Determining the Optimal Conditions for Collagenase Production by a Local Isolate of Entrobacter Cloacae Using Local Organic Waste

Background: Collagenases are proteolytic enzymes with applications in leather processing, food industries, and biomedicine. Utilizing locally sourced organic wastes as fermentation substrates offers a cost-effective and sustainable method for enzyme production. Aim: To identify the optimal physicochemical and nutritional conditions for maximal collagenase production by a native Enterobacter cloacae isolate using organic waste substrates. Methodology: Forty-eight bacterial isolates from soil, animal, and poultry wastes were screened on gelatin and collagen media to select six active strains, then narrowed to three based on performance on various organic wastes. The two highest producers were identified via cultural, microscopic, biochemical tests and the Compact VITEK2 system. One-factor-at-a-time experiments optimized pH (5.0–10.0), temperature (25–45 °C), substrate type (e.g., chicken foot waste), inoculum size (0.5–2.0 ml/100 ml), inoculum age (18–30 h), nitrogen source (peptone, yeast extract, ammonium salts), incubation mode (static vs. shaking), and incubation time (24–72 h). Collagenase activity was quantified in U/ml. Results: Both isolates were confirmed as E. cloacae. Maximum enzyme yield occurred at pH 8.0 and 35 °C using chicken foot waste. The optimal inoculum was 1 ml per 100 ml medium (1.1×10⁹ cells/ml, 24 h age). Peptone under static incubation yielded the highest activity, reaching 20.232 U/ml after 48 h. Conclusion: A local E. cloacae isolate efficiently produces collagenase on chicken foot waste under optimized conditions, demonstrating a scalable, low-cost, and eco-friendly strategy for industrial enzyme production.