抗菌药物在水产生物体内的残留与代谢变异化合物研究
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摘要
大面积和高密度集约化养殖与病害发生成正相关性。随着我国水产业的发展，病害一年比一年严重,使用药物是防治病害的必要措施。但是由于缺乏科学的指导，各种药品的滥用已达到惊人的程度，严重威胁消费者的健康。我国加入WTO后，欧盟、美国、日本等国家对我国水产品中药物的残留量提出了更高要求，我国水产品出口连续遭遇食品安全卫生“绿色壁垒”，给国家造成了巨大的损失。所以水产用药的残留代谢和次生代谢产物研究以及药物安全使用技术已成为水产业必须关注的主题。
本论文对四环素、呋喃唑酮等几种抗菌药物在水产生物体内的残留、代谢规律进行深入研究，研究其变异代谢产物的量效和时效及毒性，科学评价药物与代谢变异产物的安全性。探讨了水温等不同的养殖条件对药物残留和代谢的影响，初步研究了药物在不同水产生物体内的残留与代谢的变化。研究结果可丰富和完善药物代谢动力学和药物效应动力学理论，并对水产生物无公害安全养殖和病害防治具有重要指导价值。对保证人体健康，扩大我国水产品出口具有重要意义。具体为：
1．给吉富罗非鱼一次灌服50μg/g剂量的四环素和土霉素浑浊液，研究了不同温度下四环素和土霉素在吉富罗非鱼体内的残留与代谢。实验结果表明，肝脏中四环素和土霉素含量最高，血液中次之，肌肉中含量最低。216h后，肌肉中四环素和土霉素均低于0.1mg/kg。四环素类药物在罗非鱼体内代谢较慢，并且随着温度的降低，代谢速率明显降低，半衰期延长。23℃条件下，四环素和土霉素的半衰期分别是28℃条件下的1.79倍和1.68倍。
2. 模拟水产养殖实际，每天以剂量为50μg/g的恩诺沙星分别给吉富罗非鱼、中国对虾投喂药饵，周期为7d。通过实验首次研究发现恩诺沙星在罗非鱼和中国对虾体内的主要变异代谢产物是相同的，都为环丙沙星。但是由于动物的种属不同，恩诺沙星在罗非鱼和中国对虾体内的代谢率有明显差别，罗非鱼体内恩诺沙星的代谢率明显高于对虾。罗非鱼肌肉中恩诺沙星和环丙沙星最高含量分别为3.61、0.22μg /g ，而对虾肌肉中恩诺沙星和环丙沙星的最高含量分别为1.68、
0.066μg /g。本实验条件下，罗非鱼和中国对虾的休药期分别为22 d和12 d。
3. 建立了高效液相色谱与质谱连用技术检测呋喃唑酮及其代谢物的方法，通过HPLC/MS检测技术，首次研究发现了呋喃唑酮在罗非鱼体内的主要代谢变异产物为3﹣氨基﹣2﹣噁唑酮 （AOZ）。给罗非鱼投喂剂量为30μg /g的呋喃唑酮药饵7d，罗非鱼肌肉中呋喃唑酮和AOZ的含量分别在停药6h后和停药“零时”达到最高，分别为413.00、31.15μg/kg，24h后呋喃唑酮含量就低于检出限，而肌肉中AOZ的含量在528h后才低于1μg/kg。呋喃唑酮在罗非鱼体内代谢很快，具有很高的代谢速率，而其代谢变异产物AOZ代谢速率很低，很难消除。考虑到AOZ不容易消除，在本实验条件下，建议罗非鱼休药期至少在22d。
4. 给大鲮鲆一次口灌剂量为40mg/kg的磺胺喹噁啉，首次研究了磺胺类药物在冷水鱼类大鲮鲆体内的残留与代谢。实验结果表明，磺胺喹噁啉主要残留于肝脏中，并且由于养殖水温等因素，磺胺喹噁啉在大鲮鲆体内代谢很慢。大鲮鲆肌肉和肝脏中的药物最高含量分别为0.696、0.778 μg /g，磺胺喹噁啉在大鲮鲆体内240h后仍然有残留。在本实验条件下，建议休药期至少10d。
The aquiculture with big area and high density is positive correlations for diseases. In China, with the development of aquiculture, diseases are getting more and more serious, so antibacterial drugs were often used for prevention and cure. But all kinds of drugs were misused seriously for lack of scientific direction, and as result, the residues of drugs can do very harm to people. After China entrance to WTO, aquatic organisms exported from our country had been rejected constantly by EU, America, Japan and so on for the residues of drugs exceeded the standard. And now, it was been found that the metabolites of some antibacterial drugs in aquatic organisms have the same harm to people as the original, some even serious than the original. So it has become very important to study on the residues and metabolites of antibacterial drugs in aquatic organisms and the technique of using drugs. Our studies focus on the residues of tetracycline, furan, enrofloxacin and sulfanilamide in aquatic organisms by HPLC and HPLC-MS. The safety of antibacterial drugs and its metabolite was evaluated scientifically at the same time. We also explained that how the residues of drugs were influenced by the temperature and others. The results we obtained can enrich and perfect the theory of pharmacokinertic and residues of antibacterial drugs, what’s more it can give help to direct the aquiculture with safety and harmlessness. Main results obtained are listed as below:
The residues and elimination of tetracycline (TC) and oxytetracycline (OTC) in muscles，liver and serum of Tilapias were studied after a single oral administration at a dose of 50mg/kg in different temperature. The results show
that the residues of TC and OTC are mainly in liver, the muscle’s is the least. After 216h the concentration of TC and OTC in muscles decreased to 0.1mg/kg. The rate of eliminations of TC and OTC are very slow in Tilapias. The rate of elimination is getting more and more slowly with the falling of temperature, and the elimination half-life is prolonged. The elimination half-life of TC and OTC in 23℃ is 1.79 and 1.68 times than which in 28℃ respectively.
