Hu Xiaobin, Ye Jinyun, Zhang Rongfei, Wu Xiang, Zhang Yixiang, Wu Chenglong
a School of Life Science , Huzhou University , Huzhou , China.
J Environ Sci Health B. 2017 Oct 3;52(10):770-776. doi: 10.1080/03601234.2017.1356670. Epub 2017 Sep 22.
MC analysis of biological tissue is considered to be very difficult due to the lack of validated methods. This is the primary limiting factor for monitoring potential risks in both the flesh of aquatic organisms and the aquatic ecosystem. In this study, an effective method to determine free MCs (MC-LR and MC-RR) in the muscle and liver tissues of freshwater cultured fish was developed using solid-phase extraction (SPE) and liquid chromatography-tandem mass spectrometry (LC/MS-MS). The extraction solvent, time of extraction, eluent and purification of the extract were optimized. Various SPE cartridges were also investigated. In this optimized analytical procedure, an 85% methanol/water solution (v/v) was selected as the extraction solvent, after which the extracts were purified by removing fats and proteins; a HLB cartridge was chosen for MCs enrichment; and 90% methanol containing 0.02% formic acid/water solution (v/v) was used as the eluent. Under the optimized pretreatment conditions and instrument parameters, good recoveries of MC-LR and MC-RR were obtained at three concentrations (0.5, 1.0 and 2.0 µg g dry weight (DW)), with values ranging from 92.5 to 98.3% and 92.1 to 98.6%, respectively. The method detection limit (MDL) for muscle samples was 0.5 µg kg and 0.4 µg kg (DW) for MC-LR and MC-RR, respectively. The MDL for the liver samples was 0.8 µg kg (DW) for both MC-LR and MC-RR. The developed procedure was successfully applied to analyze MCs in the muscle and liver of fish samples collected from a Chinese freshwater aquaculture pond during bloom seasons. The MC-LR concentrations ranged from below the MDL to 4.17 µg kg and the MC-RR concentrations ranged from below the MDL to 2.64 µg kg.
由于缺乏经过验证的方法，对生物组织进行微囊藻毒素（MC）分析被认为非常困难。这是监测水生生物体内和水生生态系统潜在风险的主要限制因素。在本研究中，开发了一种使用固相萃取（SPE）和液相色谱 - 串联质谱（LC/MS - MS）测定淡水养殖鱼肌肉和肝脏组织中游离微囊藻毒素（MC - LR和MC - RR）的有效方法。对萃取溶剂、萃取时间、洗脱液和提取物的纯化进行了优化。还研究了各种SPE小柱。在这个优化的分析程序中，选择85%甲醇/水溶液（v/v）作为萃取溶剂，之后通过去除脂肪和蛋白质对提取物进行纯化；选择HLB小柱用于微囊藻毒素富集；使用含0.02%甲酸的90%甲醇/水溶液（v/v）作为洗脱液。在优化的预处理条件和仪器参数下，在三个浓度（0.5、1.0和2.0μg/g干重（DW））下获得了MC - LR和MC - RR的良好回收率，其值分别为92.5%至98.3%和92.1%至98.6%。肌肉样品中MC - LR和MC - RR的方法检测限（MDL）分别为0.5μg/kg和0.4μg/kg（DW）。肝脏样品中MC - LR和MC - RR的MDL均为0.8μg/kg（DW）。所开发的程序成功应用于分析中国淡水养殖池塘在藻华季节采集的鱼样品的肌肉和肝脏中的微囊藻毒素。MC - LR浓度范围从低于MDL到4.17μg/kg，MC - RR浓度范围从低于MDL到2.64μg/kg。
