| Phaffia rhodozyma which is considered as one of the most important sources of nature astaxathin is easly to culture and grow fastly in many media. In addition, Phaffia rhodozyma was revealed to synthesize functional fructooligosaccharide. The simutaneous production of both fructooligosaccharide and astaxanthin which would enhance the practice value of Phaffia rhodozyma is considered of great significance.This thesis which was expected to provide the foundation for further development and utilization of the astaxanthin high yielding Phaffia rhodozyma JMU-MVP14, was performed on the identification of fructooligosaccharide composition, the investigation of the regulation of fructooligosaccharide synthesis, the optimization of the process for the production of fructooligosaccharide, the exploration of coproduction of astaxthin and fructooligosaccharide in 7 L fermentor and purification of fructooligosaccharide from the cultured broth of this yeast strain. The results are showed as the followings:(1) The condition for runing the high performance liquid chromatography (HPLC) was optimzed to show the mobile phase consisting of 72% acetonitrile and 22% water was proper for the determination of the carbonhydrate in the broth of Phaffia rhodozyma JMU-MVP14 by the HPLC method in a isocratic elution procedure.(2) There were six kinds of carbonhydrate in the broth of Phaffia rhodozyma JMU-MVP14 detected by HPLC-RID and identified to be fructose, glucose, sucrose, maltose, kestose and neokestose by the analysis of LC-MS and the comparison of the carbonhydrate standards, among which the kestose and neokestos is functional frutooligosacchride.(3) Phaffia rhodozyma JMU-MVP14 specially synthesize fructooligosaccharide only in case of sucrose was utilized. The enzyme (β-Fructofuranosidase) catalyzing sucrose to fructooligosaccharide was a constitute enzyme, and its activity could detected not only on the cells but also in the broth supernatant.(4) The sucrose concentration and fermentation temperature after the addition of sucrose were the main factors effecting theproduction of astaxathin and fructooligosacchride.The steepest ascent and central composite design experiment revealed that the optimal condition for the synthesis of fructooligoacchride in the last stage of astaxathin production was sucrose concentration at 236.18 g/L and fermentation temperature at 34.5℃. (5) The two step fermentation experiment in 7 L fermentor which was consisting of the first stage for the accumulation of astaxathin by feeding glucose and the second stage for the synthesis of fructooligosacchride (neokestose and kestose) by feeding sucrose at the end of the first stage followed by increasing the fermentation temperature achieved yield high productivity of astaxathin and fructooligosacchride simultaneously. The astaxathin yield attained up to 194.08 mg/L and fructooligosacchride 40.88 g/L under optimal condition in the two-step fermentation process for the coproduction of astaxanthin and fructooligosaccharide.(6) The purification of fructooligosaccharide from Phaffia rhodozyma JMU-MVP14 broth by the chramotographies on ion-exchange, celite and active carbon column showed fructooligosacchride in the broth efficiently separated from other compounds by using the active carbon as absorbent in the fractional elution procedure at 35℃with ethanol solutions. The content of carbonhydrate in purified fructooligosacchride fraction was up to 97.65%, ash content was less than 0.80%, the content of lead was 0.053 mg/kg, cadmium was 0.003 mg/kg, the arsenic was 0.054 mg/kg, and the chrome was 0.433 mg/kg, respectivily.The thesis showed that Phaffia rhodozyma JMU-MVP14 not noly produce high concentration of astaxanthin, but also synthized fructooligosaccharide which was easy to separate during the astaxanthin fermentation. It built strong basis for further studies aiming to simutanous fermentation of astaxathin and fructooligosaccharide. |
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