To give an insight into the kinetics of microbial oil fermentation with lignocellulosic hydrolysate as substrate, lipid production by Trichosporon fermentans in rice straw hydrolysate was investigated in a 5 L fermentor. For comparison, fermentation in its simulated medium was also performed to evaluate the effect of inhibitors present in the rice straw hydrolysate on the cell growth and lipid accumulation of T. fermentans. The optimum fermentation time, maximum biomass, and lipid content of T. fermentans in the rice straw hydrolysate were 10.5 days, 28.6 g/L, and 43.9%, respectively, while the corresponding values in the simulated medium were 8.5 days, 27.0 g/L, and 65.0%, respectively, indicating that the inhibitors in the rice straw hydrolysate did show some inhibition on the cell growth and lipid accumulation of T. fermentans. Kinetic models of lipid fermentation with T. fermentans showed the maximum specific growth rate and the correlation coefficient of lipid synthesis to cell growth in the rice straw hydrolysate were 0.40 and 0.51, respectively, which are much lower than those in the simulated medium (0.58 and 0.73). To better understand the influential mechanism of inhibitors in the rice straw hydrolysate on the growth and lipid accumulation of T. fermentans, the physiological and biochemical changes of cells in the fermentation were further investigated. The reduced activities of adenosine triphosphate (ATP) citrate lyase, malic enzyme, and xylose reductase, and the elongation of cells at the beginning of fermentation, partly account for the inhibitory effect of inhibitors in the rice straw hydrolysate.