Study On Preparation And Separation Of Konjac Glucomanno-oligosaccharides With Mannase

Glucomanno-oligosaccharides have a positive effect on Bifidobacterium, and also have a variety of other physiological activitis. It is a kind of functional oligosaccharides. Konjac tubers, which has a high content of glucomannan, is an excellent raw material for preparation of glucomanno- oligosaccharides. In this paper, the preparation of konjac glucomanno-oligosaccharides by the mannase and their separation was studied on the following aspects.Konjac tubers was pre-processed by three different processed methods, granulated crushing, high shear emulsification, and high shear emulsification coupled ultra-high pressure homogenization, and then prepared for crude powder. The physical structure of konjac crude powder was observed by optical microscope and scanning electron microscope (SEM). It's found that high shear emulsification processing could destroy the physical structure of konjac and improve its fluid characters, and be more suitable for the enzyme hydrolysis. Accroding to these, the high shear emulsification method was determined as the pre-process method of the raw materials. The content of glucomannan in konjac crude powder was 47.85%, and the ratio of glucose and mannose was man: glu = 1.28:1, which was determined by High Performance Liquid Chromatography (HPLC).Konjac crude powder was used for the substrate and for preparing glucomanno-oligosaccharides by neutralβ-mannanase. Firstly, through the single-factor experiments, the four main factors in the enzyme hydrolysis and their ranges were determined, time (x1), temperature (x2), pH (x3) and enzyme-substrate ratio (E/S, x4). The four factors were optimized by response surface methodology (RSM), and the multiple regressions using the second-order polynomial model between the direct reducing sugar (DRS) content (Y1) and the four factors as following.Y1 = 3.5033 + 0.2705x1 - 0.4508x2 + 0.7536x3 - 0.2928x12 - 09113x22 - 1.5689x32 While, the optimal enzyme hydrolysis conditions were predicted as 3.4h, 41.0℃, pH 7.1 and E/S 0.49. According to validation experiment, DRS content in the enzyme hydrolysis products at the optimal reaction conditions is very close to that of the predictive value, the absolute value of relative error was less than 5%, and the total sugar yield was 64.46% relevantly.The crude sample of glucomanno-oligosaccharides prepared by neutralβ-mannose was detected by fluorophore-assisted carbohydrate electrophoresis (FACE). It's fond that the sample not only contained mannose and manno-oligosaccharides of polymeric degree 2~5, but also contained other nine oligosaccharides. The crude sample of glucomanno-oligosaccharides was separated by silica gel column chromatography and charcoal column chromatography. Four components were separated by silica gel column chromatography successfully, they were detected by TLC and HPLC. The results showed that the first component was mannobiose, the second was a kind of gluco-mannotriose, and others were compounds of the same DP of oligosaccharids. The charcoal column chromatography can only separate mannobiose, and thus it was not suitable for separation of glucomanno-oligosaccharides prepared from konjac crude powder.