t]/2u 4.705h; Vc 0.857L/kg; Vdss 1.293IAg; Clb 0.206L/kg.h; AUC 153.280 mg.h/L. The
procession after oral administration fit one compartment model, and its main pharmacokinetic parameters of Tinidazole in chickens were as follows: ka 1.915h'';k 0.099hf'; V 2.600L/kg; Clb 0.259L/kg.h ; tl/27.173h; AUC 123.446 mg.h/L;F 80.536% ;Tmax 2. 5h;Cmax 10. 720 g/ml. According to the parameters of single dose administration, the administration scheme was followed :Dosing interval 24h;Priming dose 34.484mg/kg;Maintenance dose 30mg/kg;Average steady state concentration 4.275mg/kg.Residues were determined at different interval of 12,24,48,72h after continuous oral administrations. Then collect the samples of livers, muscles, kidneys, and fats after killing every group of the chickens. The samples were determined at the conditions, which was as the same as that of plasma. At 12h, Tinidazole in the muscles is higher than any other tissues then in fats and kidneys, and the livers are the lowest. Tinidazole in muscles could be determined at 72 hours, and could not be detected in other three kinds of tissues
after 24 hours. Then suggested that the withdrawal time of Tinidazole should be 5 days.
替硝哇在鸡体内的药物动力学和残留研究
72h颈静脉放血致死，取肝脏、肌肉、肾脏、脂肪，一20℃避光保存。匀
浆后用提取液二氯甲烷:甲醇(98:Zv/v)提取，50~55℃水浴下氮气吹干
后检测。测定时，室温下解冻，匀浆、提取、HPLC分析。替硝哇在鸡组
织中的最低检测限为0.025 pg/g，组织中替硝哇浓度在0.05、0.5、8
pg/g时，日内、日l’ed变异系数分别(5%和(l既(n二3)，回收率在72.0一
96.08%之间，其平均回收率为85.044%。肌肉中替硝哇含量最高，12h时
为1.1一4士0.045 pg/g，脂肪其次为0.552士0.0一8 pg/g，肾脏再其次为
0.262士0.008 pg/g，肝脏中最低，为0.160士0.006“g/g;24h后，仅肌
肉中可检测出，为0.237士0.008协g/g，其它组织均检测不出。
结果表明，反相高效液相色谱法适用于替硝哇在鸡体内的药物动力学
和残留的研究。替硝哇在鸡体内吸收快，分布广，半衰期较短，生物利用
度高。依照给药方案投药，稳态平均血药浓度高，可达到有效治疗浓度.
组织中残留量从低至高依次为肝<肾<脂肪<肌肉，休药期建议为5天.
The studies on the Pharmacokinetics of Tinidazole (30mg/kg.b.w.) in 20 chickens after single dose of intravenous and oral administration, and on the residues of Tinidazole (30mg/kg.b.w. oral administration, five days) in tissues are reported in this paper. Plasma samples were collected from brachial vein at every different interval after administration. Tinidazole in the plasma was extracted by dichloromethane: methanol(98:2 v/v) ,and evaporated to dryness at 50-5 5 water bath. The contents were determined by reversed-phase high performance liquid chromatography. Tinidazole was separated on Spectrasystem HPLC instrument using YWG-C 18 column (250 4.6mm, 10 u m),the mobile phase consisted of a degassed mixture methandol:20% acetonitrile: water (25:25:50 v/v), the range of 0.125~64.0 g/ml (r=0.9998). The plasma calibration curve was linear in the range of 0.2~51.2 g/ml (r=0.9989) ,the average recovery was >90%,the inter-day coefficient of variance was <5%(n=3),and intra-day coefficient of variance was<10%.T
