Determination of Urine VMA, HVA and 5-HIAA by Highspeed Liquid Chmmatography

概要

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A simultaneous determination of urine VMA and HVA, as well as 5-HIAA was attempted by highspeed liquid chromatography. Urine was acidified with 6 N hydrochloric acid and extracted with ethyl acetate. The ethyl acetate layer was evaporated to dryness under reduced pressure. The residue was dissolved in 0.2 N acetic acid, and 40μl of the aliquot was injected on to the column. A highspeed liquid chromatograph was assembled from Waters ALC/GPC204 Type, the model 660 soovent programmer and packed with μBondapack C18. The mobile phase was consisted of water and methanol solution of PIC-B7. Elution was done gradiently by from 15 to 30% methanol in 10 minutes. The value of VMA was well correlated to that obtained colorimetrically. Moreover, the method proved to be very specific, reproducible with a good recovery. The technique was applied to the 10 urine specimens of normal subjects. The mean excretion of VMA was3.5±1.1 S. D. mg/day, 5-HIAA 3.3±1.3 S.D. mg/day and HVA 8.7±2.4 S.D. mg/day. In a case of phaeochromocytoma,they were 15.2mg/day, 6.2mg/day and 19.0mg/day respectively. Thus the method issimple and accurate enough for a routine simultaneous determination of urine VMA, HVA as well as 5-HIAA.