Heat Resistant Proteases Isolated from Medicinally Significant Spices: Partial Purification and Biochemical Characterization

Proteases or proteinases are essential constituents for all the existing live forms. They act as important industrial enzymes occupying about 60% of total enzyme market. In the present study proteases extracted from nine medicinally important spices; Carum copticum, Syzygium aromaticum, Cuminum cyminum, Nigella sativa, Cinnamomum verum, Foeniculum vulgare, Zingiber officinale, Cinnamomum tamala and Curcuma longa; used in our food and as household medicines on regular basis, have been investigated. Amongst these spices, the specific activity of the isolated protease enzyme was found to be significantly high in Nigella sativa (204 units/mg) and Curcuma longa (124 units/mg) and therefore, their extract was further partially purified and biochemically characterized. The crude extract of the two spices on being subjected to salt precipitation using (NH4)2SO4 as neutral salt, yielded three fractions at 0-30%, 30-60% and 60-90% saturation level. The specific activity of protease enzyme was found to be highest in 0-30% fraction, obtained from both the sources. The values of specific activity and purification-fold of enzyme from Nigella sativa and Curcuma longa were found to be 409 U/mg, 590 U/mg and 2.0-, 4.8-, respectively. The enzyme showed maximum activity at pH and temperature conditions of 5.0 and 40°C, respectively, at 20 min of incubation time, in both the cases. The pH stability values of protease from Nigella sativa ranged from 5.0 to 9.0 whereas in Curcuma longa values were from 4.0 to 8.0. The enzyme from Nigella sativa and Curcuma longa exhibited resistance against heat treatment upto 600C and 500C, respectively, increasing the industrial feasibility. The present work hereby, indicates that Nigella sativa and Curcuma longa may serve as good source of thermostable protease; an enzyme of great relevance in various chemical and bio-industries along with pharma sector.