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Olive oil production is a seasonal agricultural activity taking place around the Mediterranean Sea with high financial importance and significant environmental footprint. Unfortunately, olive milling is generating olive mill wastewater (OMWW), which is an effluent with high carbon-to-nitrogen ratio and low pH. The aim of this work is to assess the application of different high nitrogen substrates for overcoming the problem of nitrogen deficiency of the mono-digestion of olive mill wastewater. The nitrogen-rich substrates are fish meal, bone, and meat meal (BMM), blood meal (BM), feather meal (FEM), mixtures of FEM and BM, soybean meal, and corn-gluten meal, which are mixed with the olive mill wastewater in order to attain C/N ¼ 20:1. The substrates are evaluated in batch and tank reactors. According to the results, the type of protein is an insignificant process parameter and the process can be developed without inhibition phenomena to be presented with any of the co-substrates assessed. The maximum bio-methane potential registered for the mixture of olive mill wastewater and bone, and meat meal with 449 mL CH 4 g À1 volatile solids added (VS) with an organic loading rate (OLR) of 3.6 kg V À1 s À1 m À3 per day. The overall process presented remarkable tolerance during the stepwise increase of OLR from 3.6 to 5.8 kg V À1 s À1 m À3 per day with the efficiency of the process to be reduced by 15% for all co-digestion mixtures while the volumetric production increased from 1.5 to 2.1 m 3 CH 4 m À3 per day.