he single dose intravenous and oral administrations concentration-time data were analyzed by pharmacokinetic compartment analysis soft (PKANALYST). The procession after intravenous was fit the two-compartment open model, and its main pharmacokinetic parameters were followed :t1/2, 0.279h;t]/2u 4.705h; Vc 0.857L/kg; Vdss 1.293IAg; Clb 0.206L/kg.h; AUC 153.280 mg.h/L. The
procession after oral administration fit one compartment model, and its main pharmacokinetic parameters of Tinidazole in chickens were as follows: ka 1.915h'';k 0.099hf'; V 2.600L/kg; Clb 0.259L/kg.h ; tl/27.173h; AUC 123.446 mg.h/L;F 80.536% ;Tmax 2. 5h;Cmax 10. 720 g/ml. According to the parameters of single dose administration, the administration scheme was followed :Dosing interval 24h;Priming dose 34.484mg/kg;Maintenance dose 30mg/kg;Average steady state concentration 4.275mg/kg.Residues were determined at different interval of 12,24,48,72h after continuous oral administrations. Then collect the samples of livers, muscles, kidneys, and fats after killing every group of the chickens. The samples were determined at the conditions, which was as the same as that of plasma. At 12h, Tinidazole in the muscles is higher than any other tissues then in fats and kidneys, and the livers are the lowest. Tinidazole in muscles could be determined at 72 hours, and could not be detected in other three kinds of tissues
after 24 hours. Then suggested that the withdrawal time of Tinidazole should be 5 days.
引文
1．庄无忌主编．各国食品和饲料中农药兽药残留限量大全．北京：中国对外经济贸易出版社，1995．998
2．张先洲，杨克钊，蔡洪生．反相HPLC测定替硝唑的血药浓度．中国抗生素杂志，1995．6(3)：217
3．杨毓章，胡惠容，云皓．高效液相色谱法测定血浆甲硝唑浓度．药物分析杂志，1992．12(2)：96
4．竺心影．药理学．北京：人民卫生出版社，1987．376～377
5．李端．血药浓度与给药方案．上海：上海科学技术出版社，1981
6．李发美主编．医药高效液相色谱技术．北京：人民卫生出版社，1998
7．L.R.施柰德等．实用高效液相色谱法的建立．北京：科学出版社，1998
8．奚念朱主编．药物动力学．上海：上海医科大学出版社，1990
9．陆明廉．血液浓度测定．上海：上海科学技术出版社，1982
10．刘昌孝．药物代谢动力学．湖南：湖南科学技术出版社，1980．1～2
11．严衍禄主编．现代仪器分析．北京：北京农业大学出版社．1987
12．安登魁主编．药物分析．北京：人民卫生出版社，1980．288～309
13．李士敏，胡富强，俞滢．高效液相色谱法测定含服替硝唑含片后唾液中含量的药时曲线．药物分析杂志，1997．17(5)：307
14．陈少菲等．替硝唑抗厌氧菌作用的研究．浙江医科大学学报，1994．23(1)：160
15. H.Salonies, J. P. Salo. An HPLC Study of Tinidazole Hydrolysis. Chromatogeraphia, 1993. 36: 79～82
16．熊德鑫，祝小枫，盛志勇．替硝唑对厌氧菌体外抗菌活性的研究．中国抗生素杂志，1997．22：472～476
17. Boy dzhieva, N. et al. Pharmacological and microbiological investigations of Bioshik(Tinidazole), 1987. 37(4): 41～44
18．温玉麟，张瑞文．新的一类抗厌氧微生物药物—硝基咪唑类药物．医药工业，1985．16(7)：324～329
19. Sarkiala,Jarvinen,S.Valttila, et al. Pharmacokinetics of Tinidazole in dogs and cats. Journal of veterinery Pharm and Therap, 1991. 14: 257～262
20. S.Pyorala,P.Silvennoinen,U.Hanninen, et al. Pharmacokinetics of Tinidazole in Cows-a preliminary study. J. vet. Pharmacol. Therap, 1990. 13: 425～427
