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目的开展尿液和血浆中3种鹅膏毒肽和3种鬼笔毒肽测定的方法学研究。方法尿液和血浆样本经乙腈沉淀蛋白后,HLB固相萃取柱萃取净化,超高效液相色谱-串联质谱法测定α-鹅膏毒肽、β-鹅膏毒肽、γ-鹅膏毒肽、羧基二羟鬼笔毒肽、二羟鬼笔毒肽和羧基三羟鬼笔毒肽6种蘑菇毒肽,基质匹配制作工作曲线,外标法定量;对样本测定条件进行优化,减小基质干扰以提高方法的灵敏度;研究方法的适用性,考察方法的专一性、线性、灵敏度、精密度和加标回收率。结果尿液和血浆中6种毒肽的检出限均低于0.06 ng/ml;在0.2~20 ng/ml范围内,线性关系良好;相关系数R2均大于0.999 3;相对标准偏差均小于12.1%;对于上述两种基质,加标回收率在79.6%~104.8%之间。结论方法灵敏、准确、专一性好,适合于突发毒蕈中毒事件中α-鹅膏毒肽、β-鹅膏毒肽、γ-鹅膏毒肽、羧基二羟鬼笔毒肽、二羟鬼笔毒肽和羧基三羟鬼笔毒肽的确证分析,也适合对中毒患者临床治疗进行效果评价。
Objective To develop a methodological study on the determination of three amanitin peptides and three phalloidine peptides in urine and plasma. Methods Urine and plasma samples were precipitated with acetonitrile and purified by HLB SPE. The concentrations of α-amanitin, β-amanitin and γ-amanitin were determined by ultra performance liquid chromatography-tandem mass spectrometry , Carbodihydro penicillin peptide, dihvdromycin penicillin and carboxy trihydroxy penicillin peptide six kinds of mushroom toxins, matrix matching production curve, external standard method; the sample determination conditions are optimized to reduce the matrix Interference to improve the sensitivity of the method; study the applicability of the method, investigate the specificity of the method, linearity, sensitivity, precision and recovery. Results The detection limits of all six toxins in urine and plasma were lower than 0.06 ng / ml. The linearity was good in the range of 0.2-20 ng / ml. The correlation coefficients R2 were all greater than 0.999 3. The relative standard deviations were all less than 12.1 %; For the two substrates, the spiked recoveries ranged from 79.6% to 104.8%. Conclusion The method is sensitive, accurate and specific. It is suitable for alpha-amanitin, beta-amanitin, gamma-amanitin, Hydroxyapatite peptide and carboxyl trihydroxylamine penicillin peptide confirmatory analysis, but also suitable for clinical evaluation of poisoning patients.