Characteristics of galactomannanase for degrading konjac gel
Introduction
Konjac glucomannan (KGM) is a high molecular weight, water-soluble and non-ionic (neutral) polysaccharide found in roots and tubers of the Amorphophallus konjac plant. It has d-mannose (M) and d-glucose (G) in M/G molar ratio of 1.5–1.6 by β-1,4-glycosidic linkages with about 1 acetyl group in every 17–19 sugar units at C-6 position [1]. KGM has long been used in TCM (tradition chinese medicines) as an immunoregulating and health-care food, such as an indigestible dietary fiber, beneficial for digestion system. Oligosaccharides have been reported to play important roles in biological system like the health diet system of humans and farm animals by the growth of bifidobacteria and by affecting more subtly the immunology status of intestinal cell [2]. Recent study demonstrates that the function of Konjac oligosaccharide (KOS) on a diabetes model of isolated islet was evaluated and the results show one component of KOS consisted of tetrasaccharide could regulate NO level on streptozocin (STZ)-treated islets in a dose-dependent manner [3].
In general, the most practical and useful method to produce KOS is enzymatic degradation of KGM employing the relevant endo-enzymes, e.g. β-mannanase (1,4-β-d-mannan mannohydrolase, EC3.2.1.78). The β-mannanase can hydrolyze 1,4-β-d-mannopyranosyl linkages of polysaccharides (e.g. konjac powder, locust bean gum, guar gum, etc.), such as mannans, glucomannans, galactomannans and galactoglucomannans, yielding manno-oligosaccharides. This enzyme has been isolated and characterized from microbial sources [4], [5], [6], [7], [8], [9], [10]. The galactomannanase (1,4-β-d-galactan-4-mannano-hydrolase, Glmnase) is an enzyme product derived from Aspergillus niger. Glmnase, a hemicellulose-degrading and water-soluble enzyme, is generally used to remove the sediment from coffee extracts, to peel soybean and to be an anti-staling agent (especially for the presence of small amount of glucomannan or galactomannan like guar gum) in food industry. It is also used to desize fabrics made from synthetic fibers in textile industry. The interests in β-mannanase and other hemicellulose-degrading enzymes, e.g. Glmnase, have recently increased, partly because of their potential applications in the food, textile and paper or pulp industries [7], [11]. However, there is no information so far available about Glmnase from A. niger for KGM hydrolysis. In the present study, the aim was to achieve gel hydrolysis of konjac powder by Glmnase from A. niger and to report some enzymatic properties.
Section snippets
Materials
The crude Glmnase from A. niger was a gift from President C.K. Lin (Challenge Bioproducts Co., Ltd., Yun-Lin Hsien, Taiwan) and further purified by gel filtration chromatography (Bio-Gel P-30, Bio-Rad Laboratories, Inc., Hercules, CA). The partially purified Glmnase solution was dialyzed at 4 °C overnight in deionized water and then lyophilized. Konjac powder (glucomannan content ≥79%, (w/w)) was purchased from Asahi-Ya Food Co., Ltd (Taipei Hsien, Taiwan). Konjac gel was prepared by
Molecular mass of Glmnase
The data from the loaded sample (crude Glmnase) as assayed by SDS-PAGE showed two major proteins, one of about 83 kDa and one of about 45 kDa, one minor protein of about 36 kDa (Fig. 1 inset). Further purification by Bio-Gel P-30 gel filtration chromatography yielded the partially purified enzyme (about 45 kDa) preparation (Fig. 1). The Glmnase activity could be tested by hydrolyzing guar gum powder through the activity factor (AF), which is defined as AF = 2 /(T × A), where T is the time in minutes
Conclusion
In this study, the optimum activity of the purified Glmnase acting on the gel of konjac powder has been found at pH 5.0 and 60 °C and proved to be stable in the pH range 3–6 at room temperature. The enzyme was stable in the range of 30–70 °C when it was incubated at 30–80 °C for 30 min to 2 h. Nevertheless, the enzyme activity was nearly lost after 15 min at 80 °C. In fact, the complete enzymatic hydrolysis of locust bean gum galactomannan to oligomers of galactose and mannose requires the action of
Acknowledgements
This study was supported by Grant NSC93-2815-C-212-006-E from National Science Council of Taiwan. The crude galactomannanase (Glmnase) from Aspergillus niger was a kind gift from President C.K. Lin (Challenge Bioproducts Co., Ltd., Yun-Lin Hsien, Taiwan).
References (18)
- et al.
Life Sci.
(2002) - et al.
Enzyme Microbiol. Technol.
(1990) - et al.
Food Microbiol.
(1993) - et al.
J. Biotechnol.
(1998) - et al.
Meat Sci.
(2003) Phytochemistry
(1979)Methods Enzymol.
(1988)- et al.
J. Biotechnol.
(1996) - et al.
Carbohydr. Res.
(2002)
Cited by (1)
Preparation and characterization of KGM /TiO<inf>2</inf> composite membrane
2008, Materials Science Forum