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Though deemed a prospective method, the bioconversion of organic waste to biohydrogen via dark fermentation (DF) has multiple drawbacks and limitations. Technological difficulties of hydrogen fermentation may, in part, be eliminated by making DF a viable method for biohythane production. Aerobic granular sludge (AGS) is a little-known organic waste spurring a growing interest in the municipal sector; its characteristics indicate the feasibility of its use as a substrate for biohydrogen production. The major goal of the present study was to determine the effect of AGS pretreatment with solidified carbon dioxide (SCO2) on the yield of H2 (biohythane) production during anaerobic digestion (AD). It was found that an increasing dose of SCO2 caused an increase in concentrations of COD, N-NH4+, and P-PO43− in the supernatant at the SCO2/AGS volume ratios from 0 to 0.3. The AGS pretreatment at SCO2/AGS ratios within the range of 0.1–0.3 was shown to enable the production of biogas with over 8% H2 (biohythane) content. The highest yield of biohythane production, reaching 481 ± 23 cm3/gVS, was obtained at the SCO2/AGS ratio of 0.3. This variant produced 79.0 ± 6% CH4 and 8.9 ± 2% H2. The higher SCO2 doses applied caused a significant decrease in the pH value of AGS, modifying the anaerobic bacterial community to the extent that diminished anaerobic digestion performance. [ABSTRACT FROM AUTHOR]