The present investigation aims to delignify the rice straw in two different directions. In the first route, the rice straw is delignified with acidic sodium chlorite prior to sodium hydroxide while only sodium hydroxide was chosen as the delignifying agent in the next route. The effectiveness of the first stage delignification process has been studied for the comparative assessment with the second stage. Response surface methodology based on CCD (Central Composite Design) was used for optimization of process variables in both the delignification stages and to determine the optimum delignification condition. No investigation was performed so far to explore the delignification process of rice straw in successive two-stage using low quantity of chemical reagents viz., sodium chlorite, sodium hydroxide and low energy input employing statistical experimental design.