A start-up study for biohydrogen production from palm oil mill effluent (POME) is carried out in a pilot-scale up-flow anaerobic sludge blanket fixed-film reactor (UASFF). A substrate with a chemical oxygen demand (COD) of 30 g L −1 is used, starting with molasses solution for 30 days and followed by a 10% v/v increment of POME/molasses ratio. At 100% POME, a hydrogen content of 80%, hydrogen production rate of 36 L H 2 per day, and maximum COD removal of 48.7% are achieved. Bio-kinetic coefficients of Monod, first-order, Grau second-order, and Stover-Kincannon kinetic models are calculated to describe the performance of the system. The steady-state data with 100% POME shows that Monod and Stover-Kincannon models with bio-kinetic coefficients of half-velocity constant (K s) of 6000 mg COD L −1 , microbial decay rate (K d) of 0.0015 per day, growth yield constant (Y) of 0.786 mg volatile suspended solids (VSS)/mg COD, specific biomass growth rate (max) of 0.568 per day, and substrate consumption rate of (U max) 3.98 g/L day could be considered as superior models with correlation coefficients (R 2) of 0.918 and 0.989, respectively, compared to first-order and Grau's second-order models with coefficients of K 1 1.08 per day, R 2 0.739, and K 2s 1.69 per day, a = 7.0 per day, b = 0.847.