Determination of ?-hydroxy-?-methylbutyrate concentration and enrichment in human plasma using chemical ionization gas chromatography tandem mass spectrometry.

ABSTRACT: Our objective was to develop a quick and simplified method for the determination of ?-Hydroxy-?-methylbutyrate (HMB) and ?-ketoisocaproic acid (KIC) concentrations and enrichments by GC/MS/MS to determine the turnover rate of HMB in humans. In experiment 1, we provided a pulse of L-[5,5,5-2H3]leucine to younger adults in the postabsorptive state then collected blood samples over a 4h time period. In experiment 2, we provided a pulse of [3,4,methyl-13C3]HMB to older adults in the postabsorptive state then collected blood samples over a 3h time period. Plasma concentrations of KIC and HMB and MPE of KIC and HMB were determined by GC/MS/MS. Plasma enrichment of leucine was determined by LC/MS/MS. To determine plasma enrichment of [5,5,5-2H3]HMB and [3,4,methyl-13C3]HMB, samples were derivatized using pentafluorobenzyl bromide and analyzed using chemical ionization mode. The final methods used included multiple reaction monitoring of transitions 117.3>59.3 for M+0 and 120.3>59.3 for M+3. In experiment 1, peak MPE of Leu peaked at 9.76% generating a peak MPE of KIC at 2.67% and a peak HMB MPE of 0.3%. In experiment 2, the rate of appearance for HMB was? 0.66?mol/kg ffm/h. We calculated that production of HMB in humans accounts for 0.66% of total leucine turnover.

SUBMITTER: Walker DK 

PROVIDER: S-EPMC5191936 | biostudies-literature | 2017 Jan

REPOSITORIES: biostudies-literature