Research On Separation And Purification Of Soybean Oligosaccharides From Sweet Slurry In Soybean Sheet Production

Recently, with the development of science and technology researchers have paid considerable interest in biological functions of oligosaccharides. Soybean oligosaccharides are soluble carbohydrate in soybean seeds and account for 7-10 % of the total carbohydrates in soybean. Researches have found that the intake of soybean oligosaccharides have important functions on the balance of microflora in intestine, prevention of many illness, enhancing body immunity, prevention of aging, etc. This was attributed by some researchers to promotion of bifidobacterium proliferation by soybean oligosaccharides. Researches had tested that oligosaccharides led to flatus and bellyache. However moden safe tests have shown that soybean oligosaccharides is not the direct reason of flatus and it is safe to take soybean oligosaccharides.The production of soybean sheet from soybean follows a traditional process: After soaking milling, filtering, and boiling, soybean sheet is formed in the milk surface kept at certain temperature, and could be harvested at some time interval. With the harvest of soybean sheet thick and yellow slurry was left, namely the sweet slurry. Sweet slurry contains a considerable amount of soybean oligosaccharides. Normally it is used as feedstuff for animals or fed back to the soymilk pool to produce next batch of soybean sheet. However soybean sheet produced by this method was not in good merchant quality, for it was thick, easily broken, dark color with less luster. The aim of this research was to explore the method for separation and purification of soybean oligosaccharides from sweet slurry, thus to tap the potential of the sweet slurry resource and to provide alternatives for the production of soybean oligosaccharides. The research results may provide theory basis and guidence for value-added industry utilization of sweet slurry.HPLC method was used in this research to trace the flow of soybean oligosaccharides during the production of soybean sheet. The results showed oligosaccharides losses were lost in soaking, blanding and filtrating, sheet forming stages, representing 1.43 %,48.5 %,24.18 % (on weight). Soaking did not show much influence on oligosaccharides content. Surcose, raffinose and stachyose losses were 0.91 %,0.31 % and 0.21 % respectively. Blanding and filtrating was the most important step for oligosaccharides losses compared with other steps. Surcose, raffinose and stachyose losses were 21.68 %,10.76 %,16.06 % relative to the total oligosaccharides in soybean seeds. The final product soybeansheet contains 14.66 % of the total oligosaccharides. Still 11.23 % of oligosaccharides were in the sweet slurry.The component of the sweet slurry sampled from factory was analysed in this research. Results indicated that the total carbohydrate accounted for 50 % of the dry matter in sweet slurry and most of them were oligosaccharides, which represented 40 % of the total dry weight. Soluble protein accounted for 17 % of the total dry weight. Functional substance such as toal flavone and saponins concentration were 0.07 and 0.5 mg/ml respectively, representing 0.03 %,0.22 % (dry weight). SDS-PAGE result showed nine protein bands in the electrophoresis graph, and the molecule weight was 76.36,50.37s 41.35,37.44,27.14,25.78,19.94,17.62,13.64 kDa respectively. The analysis of the sweet slurry showed functional oligosaccharides content was much higher than that in soybean whey water. And the recovery percentage and total energy consumption were far lower than from the whey water.UF experiment was conducted with PES membrane. The sweet slurry must be pretreated in order to guarantee the proper function of UF. The pretreatment experiment showed that the best pretreatment condition was pH 4.5, 70℃, heating 5 min and 10 % CaCl₂ concentration. After the pretreatment the protein concentration was dropped to 3.99 mg/ml and 89 % of protein was removed. Single factor trials used MWCO 10000 kDa PES membrane. The results showed that the optimum conditions for extracting oligosaccharides from sweet slurry were: 1.75 Ba TMP, 45℃temperature and pH 7.0. Orthogonal intersection trials revealed that MWCO became the major affecting factor on oligosaccharides retention and protein removal. Membrane with MWCO 3000 kDa was best for protein removal while oligosaccharides retention and permeate flux were the lowest in all trials. The oligosaccharides retention and permeate flux attained the highest level with MWCO 10000 kDa membrane, while the protein removal was relatively low. Temperature of 50℃was good for both protein removal and oligosaccharides retention. Considering the efficiency of protein removal, oligosaccharides retention, permeate flux and membrane decline, the optimum conditions for UF were 1.5 Ba pressure, 50℃temperature, pH 7.0, MWCO 10000 kDa. After this treatment 73 % of protein was removed and more than 85 % of oligosaccharides were retained.The permeate fluid after UF contains large quantities of salts and must be removed to meet the oligosaccharides product standard. Electrodialysis (ED) was employed for the desalination process. NaCl solution was used first to test the stability of the ED apparatus. NaCl desalination trial showed that the ED setup was stable. The experience model for limited current density (LCD) of the ED apparatus employed in this experiment was I_lim=4.13 v^0.112c^0.033. Single factor trial of ED showed that the optimum conditions for desalination was 20 V operating voltage, 60 L/h flow rate and diluting 15 times. Orthogonal intersection trials revealed that 20 V operating voltage, 60 L/h flow rate and diluting 18 times were the best condition for desalination and oligosaccharides retention. Operating voltage was the major affecting factor of oligosaccharides retention, then the flow rate and finally the diluting times. After ED more than 95 % salt in the crude soybean oligosaccharides solution was removed and 90 % of oligosaccharides was retained.The crude soybean oligosaccharides solution was in dark color. Resins were used in the experiment to remove color. Static adsorption method was used to screen the best resin for decolorization from 10 resins and adsorption kinetic of coloured impurities onto resins was also studied. The results showed that more than 70 % of the pigments were removed in the final product by the macroporous adsorption resin DM-130 and AB-8 and the desorption was almost completely. Comparaed with AB-8, resin DM-130 had relatively less adsorption of sugar solution and it was the best resin for decoloration. The adsorption process was favored at lower temperature, indicating the process was exothermic and spontaneous. And room temperaure of about 20℃was better for decolorization. When the ratio of resin mass/oligosaccharides solution volumn was approximately 0.13 g/ml higher decolorization efficiency could be obtained in our experiment. Decolorization efficiency did not change much when resin mass-oligosaccharides solution volumn ratio was higher than 0.13 g/ml. The adsorption isotherm better agreed with the Freundlich model, indicating the monolayer sorption of coloured impurities onto macroporous resins and that the impurities were of multi-component nature rather than single component. Adsorption kinetic of coloured impurities onto resins fitted the pseudo-second-order kinetic model, this indicated that the adsorption process was chemical adsorption which was the speed limiting step. The intraparticle diffusion model showed that the adsorption process occurred by both surface sorption and intraparticle diffusion. The kinetic data showed the best flow speed was 1 ml/min, and the concentration of ethanol washing solution was 50 %.