摘要
目的建立一种基于液相色谱串联质谱(LC-MS/MS)系统检测人血浆中6种植物固醇的方法,并评价其性能。方法方法学建立、评价及应用。收集2023年10月至12月苏州市立医院体检中心招募的健康体检者136名,同时对苏州市立医院心血管内科就诊的5例植物固醇血症患者的血浆植物固醇含量进行验证。实验采用氘代同位素作为内标,使用Phenomenex C18(2.1×100 mm,2.6μm)色谱柱进行分离;流动相为水和甲醇,流速0.4 ml/min进行梯度洗脱,柱温25℃;使用AB Sciex 6500+三重四级杆质谱建立LC-MS/MS检测植物固醇的方法,对该方法的线性、最低检测下限、回收率、精密度和稳定性进行性能评价,并在植物固醇血症患者血浆中对检测方法进行验证。结果该方法检测的6种植物固醇批内及批间变异系数分别为1.8%~12.5%、2.6%~13.6%,符合精密度要求;回收率为85.0%~105.1%,符合正确度的要求;处理前以及处理后样本在4℃条件放置72 h均具有良好的稳定性。植物固醇血症患者血浆谷甾醇、菜油甾醇、豆甾醇、胆甾烷醇含量均明显高于健康体检组(P均<0.001)。结论建立了一种基于LC-MS/MS定量检测人血浆中6种植物固醇的方法。该方法样本前处理操作简便,通过了线性、定量下限、回收率、精密度和稳定的性能评价,可用于临床检测。
ObjectiveTo establish and evaluate the performance of a stable and sensitive liquid chromatography-tandem mass spectrometry(LC-MS/MS)method for the determination of six phytosterols in human plasma.MethodsThe methodology was established,evaluated,and applied.A number of 136 healthy subjects were recruited from the Physical Examination Center of Suzhou Municipal Hospital from October 2023 to December 2023,meanwhile plasma phytosterols′levels were validated in 5 patients with sitosterolemia.Taking deuterium isotope as internal standards,the clear separation was achieved using a Phenomenex C18(2.1×100 mm,2.6μm)column.The mobile phase consisted of water and methanol at a flow rate of 0.4 ml/min with gradient elution,and the column temperature was 25℃.The experiments were performed on AB Sciex 6500+triple quadrupole mass spectrometry,and the linearity,limit of quantification,rate of recovery,precision,and stability of the method were evaluated.The testing method was further validated in the sitosterolemia patients.ResultsThe method had in-run and between-run coefficient of variation ranging 1.8%-12.5%and 2.6%-13.6%,respectively,which accorded with the precision requirements.The rates of recovery were 85.0%-105.1%,which accorded with the accuracy requirements.The samples showed good stability before and after handling stored at 4℃for 72 hours.The plasma levels of sitosterol,campesterol,stigmasterol and cholestanol in sitosterolemia patients were apparently higher compared with those in the healthy group(P<0.001).ConclusionLC-MS/MS method for the quantitative measurement of plant sterols in human plasma has been established,which has convenient preprocessing and passed the performance evaluation including linearity,lower limit of quantification,recovery rate,precision and stability,and shown to be suitable for clinical application.
作者
章禹轩
王绿娅
万玮敏
丁虎
吴俊芳
Zhang Yuxuan;Wang Luya;Wan Weimin;Ding Hu;Wu Junfang(Department of Cardiology,Tongji Hospital,Tongji Medical College,Huazhong University of Science and Technology,Wuhan430000,China;Department of Cardiology,Beijing Anzhen Hospital,Affiliated of Capital University of Medical Sciences,Beijing100029,China;Department of Laboratory Medicine,Suzhou municipal hospital,Suzhou215002,China)
出处
《中华检验医学杂志》
北大核心
2025年第1期125-132,共8页
Chinese Journal of Laboratory Medicine
基金
国家重点研发计划(2021YFC2500600,2021YFC2500604,2021YFC2500602)
国家自然科学基金(82370397)
湖北省重点研发计划(2022BCA037)。
关键词
植物固醇
液相色谱-质谱法
血浆
谷固醇血症
Phytosterol
Liquid chromatography/mass spectrometry
Plasma
Sitosterolemia
作者简介
通信作者:吴俊芳,Email:junfang.wu@tjh.tjmu.edu.cn。