四川大学学报（医学版）2025，Vol.56Issue(5)：1226-1234,9.DOI:10.12182/20250960502
电膜萃取-液相色谱质谱法高通量检测尿液中内源性肽的研究
High-Throughput Determination of Endogenous Peptides in Urine Using Electromembrane Extraction Combined With Liquid Chromatography-Tandem Mass Spectrometry
摘要
Abstract
Objective To develop a precise method for analyzing urinary peptides based on electromembrane extraction(EME)combined with liquid chromatography-tandem mass spectrometry(LC-MS/MS),and to evaluate its potential applicability in tumor biomarker screening.Methods A total of 15 disease-associated peptides were selected as the target analytes.A supported liquid membrane(SLM)composed of n-octanol containing 5%di(2-ethylhexyl)phosphate was employed,with the donor phase being a 1∶1 mixture of urine and 100 mmol/L formic acid and urine,and the acceptor phase being 20 mmol/L formic acid containing 50%dimethyl sulfoxide(DMSO).After EME at 40 V for 15 min,the acceptor phase solution was analyzed by LC-MS/MS.Subsequently,the method,EME combined with LC-MS/MS(EME-LC-MS/MS),was preliminarily validated utilizing urine samples from 12 healthy controls and 7 patients with urinary system tumors.Results All 15 peptides exhibited excellent linearity in the range of 0.1-100.0 ng/mL(r≥0.995),with the limits of detection(LODs)being 0.01-0.50 ng/mL and the limits of quantification(LOQs)being 0.03-1.50 ng/mL.The spiked recoveries ranged from 21.0%to 71.2%,with relative standard deviations(RSDs)of 0.8%-20.0%(n=3).Small-sample analysis of clinical specimens revealed that the concentration of bradykinin 1-5 in the urine were significantly higher in tumor patients(median:0.65 ng/mL)than that in healthy controls(median:0.37 ng/mL)(P<0.05),suggesting its potential as a specific biomarker for urinary system tumors.Conclusion The EME-LC-MS/MS method established in the study features simplicity,high efficiency,and high sensitivity,enabling precise determination of trace-level peptides in urine samples.Moreover,this approach provides a reliable methodological basis for disease biomarker screening and promotes the clinical application of electromembrane extraction.关键词
电膜萃取/微萃取/肿瘤标志物/尿液/肽Key words
Electromembrane extraction/Microextraction/Tumor biomarkers/Urine/Peptides引用本文复制引用
唐勇,何俊玲,林驰量,彭绍芸,邹海民,周琛..电膜萃取-液相色谱质谱法高通量检测尿液中内源性肽的研究[J].四川大学学报（医学版）,2025,56(5):1226-1234,9.基金项目
国家自然科学基金(No.82304196)、四川省卫生健康委员会科技项目(No.24QNMP026)和成都市科技局重点研发项目(No.2024-YF05-00950-SN)资助 （No.82304196）
