A dual step statistical approach including Plackett-Burman combined with response surface methodology was used to enhance xylanase production by Bacillus sp. Using the mentioned approach, three fermentation variables (xylan, ammonium sulphate and pH), were showed significant effect on xylanase production out of all the variables studied for fermentation. The enzyme was found to be optimally active at pH 8.5 and 45 °C. A high enzyme titre of 95.46 U/ml was attained at ammonium sulphate concentration of 0.3% and pH (8.5). The enzyme may be used for hydrolysis of xylan and can be used in the bleaching and food processing industry.