. To date, no pharmacokinetic studies of BCAAs in vivo useful for the assessment
of clinical effect following daily intake has been reported. Thus in this study, an HPLC–MS/MS method
for simultaneous determination of Leu, Ile and Val in Beagle dog plasma using homoarginine as the
internal standard was developed and validated in terms of specificity, linearity, precision, accuracy, and
stability. This assay method was then applied to a pharmacokinetic study of BCAAs in dogs following oral
administration of 0.25 g/kg and 0.50 g/kg BCAAs. The HPLC–MS/MS method was found to be sensitive and
reproducible for quantification of BCAAs in dog plasma and successfully applied to the pharmacokinetic
study. All these BCAAs were well absorbed with a substantial increase in the plasma concentration after
a baseline modification. No statistical significance was identified in different gender group and no drug
accumulation was observed following multiple doses.