| Erythritol is a new kind of sweetener which has many advantages such as pure sweet taste, extremely low heat, high stability, highly tolerated by the intestinal, and prevention to dental caries. It has a broad prospect in the food industry as low energy food additive, and can be also well used in medical industry and chemical industry. An erythritol producing microorganism strain from a bee honey sample has been identified as Moniliella sp. in a previous study in our laboratory. One purpose of this study is to optimize the fermentation medium and conditions for the strain; the other purpose is to establish the separating and purifying method of erythritol from the fermentation broth.In the optimizing experiments, a 48 hours culture was determined as inolulum seeds according to the growth curve of Moniliella sp. BH010. Then fermentation factors such as carbon sources, nitrogen source, inorganic salt, NaCl concentration, initial pH value, inoculums, temperature, and fermentation time were studied by single-factor experiment. The results indicated that the best carbon source of Moniliella sp was glucose with the optimal concentration of 35 %, and the best nitrogen source was yeast extract with the optimal concentration of 1%. The addition of some mineral salt to the media such as KH2PO4, MgSO4 and CaCl2 increased the production of erythritol. The optimal initial pH was found to be 6.0, inoculum amount 5 %, fermentation temperature 30℃, and cultivate time 9 days. Then the orthogonal experiments of medium component and fermentation condition were further applied to find the optimal fermentation conditions. According to the result analyzed by SPSS , the optimal fermentation medium consisted of 30 % glucose, 1 % yeast extract, 0.2 % CaCl2. The optimal fermentation conditions were: a 48 h culture as inoculation seeds, inoculum amount 1 %, initial pH 6.0, culture temperature 30℃, and culture time 9 days. The yield of erythritol could reach 110.61 g/L under these conditions.In the experiment of separation and purification of erythritol, cells and mycelium in the broth were removed by deactivating enzyme and centrifuged. Discoloration of the broth was carried out by using activated carbon powder and the discoloring rate could reach 97.38 %.After the entraps of the large proteins and left minute carbon power by ultra filtration membrane, ions-exchange resins was used to remove the metal mineral salt, pigment, organic substances and so on. The quantity of sample was 5 mL, eluting speed 1.0 mL/min, and the collecting time 20 to 65 minues. According to the component analysis of effluent, 92.48 % of dry substance was erythritol, and the impurities were almost removed. The effluent then was concentrated by rotary evaporation to the erythritol concentration more than 60 g/100 mL. 0.05 % crystal nucleus were added while the concentrate was cooling. The concentrate stood at 10℃for 24 hours, and then the crystals formed. Crystal and bottom liquid were separated by centrifugalization, the bottom liquid was recycled and re-crystallized twice and crystals were collected and dried at 45℃until the quantity was stable. The recycle rate was calculate by quantifying the quantity of the total erythritol crystal, and the result was 74.35 % .The purity of erythritol crystal was 97.62 % evaluated by the HPLC measurement result.With single-factor experiments and orthogonal experiments, the study illustrated the optimal fermentation condition for a erythritol-producing Moniliella strain. It was found that Moniliella sp. BH010 has a high production of erythritol under simple conditions. A primary method to purify erythritol from the fermentation broth was also established. This research provided theoretical basis to the industrial production of erythritol. |
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