21. S.Pyorala,T.Kotilainen,P.Silvennoinen,et al. Pharmacokinetics of tinidazole in the horse. J. Vet. Pharmacol. Therap, 1990. 13: 76～80
22．曾衍霜，药物代谢动力学的近年进展，中国药理学会(1982)(数学药理分册)，北京：人民卫生出版社，1983．7～27
23．冯淇辉等著．兽医药物代谢动力学．北京：科学出版社．1987．33～35
24. Satu Pyorala,Stefan Soback, Vesa Rainio, et al. Pharmacokinetics of single—dose administration of tinidazole in unweaned calves. Am. J. Vet Res, 1994. 55: 831～834
25．蔡鸿生，张先洲，李荣凌等．替硝唑的药物动力学研究．中国医疗药学杂志，1995．12：531～532
26. K. B. and J. E. Patrick. High-performance liquid Chromatographic Assay for the Antiprotozoal Agent, Tinidazole in Human Plasma. J.Pharmaceutical Sciences, 1978. 68: 599
27．张登荣．鸡病学．天津：天津科技出版社，1982．147～160
28. Laufen.H,F.Scharpf,G.Bartsch,et al. Sensitive gas chromatographic assay of tinidazole in tissue and plasma. Journal of Chromatography, 1979. 163: 217～220.
29. Anthony Hobosn-Frohock and Jayne A.Reader. Determination of DMZ residues in poultry Tissue by High-performance Liquid Chromatography. Analyst, 1983. 108: 1091～1095
30. Carignan, William Skakum, Stephen Sved, et al. Dimetridazole Residues in Pork Tissue. I .Assay by Liquid Chromatography with Electrochemical Detector. J.Assoc. OFF.ANAL.CHEM, 1988. vol71:1141～1145
30. Carignan, William Skakum, Stephen Sved, et al. Dimetridazole Residues in Pock Tissue. Ⅱ.Application of Liquid Chromatographic Method to Monitor Elimination of
Drug and its Major Metabolite. J. Assoc. OFF. ANAL. CHEM, 1988. vol71: 1146～1149
31. Mallinson. Determination of Nitroimidazole Metabolites in Swine and Turkey Muscle by Liquid Chromatography. J. OF. AOAC INTERNATIONAL, 1992. 75: 790
32. Newkirk, D. R. Gas Chromatographic Determination of Incurred Dimetridazole Residues in Swine Tissues. J. Assoc. OFF. ANAL. CHEM, 1990 .vol73: 702～704
33. Renem.I,Aert,P.Soart,Irmam.Egberink,Cornelis.A.Kan,et al. Liquid Chromatographic Multicomponent Method for Determination of Residues of Ipronidazole,Ronidazole and Dimetridazole and some Relevant Metabolites in Eggs,Plasma,and Feces and ItsUse in Depletion studies in Laying Hens. J.Assoc. OFF.ANAL.CHEM, 1991. vol74: 46
34. Royal Society of Chemistry, Burlington House, Piccadilly, London WIV OBN, UK Collaborative Study of a Method for the Determination of Dimetridazole Residues in Chicken Tissue. Analyst, 1985. 110: 1391-1393
35. J. E. Riviere, et al. Interspecies allometric analysis of comparative pharmacokinetics of 44 drugs across veterinary and Caboratory animal species. J. Vet. Pharmaol. Therap, 1997. vol 20: 453～463
36. P. G. Welling and A. M. Monro. The Pharmacokinetics of Metronidazole and Tinidazole in Man. Drug Res, 1972. vol 28:2132
37. W. Cybulski,P.Larsson,H.Tjalve, et al. Disposition of metronidazole in hens (Gallus gallus) and quails (Coturnix coturnix japonica): pharmacokinetics and whole body autoradiography. J. Vet. Pharmacol. Therap, 1996. 19: 352～358