The residues of enrofloxacin and its metabolite in Jifu Tilapia and Penaeus chinensis were first studied after a single oral dose of 50mg/kg for 7 days，and the withdrawal time was made at the same time. The results show that the metabolite of enrofloxacin in Tilapia and Penaeus chinensis both is ciprofloxacin. Because of different animal, the rate of metabolizing in Tilapia is more higher than that in Penaeus chinensis, The maximal concentration of enrofloxacin and ciprofloxacin in Tilapia are 3.61μg/g, 0.22 μg/g ,but which in Penaeus chinensis are 1.68μg/g，0.066μg/g respectively. The predicted withdrawal time for Tilapia is 22d, and Penaeus chinensis is 12d.
The residues of furazolidone [3-(5-nitrofurfurylidenamino)-2-oxazolidinone] and its main metabolite in tilapias were first studied by HPLC/MS. The results show that the main metabolite of furazolidone in tilapias is 3-amina-2-oxazolldinone (AOZ). After oral dose of 30mg/kgfor 7 days, the maximum level of furazolidone in tilapias was 413.00μg/kg after 6h, but that of AOZ reached maximum 31.15μg/kg when stopping giving drug. The concentration of furazolidone was lower than 10μg/kg after 24h, but that of AOZ was just lower than 1μg/kg after 528h. The results show that AOZ is very hard to eliminate compare to furazolidone. In view of AOZ is very hard to eliminate, the predicted withdrawal time of furazolidone for tilapias was 22d at least in this condition.
The residues and elimination of Sulfaquinoxaline in muscles and liver of Scophthatmus maximus were first studied after a single oral administration at a dose of 40mg/kg. The results show that the main residues of
Sulfaquinoxaline is in the liver of Scophthatmus maximus, and The rate of elimination is very slow. The maximum level of Sulfaquinoxaline in the muscle and liver are 0.696 μg /g, 0.778 μg /g respectively. The predicted withdrawal was 10d at least in thi
本论文对四环素、呋喃唑酮等几种抗菌药物在水产生物体内的残留、代谢规律进行深入研究，研究其变异代谢产物的量效和时效及毒性，科学评价药物与代谢变异产物的安全性。探讨了水温等不同的养殖条件对药物残留和代谢的影响，初步研究了药物在不同水产生物体内的残留与代谢的变化。研究结果可丰富和完善药物代谢动力学和药物效应动力学理论，并对水产生物无公害安全养殖和病害防治具有重要指导价值。对保证人体健康，扩大我国水产品出口具有重要意义。具体为：
1．给吉富罗非鱼一次灌服50μg/g剂量的四环素和土霉素浑浊液，研究了不同温度下四环素和土霉素在吉富罗非鱼体内的残留与代谢。实验结果表明，肝脏中四环素和土霉素含量最高，血液中次之，肌肉中含量最低。216h后，肌肉中四环素和土霉素均低于0.1mg/kg。四环素类药物在罗非鱼体内代谢较慢，并且随着温度的降低，代谢速率明显降低，半衰期延长。23℃条件下，四环素和土霉素的半衰期分别是28℃条件下的1.79倍和1.68倍。
2. 模拟水产养殖实际，每天以剂量为50μg/g的恩诺沙星分别给吉富罗非鱼、中国对虾投喂药饵，周期为7d。通过实验首次研究发现恩诺沙星在罗非鱼和中国对虾体内的主要变异代谢产物是相同的，都为环丙沙星。但是由于动物的种属不同，恩诺沙星在罗非鱼和中国对虾体内的代谢率有明显差别，罗非鱼体内恩诺沙星的代谢率明显高于对虾。罗非鱼肌肉中恩诺沙星和环丙沙星最高含量分别为3.61、0.22μg /g ，而对虾肌肉中恩诺沙星和环丙沙星的最高含量分别为1.68、
0.066μg /g。本实验条件下，罗非鱼和中国对虾的休药期分别为22 d和12 d。
3. 建立了高效液相色谱与质谱连用技术检测呋喃唑酮及其代谢物的方法，通过HPLC/MS检测技术，首次研究发现了呋喃唑酮在罗非鱼体内的主要代谢变异产物为3﹣氨基﹣2﹣噁唑酮 （AOZ）。给罗非鱼投喂剂量为30μg /g的呋喃唑酮药饵7d，罗非鱼肌肉中呋喃唑酮和AOZ的含量分别在停药6h后和停药“零时”达到最高，分别为413.00、31.15μg/kg，24h后呋喃唑酮含量就低于检出限，而肌肉中AOZ的含量在528h后才低于1μg/kg。呋喃唑酮在罗非鱼体内代谢很快，具有很高的代谢速率，而其代谢变异产物AOZ代谢速率很低，很难消除。考虑到AOZ不容易消除，在本实验条件下，建议罗非鱼休药期至少在22d。
4. 给大鲮鲆一次口灌剂量为40mg/kg的磺胺喹噁啉，首次研究了磺胺类药物在冷水鱼类大鲮鲆体内的残留与代谢。实验结果表明，磺胺喹噁啉主要残留于肝脏中，并且由于养殖水温等因素，磺胺喹噁啉在大鲮鲆体内代谢很慢。大鲮鲆肌肉和肝脏中的药物最高含量分别为0.696、0.778 μg /g，磺胺喹噁啉在大鲮鲆体内240h后仍然有残留。在本实验条件下，建议休药期至少10d。
