液相色谱法同时测定水产品中孔雀石绿和结晶紫残留

Simultaneous Determination and Confirmation of Malachite Green, Crystal Violet and Their Leuco Metabolites Residues in Aquatic Product by Liquid Chromatography-Visible Detection and Tandem Mass Spectrometric Detection

用液相色谱-可见法同时测定水产品中孔雀石绿（MG）、结晶紫（CV）及其代谢物隐色孔雀石绿（LMG）和隐色结晶紫（LCV）的残留量,并用液相色谱-串联质谱法进行确证和定量.样品用乙腈提取,二氯甲烷液液分配, MCX阳离子固相萃取小柱净化,浓缩定容.以乙酸铵缓冲溶液和乙腈为流动相,经C18柱分离后,PbO2柱后衍生;用二极管阵列检测器在618 nm测定孔雀石绿和隐色孔雀石绿,在588 nm测定结晶紫和隐色结晶紫;并用串联质谱在电喷雾-多反应监测离子的模式下,进行质谱确证和定量;外标法定量,内标亮绿和氘代隐色孔雀石绿校正回收率.液相色谱-可见法的检出限为MG 0.22,LMG 0.28,CV 0.22,LCV 0.25 μg/kg;液相色谱-串联质谱法的检出限为MG 0.014,LMG 0.018,CV 0.014,LCV 0.0084 μg/kg.在2～20 μg/kg范围内,回收率为75%～95%.

The method of simultaneous determination and confirmation of malachite green, crystal violet and their leuco metabolites residues in aquatic product by liquid chromatography with visible detection and tandem mass spectrometric detection is described. The compounds were extracted from homogenized tissue with acetonitrile-buffer, partitioned against methylene dichloride, and purified with MCX cation-exchange solid-phase extraction cartridges. Samples were chromatographed isocratically using an acetonitrile- ammonium acetate buffer mobile phase on a C18 column in-line with a post-column reactor filled with lead （IV） oxide and celite followed by diode array detection （DAD） or electrospary ionization-tandem mass spectrometry （ESI-MS-MS） analysis. Chromatic and leuco malachite green were quantified by their absorbance at 618 nm, and chromatic and leuco crystal violet at 588 nm, brilliant green was used as internal standard. Analytes confirmation were performed using multiple reaction monitoring （MRM） with one precursor ion and two product ions as identitiers, employing D6-LMG as internal standard. For high performance liquid chromatography （ HPLC ）-DAD, the limit of detection （LOD） were 0.22 （MG）, 0.28 （LMG）, 0.22 （CV）, 0.25 （LCV） g/kg. And for HPLC-MS-MS, the LOD were 0.014（MG）, 0.018（LMG）, 0.014 （CV）, 0.0084（LCV） g/kg. Recoveries ranged from 75 % to 95 % at a spiking level from 2 μg/kg to 20 μg/kg.