38. Olsson. B. L, Nor. C. E. In vitro susceptibility of anaerlbic bacteria to nitroimidazoles. Scand J Infect Dis (Suppl), 1981. 26: 42
39. Sawyer.P.R.Broaden,R.M. pander, et al .Tinidazole: a review of its anstiprotozoal activity and therapeutic efficacy. Drugs, 1976. 11: 243
40. Colney Lane. Determination of Dimetridazole Residues in poultry Tissues by High-performance liguid Chromatogram aphid. Analyst, 1983. 108: 1091～1095
41．李章万，徐秀荣．色谱分析中生物样品的前处理方法．药物分析杂志，1996．16
(1)：55～58
42．朱蓓蕾主编．动物性食品药物残留．上海：上海科学技术出版社，1994．12～13
43. E. Sarkiaia, A. Jarvinen, T. Sippola, et al. Penetration of tinidazole into the gingival creviculer fluid in dogs. Research in Veterinary Science, 1992. vol 2: 52
44. Carmine, A, A. et al. Tinidazole anaerobic infections. A review of its antibacterial activity, pharmacological properties and therapeutic efficacy. Drugs, 1982. 24: 85～11
45. Hackect. L. P. Determination of metronidazole and tinidazole in luman plasma using high performance liguid chromatograply. J. Chromatogr, 1979. 175: 347
46. D. Babggot, W. D. WILSON, S. HIETALA. Clinical pharmacokinetics of metronidazole in horses. J. Vet. Pharmacol. Therap,1988. 11: 417～420
47. Ingrid, Nillsson, Ehle, et al. Liquid Chromatographic Assay for Metrondazole and Tinidazole: pharmacokinetic and Metabolic studies in Human Subjects. Antimicrobial Agents and Chemotherapy, 1981. 19: 754～760
48. Edwards.D. I. Structure-cytotoxicity relationships of nitroimidazoles in an in vitro system. Biol phys, 1982. 8(3-4): 791
49. G. L. Law, G. P. Mansfield, D. F. Muggleton, et al. Dimetridazole:A bsorption, Excretion and Metabolism in Turkeys. Nature, 1963. 97: 1024
50. Edwards ,David I. In vitro mechanisms of nitroimidazole cytotoxicity. Curr.chemother, 1981(pub, 1982). 1: 111-113
51. Knox, R. J. The mechanism of nitroimidazole damage to DNA:coulometric evidence. Int.J.Radiat. Oncol, Biol, Phys, 1984, 10(8): 1315-18
52. Laurentius. A. P, Theo. H. G. Polman, Johannes. A. van. Rhijn, et al. Biotransformation of Dimetridazole by Primary Cultures of Pig Hepatocytes. J. Agric. Food Chem, 1997. 45: 3985-3990
2．张先洲，杨克钊，蔡洪生．反相HPLC测定替硝唑的血药浓度．中国抗生素杂志，1995．6(3)：217
3．杨毓章，胡惠容，云皓．高效液相色谱法测定血浆甲硝唑浓度．药物分析杂志，1992．12(2)：96
4．竺心影．药理学．北京：人民卫生出版社，1987．376～377
5．李端．血药浓度与给药方案．上海：上海科学技术出版社，1981
6．李发美主编．医药高效液相色谱技术．北京：人民卫生出版社，1998
7．L.R.施柰德等．实用高效液相色谱法的建立．北京：科学出版社，1998
8．奚念朱主编．药物动力学．上海：上海医科大学出版社，1990
9．陆明廉．血液浓度测定．上海：上海科学技术出版社，1982
10．刘昌孝．药物代谢动力学．湖南：湖南科学技术出版社，1980．1～2
11．严衍禄主编．现代仪器分析．北京：北京农业大学出版社．1987
12．安登魁主编．药物分析．北京：人民卫生出版社，1980．288～309
13．李士敏，胡富强，俞滢．高效液相色谱法测定含服替硝唑含片后唾液中含量的药时曲线．药物分析杂志，1997．17(5)：307
14．陈少菲等．替硝唑抗厌氧菌作用的研究．浙江医科大学学报，1994．23(1)：160
15. H.Salonies, J. P. Salo. An HPLC Study of Tinidazole Hydrolysis. Chromatogeraphia, 1993. 36: 79～82
