Skip to main content
Log in

Production of milk clotting protease by a local isolate of Mucor circinelloides under SSF using agro-industrial wastes

  • Articles
  • Published:
Biotechnology and Bioprocess Engineering Aims and scope Submit manuscript

Abstract

Agro-industrial residues, a cheap source of energy have high potential in the area of fermentation for the production of enzymes. Twenty agro-industrial residues were evaluated to check the possibility of potential utilization of substrates in SSF for milk clotting enzyme protease production by Mucor circinelloides. In this study, dhal husk holds the greatest promise for cost effective production of the milk clotting enzyme. The dhal husk supported maximum milk clotting protease production, and yield was improved with the supplementation of sucrose and yeast extract as carbon and nitrogen source, respectively. Among all the physico-chemical parameters tested, the best results were obtained in a medium having moisture content of 20% at pH 7.0, when inoculated with 30% of spore suspension and incubated at 30°C for 5 days. The activity was increased further on addition of Ca2+, Cu2+, and Mg2+ ions. The purified milk-clotting protease obtained from M. circinelloides was successfully applied and compared with commercial rennet in the manufacture of a cheddar cheese.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Rao, M. B., A. M. Tanksale, M. S. Ghatge, and V. V. Deshpande (1998) Molecular and biotechnological aspects of microbial proteases. Microbiol. Mol. Biol. Rev. 62: 597–635.

    CAS  Google Scholar 

  2. Manachini, P. L. and M. G. Fortina (1998) Production in sea-water of thermostable alkaline protease by a halotolerant strain of Bacillus licheniformis. Biotechnol. Lett. 20: 565–568.

    Article  CAS  Google Scholar 

  3. Moreira, K. A., T. S. Porto, M. F. S. Teixeira, A. L. F. Porto, and J. L. Lima Filho (2003) New alkaline protease from Nocardiopsis sp.: partial purification and characterization. Process Biochem. 39: 67–72.

    Article  CAS  Google Scholar 

  4. Tunga, R., B. Shrivastava, and R. Banerjee (2003) Purification and characterization of protease from solid state culture of Aspergillus parasiticus. Process Biochem. 38: 1553–1558.

    Article  CAS  Google Scholar 

  5. Jarai, G., H. van den Hombergh, and F. B. Buxton (1994) Cloning and characterization of the pepE gene of Aspergillus niger encoding a new aspartic protease and regulation of pepE and pepC. Gene 145: 171–178.

    Article  CAS  Google Scholar 

  6. Young, J. W., A. Wadeson, D. J. Glover, R. V. Quincey, M. J. Butlin, and E. A. Kamei (1996) The extracellular acid protease of Yarrowia lipolytica: sequence and pH-regulated transcription. Microbiol. 142: 2913–2921.

    Article  CAS  Google Scholar 

  7. vankuyk, P. A., B. F. Cheetham, and M. E. Kate (2000) Analysis of two Aspergillus nidulans genes encoding extracellular proteases. Fungal Genet. Biol. 29: 201–210.

    Article  CAS  Google Scholar 

  8. Sousa, M. J. and F. X. Malcata (2002) Advances in the role of a plant coagulant (Cynara cardunculus) in vitro and during ripening of cheeses from several milk species. Lait 82: 151–170.

    Article  CAS  Google Scholar 

  9. Tsugo, T., U. Yoshino, K. Taniguchi, A. Ozawa, Y. Miki, S. Iwasaki, and K. Arima (1964) Cheese-making by using the milk-clotting enzyme of Mucor pusillus. Lindt. 1. Rennet properties of the enzyme. Jap. J. Zootech. Sci. 35: 221–228.

    Google Scholar 

  10. Abdel-Fattah, A. F., A. S. Ismail, and S. A. El Aassar (1984) Production of rennin like enzyme by Absidia cylindrospora. Agric. Wastes. 11: 125–131.

    Article  CAS  Google Scholar 

  11. Ismail, A. M. S., S. A. El Aassar, and A. F. Abdel-Fattah (1984) Production of milk clotting and proteolytic enzymes by fungi. Agric. Wastes. 10: 95–102.

    Article  CAS  Google Scholar 

  12. Hashem, A. M. (2000) Purification and properties of a milk-clotting enzyme produced by Penicillium oxalicum. Bioresour. Technol. 75: 219–222.

    Article  CAS  Google Scholar 

  13. Cavalcanti, M. T. H., M. F. S. Teixeira, J. L. Lima Filho, and A. L. F. Porto (2004) Partial purification of new milk-clotting enzyme produced by Nocardiopsis sp. Bioresour. Technol. 93: 29–35.

    Article  CAS  Google Scholar 

  14. Tubesha, Z. A. and K. S. Al Delaimy (2003) Renin like milk coagulant enzyme produced by a local isolate of Mucor. Int. J. Dairy Technol. 56: 237–341.

    Article  CAS  Google Scholar 

  15. Couto, S. R. and M. A. Sanroman (2006) Application of solid-state fermentation to food industry-a review. J. Food Eng. 76: 291–302.

