This study evaluated the impacts of steam explosion pretreatment of lignin-rich macrophyte (Potamogeton maackianus) on semi-continuous anaerobic digestion. The pretreatment accelerated hydrolysis efficiency from 9% to 19% (untreated) to 38%–48% (pretreated), which considerably enhanced the methane yield from 108 ± 31 to 200 ± 36 mL g-VS−1. As a negative impact, the soluble COD concentration in the effluent in the pretreatment condition was high (10,000–12,000 mg L−1) due to the accumulation of steam explosion by-products (soluble lignin and phenolic compounds). This high concentration of by-products in the digestate can destabilise subsequent biological treatment processes; therefore, their removal from the digestate will be necessary. Although the concentration of inhibitory phenolic compounds in the pretreated P. maackianus was high, it was continuously removed (38 ± 10%) in the reactor, which ensured process stability. As a result, this study demonstrated for the first time the effectiveness of direct anaerobic digestion treatment of steam-exploded lignocellulose biomass over a long-term continuous operation. By summarising the results of previous studies and our study, we found that the improvement in CH4 production (y, %) by steam explosion pretreatment is regulated by lignin content in the untreated biomass (x, %-TS) and expressed by the equation, y = 18.1 e0.071x (p = 0.0001). This empirical formula indicates that the effectiveness of steam explosion pretreatment increases exponentially with an increase in the lignin content.