In this study alternative agricultural substrates are investigated as potential substitutes of maize for biogas production in the region of South Europe. Crop silages of cardoon, maize, milk thistle and sorghum as well as bedding straw from cattle farm were examined in the anaerobic co-digestion procedure with cattle manure. Milk thistle crop was further investigated in a naturally sun dried form and the effect of mechanical, thermal and thermo-chemical pretreatments on fiber composition and methane yield was evaluated. Pretreatment with NaOH increase the solubilization by 77.7%. The co-digestion experiment was carried out in 28 batch reactors at 37 degrees C. The highest methane yields of 308, 271 and 267 L CH4 kg (1) of volatile solids were obtained by co-digestion of cattle manure with cardoon silage, thermo-chemical pretreated milk thistle stalks with NaOH and maize silage, respectively. Furthermore, co-digestion of bedding straw and cattle manure had similar methane yield with maize silage. (C) 2014 Elsevier Ltd. All rights reserved.