Screening, isolation and purification of amylase produced by a bacterium from deteriorating yam
| dc.contributor.author | Aladejana, Oluwatoyin | |
| dc.date.accessioned | 2020-02-06T09:29:02Z | |
| dc.date.available | 2020-02-06T09:29:02Z | |
| dc.date.issued | 2014 | |
| dc.description | xix,93 | en_US |
| dc.description.abstract | This study screened and isolated an α-amylase producing bacterium from decayed yam, optimized α-amylase production conditions and characterized the purified extracellular α- amylase produced by the bacterium. This was with a view to obtaining α- amylase from a cheaper source. solates were obtained using pour plate method. They were purified and kept on nutrient agar slant stored at 4oC. The isolates were cultured on 1% (w/v) starch agar and screened for starch hydrolysis using Gram’s iodine solution. Isolate(s) with ability to hydrolyze starch was chosen as good candidates for the production of amylase and was identified to species level using morphological, biochemical characteristics and 16SrRNA gene sequencing technique. Alpha-amylase production was carried out in a production medium with pH adjusted over a range of 6.0 to 8.0. The medium was incubated in a shaker at 45oC and 150 rpm for 48 hours. Cell free supernatant containing crude α-amylase was obtained and the activity of the crude α-amylase was assayed following standard methods. Purification was carried out using ion exchange and gel filtration chromatography. The molecular weight and kinetic parameters were determined using established methods. Thirty six (36) bacteria colonies were isolated, seven (7) of the isolates showed amylolytic property and were denoted as Y4, Y22, Y23, Y25, Y31, Y35 and Y36 respectively. Four genera were identified. The genera are Staphylococcus, Lactobacillus, Neisseria and Bacillus. The bacterium with the highest amylolytic activity was Bacillus subtilis (Y25). The conditions for optimum α-amylase production by Bacillus subtilis (Y25) were incubation time of 38 hours in the presence of 1% starch, peptone as nitrogen source, temperature of 45 ºC and pH of 8.0. The specific activity of the amylase obtained was 16.4 Units/min with a yield of 69 %. The Km and V max obtained were 230 mg/ml and 166.7 Units/mg protein respectively. The optimum temperature and pH determined for the partially purified α-amylase were 60oC and 8.0 respective. About 65% activity was retained at 340 mM of Na+ while Ca2+ enhanced the purified α-amylase activity up to 40 mM. Magnesium ion enhanced its activity only up to 40 mM while Al3+ was found to inhibit the activity of the enzyme steadily up to 60 mM. EDTA showed residual activity up to 40 mM. The molecular weight of the partially purified α-amylase from Bacillus subtilis (Y25) was estimated to be 58 KDa. The study concluded that Bacillus subtilis Y25 produced an alkaline α-amylase. | en_US |
| dc.identifier.citation | Aladejana, O. (2014). Screening, isolation and purification of amylase produced by a bacterium from deteriorating yam. Obafemi Awolowo University | en_US |
| dc.identifier.uri | https://ir.oauife.edu.ng/handle/123456789/5147 | |
| dc.language.iso | en | en_US |
| dc.publisher | Microbiology, Obafemi Awolowo University | en_US |
| dc.subject | Screening | en_US |
| dc.subject | Isolation | en_US |
| dc.subject | Purification | en_US |
| dc.subject | Amylase Produced | en_US |
| dc.subject | Deteriorating Yam | en_US |
| dc.subject | Amylase | en_US |
| dc.subject | Yam | en_US |
| dc.title | Screening, isolation and purification of amylase produced by a bacterium from deteriorating yam | en_US |
| dc.type | Thesis | en_US |