16．熊德鑫，祝小枫，盛志勇．替硝唑对厌氧菌体外抗菌活性的研究．中国抗生素杂志，1997．22：472～476
17. Boy dzhieva, N. et al. Pharmacological and microbiological investigations of Bioshik(Tinidazole), 1987. 37(4): 41～44
18．温玉麟，张瑞文．新的一类抗厌氧微生物药物—硝基咪唑类药物．医药工业，1985．16(7)：324～329
19. Sarkiala,Jarvinen,S.Valttila, et al. Pharmacokinetics of Tinidazole in dogs and cats. Journal of veterinery Pharm and Therap, 1991. 14: 257～262
20. S.Pyorala,P.Silvennoinen,U.Hanninen, et al. Pharmacokinetics of Tinidazole in Cows-a preliminary study. J. vet. Pharmacol. Therap, 1990. 13: 425～427
21. S.Pyorala,T.Kotilainen,P.Silvennoinen,et al. Pharmacokinetics of tinidazole in the horse. J. Vet. Pharmacol. Therap, 1990. 13: 76～80
22．曾衍霜，药物代谢动力学的近年进展，中国药理学会(1982)(数学药理分册)，北京：人民卫生出版社，1983．7～27
23．冯淇辉等著．兽医药物代谢动力学．北京：科学出版社．1987．33～35
24. Satu Pyorala,Stefan Soback, Vesa Rainio, et al. Pharmacokinetics of single—dose administration of tinidazole in unweaned calves. Am. J. Vet Res, 1994. 55: 831～834
25．蔡鸿生，张先洲，李荣凌等．替硝唑的药物动力学研究．中国医疗药学杂志，1995．12：531～532
26. K. B. and J. E. Patrick. High-performance liquid Chromatographic Assay for the Antiprotozoal Agent, Tinidazole in Human Plasma. J.Pharmaceutical Sciences, 1978. 68: 599
27．张登荣．鸡病学．天津：天津科技出版社，1982．147～160
28. Laufen.H,F.Scharpf,G.Bartsch,et al. Sensitive gas chromatographic assay of tinidazole in tissue and plasma. Journal of Chromatography, 1979. 163: 217～220.
29. Anthony Hobosn-Frohock and Jayne A.Reader. Determination of DMZ residues in poultry Tissue by High-performance Liquid Chromatography. Analyst, 1983. 108: 1091～1095
30. Carignan, William Skakum, Stephen Sved, et al. Dimetridazole Residues in Pork Tissue. I .Assay by Liquid Chromatography with Electrochemical Detector. J.Assoc. OFF.ANAL.CHEM, 1988. vol71:1141～1145
30. Carignan, William Skakum, Stephen Sved, et al. Dimetridazole Residues in Pock Tissue. Ⅱ.Application of Liquid Chromatographic Method to Monitor Elimination of
Drug and its Major Metabolite. J. Assoc. OFF. ANAL. CHEM, 1988. vol71: 1146～1149
31. Mallinson. Determination of Nitroimidazole Metabolites in Swine and Turkey Muscle by Liquid Chromatography. J. OF. AOAC INTERNATIONAL, 1992. 75: 790
32. Newkirk, D. R. Gas Chromatographic Determination of Incurred Dimetridazole Residues in Swine Tissues. J. Assoc. OFF. ANAL. CHEM, 1990 .vol73: 702～704
33. Renem.I,Aert,P.Soart,Irmam.Egberink,Cornelis.A.Kan,et al. Liquid Chromatographic Multicomponent Method for Determination of Residues of Ipronidazole,Ronidazole and Dimetridazole and some Relevant Metabolites in Eggs,Plasma,and Feces and ItsUse in Depletion studies in Laying Hens. J.Assoc. OFF.ANAL.CHEM, 1991. vol74: 46
34. Royal Society of Chemistry, Burlington House, Piccadilly, London WIV OBN, UK Collaborative Study of a Method for the Determination of Dimetridazole Residues in Chicken Tissue. Analyst, 1985. 110: 1391-1393