The aquiculture with big area and high density is positive correlations for diseases. In China, with the development of aquiculture, diseases are getting more and more serious, so antibacterial drugs were often used for prevention and cure. But all kinds of drugs were misused seriously for lack of scientific direction, and as result, the residues of drugs can do very harm to people. After China entrance to WTO, aquatic organisms exported from our country had been rejected constantly by EU, America, Japan and so on for the residues of drugs exceeded the standard. And now, it was been found that the metabolites of some antibacterial drugs in aquatic organisms have the same harm to people as the original, some even serious than the original. So it has become very important to study on the residues and metabolites of antibacterial drugs in aquatic organisms and the technique of using drugs. Our studies focus on the residues of tetracycline, furan, enrofloxacin and sulfanilamide in aquatic organisms by HPLC and HPLC-MS. The safety of antibacterial drugs and its metabolite was evaluated scientifically at the same time. We also explained that how the residues of drugs were influenced by the temperature and others. The results we obtained can enrich and perfect the theory of pharmacokinertic and residues of antibacterial drugs, what’s more it can give help to direct the aquiculture with safety and harmlessness. Main results obtained are listed as below:
The residues and elimination of tetracycline (TC) and oxytetracycline (OTC) in muscles，liver and serum of Tilapias were studied after a single oral administration at a dose of 50mg/kg in different temperature. The results show
that the residues of TC and OTC are mainly in liver, the muscle’s is the least. After 216h the concentration of TC and OTC in muscles decreased to 0.1mg/kg. The rate of eliminations of TC and OTC are very slow in Tilapias. The rate of elimination is getting more and more slowly with the falling of temperature, and the elimination half-life is prolonged. The elimination half-life of TC and OTC in 23℃ is 1.79 and 1.68 times than which in 28℃ respectively.
The residues of enrofloxacin and its metabolite in Jifu Tilapia and Penaeus chinensis were first studied after a single oral dose of 50mg/kg for 7 days，and the withdrawal time was made at the same time. The results show that the metabolite of enrofloxacin in Tilapia and Penaeus chinensis both is ciprofloxacin. Because of different animal, the rate of metabolizing in Tilapia is more higher than that in Penaeus chinensis, The maximal concentration of enrofloxacin and ciprofloxacin in Tilapia are 3.61μg/g, 0.22 μg/g ,but which in Penaeus chinensis are 1.68μg/g，0.066μg/g respectively. The predicted withdrawal time for Tilapia is 22d, and Penaeus chinensis is 12d.
The residues of furazolidone [3-(5-nitrofurfurylidenamino)-2-oxazolidinone] and its main metabolite in tilapias were first studied by HPLC/MS. The results show that the main metabolite of furazolidone in tilapias is 3-amina-2-oxazolldinone (AOZ). After oral dose of 30mg/kgfor 7 days, the maximum level of furazolidone in tilapias was 413.00μg/kg after 6h, but that of AOZ reached maximum 31.15μg/kg when stopping giving drug. The concentration of furazolidone was lower than 10μg/kg after 24h, but that of AOZ was just lower than 1μg/kg after 528h. The results show that AOZ is very hard to eliminate compare to furazolidone. In view of AOZ is very hard to eliminate, the predicted withdrawal time of furazolidone for tilapias was 22d at least in this condition.
