通过式固相萃取-高效液相色谱-串联质谱法快速检测发酵乳中6种生物胺

Rapid determination of 6 kinds of biogenic amines in fermented dairy products by high performance liquid chromatography-tandem mass spectrometry with pass-through solid-phase extraction

中文关键词:  生物胺  发酵乳  通过式固相萃取  高效液相色谱-串联质谱

英文关键词:biogenic amines  fermented dairy products  pass-through solid-phase extraction  high performance liquid chromatography-tandem mass spectrometry

中文摘要:

      目的 建立通过式固相萃取-高效液相色谱-串联质谱法(high performance liquid chromatography-tandem mass spectrometry, HPLC-MS/MS)快速检测发酵乳中6种生物胺的方法。方法 样品经2%甲酸/乙腈溶液提取后, 采用Oasis PRiME HLB固相萃取柱进行净化, 有效去除蛋白质和磷脂等干扰物。色谱分离采用亲水作用色谱柱, 质谱检测在多反应监测模式下进行, 以内标法定量。结果 6种生物胺在一定浓度范围内线性关系良好(r2≥0.996), 检出限为0.20~1.80 μg/kg, 定量限为0.67~6.00 μg/kg, 加标回收率为76.0%~117.8%, 相对标准偏差小于13%。对市售发酵乳样品分析结果表明, 酪胺为最主要的生物胺, 浓度范围为在0.80~1.70 mg/kg, 而组胺等与安全风险相关的生物胺含量极低或未检出。结论 本研究建立的方法, 通过引入Oasis PRiME HLB净化策略, 有效解决了乳品中磷脂干扰的问题, 具备前处理简便快速、灵敏度高、抗干扰能力强等优点, 适用于发酵乳中多种生物胺的高通量测定, 为乳制品质量控制与安全风险评估提供有力的技术支撑。

英文摘要:

      Objective To establish a method for the rapid determination of 6 kinds of biogenic amines in fermented milk by pass-through solid-phase extraction coupled with high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). Methods After extraction with a 2% formic acid/acetonitrile solution, samples were purified using an Oasis PRiME HLB solid-phase extraction cartridge to efficiently remove interferences such as proteins and phospholipids. Chromatographic separation was performed on a hydrophilic interaction chromatography column. Mass spectrometric detection was carried out in multiple reaction monitoring mode, and quantification was achieved by the internal standard method. Results The method was validated and demonstrated good linearity (r2≥0.996) for the 6 kinds of biogenic amines within a certain concentration range, the limits of detection were 0.20 μg/kg–1.80 μg/kg, the limits of quantitation were 0.67–6.00 μg/kg. Spiked recoveries were 76.0%–117.8%, with relative standard deviations below 13%. Analysis of commercial fermented dairy samples revealed that tyramine was the predominant biogenic amine, with concentrations ranging from 0.80 to 1.70 mg/kg, while safety-risk-associated biogenic amines such as histamine were detected at extremely low levels or were not detected. Conclusion The established method, by incorporating the Oasis PRiME HLB purification strategy, effectively resolves the critical issue of phospholipid interference in dairy products. It features advantages such as simple and rapid sample preparation, high sensitivity and strong anti-interference capability. The method is suitable for the high-throughput determination of multiple biogenic amines in fermented dairy products, providing robust technical support for the quality control and safety risk assessment of these products.