A common approach to increasing biogas production is enhancing the hydrolysis of wheat straw by removing the lignin-using chemicals. However, partial organic matter will also dissolve into chemical solutions during pretreatment, which leads to the loss of organic matter for the anaerobic digestion process. The enzymatic hydrolysis of wheat straw and loss of total organic carbon increased with NaOH concentration and pretreatment time. The results suggest that when evaluating a pretreatment method, not just the hydrolysis but the loss of organic matter should be taken into consideration.