A novel alkaline keratinase from Bacillus subtilis DP1 with potential utility in cosmetic formulation

• The purified enzyme was active between pH 8.0 and 12.0 with an optimum pH 10.0.
• A CD spectrum implies that purified keratinase is β-pleated sheet rich protein.
• Compability studies of constituents used in formulation by FTIR studies.
• Preparation of biological hair removal depilatory using purified keratinase from Bacillus subtilis DP1.
• Prepared formulation efficiently removes hair visualized by zone of clearance.

The Bacillus subtilis DP1 was isolated from poultry farm soil at Anand district, India. The highest enzyme production (379.65 U/ml) was obtained at pH 10.0, a temperature of 37 °C and a growth period of 72 h. The extracellular keratinase was purified by gel filtration chromatography with 27.98 purification fold. Purity was also confirmed by High-Performance Liquid Chromatography (HPLC) analysis, where a major peak having retention time of 2.5 min was obtained on C18 column using photo diode array detector. Purified keratinase was stable in a broad range of pH (8–12) and temperature (20–50 °C) with optimum at pH 10.0 and 37 °C. The metallic ions, Ca2+ and Mg2+ enhance keratinase activity. Secondary structure from Circular Dichroism (CD) spectra implies that purified keratinase is largely β-pleated sheet rich protein. For preparation of dehairing cream formulation, compatibility studies of excipients were carried out. Fourier transform infrared spectroscopy (FTIR) spectra of sodium stearate, calcium carbonate and sodium lauryl sulphate shows no reactivity of functional groups and hence mixture was compatible for formulation of keratinase dehairing cream. Prepared biological depilatory was able to remove hair more efficiently compared to marketed formulations.