Improvement of beta-glucan production in Saccharomyces cerevisiae using some additives

Abstract

After screening of S. cerevisiae strains, S. cerevisiae Angel® was selected as the test culture to study the effect of additives on quantity and quality of yeast β-glucan on the basis of high net β-glucan yield. EDTA, SDS and NaCl were supplemented into the medium at various concentrations. Cells were cultivated at pH 4, 30 °C with a reciprocal shaking at 200 rpm for 48 h. Sigmoid curve of yeast growth was analyzed using non-linear regression and presented as a function of shifted logistic equation. Treatments with SDS at 100 ppm and 200 ppm gave higher growth level than control. On the other hand, 50 and 100 ppm EDTA resulted in reduction of growth level. Similarly, the growth level of yeast cell was reduced in 30,000 ppm and 60,000 ppm NaCl. In addition, the time to reach the highest growth rate was longer in the presence of additives. Conditions with no growth and cell death were found in treatments with high concentrations and combination of additives. Based on this observation, the optimum concentration of each additive supplemented solely was 50 ppm EDTA, 100 ppm SDS and 30,000 ppm NaCl. Using computer software for estimation, 4 suitable combination of additives were EDTA 5 ppm and SDS 10 ppm; EDTA 5 ppm and NaCl 3,000 ppm; SDS 20 ppm and NaCl 3,000 ppm; and the combination of EDTA 5 ppm, SDS 10 ppm and NaCl 3,000 ppm. For further investigation, eight treatments (four single additives and four combination of additives) were set to study the effect of additives on cell shape, wall surface, β-glucan production and composition of cell in S. cerevisiae Angel®. Yeast cultured in medium supplemented with 100 ppm SDS exhibited rounder cell shape (the difference between major axis and minor axis length is 0.84 μm) with highest number of bud scars and highest β-glucan content (8.15 %w/w). Whilst, the second group was long oval cell shape and the longest cell was exhibited in treatment with combination of 5 ppm EDTA and 3,000 ppm NaCl (the difference between major axis and minor axis length is 2.00 μm). From FTIR spectra of yeast cell, treatments with additives showed slightly higher amount of polysaccharide region compared to control, with the exception in the presence of combination of 5 ppm EDTA and 3,000 ppm NaCl. Interestingly, these results implied that additives could activate β-glucan formation and cell wall synthesis. The ratio of protein region when yeast was cultivated with 100 ppm SDS was lower than control and other treatments. The highest protein was found when yeast was cultured with three combination of additives. For the β-(1,6) : β-(1,3) in yeast cell, calculation showed that the ratios were higher than control when yeast was cultivated with additives, with the exception of treatment with 50 ppm EDTA. However, after β-glucan extraction, the β-(1,6) : β-(1,3) was lower in treatments with additives. The higher weight average molecular weight than control were found when cultivated yeast with combination of 20 ppm EDTA, 3,000 ppm NaCl and three combination of EDTA, SDS and NaCl. For immunomudulatory properties, the β-glucan was derivatized to carboxymethylglucan (CM-glucan) before test. It was found that degree of substitution (DS) and concentration of CM-glucan seemed to affect significantly stimulation activity of macrophage cell as well as their cytotoxicity. The higher DS and concentration, the higher activity tended to be.