Papain is a common protease exhibiting antioxidant and anti-inflammatory activities. Papain can be extracted from seeds, leaves or latex of papaya (Carica papaya L.). It is involved in accelerating wound healing process. Hydrogels are widely used in biomedical applications such as tissue engineering, drug delivery system, and wound healing. Cross-linked hydrogels containing papain are more aqueous, durable and promising devices to minimize recovering time of lesions. Ingredients of HPMC hydrogels were investigated and immobilized with papain. The hydrogel thickness, moisture content, Fourier transform infrared, drug content and in vitro papain release were characterized. The formula of 0.5 g papain, 5 g HPMC, 5 g CA, 7.5 g glycerin, 5 g P407, 0.3 g nipagin M and EtOH quantum satis (q.s) 100 g resulted in hydrogels 0.8 mm of thickness, 3.83 % of moisture. FTIR analysis confirmed papain incorporation into prepared hydrogels. The FE-SEM examination revealed a good distribution of papain on the hydrogel and size of papain was (5-10) µm. In addition, from 2.6652 % to 5.1021 % of papain was released from the hydrogel within 60 minutes