The residues and elimination of Sulfaquinoxaline in muscles and liver of Scophthatmus maximus were first studied after a single oral administration at a dose of 40mg/kg. The results show that the main residues of
Sulfaquinoxaline is in the liver of Scophthatmus maximus, and The rate of elimination is very slow. The maximum level of Sulfaquinoxaline in the muscle and liver are 0.696 μg /g, 0.778 μg /g respectively. The predicted withdrawal was 10d at least in thi
引文
1. 农业部，农业部发布关于食品动物禁用的兽药及其他化合物清单。中国兽药杂志，2002，36（4）：11-12。
Ngata，Seki．Simultaneous determination of thiampenicol，florfenico and chloramphenicol residues in muscles of animals and cultured fish by liquid chromatography[J] ．J quid chromatography，1992，15（12）：2045-2056．
Pfenning A P， Roybal J E，Rupp H S．et al． Simultaneous determination residues of chloramphenicol ，florfenicol，florfenicol amine，and thiamphenicol in shrimp tissue by gas chromatography with electron capture detection[J]．J AOAC Int，2000，83(1)：26-30．
李兰生 王勇强，对虾体内氯霉素含量测定[J] 。青岛海洋大学学报，1995，25（3）。
李爱华。氯霉素在草鱼和复合四倍体异育银鲫体内的比较药代动力学研究[C]。 海洋与湖沼年会论文集，1996。
王建华等。同时测定鱼肉中氯霉素和甲砜霉素残留量的毛细管气相色谱法 [J]。分析测
试学报，2001，20（3）。
焦彦朝等。微生物检定法测定鲤鱼肌肉组织残留氯霉素。中国饲料，2000，5：22-24。
8. Samuelsen O B．Simultaneous determination of Ormethoprim and Sulfadimethoxine in plasma and muscle of Alantic Salmon (Salmo-Salar)，Journal of chromatographyb-biomedical applications，1994，660(2)：412-417．
9. Samuelsen O B． Absorption,tissue distribution，metabolism and excretion of Ormethoprim and Sulfadimethoxine in Alantic Salmon (Salmo-Salar) after intravenous and oral-administration of romet(30) ．Xenobiotica，1995，25(11)：1169-1180．
10. Hormazabal V．et al．Simultaneous extraction and determination of sulfadiazine and trimethoprim in medicated fish feed by High-Performance Liquid-Chromatography[J] ．Journal of Chromatography，1993，648(1)：183-186．
11. Walker C C．et al．Extraction and liquid chromatographic analysis of sulfadimethoxine and 4-N-acetyl- Sulfadimethoxine residues in channel catfish (Ictalurus Punctatus) muscle and plasma [J]． Journal of AOAC Internatioanal，1994，77(6)：1460-1466．
12. Gehring T A．et al． Liquid chromatographic determination of sulfadiazine by postcolum deribvatization and fluorescence detection．[J] Journal of AOAC Internatioanal，1995，78(5)：1161~1164．
13. Uno K．Pharmacokinetics and metabolism of Sulphamonomethoxine in Rainbow Trout(oncorhynchus Mykiss) and yellowtail(Seriola Quinqueradiata) following Bolus Intravascular administration[J] ．Aquaculture，1997，153： 1～8．
14. Uno K． Pharmacokinetics of Sulphamonomethoxine and Sulphadimethoxine following oral administration to Cultured Rainbow Trout (oncorhynchus Mykiss) [J] ．Aquaculture，1993，115：209-219．
15. Park E D， Donald V，Lightner．et al．Exploratory bioavailability and pharmacokinetic studies of sulphadimethoxine and ormetoprim in the penaeid shrimp，Penaeous vannamei． [J] Aquaculture，1995，130：113~128．
16. Samuelsen O B，et al．Multiple-dose pharmacokinetic study of Romet30 in Alantic Salmon (Salmo-Salar) and in vitro antibacterial activity against Aeromonas salmonicida． Aquaculture，1997，152：13-24．
17. Roybal J E，Pfenning P，Turnipseed S B．et al． Application of size-exclusion chromatography
to the analysis of shrimp for sulfonamide residues． Analytic chimica acta．2003，483：147-152．
18. 农业部。 动物源食品中磺胺类药物残留检测方法--高效液相色谱法。中国兽药杂志，2002，36（6）：12-13。
19. 王群，马向东。 HPLC法研究磺胺类药物在水产动物体内的代谢和残留。海洋湖沼通报，2002，2：63-70。
20. 艾晓辉，陈正望。 磺胺二甲基嘧啶在银鲫体内的药动学及组织残留研究。淡水渔业，2001，31（6），52-54。
21. 朱秋华。 3种药物在甲鱼体内的残留研究，中国水产科学，2001，8(3)，50-53。
22. Carignan G． Assay of oxytetracycline residues in Salmon muscle by liquid chromatography with ultraviolet detection [J] ． Journal of AOAC international，1993，76(2)：325-328．
23. Macneil D．et al． Ch1ortetracycline，Oxytetracycline， and Tetracycline in edible animal tissues， liquid chromatographic method：collaborative study [J] ．Journal of AOAC international，1996，79(2)：405-416．
