The optimal environmental and nutritional conditions for chitosan production by two strains of R.nigricans (R6 and R22) were studied. Also, molasses and corn steep liquor were used for maximization chitosan production. Results showed that the highest records of chitosan production by both strains of R.nigricans were observed when sucrose (2.5%) and peptone were used as a sole carbon and nitrogen sources, respectively. Concerning the effect of incubation temperatures ,the results emphasized that increasing the incubation temperature led to gradual increase in biomass and chitosan yield up to 28 ˚C for both strains of R. nigricans. Moreover, results indicated that the highest values of chitosan production were obtained when R.nigricans was incubated at pH 5 for 7 days and decreased thereafter. Results showed that chitosan production by the two Rhizopus strains increased with the increasing of corn steep liquor to reach their maximum at 150 g/l. Also, the two Rhizopus strains produced higher amounts of chitosan when cane molasses was used as a carbon source rather than beet molasses .Moreover, maximum values of biomass and chitosan were observed when mixture of cane and beet molasses was used in the fermentation medium as a sole carbon source.