Aureobasidium pullulans ile Liamocin (Ağır Yağ) Üretiminin İncelenmesi

Abstract

In this thesis, the effects of various fermentation parameters on liamocin production of Aureobasidium pullulans NBRC 100716 were investigated. In the study, all fermentation experiments were carried out in batch system, using a water bath with controllable shaking speed and temperature. Primarily, the liamocin production of six A. pullulans strains was macroscopically examined with a pre-screening test using an agar medium in which three different sugars were tested separately and it was determined that all strains could grow on agar medium and produce an oil layer around the colony as an indicator of liamocin production. Afterwards, liamocin production of six A. pullulans strains was compared with the aim of identifying a strain that produced high concentrations of liamocin, and it was determined that A. pullulans NBRC 100716 produced higher concentrations of liamocin (4.20 g/L) than the other strains studied. Then, liamocin production of this selected strain was investigated in fermentation media containing different carbon sources. In the experiments carried out for this purpose, when glucose, fructose, maltose and sucrose were used as carbon sources in the fermentation medium, it was observed that the highest liamocin concentration (5.50 g/L) was obtained in the experiments using fructose. For this reason, it was decided to continue the study with a fermentation medium containing fructose as a carbon source. In the trials where the effects of different nitrogen sources used in the fermentation medium on the liamocin production of the tested strain were investigated; peptone and yeast extract were used as organic nitrogen sources and (NH4)2SO4 was used as inorganic nitrogen source. In experiments where peptone and yeast extract were used together as nitrogen sources, it was determined that the production of liamocin was at a higher concentration (4.26 g/L) than in the fermentation medium where other nitrogen sources were used. In the next part of this study, optimization of liamocin production of A. pullulans NBRC 100716 strain was performed by response surface method. For this purpose, in the Design Expert®12 (Stat-Ease Inc., Minneapolis, ABD) software; rotatable central composite design method was used. Within the scope of the experimental design plan created by the program, 20 experiments, 6 parallels at the central point, were carried out. While the independent variables selected in the design were the initial fructose concentration of the fermentation medium, initial pH and temperature, the dependent variables were the maximum liamocin concentration, the microorganism specific growth rate and the maximum specific liamocin formation rate. Multiple regression analysis was applied to the data obtained as a result of the experiments carried out under the conditions given in the design plan, with the help of the program, and model equations showing the effects of independent variables on dependent variables were derived. The convenience of the created model equations was evaluated by analysis of variance (ANOVA). An experiment was carried out under these conditions by choosing a solution with a desirability function was 0.825, initial fructose concentration of fermentation medium was 93.47 g/L, initial pH was 4.92 and temperature of 27.2°C from 58 solutions created by the program during the optimization. In the experiment performed under the selected conditions, the maximum liamocin concentration, the specific growth rate of A. pullulans NBRC 100716 strain and the maximum specific liamocin formation rate were found as; 3.56 g/L, 0.0670 h-1 and 0.00450 [g liamocin/(g mo.h)], respectively. It has been verified that the obtained results are within 95% confidence intervals. In this thesis, molecular confirmations of some liamocin samples produced by the tested strain were performed by MALDI-TOF-MS. In addition, the fatty acid profile of liamocin produced by the tested strain was also determined by GC-FID analysis. In this study, some tests were also performed to determine the biosurfactant activity of liamocin samples produced by the tested strain. The oil spreading diameter of the 1.08 mg/mL liamocin sample was measured as 8.10 cm. The surface tension value of the liamocin sample was determined as 31.6 mN/m using the pendant drop method.