24. Cravedi J P，Choubert G，Delous G． Digestibility of chloramphenicol，oxolinic acid and oxytetracycline in rainbow trout and influence of these antibiotics on lipid digestability．Aquaculture，1987，60：133-141．
25. Rogstad A．et al． Pharmacokinetics study of oxytetracycline in fish， Absorption，Distribution and accumulation in rainbow trout in freshwater[J] ． Aquaculture，1991，96：219-226．
26. Bruno D W． An investigation into oxytetracycline residues in Atlantic salmon，Salmo salar ．J. Fish Diseases，1989，12：77-86．
27. Bjorklund H V，Bylund G． Temperature-related absorption and excretion of oxytetracycline in rainbow trout(Salmo gairdneri R.) ． Aquaculture，1990，84(3/4)：363-372．
28. Bjorklund H V． Residues of oxolinic acid and oxytetracycline in fish and sediments from fish farms． Aquaculture，1991，97(1)：85-86．
29. Bjorklund H V． Residues of oxytetracycline in wild fish and sediments from fish farms．Aquaculture，1990， 84(3/4)：359-367．
30. Mohney L L， Rodney R，Thomas A et al．Residues of oxytetracycline in cultured juvenile blue shrimp，penaeous stylirostris (Cyustacea:Decapod)，fed medicated feed for 14 days．Aquaculture，1997，149：193~202．
31. Jacobsen M D．Withdrawal times of freshwater rainbow trout，Salmo gairdneri Richardson，after treatment with oxolinic acid ， oxytetracycline and trimethoprim．Fish Diseases，1989，12:29-36．
32. Kerry J，Hiney M．Frequency and distribution of resistance to oxytetracycline in micro-organisms isolated from marine fish farm sendiments following therapeutic use of oxytetracycline．Aquaculture，1994，123:43-54．
33. Kerry J，Hiney M． Fish feed as a source of oxytetracycline-resistant bacteria in the sendiments under fish farm ．Aquaculture，131：101-113．
34. Cloudio D，Miranda．Bacterial resistance to oxytetracycline in Chilean salmon farming．Aquaculture，2002，212：31-47．
35. Julie B W ，Graham B，Mary C C． Oxytetracycline residues in a freshwater recirculating system．Aquaculture，2002，205：221-230．
36. Marc C，Mario S，Siska C，et al． Quantitative multi-residues analysis of tetracyclines and their 4-epimers in pig tissues by high-performance liquid chromatography combined with positive-ion electrospray ionization mass spectrometry． Analytic chimica acta，2003，492：199-213．
37. 祝仕清等。 高效毛细管电泳分离四环素类抗生素及测定四环素中杂质[J]。药物分析杂志，1998，18（4）。
38. 徐冬梅。 高效液相色谱法同时测定动物组织中3种抗生素残留量。中国公共卫生，2002，18（2）。
39. 李美同，郭文林，郑国兴等。土霉素在鳗鲡组织中残留的消除规律。水产学报，1997，21（1）：39-43。
40. 王群等。土霉素在黑鲷体内的药物代谢动力学研究。海洋水产研究，2001，22（1）。
41. Conneely A，Nugent A，O’keeffe M． Use of solid phase extraction for the isolation and clean-up of a derivatised furazolidone metabolite from animal tissues．Analyst，2002，127(6)：705-709．
42. Rupp H S．et al． Simultaneous determination of nitrofurazone and furazolidone in shrimp (penaeus vannamei) muscle tissue by Liquid Chromatography with UV detection [J] ．Journal of AOAC Internatioanal，1993，77(4)：901-904．
43. 农业部。 动物源食品中呋喃唑酮残留检测方法--高效液相色谱法。中国兽药杂志，2002，
36（2）：13-14。
44. 葛宝坤，王云凤，常春艳等。 测定鸡肉、水产品中四种硝基呋喃类药物残留量的固相萃取-液相色谱法 [J]。 分析测试学报， 2003，22（5）：91-93。
45. Horie M，Nakazawa H． Determination of miloxacin and its metabolite in fish by High-Performance Liquid-Chromatography with fluorescence and UV detection [J] ．Journal of Liquid Chromatography，1992，15(12)：2057-2070．
46. Rogstad A．et al． Extraction and High-Performance Liquid-Chromatography determination of endofloxacin in fish serum and tissues [J] ．Journal of Liquid Chromatography，1991，14(3)：521-531．
47. Hormazabal V．et al． Rapid assay for monitoring residues of enrofloxacin and sarafloxacin in fish-tissues by High-Performance Liquid-Chromatography [J] ． Journal of Liquid Chromatography，1991，14(8)：1605-1614．
48. Steffenak I， Hormazabal V，Yndestad M． Rapid assay for the simultaneous determination of residues of oxolinic acid and flumequine in fish-tissues by High-Performance Liquid-Chromatography [J] ． Journal of Liquid Chromatography，1991，14(1)：61-70．
