A series of batch assays have been conducted to investigate the optimal factors that can be adopted to improve the anaerobic digestion (AD) performance of Phragmites australis and increase biogas production. The assays were carried out using 125 mL microcosm reactors with a working volume of 80 mL and incubated at mesophilic conditions (37 ± 1ºC). The effect of particle size (10, 5, 2, and < 1 mm) and alkaline pre-treatment of P. australis using various concentrations of sodium hydroxide (0.5, 1, 2, and 4%) on biogas production was examined. Furthermore, the best pre-treatment incubation time (12, 24, 48, 72, 96, and 120 h) and the optimal inoculum to substrate ratio (ISR: 4:1, 2:1, 1:1, 1:2 and 1:4) were also assessed. The results revealed that the highest biogas production from P. australis was achieved at particle size < 1 mm (27.97 ± 0.07 and 16.67 ± 0.09 mL/g VS added, for pre-treated and untreated P. australis respectively); 2% and 4% NaOH concentration for pre-treatment (70.01 ± 3.75 and 76.14 ± 2.62 mL/g VS added, respectively); pre-treatment incubation time of 72, 96, and 120 h (71.18 ± 1.79, 72.46 ± 1.08, and 73.78 ± 1.87 mL/g VS added, respectively); and ISR of 1:2 for pre-treated P. australis (78.21 ± 0.36 mL/g VS added) and ISR 1:4 for untreated P. australis (28.93 ± 1.55 mL/g VS added). Determining optimal parameters in this work would guide further development of process configurations, such as continuous AD systems.