    Article  CAS  Google Scholar 

  16. Holker, U. and J. Lenz (2005) Solid-state fermentation — are there any biotechnological advantages? Cur. Opin. Microbiol. 8: 301–306.

    Article  CAS  Google Scholar 

  17. Krishna, C. (2005) Solid-state fermentation systems-an overview. Cri. Rev. Biotechnol. 25: 1–30.

    Article  CAS  Google Scholar 

  18. Nagampoothiri, K. M. and A. Pandey (1996) Solid state fermentation for L-glutamic acid production using Brevibacterium sp. Biotechnol. Lett. 18: 199–204.

    Article  Google Scholar 

  19. Roukas, T. (1999) Citric acid production from pod by solid-state fermentation. Enz. Microbiol. Biotechnol. 24: 54–59.

    Article  CAS  Google Scholar 

  20. Vanderberghe, L. P. S., C. R. Soccol, A. Pandey, and J. M. Lebeault (2000) Citric acid production by Aspergillus niger in solid state fermentation. Bioresour. Technol. 74: 175–178.

    Article  Google Scholar 

  21. Pandey, A., P. Selvakumar, C. R. Soccol, and P. Nigam (1999) Solid state fermentation for the production of industrial enzymes. Cur. Sci. 77: 149–162.

    CAS  Google Scholar 

  22. Selvakumar, P. and A. Pandey (1999) Solid state fermentation for the synthesis of inulinase from Staphylococcus sp. and Kluyveromyces marxianus. Process Biochem. 34: 851–855.

    Article  CAS  Google Scholar 

  23. Balakrishna, K. and A. Pandey (1996) Production of biologically active secondary metabolites in solid state fermentation. J. Sci. Ind. Res. 55: 365–372.

    Google Scholar 

  24. Ohno, A., T. Ano, and M. Shoda (1996) Use of soybean curd residue, okara, for the solid state substrate in the production of lipopeptide antibiotic, iturin A by Bacillus subtilis NB 22. Process Biochem. 31: 801–806.

    Article  CAS  Google Scholar 

  25. Yang, S. S. and W. J. Swei (1996) Oxytetracycline production by Streptomyces rimosus in solid state fermentation of corn-cob. W. J. Microbiol. Biotechnol. 12: 43–46.

    Article  CAS  Google Scholar 

  26. Kota, K. P. and P. Sridhar (1999) Solid state cultivation of Streptomyces clavuligerus for cephamycin C production. Process Biochem. 34: 325–328.

    Article  CAS  Google Scholar 

  27. Sekhar, C. and K. Balaraman (1998) Optimization studies on the production of cyclosporin A by solid state fermentation. Bioprocess Eng. 18: 293–296.

    Article  Google Scholar 

  28. Ramana Murthy, M. V., E. V. S. Mohan, and A. K. Sadhukhan (1999) Cyclosporin A production by Tolypocladium inflatum using solid state fermentation. Process Biochem. 34: 269–280.

    Article  Google Scholar 

  29. Tunga, R., R. Banarjee, and B. C. Bhattacharyya (1998) Optimizing some factors affecting protease production under solid state fermentation. Bioprocess Eng. 19: 187–190.

    Article  CAS  Google Scholar 

  30. Elibol, M. and A. R. Moreira (2005) Optimizing some factors affecting alkaline protease production by a marine bacterium Teredinobacter turnirae under solid substrate fermentation. Process Biochem. 40: 1951–1956.

    Article  CAS  Google Scholar 

  31. Prakasham, R. S., C. H. Subba Rao, and P. N. Sarma (2006) Green gram husk-an inexpensive substrate for alkaline protease production by Bacillus sp. in solid-state fermentation. Bioresour. Technol. 97: 1449–1454.

    Article  CAS  Google Scholar 

  32. Palaniswamy, M., B. V. Pradeep, R. Sathya, and J. Angayarkanni (2008) Isolation, identification, and screening of potential xylanolytic enzyme from litter degrading fungi. Afri. J. Biotech. 7: 1978–1982.

    CAS  Google Scholar 

  33. Arima, K., J. Yu, and S. Iwasaki (1970) Methods in Enzymology. pp. 446–459. In: G. Perlmann, L. Lorand (eds.). Milk clotting enzyme from Mucor pusillus var. Lindt. Academy Press, NY, USA.

    Google Scholar 

  34. Pandey, A., C. R. Soccol, P. Nigam, D. Brand, R. Mohan, and S. Roussos (2000) Biotechnological potential of coffee pulp and coffee husk for bioprocesses. Biochem Eng. J. 6: 153–162.

    Article  CAS  Google Scholar 

  35. Shata, H. M. A. (2005) Extraction of Milk- clotting enzyme produced by solid state fermentation of Aspergillus oryzae. Polish J. Microbiol. 54: 241–247.

    CAS  Google Scholar 

  36. Preetha, S. and R. Boopathy (1994) Influence of culture conditions on the production of milk clotting enzyme from Rhizomucor. W. J. Microbiol. Biotech. 10: 527–530.

