Screening Of Strains With High Yield Of Hdmf And Optimization Of The Fermentation Conditions

4-hydroxy-2,5-dimethyl-3(2H)-furanone(HDMF) which widely exists in nature products, is an aroma compound exhibiting pineapple, strawberry-like flavor in dilute solutions and caramel-like flavor in concentrates. HDMF has great commercial value as an excellent flavoring agent. Presently, HDMF is widely used in many food products, such as bread, candy, meat, milk, beverages and wine. It is the future trend to produce HDMF by biotechnology because of its pleasant note, resourse conservation and high economic returns. This article was to screen the strains with higher yields of HDMF, exploring the bio-technologies in producing HDMF by mutagenesis and optimization of fermentation conditions. The main contents are as follow.Spread plate method was used to separate yeasts strain from fruits, such as pineapple, strawberry, apple; HPLC was used to determine the yield of HDMF by yeasts. Of the 46 wild yeast strains and 13 kwon strains reserved in the lab, 13 strains had the ability of producing HDMF, of which 4 strains had higher yields, they were C5(isolated from strawberry), P3(from apple), Zygosaccharomyces rouxii , Saccharomycodes ludwigii. P3 was identified as Pichia capsulata and C5 was Hanseniaspora sp by molecular biological identification and the traditional identification method.6 kinds of sugar, including rhamnose, fructose, sucrose, were used as the precursors producing of HDMF, formulating medium to observe the situations of the 4 stains in exploiting sugar. The results showed that rhamnose, fructose, maltose were the effective precursor of the 4 stains in producing HDMF. P3 strain had the best effect in producing HDMF using fructose; it had the highest yield, 10.96mg/L, when the fructose concentration in medium was 120g/L. Therefore, P3 was selected as the original strain.The strains P3-UV2-13 and P3-UV2-17 which were after the original strain P3 being treated by DES had higher yields of HDMF, then were used as original strains for further treating by 1% DES (v/v), and one strain had genetic stability while with higher yield of HDMF was obtained, named P3-UV-DES-12, which had stability in passaging 5 generations, and the yield of HDMF was 2.12 times of that of the original strain. Response surface analysis was used to optimize the fermentation conditions, components of medium determined finally were fructose 141.65g/L, casein peptone 15mg/L, K₂HPO₄ 4 g/L, MgSO₄.7H₂O 5g/L, initial pH 3.5; the best culture conditions were liquid volume 10/100mL, seed age 18h, inoculation 3%(vol/vol), rotatory shaker at 150 r/min, culture temperature 35℃, fermentation for 144h, the yield of HDMF was up to 39.43mg/L.