49. Hormazabal V，et al． A rapid and time-effective assay for determination of oxolinic acid and flumequine in fish-tissues by HPLC[J] ．Journal of Liquid Chromatography，1994，17(13)：2911-2917．
50. Turnipseed S B，et al． Confirmation of fluoroquinalones in catfish muscle by electrospray liquid chromatography/mass spectrometry [J] ．Journal of AOAC International，1998，81(3)：554-562．
51. Hustvedt S O，et al． Absorption distribution and elimination of oxolinic acid in Atlantic salmon (Salmo salar L.)after vairous routes of administration．Aquaculture，1991，95：193-199．
52. Hustvedt S O，et al． Distribution and elimination of oxolinic acid in rainabow trout(Oncorhynchus mykiss Walbaum)after a single intravascular injection．Aquaculture，1991，92：297-303．
53. Bjorklund H V，et al． Temperature-related absorption and excretion of oxolinic acid in rainabow trout(Oncorhynchus
mykiss Walbaum) ．Aquaculture，1992，102:17-27．
54. Sohlberg S，et al． Temperature-dependent absorption and elimination of flumequine in rainbow trout (Oncorhynchus mykiss Walbaum) in fresh water．Aquaculture，1994，119：1-10．
55. Samuelsen O B，Ervik A． Single dose pharmacokinetic study of flumequine after intravenous，intraperitoneal and oral administration to Atlantic halibut(Hipoglossus hipoglossus) held in seawater at 9℃．Aquaculture，158：215-227．
56. Malvisi J， Rocca G D，Anfossi P ，et al． Tissue distribution and depletion of flumequine after in-feed administration in sea bream (Sparus aurata) ．Aquaculture，1997，157：197-204．
57. Boon J H，et al． Disposition of flumequine plasma of European eel(Anguilla anguilla) after a single intra-muscular injection．Aquaculture，1991，99：213-223．
58. Pascual-Reguera M I，Gertrudis P P，Antonio M D． Solid-phase UV spectrophotometric method for determination of ciprofloxacin． Microchemical journal，2004，44：79-84．
59.Pascual-Reguera M I，Gertrudis P P，Antonio M D. Solid-phase UV spectrophotometric method for determination of ciprofloxacin. Microchemical journal，2004,44:79-84.
60. 张雅斌，张祚新，郑伟等。 不同给药方式下鲤对诺氟沙星的药代动力学研究 [J]。水产学报，2000，24(6)：559-563。
61. 陈文银等。 诺氟沙星在中华鳖体内的药代动力学研究 [J]。水产学报，1997，21(4)：434-436。
62. Hormazabal V，et al． A time cost-effective assay for the determination of residues of Malachite Green in fish-tissues by HPLC [J] ．Journal of Liquid Chromatography，1992，15(12)：2035-2044．
Ngata，Seki．Simultaneous determination of thiampenicol，florfenico and chloramphenicol residues in muscles of animals and cultured fish by liquid chromatography[J] ．J quid chromatography，1992，15（12）：2045-2056．
Pfenning A P， Roybal J E，Rupp H S．et al． Simultaneous determination residues of chloramphenicol ，florfenicol，florfenicol amine，and thiamphenicol in shrimp tissue by gas chromatography with electron capture detection[J]．J AOAC Int，2000，83(1)：26-30．
李兰生 王勇强，对虾体内氯霉素含量测定[J] 。青岛海洋大学学报，1995，25（3）。
李爱华。氯霉素在草鱼和复合四倍体异育银鲫体内的比较药代动力学研究[C]。 海洋与湖沼年会论文集，1996。
王建华等。同时测定鱼肉中氯霉素和甲砜霉素残留量的毛细管气相色谱法 [J]。分析测
试学报，2001，20（3）。
焦彦朝等。微生物检定法测定鲤鱼肌肉组织残留氯霉素。中国饲料，2000，5：22-24。
8. Samuelsen O B．Simultaneous determination of Ormethoprim and Sulfadimethoxine in plasma and muscle of Alantic Salmon (Salmo-Salar)，Journal of chromatographyb-biomedical applications，1994，660(2)：412-417．
9. Samuelsen O B． Absorption,tissue distribution，metabolism and excretion of Ormethoprim and Sulfadimethoxine in Alantic Salmon (Salmo-Salar) after intravenous and oral-administration of romet(30) ．Xenobiotica，1995，25(11)：1169-1180．
10. Hormazabal V．et al．Simultaneous extraction and determination of sulfadiazine and trimethoprim in medicated fish feed by High-Performance Liquid-Chromatography[J] ．Journal of Chromatography，1993，648(1)：183-186．
11. Walker C C．et al．Extraction and liquid chromatographic analysis of sulfadimethoxine and 4-N-acetyl- Sulfadimethoxine residues in channel catfish (Ictalurus Punctatus) muscle and plasma [J]． Journal of AOAC Internatioanal，1994，77(6)：1460-1466．
12. Gehring T A．et al． Liquid chromatographic determination of sulfadiazine by postcolum deribvatization and fluorescence detection．[J] Journal of AOAC Internatioanal，1995，78(5)：1161~1164．
