Paenibacillus sp. Isolate 2S-3與2S-6菌種特性分析與紙漿漂白前處理應用

Abstract

Paenibacillus sp. isolate 2S-3 與2S-6是由製漿漂前黑液中所分離出的菌種。本研究的黑液是由中華紙漿公司收集得來，2S-3以及2S-6皆具有木聚醣酶活性，並利用16S rDNA定序鑑定出與其他菌種的親緣關係。 分析結果顯示2S-3產生的木聚醣酶大小為40kDa，2S-6會因SDS與sample buffer而變性，所以利用膠體層析分析2S-6產生的木聚醣酶大小為37.65kDa。菌株2S-3、2S-6所產生的粗酵素液最適合的溫度與pH為pH 7、溫度分別在70oC、60oC，利用碳源木聚醣(birchwood xylan)、米糠(rice bran)、米殼(rice husk)與氮源0.5%酵母抽出物(yeast extract)加0.5%蛋白棟(peptone)、大豆粉(soy bean)可誘導產生木聚醣酶，2S-3與2S-6分別以基質2%木聚醣、0.5%酵母抽出物、0.5%蛋白棟培養會有最高活性6.1 IU/ml和5.6 IU/ml。 將硫酸鹽尤加利紙漿以粗木聚醣酶酵素液每克絕乾漿5 IU作前處理後，再進行C70/D30-Eo-D漂白，菌株2S-3和2S-6所分泌之木聚醣酶前處理漂白後所測得的白度90.1，相較於對照組88.2皆有明顯的提升，由結果發現木聚醣酶前處理能節省漂白藥劑、增進漂白效果，足證此等木聚醣酶具有進一步研究應用的潛力。

Paenibacillus sp. isolate 2S-3 and 2S-6 were isolated by screening bacterial strains from the thermoalkaline black liquor. The black liquor was collected from Chung Hwa Pulp Corporation. Strains 2S-3 and 2S-6 exhibited xylanase activities and its’ phylogenetic relationship with other bacteria were identified by 16S rDNA sequencing. The xylanase of strain 2S-3 and 2S-6 have been analyzed. The xylanase produced from 2S-3 was analyzed in zymogram and showed a molecular mass of about 40 kDa. 2S-6 xylanase could not analyze in zymogram because xylanase would denature with SDS and sample buffer. Using gel chromatography column to determine molecular mass of crude xylanase was the suitable way and showed the molecular mass of about 37.65 kDa. The optimum temperature and pH for crude enzyme of 2S-3 and 2S-6 were 70oC and pH 7, 60oC and pH 7. The results showed Paenibacillus sp. isolate 2S-3 and 2S-6 used different carbon and nitrogen sources to induce xylanase. The carbon source included 2% xylan, rice bran, rice husk and nitrogen source contained 0.5% yeast extract plus 0.5% peptone, soy bean, corn steep liquor (C .S .L.), (NH4)2SO4. The highest xylanase activities of 2S-3 and 2S-6 were 6.1 IU/ml and 5.6 IU/ml with medium contained 2% xylan, 0.5% yeast extract and 0.5% peptone. After pretreated by 2S-3 and 2S-6 xylanases at 5 IU per gram oven dry pulp (o. d. p), eucalypt kraft pulp was bleached by C70/D30-Eo-D sequence. In 2S-3 and 2S-6, 90.1% brightness were achieved by both xylanase pretreatment, compared by 88.2% from controls. Result showed xylanase pretreatment can be integrated into different bleaching sequences. It can save bleaching chemical usages and enhance brightness of final products. The results validated the efficiency of xylanase and warranted studies for further application.