title: Determination of β-hydroxy-β-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 β-Hydroxy-β-methylbutyrate (HMB) and ɑ-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  0.66μmol/kg ffm/h. We calculated that production of HMB in humans accounts for 0.66% of total leucine turnover.
authors.affiliation: Center for Translational Research in Aging & Longevity, Department of Health & Kinesiology, Texas A&M University, College Station, TX 77843, USA. Electronic address: dk.walker@tamu.edu.
authors.forename: Dillon K
authors.initials: DK
authors.lastname: Walker
authors.affiliation: Center for Translational Research in Aging & Longevity, Department of Health & Kinesiology, Texas A&M University, College Station, TX 77843, USA. Electronic address: jj.thaden@ctral.org.
authors.forename: John J
authors.initials: JJ
authors.lastname: Thaden
authors.affiliation: Center for Translational Research in Aging & Longevity, Department of Health & Kinesiology, Texas A&M University, College Station, TX 77843, USA. Electronic address: a.wierzchowska@ctral.org.
authors.forename: Agata
authors.initials: A
authors.lastname: Wierzchowska-McNew
authors.affiliation: Center for Translational Research in Aging & Longevity, Department of Health & Kinesiology, Texas A&M University, College Station, TX 77843, USA. Electronic address: mpkj.engelen@ctral.org.
authors.forename: Marielle P K J
authors.initials: MPKJ
authors.lastname: Engelen
authors.affiliation: Center for Translational Research in Aging & Longevity, Department of Health & Kinesiology, Texas A&M University, College Station, TX 77843, USA. Electronic address: nep.deutz@ctral.org.
authors.forename: Nicolaas E P
authors.initials: NEP
authors.lastname: Deutz
mesh_terms: D000328:Adult; D000368:Aged; D008401:Gas Chromatography-Mass Spectrometry / Q000191:economics / Q000379:methods*; D006801:Humans; D007651:Keto Acids / Q000097:blood*; D057230:Limit of Detection; D008875:Middle Aged; D053719:Tandem Mass Spectrometry / Q000191:economics / Q000379:methods*; D014631:Valerates / Q000097:blood*; D055815:Young Adult
chemical_list: D007651:Keto Acids; D014631:Valerates; C004961:beta-hydroxyisovaleric acid; C013082:alpha-ketoisocaproic acid
keywords: Chemical ionization; Gas chromatography; HMB; Tandem mass spectrometry; Turnover