13. Uno K．Pharmacokinetics and metabolism of Sulphamonomethoxine in Rainbow Trout(oncorhynchus Mykiss) and yellowtail(Seriola Quinqueradiata) following Bolus Intravascular administration[J] ．Aquaculture，1997，153： 1～8．
14. Uno K． Pharmacokinetics of Sulphamonomethoxine and Sulphadimethoxine following oral administration to Cultured Rainbow Trout (oncorhynchus Mykiss) [J] ．Aquaculture，1993，115：209-219．
15. Park E D， Donald V，Lightner．et al．Exploratory bioavailability and pharmacokinetic studies of sulphadimethoxine and ormetoprim in the penaeid shrimp，Penaeous vannamei． [J] Aquaculture，1995，130：113~128．
16. Samuelsen O B，et al．Multiple-dose pharmacokinetic study of Romet30 in Alantic Salmon (Salmo-Salar) and in vitro antibacterial activity against Aeromonas salmonicida． Aquaculture，1997，152：13-24．
17. Roybal J E，Pfenning P，Turnipseed S B．et al． Application of size-exclusion chromatography
to the analysis of shrimp for sulfonamide residues． Analytic chimica acta．2003，483：147-152．
18. 农业部。 动物源食品中磺胺类药物残留检测方法--高效液相色谱法。中国兽药杂志，2002，36（6）：12-13。
19. 王群，马向东。 HPLC法研究磺胺类药物在水产动物体内的代谢和残留。海洋湖沼通报，2002，2：63-70。
20. 艾晓辉，陈正望。 磺胺二甲基嘧啶在银鲫体内的药动学及组织残留研究。淡水渔业，2001，31（6），52-54。
21. 朱秋华。 3种药物在甲鱼体内的残留研究，中国水产科学，2001，8(3)，50-53。
22. Carignan G． Assay of oxytetracycline residues in Salmon muscle by liquid chromatography with ultraviolet detection [J] ． Journal of AOAC international，1993，76(2)：325-328．
23. Macneil D．et al． Ch1ortetracycline，Oxytetracycline， and Tetracycline in edible animal tissues， liquid chromatographic method：collaborative study [J] ．Journal of AOAC international，1996，79(2)：405-416．
24. Cravedi J P，Choubert G，Delous G． Digestibility of chloramphenicol，oxolinic acid and oxytetracycline in rainbow trout and influence of these antibiotics on lipid digestability．Aquaculture，1987，60：133-141．
25. Rogstad A．et al． Pharmacokinetics study of oxytetracycline in fish， Absorption，Distribution and accumulation in rainbow trout in freshwater[J] ． Aquaculture，1991，96：219-226．
26. Bruno D W． An investigation into oxytetracycline residues in Atlantic salmon，Salmo salar ．J. Fish Diseases，1989，12：77-86．
27. Bjorklund H V，Bylund G． Temperature-related absorption and excretion of oxytetracycline in rainbow trout(Salmo gairdneri R.) ． Aquaculture，1990，84(3/4)：363-372．
28. Bjorklund H V． Residues of oxolinic acid and oxytetracycline in fish and sediments from fish farms． Aquaculture，1991，97(1)：85-86．
29. Bjorklund H V． Residues of oxytetracycline in wild fish and sediments from fish farms．Aquaculture，1990， 84(3/4)：359-367．
30. Mohney L L， Rodney R，Thomas A et al．Residues of oxytetracycline in cultured juvenile blue shrimp，penaeous stylirostris (Cyustacea:Decapod)，fed medicated feed for 14 days．Aquaculture，1997，149：193~202．
31. Jacobsen M D．Withdrawal times of freshwater rainbow trout，Salmo gairdneri Richardson，after treatment with oxolinic acid ， oxytetracycline and trimethoprim．Fish Diseases，1989，12:29-36．
32. Kerry J，Hiney M．Frequency and distribution of resistance to oxytetracycline in micro-organisms isolated from marine fish farm sendiments following therapeutic use of oxytetracycline．Aquaculture，1994，123:43-54．
33. Kerry J，Hiney M． Fish feed as a source of oxytetracycline-resistant bacteria in the sendiments under fish farm ．Aquaculture，131：101-113．
34. Cloudio D，Miranda．Bacterial resistance to oxytetracycline in Chilean salmon farming．Aquaculture，2002，212：31-47．
35. Julie B W ，Graham B，Mary C C． Oxytetracycline residues in a freshwater recirculating system．Aquaculture，2002，205：221-230．
36. Marc C，Mario S，Siska C，et al． Quantitative multi-residues analysis of tetracyclines and their 4-epimers in pig tissues by high-performance liquid chromatography combined with positive-ion electrospray ionization mass spectrometry． Analytic chimica acta，2003，492：199-213．
37. 祝仕清等。 高效毛细管电泳分离四环素类抗生素及测定四环素中杂质[J]。药物分析杂志，1998，18（4）。
