液质联用法检测备孕及妊娠妇女血液中9种叶酸及其代谢物  

Determination of nine folates and its metabolites in the blood of preconception and pregnant women by LC-MS/MS

作　　者：李海娟 顾玲琪 陈万杰 施益金 秦燕[1] 张振宇 顾锦华[1,3,4] LI Haijuan;GU Lingqi;CHEN Wanjie;SHI Yijin;QIN Yan;ZHANG Zhenyu;GU Jinhua(Department of pharmacy,Affiliated Maternity and Child Health Care Hospital of Nantong University(Nantong Children’s Hospital),Jiangsu Nantong 226001,China;Department of Clinical Laboratory,Affiliated Maternity and Child Health Care Hospital of Nantong University(Nantong Children’s Hospital),Jiangsu Nantong 226001,China;Department of Clinical Pharmacy,School of Pharmacy,Nantong University,Jiangsu Nantong 226019,China;Nantong Institute of Genetics and Reproductive Medicine,Jiangsu Nantong 226001,China)

机构地区：[1]南通大学附属妇幼保健院(南通市儿童医院)药学部,江苏南通226001 [2]南通大学附属妇幼保健院(南通市儿童医院)检验科,江苏南通226001 [3]南通大学药学院临床药学系,江苏南通226019 [4]南通市遗传与生殖医学研究所,江苏南通226001

出　　处：《中国医院药学杂志》2025年第3期287-294,共8页Chinese Journal of Hospital Pharmacy

基　　金：南通市科技计划项目(编号:MSZ2022097);江苏卫生创新团队项目(编号:CX2023006)。

摘　　要：目的:建立并验证基于LC-MS/MS定量检测人体内9种叶酸及其代谢物的方法,并应用于备孕及妊娠群体。方法:对血液样本进行抗氧保护、蛋白解离及有机溶剂蛋白沉淀预处理;色谱柱选用Kinetex F5,流动相为含0.1%甲酸-水(A)和甲醇(B),以300μL·min-1的流速梯度洗脱8 min;采用电喷雾离子源,正离子多反应监测模式。对2022年5月至2023年9月南通市妇幼保健院918例备孕期及妊娠期妇女血液样本检测结果进行统计分析,比较不同阶段叶酸代谢物的差异并总结其分布情况。结果:9种叶酸代谢产物[叶酸FA、5-甲基四氢叶酸(5-MTHF)、5-甲酰基四氢叶酸(5-FTHF)、同型半胱氨酸(Hcy)、S-腺苷甲硫氨酸(SAM)、S-腺苷同型半胱氨酸(SAH)、维生素B12、维生素B2、维生素B6和红细胞中5-MTHF]分别在一定范围内呈线性相关,相关系数r2≥0.9928;准确度87.61%~109.72%,RSD≤7.97%;批内和批间精密度RSD≤18.18%;最大稀释倍数、仪器残留符合相关要求;样品在进样器中存放24 h后稳定性良好。随着孕周增加,血清和红细胞中5-MTHF呈上升趋势,Hcy和维生素B12浓度呈下降趋势,SAM/SAH比例呈下降趋势。结论:该方法操作简便,具有良好的准确度、灵敏度和重复性,可同时测定备孕期及妊娠期妇女血液中9种叶酸代谢物的水平。临床实践证明,该方法适用于备孕期及妊娠期妇女叶酸的精准补充,可用于建立本地区叶酸参考范围。OBJECTIVE To establish and validate a liquid chromatography tandem mass spectrometry(LC-MS/MS)method for quantification of nine folate metabolites in biological samples,and to apply it to the preconception and pregnancy cohorts.METHODS The blood samples were pre-processed by antioxidant protection,protein dissociation,and organic solvent protein precipitation.Using the Kinetex F5 column,0.1%formic acid aqueous solution and methanol as the mobile phase at a flow rate of 300μL·min-1 for an 8-minute gradient elution,LC-MS/MS was conducted.Electrospray ionization was in positive ion multiple reaction monitoring mode.A total of 918 blood samples collected from Nantong Maternal and Child Health Hospital from May 2022 to September 2023 were analyzed for the differences in folate metabolites at different stages.Their distribution patterns were summarized.RESULTS Nine folate metabolites,including folic acid(FA),5-methyltetrahydrofolate(5-MTHF),5-formyltetrahydrofolate(5-FTHF),homocysteine(Hcy),S-adenosylmethionine(SAM),S-adenosylhomocysteine(SAH),vitamin B12(VB12),vitamin B2(VB2),vitamin B6(VB6),and 5-MTHF in red blood cells,exhibited linear correlations within a defined range,with correlation coefficients≥0.9928.The accuracy was 87.61%-109.72%,and the precision(relative standard deviation[RSD])≤7.97%.The intra-and inter-batch precision(RSD)was lower than 18.18%.The maximum dilution and instrument residues met relevant requirements,and sample stability was good after 24 hours in the autosampler.As pregnancy progressed,serum and red blood cell levels of 5-MTHF increased,while concentrations of Hcy and VB12 decreased.Additionally,the ratio of SAM to SAH(SAM/SAH)declined.CONCLUSION The LC-MS/MS method is simple to operate,with good accuracy,sensitivity,and repeatability,allowing for simultaneous determination of 9 folate metabolites.Clinical practice evidence validates its effective application to precise supplementation of folates in women during preconception and pregnancy,enabling the establishment of local referenc

关 键 词：液质联用法 叶酸及其代谢物 备孕期及妊娠期群体 精准补充 

分 类 号：R927.2[医药卫生—药学]