A single-run reverse phase-high performance liquid chromatography method for the quantification of humulinones, α-acids, iso-α-acids and reduced iso-α-acids (where present) in commercial beer samples is presented. The method utilizes a binary solvent system consisting of (A) 1% v/v acetic acid and (B) 0.1% v/v orthophosphoric acid in acetonitrile. Separation was achieved on a Purospher® star RP-18 column (250 × 46 mm, 3 µm) with a flow rate of 0.5 mL/min. The compounds of interest eluted within 32 min. The method was fully validated according to International Conference on Harmonization guidelines and subsequently applied to monitor degradation of hop acids in a storage trial where four lager beers were aged at 28 and 38 °C for 70 and 60 days, respectively. Results confirmed the widely reported degradation through storage of trans-iso-α-acids whilst demonstrating that the HLPC method was sufficiently sensitive to monitor and model this degradation. One beer exhibited a significantly lower ( P < 0.05) rate of trans-iso-α-acid degradation than the other conventionally hopped beers in the study, which might have been linked to its higher pH (4.71 vs 4.36). The relative stability of reduced iso-α-acids during ageing was also confirmed. [ABSTRACT FROM AUTHOR]