38. 徐冬梅。 高效液相色谱法同时测定动物组织中3种抗生素残留量。中国公共卫生，2002，18（2）。
39. 李美同，郭文林，郑国兴等。土霉素在鳗鲡组织中残留的消除规律。水产学报，1997，21（1）：39-43。
40. 王群等。土霉素在黑鲷体内的药物代谢动力学研究。海洋水产研究，2001，22（1）。
41. Conneely A，Nugent A，O’keeffe M． Use of solid phase extraction for the isolation and clean-up of a derivatised furazolidone metabolite from animal tissues．Analyst，2002，127(6)：705-709．
42. Rupp H S．et al． Simultaneous determination of nitrofurazone and furazolidone in shrimp (penaeus vannamei) muscle tissue by Liquid Chromatography with UV detection [J] ．Journal of AOAC Internatioanal，1993，77(4)：901-904．
43. 农业部。 动物源食品中呋喃唑酮残留检测方法--高效液相色谱法。中国兽药杂志，2002，
36（2）：13-14。
44. 葛宝坤，王云凤，常春艳等。 测定鸡肉、水产品中四种硝基呋喃类药物残留量的固相萃取-液相色谱法 [J]。 分析测试学报， 2003，22（5）：91-93。
45. Horie M，Nakazawa H． Determination of miloxacin and its metabolite in fish by High-Performance Liquid-Chromatography with fluorescence and UV detection [J] ．Journal of Liquid Chromatography，1992，15(12)：2057-2070．
46. Rogstad A．et al． Extraction and High-Performance Liquid-Chromatography determination of endofloxacin in fish serum and tissues [J] ．Journal of Liquid Chromatography，1991，14(3)：521-531．
47. Hormazabal V．et al． Rapid assay for monitoring residues of enrofloxacin and sarafloxacin in fish-tissues by High-Performance Liquid-Chromatography [J] ． Journal of Liquid Chromatography，1991，14(8)：1605-1614．
48. Steffenak I， Hormazabal V，Yndestad M． Rapid assay for the simultaneous determination of residues of oxolinic acid and flumequine in fish-tissues by High-Performance Liquid-Chromatography [J] ． Journal of Liquid Chromatography，1991，14(1)：61-70．
49. Hormazabal V，et al． A rapid and time-effective assay for determination of oxolinic acid and flumequine in fish-tissues by HPLC[J] ．Journal of Liquid Chromatography，1994，17(13)：2911-2917．
50. Turnipseed S B，et al． Confirmation of fluoroquinalones in catfish muscle by electrospray liquid chromatography/mass spectrometry [J] ．Journal of AOAC International，1998，81(3)：554-562．
51. Hustvedt S O，et al． Absorption distribution and elimination of oxolinic acid in Atlantic salmon (Salmo salar L.)after vairous routes of administration．Aquaculture，1991，95：193-199．
52. Hustvedt S O，et al． Distribution and elimination of oxolinic acid in rainabow trout(Oncorhynchus mykiss Walbaum)after a single intravascular injection．Aquaculture，1991，92：297-303．
53. Bjorklund H V，et al． Temperature-related absorption and excretion of oxolinic acid in rainabow trout(Oncorhynchus
mykiss Walbaum) ．Aquaculture，1992，102:17-27．
54. Sohlberg S，et al． Temperature-dependent absorption and elimination of flumequine in rainbow trout (Oncorhynchus mykiss Walbaum) in fresh water．Aquaculture，1994，119：1-10．
55. Samuelsen O B，Ervik A． Single dose pharmacokinetic study of flumequine after intravenous，intraperitoneal and oral administration to Atlantic halibut(Hipoglossus hipoglossus) held in seawater at 9℃．Aquaculture，158：215-227．
56. Malvisi J， Rocca G D，Anfossi P ，et al． Tissue distribution and depletion of flumequine after in-feed administration in sea bream (Sparus aurata) ．Aquaculture，1997，157：197-204．
57. Boon J H，et al． Disposition of flumequine plasma of European eel(Anguilla anguilla) after a single intra-muscular injection．Aquaculture，1991，99：213-223．
58. Pascual-Reguera M I，Gertrudis P P，Antonio M D． Solid-phase UV spectrophotometric method for determination of ciprofloxacin． Microchemical journal，2004，44：79-84．
59.Pascual-Reguera M I，Gertrudis P P，Antonio M D. Solid-phase UV spectrophotometric method for determination of ciprofloxacin. Microchemical journal，2004,44:79-84.
60. 张雅斌，张祚新，郑伟等。 不同给药方式下鲤对诺氟沙星的药代动力学研究 [J]。水产学报，2000，24(6)：559-563。
61. 陈文银等。 诺氟沙星在中华鳖体内的药代动力学研究 [J]。水产学报，1997，21(4)：434-436。
62. Hormazabal V，et al． A time cost-effective assay for the determination of residues of Malachite Green in fish-tissues by HPLC [J] ．Journal of Liquid Chromatography，1992，15(12)：2035-2044．