Study On The Preparation,Modification And Solution Conformation Of β-glucans In Saccharomyces Cerevisiae

β-glucan from Saccharomyces cerevisiae had many bioactivities, such as immunity-enhancement,antitumor activity, antioxidation, and so on and had been attracted the attention of researchers at homeand abroad. However, water insolubility limited its application in food, cosmetic and pharmaceuticalindustries. In this research, the preparation and modification of yeast β-glucan had been studied. Therheological properties and solubility properties of yeast β-glucan and water soluble glucan had beencompared, and the structure and solution conformation of water soluble gluan had been investigated atthe same time. The main conclusions were summarized as follows:We developed a method including induced autolysis, high pressure microfluidization （HPM） andcompound enzymolysis to prepare β-glucan from Saccharomyces cerevisisae. The optimum conditionswere HPM1:210MPa, concentration of20%in wet weight （w/w）, passed3times, compoundenzymolysis:50mins, enzyme concentration1.0%（w/w）,55℃, pH7.0, ratio of wet weight material toliquid1:4（g/ml）, HPM2:210MPa, passed3times.The purity of the yeast β-glucan extraction was81.070.82%and the yield was14.73%. The new method of yeast β-glucan preparation had many advantages,such as mild working conditions, environment friendly and simple work-up procedure. Among all thepreparation process, HPM1, compound enzymolysis and HPM2processes had significant effect on thetotal sugar content and purity of β-glucan which increased from70.17%to91.60%and48.85%to81.07%, respectively. However, the total nitrogen content decreased from51.18%to3.26%during thepreparation processes and the oval-shaped structure of yeast had been destroyed gradually and changedinto unshaped tabular form.Compared three kinds of ionic liquids and chose the one which could completely dissolve yeastβ-glucan in the shortest time for further modification. On this basis, the modification technology ofyeast β-glucan under high pressure microfluidizaion （HPM） in ionic liquid had been studied and theoptimum process conditions of HPM were pressure175MPa, residue time9min for200mL ionicliquid, concentration of0.5%（w/w）. The yield of water soluble glucan （WSG） reached79.25%with thetotal sugar content of97.22±0.54%. Furthermore, the effect of recycle times of ionic liquid on the yieldof water soluble glucan had been researched. It was found that the yield of water soluble glucandecreased when the ionic liquid recycling times increased. The structure of ionic liquid before and afterrecycle had been studied with FTIR and1H NMR techniques, and it could be concluded that HPMtreatment wouldn’t destruct ionic liquid structure.The rheological analysis showed that both yeast β-glucan and WSG were shear thinning fluidunder various conditions, such as different polysaccharides concentration, pH, NaCl concentration andtemperature. At lower share rate （0.1s-1）, the apparent viscosity of yeast β-glucan had lower dependenceon concentration, but it was sensitive to strong acid （pH2） and alkaline （pH12） conditions which makesthe apparent viscosity decrease dramatically. Low concentration of NaCl （1.0%） could contribute toviscosity increasing, however, the excessive concentration of NaCl （＞5.0%） would decrease theviscosity. The apparent viscosity of WSG had highly correlation with concentration at lower share rate (0.1s^-1) and was stable in a wild pH range l, but only sensitive to strong alkaline （pH12） condition. Theviscosity of WSG increased along with the enhanced NaCl concentration （0⁵.0%）, but excessive NaClconcentration （＞5.0%） could reduce the viscosity. The relationship between storage moss （G’） and lossmoss （G’’） of yeast β-glucan solution were G’’＞G’ at25℃, and both of them increased while thetemperature increase. But G’ and G’’ had no intersection point all the time. The relationship betweenstorage modulus （G’） and loss modulus （G’’） of WSG solution were G’＞G’’ at25℃. G’ decreasedsharply and G’’ decreased slowly while the temperature increase, and there was one interaction point ofG’and G’’at about45℃, which indicated that there was gel structure formation of WSG.The solubility of yeast β-glucan and WSG in ethanol, acetone, DMSO and ionic liquid （EmimAc）have not changed. Both of them were insoluble in ethanol and acetone and soluble in DMSO andEmimAc, but the water solubility have improved significantly after modification. Before modification,only0.4%of yeast β-glucan could dissolve in water, and after modification, there were about80%ofWSG could dissolve in water.The structure and solution conformation of water soluble glucan have been studied using FTIR,¹³CNMR and HPSEC-MALLS. Both yeast β-glucan and WSG are polysaccharides with β-（1-3） glycosidicbonds. Compared with yeast β-glucan, more replacements on C-6were found in WSG. The chainconformation of water-soluble glucan in0.1M NaNO₃aqueous solutions at25°C was studied withHPSEC-MALLS and viscometry. The Mark-Houwink equation was established to be [η]=2.09×10^-2Mw^0.63（cm³g（-1））. On the basis of YFY wormlike cylinder model, the conformation parameters werecalculated to be M_L=752nm^-1, q=3.7nm, which indicated that water soluble glucan existed assemi-stiff chain in aqueous solution.