الفهرس 
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Abstract
Proteases are the hydrolase enzymes that catalyze the hydrolysis of the peptide bonds in the primary structure of proteins and peptides .Proteases can be sourced from animals, plants, viruses, bacteria and fungi. Microbial proteases are more desired for industrial applications due to the flexibility of microbes to genetic manipulation and the fact that microbial proteases possess almost all the characteristics needed for biotechnological application.
Microbial proteases are the most important enzymes in the industry, accounting for 60% of the total enzyme scales in the world. The proteases of the lactic acid bacteria (LAB) have received special attention because of their importance in the food and dairy industry.
In the present study four lactic acid bacterial (LAB) isolates were screened for extracellular protease production in submerged fermentation medium using the dairy waste (whey) as a substrate .Different fermentation parameters such as pH, incubation temperature,inoculum volume, incubation time, carbon source and nitrogen source were studied for maximum production of protease enzyme.. Two isolated lactic acid bacteria (LAB) showed maximum proteolytic activity and were identified as Lactobacillus. plantarum and Enterococcus .faecium using Biolog system. The highest protease activity of L.plantarum was (8.72 Unit/ml), protein content (4.13mg/ml)and specific activity (2.11Unit/mg) at optimum pH- 5 and 30ºC after 36hrs of incubation with inoculum size of 5%, fructose as carbon source and yeast extract as nitrogen source , where the protease activity was (10.2Unit/ml), protein content (4.8mg/ml)and specific activity (2.13Unit/mg)at optimum pH-5.5 and 45ºC after 36hrs of incubation with inoculum size of 7%, sucrose as carbon source and yeast extract as nitrogen source for Enterococcus .faecium .
Two protease enzymes were partially purified using ammonium sulphate precipitation followed by sephadex G-100 column chromatography and then characterized.
The crude extract and partially purified protease enzyme produced from selected isolated bacterial strain identified as Lactobacillus plantarum were used in the ripening and flavour improvement of soft white cheese (Domiati -type).
The effect of protease on the chemical and sensory properties of Domiati cheese during storage period was studied. The obtained results showed that, pH value, moisture and protein contents of all domiati cheese treatments were decreased with protease addition and increasing of storage period, whereas the soluble nitrogen, tyrosine and tryptophan and fat contents for all cheese treatments increased with protease addition as storage period advanced compared to control .
It could be seen from the results that, most free fatty acids (FFA) contents were similar between control and cheese treatments made using protease enzyme(crude and purified) during storage period. Whereas the free amino acids (FAA) contents for all cheese treatments with protease were increased with increasing of storage period.
The results indicated that, the predominated free fatty acids in domiati cheese at the end of the ripening were palmitic acid was the most abundant, followed by oleic acid, stearic and myristic acids .The free amino acids glutamic acid, proline, leucine ,aspartic, lysine , serine and valine were present at higher concentrations and represented more than 50% of total amino acids at the end of the ripening period in all domiati cheese samples .
The free amino acid contents of treatment (T1)made using 1% crude protease increased with increasing of storage period till 30 days then decreased In cheese treatment T2 with 2% of crude protease enzyme, it was clear that, free amino acid contents increased till end of the storage period .For cheese treatments T3 with 1% and T4 with 2% of purified protease enzyme, it was found that free amino acid contents decreased during storage period than control, The highest levels of most free amino acids( aspartic, therionine, serine, glutamic, alanine, isoleucine, arginine and proline)were observed with cheese treatment (T1) made using 1% crude protease enzyme at 30 days of storage period.
For sensory evaluation, the results indicated that , as the ripening period progress, the flavor characteristics were gradually increased leading to improvement of organoleptic properties, compared with control one. It also was noticed that, the crude protease enzyme cheese treatments (T1&T2) gained higher body and texture scores than those made with purified protease enzyme (T3&T4) during the storage.. It was found that, cheese samples treated with protease enzyme extract gave better flavor as compared to control without protease during the ripening process. The results showed that, at end of the storage period, the cheese treatment (T2) with 2% crude enzyme recorded the highest flavor score (46.7). Also, the results indicated that, addition of 2% of crude protease enzyme (T2) accelerate the ripening process of domiati cheese through 60 days without any defects in its properties.
Key words: protease, Lactic acid bacteria, Whey, submerged fermentation, sephadex , Domiati cheese , Ripening process, Free amino acid, Free fatty acid , Sensory evaluation and flavor.