35. J. E. Riviere, et al. Interspecies allometric analysis of comparative pharmacokinetics of 44 drugs across veterinary and Caboratory animal species. J. Vet. Pharmaol. Therap, 1997. vol 20: 453～463
36. P. G. Welling and A. M. Monro. The Pharmacokinetics of Metronidazole and Tinidazole in Man. Drug Res, 1972. vol 28:2132
37. W. Cybulski,P.Larsson,H.Tjalve, et al. Disposition of metronidazole in hens (Gallus gallus) and quails (Coturnix coturnix japonica): pharmacokinetics and whole body autoradiography. J. Vet. Pharmacol. Therap, 1996. 19: 352～358
38. Olsson. B. L, Nor. C. E. In vitro susceptibility of anaerlbic bacteria to nitroimidazoles. Scand J Infect Dis (Suppl), 1981. 26: 42
39. Sawyer.P.R.Broaden,R.M. pander, et al .Tinidazole: a review of its anstiprotozoal activity and therapeutic efficacy. Drugs, 1976. 11: 243
40. Colney Lane. Determination of Dimetridazole Residues in poultry Tissues by High-performance liguid Chromatogram aphid. Analyst, 1983. 108: 1091～1095
41．李章万，徐秀荣．色谱分析中生物样品的前处理方法．药物分析杂志，1996．16
(1)：55～58
42．朱蓓蕾主编．动物性食品药物残留．上海：上海科学技术出版社，1994．12～13
43. E. Sarkiaia, A. Jarvinen, T. Sippola, et al. Penetration of tinidazole into the gingival creviculer fluid in dogs. Research in Veterinary Science, 1992. vol 2: 52
44. Carmine, A, A. et al. Tinidazole anaerobic infections. A review of its antibacterial activity, pharmacological properties and therapeutic efficacy. Drugs, 1982. 24: 85～11
45. Hackect. L. P. Determination of metronidazole and tinidazole in luman plasma using high performance liguid chromatograply. J. Chromatogr, 1979. 175: 347
46. D. Babggot, W. D. WILSON, S. HIETALA. Clinical pharmacokinetics of metronidazole in horses. J. Vet. Pharmacol. Therap,1988. 11: 417～420
47. Ingrid, Nillsson, Ehle, et al. Liquid Chromatographic Assay for Metrondazole and Tinidazole: pharmacokinetic and Metabolic studies in Human Subjects. Antimicrobial Agents and Chemotherapy, 1981. 19: 754～760
48. Edwards.D. I. Structure-cytotoxicity relationships of nitroimidazoles in an in vitro system. Biol phys, 1982. 8(3-4): 791
49. G. L. Law, G. P. Mansfield, D. F. Muggleton, et al. Dimetridazole:A bsorption, Excretion and Metabolism in Turkeys. Nature, 1963. 97: 1024
50. Edwards ,David I. In vitro mechanisms of nitroimidazole cytotoxicity. Curr.chemother, 1981(pub, 1982). 1: 111-113
51. Knox, R. J. The mechanism of nitroimidazole damage to DNA:coulometric evidence. Int.J.Radiat. Oncol, Biol, Phys, 1984, 10(8): 1315-18
52. Laurentius. A. P, Theo. H. G. Polman, Johannes. A. van. Rhijn, et al. Biotransformation of Dimetridazole by Primary Cultures of Pig Hepatocytes. J. Agric. Food Chem, 1997. 45: 3985-3990