Lignocellulosic biomass is the most abundant renewable resource in nature. However, low-temperature fuel cell technology has not yet been able to use it as an efficient fuel. In this research study, a low temperature direct biomass fuel cell, which did not use enzymes or microorganisms as catalysts, was demonstrated using barley straw as the fuel, methyl violet as an electron mediator, nickel foam as an anode, and carbon cloth supported 0.15mg/cm2 of 10wt% Pt/C doping graphene as a cathode. The effects of the particle sizes of the raw materials on the performance of the fuel cell were examined using 3% (w/v) different particle sizes of barley straw powder in 5M NaOH and 0.02M MV solutions. It was found that the smaller the particle size of barley straw powder was, the higher the electrochemical performance of the fuel cell would be. In addition, this study also examined the effects of several dissolution systems on the electrochemical performance of the fuel cell, including a 10wt% LiCl/DMAC, 5M NaOH, 7wt% NaOH/12wt% Urea, and 65wt% ZnCl2. It was found that the fuel cell in the 65wt% ZnCl2 solvent had the best electrochemical performance, and when the MV concentration increased, the electrochemical performance of the fuel cell also increased accordingly. The maximum power density of the fuel cell reached to 0.3mW/cm2 when the concentration of MV increased to 0.08M, which was nearly 40 times higher than the microbial fuel cells fed with cellulose. Meanwhile, the open circuit voltage was 541mV, and the limit current density was 0.3mA/cm2. This allowed the raw materials of the fuel cell to take a further step toward a more complex lignocellulosic biomass.