HPLC Separation of β‑Hydroxy-β-methylbutyric Acid, 2-Hydroxyisocaproic Acid and Leucine on Primesep 100 Column

Both β-hydroxy-β-methylbutyric acid (HMB) and 2-hydroxyisocaproic acid (HICA) are metabolites of the essential dietary branched chain amino acid leucine.


Column Primesep 100, 4.6x150 mm, 5 µm, 100A
Mobile Phase MeCN - 5%, 3 min hold, Gradient MeCN 5-30% 12 min
Buffer H2SO4 - 0.05 % 3 min hold, GradientH2SO4 0.05-3% 12 min
Flow Rate 1.0 ml/min
Detection UV, 210 nm


Class of Compounds Acid, Hydrophilic,  Ionizable, Normal human metabolite, Dietary supplement, Essential amino acid, Branched-chain amino acid (BCAA)
Analyzing Compounds β‑Hydroxy-β-methylbutyric (HMB), 2-Hydroxyisocaproic Acid (HICA), Leucine

Application Analytes:

2-Hydroxyisocaproic Acid (HICA)
β‑Hydroxy-β-methylbutyric Acid (HMB)