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目的:考察EDTA-K2和肝素锂抗凝血浆不同前处理方法和液相条件对于高效液相色谱质谱联用法检测莎巴比星及其代谢产物M3基质效应的影响。方法:以含不同抗凝剂的空白血浆配制低、中、高浓度样品,乙酸乙酯或氯仿液液萃取,流动相:A:0.1%甲酸水,B:乙腈,等度洗脱或梯度洗脱。莎巴比星、M3和内标多柔比星检测离子对分别为:m/z 644→130,m/z 646→333.2,m/z 544→360。采用提取后加入法,评价各浓度莎巴比星和M3在2种抗凝剂血浆中基质效应的差异以及方法优化对基质效应的影响。结果:低浓度时,EDTA-K2组莎巴比星和M3基质效应分别为123%和110%,肝素锂组分别为143%和160%。基质效应在不同抗凝剂组间差异有统计学意义(莎巴比星:P<0.05;M3:P<0.05)。氯仿萃取肝素锂抗凝血浆中莎巴比星低、中、高浓度基质效应分别为142%,94%和78%,M3分别为123%,102%和89%。梯度洗脱莎巴比星低、中、高浓度基质效应分别为108%,81%和73%,而M3分别为93%,83%和73%。最终低浓度莎巴比星和M3基质效应分别为85.4%和98.7%,高浓度分别为88.5%和96.0%。结论:肝素锂抗凝血浆中莎巴比星及其代谢物M3的基质增强效应更强,且在不同浓度存在较大差异,优化前处理方法以及液相方法后基质效应得到显著改善。
OBJECTIVE: To investigate the effect of different pretreatment methods and liquid phase conditions of EDTA-K2 and lithium heparin anticoagulated plasma on the matrix effects of shababixix and its metabolite M3 by high performance liquid chromatography-mass spectrometry. Methods: Samples with low, medium and high concentrations of blank plasma containing different anticoagulants were prepared and extracted with ethyl acetate or chloroform. The mobile phase consisted of A: 0.1% formic acid water, B: acetonitrile, isocratic elution or gradient elution Off Sabourad, M3 and the internal standard doxorubicin detection ion pairs were: m / z 644 → 130, m / z 646 → 333.2, m / z 544 → 360. Addition method was used to evaluate the difference of matrix effects between sharbix and M3 in two kinds of anticoagulant plasma and the effect of the method on matrix effect. RESULTS: At low concentrations, the matrix effects of shababixin and M3 in EDTA-K2 group were 123% and 110%, respectively, and 143% and 160% in heparin group, respectively. There was significant difference in matrix effect between different anticoagulant groups (shabavirine: P <0.05; M3: P <0.05). The matrix effects of low, medium and high concentrations of sharbixin in chloroform-extracted lithium heparin anticoagulated plasma were 142%, 94% and 78%, respectively, and M3 was 123%, 102% and 89%, respectively. The matrix effects of low, medium, and high concentration of shaburabine were 108%, 81% and 73%, respectively, while those of M3 were 93%, 83% and 73%, respectively. The final low concentration of Sabourad and M3 matrix effects were 85.4% and 98.7%, respectively, with high concentrations of 88.5% and 96.0%, respectively. CONCLUSIONS: The enhancement effect of shababixin and its metabolite M3 in lithium heparin anticoagulated plasma is stronger and different at different concentrations. The pretreatment method and liquid-phase method have significantly improved the matrix effect.