BACKGROUND:The current study demonstrated the possibility of statistical design tools combination with computational tools for optimization of fermentation conditions for enhanced fibrinolytic protease production.METHODS:The effects of using different carbon and nitrogen sources for protease production by Streptomyces radiopugnans_VITSD8 were examined by a full factorial design method.The incubation time,temperature,pH of the medium,and RPM were assessed by the predictable one factor at a time (OFAT) method.Optimization was carried out using starch and oat meal as carbon source,nitrogen source as peptic and malt extract using Fractional Factorial Design (FFD).The analysis was further continued for medium volume,temperature,initial medium pH,inoculum concentration,high determination co-efficient as (R’-0.965),and lower determination co-efficient of variation (CV-8.19％),which defines a reliable and accurate experimental value.RESULTS:Analysis of variance by the fixed slope effect by temperature and starch;temperature and L-aspargine,temperature and oat meal,temperature and peptic extracts,temperature and pH,temperature and duration of incubation were more vital for protease production at an interactive level.Response surface plots revealed that temperature,starch,and peptic extracts affix critical concing in temperature.Programming estimated a 28％ increase in protease production.Incubation temperature and medium volume portrayed extreme impact among all factor.Starch,peptic and temperature play an important regulatory role in protease production.Optimium temperature for protease production was 33℃.The ratio of carbon and nitrogen sources and pH were the major regulatory factors in protease production by Streptomyces radiopugnans_VITSD8.It demonstrated a 4％ noteworthy change in condition.CONCLUSION:Among all the selected parameters,temperature was the most intuitive factor,demonstrating a notable connection with the type of media and pH,while inoculum fixation had a direct impact on protein production.