The determination of organic acids in urine with gas chromatography-mass spectrometry and its application in clinical diagnosis / 中华检验医学杂志

ABSTRACT

Objectivo To analyze urine organic acids in the urine using gas chromatography-mass spectrometry(GC/MS)for diagnosis of inherited metabolic diseases,especially for organic acids metabolic disorders.Methods 195 clinical urine samples from the patients with suspected organic acids metabolic disorders and 5 normal urine from adults were collected.After mixing some urine with intemal standards according to the concentration creatinine and adding hydroxylamine hydrochloride to mixture,the organic acids with hydroxyl group were oximated to the ketobodies.Organic acids were extracted twice with ethyl acetate and ethyl ether and derivatized with BSTFA-TMCS.An the organic acids were determined with Agilent GC/MS 6890/5973i with scan model.the mass-to-charge ratio range is 50-550 m/z,all data were nalyzed with Agilent GCMSD ChemStationG1701DA.We also investigated the linearity, accurate,precision.recovery and Carry-over by determining the internal standards in normal samples and positive organic acids in spiked control samples.Results More than one hundred kinds of organic acids in urine samples can be analyzed with this method.According to the two internal standards in normal urine samples,the minimal detection limit MMA and 2.PA was 2.5-2.8 μmol/L.Intra-and interassay coefficient of variation for MMA and 2-PA are both less than 10%.Pre-processing Interassay coefficient by sequential preparations of the same sample was 14%.The recoveries of the spiked samples were 95%-105%.Carryover analysis was less than 1%.All the parameters meet the requirement for clinical diagnosis.12 samples demonstrated positive including 6 cases of methylmalonic acidemia,1 case of propionic acidemia,3 cases of tyrosinemia Ⅰ,1 case of maple syrup urine disease and 1 cases of ketosis.Conclusions The method for the determination of organic acids in urine by GC/MS has been successfully established.It can be used for clinical screening and diagnosis for inherited genetic metabolic diseases.