Effect Of Carbon Sources On The Structure And Biological Activity Of Exopolysaccharides From Two Macro Fungi

As the main source of polysaccharide sugar units, carbon sources in medium may affect the molecular structure of exopolysaccharide(EPS), and thus affect the biological activities associated with its structure. In order to investigate the effects of different carbon sources on the EPS molecular structure and biological activity, this paper use Phellinus vaninii Ljup and Scleroderma areolatum Ehrenb as the object, the flask fermentation conditions were optimized, the morphological changes under optimal fermentation conditions were investigated. Five kinds of preferred carbon sources were selected to prepare the crude EPS using fermentation tanks. The EPS was purified after removing the protein and pigments and subjected to gel filtration chromatography. The molecular weight of purified EPS was determined by gel filtration, and its molecular structure was further characterized by FT-IR, gas chromatography mass spectrometry(GC/MS), size exclusion chromatography(SEC) and multi-angle laser light scattering detector(MALLS) associated with differential refractive index detector(RI), and viscosity method respectively. Finally, the scavenging activity of EPS to ?OH and ?DPPH was determined. The main results are listed as follows:(1) The optimal culture conditions for EPS production of P. vaninii were 40 g/L sucrose, 4 g/L corn steep powder, 4 mmol/L KH2PO4, 28℃ and p H 7, 160 r/min. The maximum EPS yield was 0.352 g/L under the optimal conditions. The optimum culture conditions for EPS production of S. areolatum were 20.0 g/L fructose, 5.0 g/L soybean meal, 2.5 mmol/L KH2PO4, 2.5 mmol/L Mg SO4, 26℃, p H 8, and 160 r/min; the maximum EPS yield was 2.627 g/L. The average diameter and density of the mycelial pellets of P. vaninii increased along with the prolong of culture time increases, however, the roundness of the pellets increased first, then decreased sharply. The roughness was negatively correlated with the roundness. The mycelia of S. areolatum was branched growth, the population was lamellar rather than spherical as most filamentous fungi. The viscosity was positively related with the culture period, and the p H value was negatively relevant to the culture time.(2) The relative molecular weight of EPS Fr-I of P. vaninii produced using glucose, fructose and sucrose, maltose as carbon sources was 627.5 and 1153.5 KDa, respectively.However,the relative molecular weight of EPS Fr-II produced using glucose and sucrose, fructose and maltose as carbon sources was 55.0, 74.5 and137.0 KDa, respectively. The relative molecular weight of EPS produced using lactose as carbon source was 101.0 KDa.The relative molecular weight of EPS Fr-I of S. areolatum produced using glucose, fructose and sucrose, and maltose and lactose as carbon sources was 627.5 and 1563.9 KDa. The relative molecular weight of EPS Fr-II produced using glucose, sucrose, fructose, maltose, and lactose as acarbon sources was 4.82 KDa.(3) The FT-IR assay showed that the EPS Fr-I of P. vaninii produced using glucose as carbon source was β-pyranoside acidic polysaccharides, EPS Fr-I fermented use fructose and maltose as carbon sources were α-mannopyranoside acidic polysaccharides, EPS Fr-I produced using sucrose as carbon source was α- and β-mannopyranoside acidic polysaccharides; Moreover, EPS Fr-II produced using maltose as carbon source was β-mannopyranoside acidic polysaccharides, EPS Fr-II fermented use glucose, fructose and sucrose as carbon sources were α-mannopyranoside acidic polysaccharides, while the EPS produced using lactose as carbon source was mannose acidic polysaccharides. The purified EPS from five kinds of carbon sources contained a large number of glucuronic acid and galacturonic acid, and the most abundant monosaccharide component was mannose, followed by glucose, and the content of rhamnose and ribose was least. SEC/MALLS analysis suggested that the polydispersion and solubility of the EPS was very low, and the EPS kept spherical conformation in aqueous solution. The EPS was a highly compact aggregates with polysaccharidal branches. The k’ and ρ values in viscosity determination indicated that the solubility of purified EPS was very low in aqueous solution. The EPS exhibited highly compact aggregates with polysaccharidal branches.EPS Fr-I of S. areolatum produced using five kinds of carbon sources were α-pyranoside acidic polysaccharides, and showed mannose characterized absorption peaks. EPS Fr-II produced using glucose, fructose, lactose and maltose as carbon sources were α-mannopyranoside acidic polysaccharides, while the EPS Fr-II fermentated use sucrose as carbon source was β-mannopyranoside acidic polysaccharides. The monosaccharide compositions of purified EPS included arabinose, rhamnose, ribose, xylose, glucuronic acid, galactose, glucose and mannose, with galacturonic acid and glucuronic acid as the common compositions. However, the type and content of the composition were different significantly. SEC/MALLS analysis showed that the polydispersion and solubility of the EPS was also very low, and the EPS kept spherical conformation in aqueous solution. The EPS was a highly compact aggregate with polysaccharidal branches. The k’ and ρ values in viscosity assay showed that the solubility of purified EPS was very low in aqueous solution. The EPS exhibited a highly compact aggregate with polysaccharidal branches.(4) The EPS of P. vaninii produced using sucrose, fructose, lactose, glucose and maltose as carbon sources showed the strongest scavenging activity to ?OH at the concentration of 10 mg/m L, and the scavenging rate was 38.58%, 34.50%, 30.18%, 26.60% and 20.68%, respectively. The EPS showed the strongest scavenging activity to ?DPPH at the concentration of 5 mg/m L. The scavenging rate was 59.17%, 44.38%, 34.32%, 22.78% and 33.43%, respectively. The EPS of S. areolatum produced using fructose, lactose, sucrose, maltose and glucose as carbon sources showed the strongest scavenging activity to ?OH at the concentration of 10 mg/m L, the rate wase 22.65%, 18.87%, 16.69%, 14.86% and 13.66%, respectively. The EPS showed the strongest scavenging activity to ?DPPH at the concentration of 5 mg/m L, the rate was 41.32%, 33.53%, 31.10%, 20.23% and 25.77%, respectively. The study suggested that EPS with moderate molecular weight displayed the highest antioxidant activity, however, the EPS with higher viscosity showed lower antioxidant activity. Furthermore, the composition of mannan may influence the antioxidant activity of EPS.The EPS with β- configurationexhibited higher antioxidant activity. The type and content of monosaccharide component, especially those of the mannosyl, also showed important infulence on the antioxidant activity of EPS.