Two methods for the determination of alpha and iso-alpha bitter acids composition by HPLC, are validated in the Food Safety and Quality Testing Laboratory (LICSA) of USAMV Cluj-Napoca in order to asses the capability of the laboratory to assure quality results. In the validation study, the following quality parameters were determined: specificity, linearity, precision and accuracy of the method. The method is selective for: co-isohumulone, isohumulone, Ad-isohumulone, Cohumulone and Colupulone and isn’t selective for the separation of normal-humulone from ad-humulone, which appear together as a single peak. The same thing goes for the normal-lupulone and ad-lupulone peak.