Batch experiments were conducted to evaluate the impact of inoculum sources, inoculum to substrate (IS) ratio and storage conditions on the potential and production rate of methane (CH4) from different substrates: wheat straw, whole crop maize, cattle manure, grass and cellulose. The results of the test with four inocula and four substrates indicated that inoculum source could have a significant impact on both CH4 potential (BMP) and the kinetics parameters of different substrates. The two inocula showing the highest BMP and production rates in each period were those coming from a feeding with more than 70% of animal manure under thermophilic conditions. The impact of the IS ratio in the range 0.25-2.5, in terms of g volatile solids (VS) substrate/g VS inoculum, depended on substrate type. Maize silage was more affected to changes in the IS ratio than wheat straw. The optimal IS ratio range for maize was 1.0-1.5, however, a wider IS range can be used in wheat straw (0.5-2.5). The impact of freezing and drying depended on biomass type. Freezing, drying and ensiling of grass increased the CH4 yield compared to fresh grass. Drying of maize had no impact while freezing reduced the CH4 potential. Drying and freezing had no impact on straw. (C) 2015 Elsevier Ltd. All rights reserved.