The objective of this study was to evaluate the effects of osmoprotectants on the high salinity anaerobic digestion in engineering and microbial aspects with a special focus on syntrophic microbial populations. The experimental results indicated that glycine betaine (GB) exerted some differentiated influences on anaerobic syntrophic microbial populations at high salinity conditions. It was found that glycine betaine was beneficial for acetic acid producers, aceticlastic methanogens, n-butyrate utilizers, and comparatively not so effective for the propionate producers and utilizers. Moreover, although the addition of GB in advance could not prevent the imbalance of anaerobic process caused by the high salt shock, it was helpful for the anaerobes to recover the methanogenic activity following salt inhibition. Repeated feeding experimental results indicated that the osmoprotectant played a prolonged positive role on high saline anaerobic digestion, which was proved to be closely related to the extracellular osmoprotectants. These findings could provide a deep insight on the microbiological aspect on osmoprotectants enhancing anaerobic digestion, and some operating strategies could be developed based on it. (C) 2015 Elsevier B.V. All rights reserved.