A Research On The Production Of Kojic Acid By Aspergillus Oryzae Strain Breeding, Fermentation Condition And Bacteriostasis

kojic acid is a weakly acidic organic acid which is produced by some aerobic eumycetes. It is antibacterial, antioxidant, non-toxic, color-protecting and inhibiting polyphenol oxidase, which allows the kojic acid to be widely used in food, cosmetic, medicine, agriculture and other fields. As applications in market continue to expand, particularly in food additives and cosmetics, kojic acid will maintain its favorable position in price and demand.A kojic acid producing strain is isolated from a naturally fallen and moldy orange peel. After analyzing the cultural characteristics, micro-morphology and typical conidial structure, it is identified as Aspergillus oryzae and named as LYJ-1. A genetically stable mutant strain UVDH-14is obtained by compound mutation of UV, DES and MW, with Aspergillus oryzae LYJ-1as the parent strain. By liquid-state fermentation, the kojic acid production of the UVDH-14reached54.20g/L, which is2.67times as that of the parent strain.Using the single factor experiments, the effects from different fermentation conditions on kojic acid production of the mutant strain UVDH-14were studied, such as carbon sources, nitrogen sources, alcohol adding, initial pH, fermentation temperature, liquid volume, inoculation amount, seed age, fermentation time. On the basis of previous results, the major fermentation conditions are identified and optimized by response surface method, a quadratic regression equation model of initial pH value, fermentation temperature, liquid loading and fermentation time is established. The results prove to be highly reliable, where the model could explain90.51%of the variation in the response value and Adj R-Squared is0.905, Adeq precision is15.783, C.V is1.50%. The results showed that the optimal fermentation medium are as follows:corn starch5%, soybean-cake powder0.5%, absolute alcohol4%, yeast extract0.2%, K2HPO40.1%, MgSO4·7H2O0.05%, KCl0.05%, and the optimum fermentation conditions are:initial pH5.4, fermentation temperature35℃, inoculation amount10%(V/V), fermentation time9.5d, liquid loading92mL/250mL for triangle flask. Under these conditions, the kojic acid average production is93.51g/L, with the theoretical expectation of93.8685g/L, and the relative error is only0.382%. The model is proved to be capable of predicting the actual kojic acid production of the mutant strain UWDH-14, which is of great significance in industry.The extraction process is studied and the decolorizing agent and decoloring technique are selected. The bacteriostasis of kojic acid is preliminarily studied. The results, with activated clay as decolorizing agent, show that the decolorizing rate is97.26%and the loss rate of kojic acid is22.93%under the following conditions: decolorizing temperature60℃, decolorizing time1.5h and natural pH. The antibacterial effect is studied, using the oxford cup method and agar dilution method. The results show that the inhibiting effect of kojic acid is better on bacterial, but could not inhibit Candida utilis and Saccharomyces cerevisia. Kojic acid has an inhibitory action on phytopathogenic eumycetes, especially Alternaria longipes, with the inhibition ratio of65.34%.