超高效液相色谱-质谱法测定蘑菇中5种毒肽

UHPLC-MS Determination of 5 Poisonous Peptides in Mushrooms

采用超高效液相色谱-质谱法测定蘑菇中α-鹅膏毒肽、β-鹅膏毒肽、γ-鹅膏毒肽、二羟鬼笔毒肽和羧基二羟鬼笔毒肽等5种毒肽的含量。均质后的样品用甲醇超声提取15min,离心后,上清液置于50℃水浴中用氮气吹干,残留物用甲醇(1+9)溶液溶解并定容至3mL,再用0.2μm滤膜过滤,滤液经HLB固相萃取柱净化。以Acquity UPLC HSS T3色谱柱为固定相,以不同体积比的含5mmol·L^(-1)乙酸铵的0.1%(体积分数)甲酸溶液和甲醇混合液为流动相进行梯度洗脱,串联质谱分析中采用电喷雾正离子源和多反应监测模式。5种毒肽的质量浓度均在0.05～1.00mg·L^(-1)内与其对应的峰面积呈线性关系,检出限(3S/N)为0.100～0.300mg·kg^(-1)。加标回收率为54.9%～106%,测定值的相对标准偏差(n=6)均小于7.5%。

UHPLC-MS was applied to the determination of 5 poisonous peptides,i.e.α-amatoxins,β-amatoxins,γ-amatoxins, dihydroxy-phallotoxins and carboxyl dihydroxy-phallotoxins in mushrooms.The homogenized sample was extracted ultrasonically for 15 min with methanol.After centrifugation,the supernatant was evaporated to dryness by N2-blowing in a water bath at 50℃and then the residual was dissolved and made up to 3 mL with methanol(1+9)solution.After filtering on 0.2μm filtering membrane,the filtrate was purified with solid phase extraction column.Acquity UPLC HSS T3 chromatographic column was used as stationary phase,and the mixture of 0.1%(φ)formic acid solution(containing 5 mmol·L-1 ammonium acetate)and methanol mixed in different ratios was used as mobile phase in gradient elution.ESI+and multi-reactions monitor mode were adopted in MS/MS.Linear relationships between values of peak area and mass concentration of the 5 poisonous peptides were kept in the same range of 0.05-1.00 mg·L-1,with detection limits(3 S/N)in the range of 0.100-0.300 mg·kg-1.Values of recovery found by standard addition method were in the range of 54.9%-106%,and values of RSDs(n=6)were less than 7.5%.