Screening Of Fungi With PUFAs And Study On Nutritional Physiology

Polyunsaturated fatty acids (PUFAs) have very important physiological functions and are generally obtained from natural plants and deep-sea fishes. However, the production cannot satisfy the market demands. In recent years, microorganism fermentation to produce PUFAs gradually becomes a more and more popular method. A method analysing PUFAs in mycelium rapidly and accurately is established: mycelium is dried in high temperature,the petroleum ether and aether are extraction solvents,the extraction pH is about 5.0,the optimal extraction combination is 5.0g dried mycelium/100mL petroleum ether,the extraction time is 16 hours and the operational temperature is under 50℃. Prelimimary screening with microscope and plate culture, and re-screening in liquid media, a strain of fungi was obtained, numbered AG, and was identified as Monoblepharis polymorpha Cornu. Radiated at wavelength of 532nm under 10mw for 10 minutes using PIV400 Nd:YAG laser, a highly productive mutate fungi was selected, named AGm59. Its targeted characteristics are improved quite a lot. After six days'culturing in potato media, the biomass, lipid%, GLA, AA, EPA, DHA are 8g/L, 53.4%, 343.1mg/L, 1209.0mg/L, 125.1mg/L, and 119.4mg/L, respectively. In other words it is 2.5 times, 9.7 times, 18.0 times, and 3.8 times of tha of the control strains. Therefore, laser mutation is an effective method to obtain high quantity of PUFAs especially with the ω-3 series. By studying the nutritional physiology of AGm59,the results indicate:the mixed carbon sources of glucose and sucrose were the best carbon sources,and KNO₃ is the best nitrogen source; the pH(neutral) in which Fungi grows healthily is not consistent the optimal fermentation pH(5.5); the quantities of PUFAs increase by adding Fe²⁺; the inoculation days of producing different PUFAs are not steady; and its growth periods is long,attaining 20 days.The following optimal fermentation conditions are obtained:The fungi was maintained on slant tubes of 2% glucose and 3% agar for 20 days; the inoculation culture was created by adding some of culture grown on PDA to 200mL of 2% glucose and shaking at 25℃for 4 days ( 150r/min ) ; the culture medium of fermentation was created by adding glucose(54.0g/L), sucrose(6.0g/L), KNO3(3.198g/L) and FeSO4.7H2O(0.2-0.4g/L); the culture medium was adjusted to pH 5.5 with 0.2mol/L HCl; the biomass of Monoblepharis polymorpha Cornu is up to17.25g/L ,the lipid component of the biomass is 57.7% and the total quantity of PUFAs is up to 7670.8mg/L in 10-12 days'cultivation at 25℃.The quantity of the GLA, AA, EPA, and DHA in the lipid is 674.9mg/L, 3330.6mg/L , 327.6mg/L , 352.3mg/L.