Characterization of Extracellular Enzyme from Paenibacillus sp. for Hexenuronosyl-xylan Degradation in Pulp Bleaching

摘要

Presence of HexA in unbleached kraft may cause brightness reversion and excessive consumption of bleaching chemicals. HexA removal can be accomplished by acid hydrolysis with ECF and TCF bleaching, but process selectivity and bleaching efficiency arelow, otherwise toxic organic chlorinated compounds will generate. An effort and some process modification, such as enzymatic strategic, are needed for HexA removal. Paenibacillus sp., is known to utilize hexenuronosyl-xylotriose (A-X3) as carbon source.Objective of this research is to characterize extracellular and intracellular enzyme from the strain. First, A-X3 was produced from Eucalyptus LOKP by enzymatic hydrolysis using two commercial enzymes sequentially. The hydrolysate then purified using activated carbon column chromatography. A-X3 content was determined using Ion Chromatography (HPAEC-PAD). Enzyme was produced from the bacteria using medium contains 0.5% birchwood xylan as a carbon source. The culture was incubated at 37°C for 24 h. Culture supernatant and bacterial pellet were separated by centrifugation. Obtained culture supernatant was used as extracellular enzyme. Meanwhile, the pellet was suspended in phosphate buffer saline, pH 7 then disrupted using sonicator and used as intracellular enzyme. Crude enzyme activity was performed using A-X3 as a substrate in 50 mM sodium acetate buffer, pH 6 at 50°C, reaction product was analyzed by HPAEC-PAD. Chromatogram result showed that xylose, A-X3 and A-X2 peak were appeared when intracellular enzyme from Paenibacillus was used to hydrolysis the substrate. Meanwhile, xylotriose appeared when extracellular enzyme was used to hydrolyze the substrate. These result suggested that intracellular enzyme of Paenibacillus was able to hydrolyze xylosidic linkages at the reducing-ends side of A-X3, while the extracellular enzyme was having the hexenuronic acid degrading enzyme.