Full Length Research Paper

Milk-clotting fungus enzymes production: A sustainable development approach based on whey recycling

Aïcha Benlounissi¹*, Aïcha Mechakra-Maza¹, Zoubida Gheribi³, Meriem Mahfouz², Loïc J. Blum⁴ and Christophe A. Marquette⁴

1Laboratory of Environmental Biology, Department of Biochemistry-Microbiology, Faculty of Natural Science and Life, University Mentouri, 25017 Constantine, Algeria.
2Laboratory of Mathematics, University Claude Bernard Lyon 1, Central Scool and INSA, Institut Camille Jordan UMR 5208, Bat Braconnier, Bureau 228, 21 Avenue Claude Bernard, Domaine de la Doua, 69366 Villeurbanne, France.
3Laboratory of research in applied mathematics and modelling, Campus Ahmed Hamani, Road Aïn El Bey 25017 Constantine, Algérie.
4Enzyme Engineering Team, Biomimetic Membranes and Supramolecular Assemblies, Institute of Chemistry, Molecular and Supramolecular Biochemistry, University Lyon 1 - CNRS 5246 ICBMS, Building CPE - 43, bd du 11 Novembre 1918 - 69622 Villeurbanne, Cedex, France.

*Corresponding author. E-mail: benlouni@hotmail.com.

Accepted 22 July, 2011

   Abstract

Five species of Aspergillus fungus: Aspergillus niger, Aspergillus flavus, Aspergillus awamori, Aspergillus tubingensis and Aspergillus tamarii, and one Penicillium: Penicillium pinophilum were isolated from Algerian soil and identified using both microscopic observation and ICT sequence identification. Their potential milk-clotting activity was demonstrated and attributed to an acidic protease activity. For the culture and enzyme production from the six selected fungi, an approach compatible with sustainable development was developed. A whey (cheese industry secondary product) based culture medium was designed and optimised for each species using first Plackett–Burman and then Box and Wilson statistical experiments. The studied factors were pH, stirring, lactose, yeast extract, peptone, CaCl2 and salts: MgSO4 and FeSO4. Once the selected factors were optimised, the whey based fermentation mediums were used for the production of acidic protease. All molds were shown to be able to produce acidic protease directly in their culture medium and to generate milk clotting within 15 min. The only exception was A. awamori, which exhibits a good acidic protease activity but no clotting capability.

Key words: Acidic protease, Aspergillus, experimental design, milk-clotting enzyme, optimization, Penicillium.