    Article  CAS  Google Scholar 

  37. Sannabhadti, S. S. and R. A. Srinivasan (1977) Milk clotting enzymes from Abisidia ramosa. Part 1. Factors influencing production. Ind. J. Dairy Sci. 30: 331–335.

    CAS  Google Scholar 

  38. Thakur, M. S., N. G. Karanth, and N. Krishna (1990) Production of fungal rennet by Mucor meiehei using solid state fermentation. Appl. Microbiol. Biotechnol. 32: 409–413.

    Article  CAS  Google Scholar 

  39. Nehra, K. S., S. Dhillon, K. Chaudhary, and S. Randir (2002) Production of alkaline protease by Aspergillus sp. under submerged and solid substrate fermentation. Ind. J. Microbiol. 42: 43–47.

    Google Scholar 

  40. Moon, S. H. and S. J. Parulekar (1991) A parametric study of protease production in batch and fed batch cultures of Bacillus firmus. Biotechnol. Bioeng. 37: 467–483.

    Article  CAS  Google Scholar 

  41. Yu, P. J. and C. C. Chou (2005) Factors affecting the growth and production of milk clotting enzyme by Amylomyces rouxii in rice liquid medium. Food Technol. Biotechnol. 43: 283–288.

    CAS  Google Scholar 

  42. D’souza, T. M. and L. Pereira (1982) Production and immobilization of a bacterial milk clotting enzyme. J. Dairy Sci. 65: 2074–2081.

    Article  Google Scholar 

  43. Ghareib, M., H. S. Hamdy, and A. A. Khalil (2001) Production of intracellular milk clotting enzyme in submerged cultures of Fusarium subglutinans. Acta Microbiol. Polon. 50: 139–147.

    CAS  Google Scholar 

  44. Su, Y. C. and W. P. Chen (1970) Studies on milk clotting enzymes from microorganisms. Part 1. Screening tests and production of the enzymes. J. Chin. Agric. Chem. Soc. 8: 73–83.

    CAS  Google Scholar 

  45. Hung, Y. C. and C. C. Chou (1997) Growth and milk clotting enzyme production in submerged culture of Amylomyces rouxii. J. Chin. Agric. Chem. Soc. 35: 422–432.

    CAS  Google Scholar 

  46. Nigam, J. M., K. R. Pillai, and J. N. Baruah (1981) Effect of carbon and nitrogen sources on neutral proteinase production by Pseudomonas aeruginosa. Folia Microbiol. 26: 358–363.

    Article  CAS  Google Scholar 

  47. Zandrazil, F. and H. Brunert (1981) Investigation of physical parameters important for solid-state fermentation of straw by white rot fungi. Eur. J. Appl. Microbiol. Biotechnol. 11: 183–188.

    Article  Google Scholar 

  48. Lekha, P. K. and B. K. Lonsane (1994) Comparative titres, location, and properties of tannin acyl hydrolase produced by Aspergillus niger PKL104 in solid state, liquid surface, and submerged fermentation. Process Biochem. 29: 497–503.

    Article  CAS  Google Scholar 

  49. Chu, I. M., C. Lee, and T. S. Li (1992) Production and degradation of alkaline protease in batch cultures of Bacillus subtilis ATCC 14416. Enz. Microbiol. Technol. 14: 755–761.

    Article  CAS  Google Scholar 

  50. Gupta, R., Q. K. Beg, S. Khan, and B. Chauhan (2002) An overview on fermentation, downstream processing and properties of microbial proteases. Appl. Microbiol. Biotechnol. 60: 381–395.

    Article  CAS  Google Scholar 

  51. Hesseltine, C. W., M. Smith, and H. L. Wang (1976) Product of fungal spore as inocula for oriental fermented food. Dev. Ind. Microbiol. 17: 101–115.

    Google Scholar 

  52. Hashem, A. M. (1999) Optimization of milk-clotting enzyme productivity by Penicillium oxalicum. Bioresour. Technol. 70: 203–207.

    Article  CAS  Google Scholar 

  53. Abdel-Fattah, A. F. and S. A. Saleh (1979) Production and isolation of milk clotting enzyme from Aspergillus versicolor. Zbl. Bakt. II Abt. 134: 547–550.

    CAS  Google Scholar 

  54. Kobayashi, F., M. Yabuki, K. Hoshino, and M. Sakamoto (1975) Isolation and characterization of Trametes ostreiformis K-1, and purification and properties of milk clotting enzyme produced by fungus. J. Agric. Chem. Soc. Jpn. 49: 81–92.

    CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to M. Palaniswamy.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Sathya, R., Pradeep, B.V., Angayarkanni, J. et al. Production of milk clotting protease by a local isolate of Mucor circinelloides under SSF using agro-industrial wastes. Biotechnol Bioproc E 14, 788–794 (2009). https://doi.org/10.1007/s12257-008-0304-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12257-008-0304-0

Keywords

Navigation