利用高效液相層析法建立禽畜水產品中動物用藥Quinolones之多重殘留分析方法   全文下載

Simultaneous Determination of Quinolones in Livestock and Marine Products by High Performance Liquid Chromatography

本研究建立以高效液相層析法（high performance liquid chromatography, HPLC）同時檢測禽畜水產品中nalidixic acid、flumequine、oxolinic acid、piromidic acid、danofloxacin、enrofloxacin及sarafloxacin七種quinolones抗菌劑之分析方法。檢體以03%偏磷酸：乙?（1:10, v／v）萃取後，經Bond Elut C18萃取匣淨化，再以HPLC分析。結果發現利用Cosmosil 5C18-AR-II （5 mm, 4.6 mm i.d. × 250 mm）層析管柱，以含35 mM sodium dodecyl sulfate之乙?：0..05M NaH2PO4 （pH 2.5 ） （35:65, v／v）移動相溶液，於光二極體列陣檢出器及螢光檢出器（波長採time program）偵測下，quinolones之分離效果及重複性皆佳。經以0.05-10.0 μg／mL （danofloxacin為0.005~2.0 μg／mL）之七種quinolones混合標準溶液所製作之標準曲線之相關係數皆大於0.999。檢體中添加七種quinolones 0.01、0.1 、0.4及2.0 ppm（danofloxacin為0.001、0.01、0.04及0.20 ppm） 之平均回收率，除了低濃度者介於74.3~85.5%外，其餘介於80.1~99.9%，且變異係數皆小於5.8%。本方法之最低檢出限量，紫外光檢測介於0.01~0.04 ppm，螢光檢測則為0.0006-0.05 ppm。同日內及異日間之變異係數分別小於3.29%及5.23%，顯示本方法之精密度可接受。進而利用本方法分析市售禽畜水產品63件，結果有9件烏骨雞肉（wu ku chicken muscles）檢出enrofloxacin 0.08~4.04 ppm，4件烏骨雞肝檢出enrofloxacin 0.01~0.27 ppm，3 件香魚檢出0.13~0.35 ppm之oxolinic acid，與規定不符者達25.4%。雞肉中殘留之enrofloxacin安定性高，冷藏儲存時不易消退。對檢出不符規定者，衛生署已責成地方衛生機關依食品衛生管理法處辦。另因食品中動物用藥殘留問題之發生，係因農民於畜養階段違反動物用藥管理法所攻，故衛生署亦同時通知農業主管機關，加強農民輔導與管理。"

A high performance liquid chromatographic (HPLC) method was developed for the simultaneous determination of nalidixic acid, flumequine, oxolinic acid, piromidic acid, danofloxacin, enrofloxacin and sarafloxacin in chicken, pork and fish. These seven quinolone antibacterials were extracted with 0.3% metaphosphoric acid: acetonitrile (1:10, v/v), followed by a Bond Elut C18 cartridge clean up procedure. The HPLC separation was achieved on a Cosmosil 5C18-AR-II column (5 µm, 4.6 mm i.d. ¥ 250 mm) with acetonitrile: 0.05M NaH2PO4 (pH 2.5)(35:65, v/v) containing 3.5 mM sodium dodecyl sulfate as a mobile phase, and detection was performed with photodiode array and fluorescence detector (by using wavelength programming). Good linearity was observed from the calibration plot at concentrations from 0.05 to 10.0 mg/mL (0.005~2.0 µg/mL for danofloxacin). Recovery studies of the analytes were performed at 0.01, 0.1, 0.4 and 2.0 ppm (0.001, 0.01, 0.04 and 0.20 ppm for danofloxacin) spiked levels. Average recoveries of low concentration ranged from 74.3 to 85.5% and those of the rest ranged from 80.1 to 99.9% with coefficients of variation less than 5.8%. The detection limits of quinolones were 0.01~0.04 ppm with photodiode array detection and 0.0006~0.05 ppm with fluorescence detection. The coefficients of variation of intra-day and inter-day assays were lower than 3.29% and 5.23%, respectively. These results indicated that the developed method had an acceptable precision. Using this method to detect quinolones in sixty three samples purchased from various markets in Taipei, we found that nine wu ku chicken muscles contained enrofloxacin residues ranged from 0.08 to 4.04 ppm, four wu ku chicken liver muscles contained enrofloxacin residues ranged from 0.01 to 0.27 ppm, and oxolinic acid residues in three sweet fish samples ranged from 0.13 to 0.35 ppm. The results indicated that 25.4% of samples violated the regulation set by the Department of Health. The enrofloxacin residue in wu ku chicken muscle is very stable even after a long period of refrigeration at 4℃. 本研究建立以高效液相層析法（high performance liquid chromatography, HPLC）同時檢測禽畜水產品中nalidixic acid、flumequine、oxolinic acid、piromidic acid、danofloxacin、enrofloxacin及sarafloxacin七種quinolones抗菌劑之分析方法。檢體以03%偏磷酸：乙腈（1:10, v／v）萃取後，經Bond Elut C18萃取匣淨化，再以HPLC分析。結果發現利用Cosmosil 5C18-AR-II （5 mm, 4.6 mm i.d. × 250 mm）層析管柱，以含35 mM sodium dodecyl sulfate之乙腈：0..05M NaH2PO4 （pH 2.5 ） （35:65, v／v）移動相溶液，於光二極體列陣檢出器及螢光檢出器（波長採time program）偵測下，quinolones之分離效果及重複性皆佳。經以0.05-10.0 µg／mL （danofloxacin為0.005～2.0 µg／mL）之七種quinolones混合標準溶液所製作之標準曲線之相關係數皆大於0.999。檢體中添加七種quinolones 0.01、0.1 、0.4及2.0 ppm（danofloxacin為0.001、0.01、0.04及0.20 ppm） 之平均回收率，除了低濃度者介於74.3～85.5%外，其餘介於80.1～99.9%，且變異係數皆小於5.8%。本方法之最低檢出限量，紫外光檢測介於0.01～0.04 ppm，螢光檢測則為0.0006-0.05 ppm。同日內及異日間之變異係數分別小於3.29%及5.23%，顯示本方法之精密度可接受。進而利用本方法分析市售禽畜水產品63件，結果有9件烏骨雞肉（wu ku chicken muscles）檢出enrofloxacin 0.08～4.04 ppm，4件烏骨雞肝檢出enrofloxacin 0.01～0.27 ppm，3 件香魚檢出0.13～0.35 ppm之oxolinic acid，與規定不符者達25.4%。雞肉中殘留之enrofloxacin安定性高，冷藏儲存時不易消退。對檢出不符規定者，衛生署已責成地方衛生機關依食品衛生管理法處辦。另因食品中動物用藥殘留問題之發生，係因農民於畜養階段違反動物用藥管理法所攻，故衛生署亦同時通知農業主管機關，加強農民輔導與管理。