反相高效液相色谱法检测鲫鱼组织中的节球藻毒素

Determination of Nodularin in Carassius auratus Tissues by Reversed Phase High Performance Liquid Chromatography

通过对鲫鱼各组织前处理条件和反相高效液相色谱条件的优化,建立了反相高效液相色谱法检测鲫鱼肝胰脏、肾脏、血浆及肌肉中节球藻毒素含量的方法。样品利用90%甲醇超声萃取,CNW Bond C18固相萃取柱净化除杂,反相高效液相色谱配置紫外检测器检测。C18柱净化步骤依次为活化-上样-淋洗-洗脱,洗脱剂为含0.1%三氟乙酸（TFA）的90%甲醇水溶液。RP-HPLC检测以0.1%TFA-甲醇（42∶58）为流动相,流速为1.0 m L/min,柱温为30℃,进样量为20μL,检测波长为238 nm。结果表明：在0.05~5.00mg/L浓度范围内,4种鲫鱼组织中节球藻毒素的质量浓度与其峰面积的线性关系良好（r2〉0.99）。在0.08,0.40,2.50 mg/L加标水平下,回收率为79.5%~106.4%,批内相对标准偏差（RSD）为0.54%~5.3%。肌肉中方法检出限为11.30μg/kg,血浆中方法检出限为24.00μg/L。本方法结果准确可靠,可应用于鲫鱼各组织中节球藻毒素含量的动态分布以及残留检测。

A reversed phase high performance liquid chromatographic（ RP- HPLC） method was developed for the determination of nodularin in hepatopancreas,kidney,muscle and plasma tissues of Carassius auratus（ C. auratus） by optimizing conditions of pretreament and chromatography. Samples were extracted with 90% methanol solution by ultrasonic,followed by purification with a CNW Bond C18solid-phase extration column,and then determined by RP-HPLC equipped with ultraviolet（ UV）detector. The steps of purification included activating columns, sample absorbing onto columns,flushing the columns and eluting the columns. The eluent was 90% methanol containing 0. 1% trifluoroacetic acid（ TFA）. The detection of target compound was performed with RP- HPLC using 0. 1%trifluoroacetic acid（ TFA）- methanol（ 42 ∶ 58） as mobile phase at a constant flow of 1. 0 m L/min.The column temperature was set at 30 ℃ and the wavelength of UV detector was set at 238 nm while the injection volume was 20 μL. The experimental results showed that the concentration of nodularin in four crucian tissues showed good linearity（ r2〉 0. 99） with peak area in the range of 0. 05- 5. 00 mg/L. The recoveries of nodularin from crucian tissues at three spiked concentration levels of 0. 08,0. 40,2. 50 mg/L ranged from 79. 5% to 106. 4%,with RSDs of 0. 54%- 5. 3%. The method detection limit was 11. 30 μg/kg for muscle and 24. 00 μg/L for plasma sample. The method was accurate and reliable,and could be applied in the dynamic distribution and residues determination of nodularin in C. auratus,providing an effective tool for the detection of nodularin in other aquatic products and for the safety control of aquatic products quality.