Production Of Microbial Rennet

Rennet production S.Shanmuga Priya BTF-07-023

Rennet • • Rennet is in use for cheese making at least since 2000 B.C. • An enzyme extracted from gastric mucous membrane of bovines • The production of countless different types of cheese use proteolytic enzyme, rennin the one most widely used

Cheese §Cottage cheese §Roquefort §Stilton §Blue cheese §Gorgonzola §Camembert §Brie §Swiss §Baby Swiss §Gouda §Edam

§Limburger §Traditional brick cheese §Cheddar §Colby §Sweet Brick §Muenster §Havarti §Mozzarella §Provolone §Parmesan §Romano

Alternatives source of rennet • Global increase in cheese production • Shortage of calf rennet • Demand for meat made more calves reared to maturity • Let to high cost of natural calf rennet and find alternate proteolytic enzymes that would duplicate the action of calf rennet in cheese vat. •

Microbial rennet • Some molds such as Rhizomucor miehei, Mucor miehei, Mucor pusillis and Endothia parasiticaare able to produce proteolytic enzymes • The flavor and taste of cheeses produced with microbial rennet's tend towards some bitterness, especially after longer maturation periods.

Genetically Engineered Rennet

• Calf genes to modify some bacteria, fungi or yeasts to make them produce chymosin • Recombinant technique Recombinant chymosin • the first artificially produced enzyme registered and allowed by the U.S. Food and Drug Administration •

Conti…. • 60% of U.S. hard cheese was made with genetically engineered chymosin • it has up to 80% of the global market share for rennet.

• the most widely used genetically engineered rennet is produced by the fungus Aspergillus niger • Other micro organisms are Escherichia coli, Kluyveromyces lactis • Escherichia coli – not regarded as GRAS (Generally Regarded As Safe) – recombinant proteins are frequently synthesized as intracellular inclusion bodies process cost increases

• Cheese production with chymosin is similar to production with natural calf rennet • Preparation of calf rennet has two forms of chymosin A in proportion 40% & B 60 % • Genetic rennet contains only one of the known main chymosin types, either type A or type B • Other do not exist

Conti….. • genetically engineered chymosin and natural pepsin is mixed to imitate the complexity of natural rennet and to get the same results in coagulation and in development of flavor and taste • suitable for vegetarians if there was no animal based alimentation used during the production in the fermenter

Aspergillus niger • Aspergillus niger is a fungus and one of the most common species of the genus Aspergillus • a common contaminant of food • A. niger is cultured for the industrial production of many substances. • Various strains of A. niger are used in the industrial preparation of citric acid (E330) and gluconoic acid (E574) • A. niger fermentation is "generally regarded as safe“ by the FDA.

Fermentation

The production organism is grown in several stages to build up the inoculums for large-scale production

After inoculation the cells grown aerobically under the proper conditions of pH, temperature, nutrient composition After chymosin reaches a desired level in the fermentation broth, the fermentation is stopped and the fungal cells are inactivated and separated from the liquid.

• Chymosin recovered from the broth two methods: • the broth is filtered, followed by chromatographic purification and concentration of chymosin • the chromatographic step is preceded by extraction of chymosin from the fermentation broth • Chymosin eluted from the chromatographic column is formulated to the commercial

Rennet producing industries • Gist brocades – maxiren – K.lactis • Genecor / Chr. Hansen – chymogen – A.niger • Pfizer – chymax- E.coli • Iranian industrial enzymes company – rennilase