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Abstract

A COMPARATIVE STUDY IN THE CONFIRMATORY METHOD DEVELOPMENT AND VALIDATION OF FLUBENDAZOLE IN FISH AND SHRIMP MUSCLES BETWEEN HPLC-MS/MS (Q MICRO) AND UPLC-MS/MS (TQD)

Md. Ashraful Alam*, Talukdar Muhammad Waliullah, Akter Mst Yeasmin,
Saleh Ahmed and Md. Manik Mia

Abstract

A confirmatory method for determination of flubendazole wasdeveloped in ultra performance liquid chromatography–tandemquadrupole mass spectrometry (AQUITY UPLC–TQD) and in HPLCMS/MS (Quattro micro) at same time. The method was validated inBagda shrimp (Penaeus monodon) and Tilapia (Oreochromis niloticus)fish. The liquid chromatographic separation was done with gradientelution by using (Atlantis® dC18, 3μm, 4.6x100 column in case ofHPLC system whereas AQUITY UPLC BEH C18, 2.1x50mm, 1.7μm,Column in UPLC system) same mobile phase of 0.1% formic acid inwater and 0.1% formic acid in acetonitrile. Mass spectral acquisitionwas done using electrospray ionization in the positive ion mode applying multiple reactionmonitoring (MRM) of two diagnostic transition reactions for flubendazole and forflubendazole_d3. Shrimp and fish samples were extracted with ethyl acetate and evaporatedto dryness and finally reconstituted with acetonitrile. The method validation was carried outaccording to the criteria of Commission decision 2002/657/EC. The calibration curveshowed a good linearity in the concentration range from 1 to 20 ng/g with the correlationcoefficient of >0.996 in all cases. In case of shrimp samples the decision limit (CCα) and thedetection capability (CCβ) were 5.8μg/kg and 6.5μg/kg respectively in system1 and 5.6μg/kgand 6.2μg/kg in system2. In case of fish sample CCα values were 6.00μg/kg and 5.4μg/kg insystem1 and system2 and CCβ values were 7.1μg/kg and 5.8μg/kg respectively for flubendazole. The precision of the method, expressed as RSD values for the within laboratoryreproducibility at the three levels of fortification (2.5, 5.0 and 7.5) μg/kg was less than 15%.The mean recoveries were in the range of 90–107% for shrimp and that of 90-109% for fish.

Keywords: UPLC-TQD, HPLC-Quattro micro, Flubendazole, Shrimp, Fish and Validation.


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