Browsing by Author "Gessesse, Amare"

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    Alkaline and Thermostable Xylanases of Potential Industrial Importance from Alkaliphiles
    (Addis Ababa University, 1998-12) Gessesse, Amare; Mattiasson, Bo (Professor)
    Three xylanase producing alkaliphilic bacterial strains designated AR-009, AR-135 and AR-22-1, were isolated from Lake Arenguadie, an alkaline soda lake in Ethiopia. AR- 009 and AR-135 were identified as strains of the genus Bacillus and Micrococcus, respectively, while AR-22-1 remains unidentified. The enzyme from all the 3 strains were purified following standard protein purification procedures, and the molecular weight of each enzyme was estimated using SDS-PAGE. Two xylanases designated as xylA and xylB, having molecular weights of 23 and 48 kD, respectively, were purified from the cell free culture supernatant of Bacillus sp. AR-009. XylA was optimally active at pH 9 while xylB showed optimum activity in the pH range of 9 to 10 and stable from pH 5 to 11. The optimum temperature for the activity of xylA was 70°C at pH 8 and 60°C at pH 9. On the other hand xylB was optiImilly active at 70°C at pH 8 and 75°C at pH 9. Both enzymes showed good stability at 60°C and at a pH of 8 and 9. AR-135 xylanase was optimally active at 55°C and a pH of 7.5 - 9.0. Over 60% of the maximum activity was displayed at pH 11. Its thermal stability above 40°C was very low. The optimum temperature and pH for the activity of AR-22-1 xylanase was 70°C and 8.0 to 9.5, respectively. The enzyme was stable in a broad pH range and showed good stability up to 60°C at pH 8 and 9 Agar immobilized cells of Bacillus sp. AR-009 were used for xylanase production with a view of finding cheap ways of enzyme production. In a batch culture maximum enzyme production was observed after 48 h and remained high up to 72 h. In repeated batch cultivation immobilized cells produced appreciable level of xylanase activity in 7 consecutive batches with out any significant decline in productivity. For continuous xylanase production immobilized cells were packed in a jacketed glass coluIlUl and sterile medium was continuously pumped. A stable continuous production of xylanase was observed over a period of one month. The volumetric productivity of the continuous culture was 17 fold higher than the batch culture using free cells. Another method of enzyme production investigated was the use of solid state fennentation. Bacillus sp. AR-009 produced high level of xylanase activity when grown using solid state fermentation (SSF) with wheat bran serving as a substrate. Xylanase production was highest at a wheat bran to moisture ratio of between 1:0.5 to 1: 1.5 and a Na2COJ concentration of 10% (w/w). No significant effect was observed on xylanase production when wheat bran is supplemented with peptone, try one, and yeast extract, thus avoiding the need to supplement wheat bran with expensive media. The ability of the organism to produce high titre xylanase activity at alkaline pH and lower wheat bran to moisture ratio could have a potential advantage in minimising the risk of contamination. In addition, because the enzyme can be extracted using minimum volume of liquid, the cost of down stream processing during product upgrading and the cost of waste treatment steps can be greatly reduced. The use of SSF for the production of xylanase by Bacillus sp. AR-009 could therefore lead to substantial reduction in the over all cost of enzyme production.
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    Production of Microbiological Peptone From Hydrolysis of Slaughterhouse Offal Using Bacterial Protease
    (Addis Ababa University, 2018-01-05) Teshome, Medhanit; Gessesse, Amare
    Proteases are the most important class of industrial enzymes accounting for 60% of the global industrial enzyme market. Microorganisms are the major source of these enzymes. Production of hyrolysates from different protein sources is among the different application of proteases. Protein hydrolysates have a variety of food and non-food applications. Although different proteases are available in the market, there is always a need for the development of new enzymes from bacterialsources. This is especially important in countries like Ethiopia where there are no local enzyme producers. The aim of this study was, therefore, to isolate new protease producing bacterial isolates to be used for the hydrolysis of slaughterhouse offal, optimize enzyme production and hydrolysis conditions, and test hydrolysates as a microbiological growth media. Based on screening data on solid and liquid media, one bacterial isolate designated as aau5 was selected for further study. The isolate grew under solid-state fermentation (SSF) and produced up to 5,773 U/g of enzyme. Enzyme production was optimal when the solid to moisture ratiowas 1:2(66.7 % moisture content) and in the presence of organic nitrogen sources. Protease aau5 was optimally active at pH 7.5 and temperature of 55°C. After one hour incubation, the enzyme retained up to 66% and 41% of its original activity at 50°C and 55°C, respectively. Protease aau5 was used for the hydrolysis of slaughter house offal (lung and bone) and soybean protein. The hydrolysate (peptone) was then tested as a microbiological media for the growth of different bacterial species. Compared to commercial peptone, hydrolysate obtained from lung (LPA) and bone extracted protein (BPA) supported better growth of the test organisms. So, by using waste and by products of slaughter houses, beneficial hydrolysate like peptone can be produced through enzymatic hydrolysis