禽類產品運用EMR-QuEChERS前處理技術以液相層析串聯式質譜儀分析多重動物用藥殘留的方法建立與確效

在大量飼養家禽下，業者需使用動物用藥治療及預防禽類疾病並提升其產量。為確保食品安全，此類產品上市前後分別由農政單位與衛政單位抽樣監控。動物用藥種類各有不同特性，對應之檢驗方法亦多元，故需花費較多人力與物力來執行檢驗。本研究建立一種檢驗禽類產品中磺胺劑類、喹諾酮類抗生素、巨環內酯類抗生素及其代謝物、氯黴素類抗生素、抗原蟲劑類及離子型抗球蟲藥類之方法，並執行其確效相關工作。前處理過程使用Enhanced Matrix Removal (EMR-lipid)替代固相萃取法與液-液萃取法，再以液相層析串聯式質譜儀檢驗並以基質添加檢量線法定量。實驗結果顯示於79品項藥劑中salinomycin及narasin在EMR-lipid處理之檢液混合後出現明顯降解情形；florfenicol amine因在C18管柱中不易滯留，無法有良好的線性關係，故本研究並未整併此3品項。標準品穩定性測試結果顯示，在10oC溫控的環境下，溶解於溶劑部分品項於第54小時檢驗訊號面積與第0小時起始訊號面積相比已低於50%，主要為喹諾酮類抗生素。若測試藥品添加含有經EMR-lipid處理之禽肉及蛋基質檢液，則出現磺胺劑類降解而喹諾酮類抗生素反而穩定現象。使用禽肉、蛋及內臟基質之添加試驗結果顯示，添加10 ppb標準品之禽肉類、蛋類及內臟類的回收率分別為65%-116%、76%-144%及69%-163%；添加25 ppb者分別為87%-118%、70%-127%及60%-133%；而添加50 ppb者則分別為67%-105%、82%-133%及60%-127%。10 ppb添加試驗之相對標準偏差(RSD%)於禽肉類、蛋類及內臟類分別為2%-26%、1%-30%及0%-40%；添加25 ppb者分別為2%-25%、1%-16%及3%-36%；而添加50 ppb者則分別為3%-23%、2%-37%及1%-30%。本研究方法所獲得之定量極限與公告檢驗方法相比，erythromycin, clindamycin, lincomycin及metronidazole (內臟基質)等4品項未能符合要求，其餘72品項皆可符合要求。最後本研究實際測試2件真實樣品，此結果與公告檢驗方法之結果相符，故本方法後續可進行作為篩檢用途之公告檢驗方法可行性評估。

關鍵字

多重動物用藥殘留； Enhanced Matrix Removal technique (EMR-QuEChERS) ；液相層析串聯式質譜；基質添加檢量線法

並列摘要

Under the large-scale breeding of poultry, the operators need to use animal medicines to treat and prevent poultry diseases to increase their production. In order to ensure food safety, such products are sampled and monitored by Council of Agriculture and Taiwan Food and Drug Administration, respectively, before and after they are put on the market. Since the types of veterinary drugs have different characteristics and the corresponding inspection methods are also varied, it takes a lot of manpower and material resources to carry out these tests. In this study, a multi-residues testing method for poultry products has been developed and validated, including sulfonamides, quinolone antibiotics, macrolide antibiotics, antiprotozoal drugs, chloramphenicol antibiotics, and ionophore coccidiostats. In the pretreatment process, a commercial kit-Enhanced Matrix Removal-lipid (EMR lipid) was used to replace the traditional solid-phase extraction and liquid-liquid extraction, and subsequently, liquid chromatography tandem mass spectrometer was used to detect multiple animal drug residues, and quantified by procedural standard calibration. The experimental results showed that salinomycin and narasin degraded obviously in the EMR-lipid treated solution, and the poor repeatability and linearity of the calibration curve of florfenicol amine due to the poor retention ability in the C18 column. Therefore, the three drugs could not be integrated into the testing method in this study. The stability test of standards showed that the peak area of 11 quinolone drugs decreased about 50% after 54 hours in the pure solvent solution under the temperature-controlled environment of 10°C. The sulfonamides decreased significantly and the quinolone drugs were stable in the EMR-lipid treated chicken and the egg-matrix test solution. The recovery rates of 10 μg/kg fortified tests were 65%-116%, 76%-144%, and 69%-163% for chicken, egg, and viscera, respectively; those of 25 μg/kg fortified tests were 87%-118%, 70%-127%, and 60%-133%, respectively; and those of 50 μg/kg fortified tests were 67%-105%, 82%-133% and 60%-127% for chicken, egg, and viscera, respectively. The relative standard deviation (RSD%) of 10 μg/kg fortified tests were 2%-26%, 1%-30%, and 0%-40% for chicken, egg, and viscera, respectively; those of 25 μg/kg fortified tests were 2%-25%, 1%-16%, and 3%-36%, and those of 50 μg/kg fortified tests were 3%-23%, 2%-37% and 1%-30%, respectively. The limit of detection of this method was then compared with those of the official method. Four veterinary drugs including erythromycin, clindamycin, lincomycin and metronidazole (in visceral matrix) failed to meet the requirements, and the other 72 veterinary drugs could meet the requirements. In this study, two real samples were tested, and the results were consistent with those of the official methods. The feasibility of using the method developed in this study as an alternative for the official methods can be evaluated in the future.