Research On Coenzyme Q10 Traditional Extraction Process And Fermentation Production Of Coenzyme Q10 High-yield Strains Breeding

Background:Coenzyme Q10(CoQl0),also known as Ubiquinone,is a kind of similar to vitamin K structure of fat soluble quinone compounds,is one of the most important coenzyme in the cell.As an essential component of the respiratory chain of eukaryotic cells,it is an activator of cell metabolism and cell respiration.It has the functions of promoting the metabolism of the body,improving the non-specific immunity,scavenging free radicals,enhancing the antioxidant capacity,stabilizing the cell membrane structure,inhibiting the apoptosis,Anti-aging and protect the heart and other multiple effects,widely used in medical,dietary supplement,health care products,cosmetics and other fields.Related research is expected in 2015-2020 period,coenzyme Q10 market demand for compound annual growth rate will reach more than9%.China's development and utilization of coenzyme Q10 relatively late,its main consumer market in Europe and the United States.With the coenzyme Q10 health care effect is widely known by Chinese people,domestic demand began to surge.In this paper,we studied the process of extraction of coenzyme Q10 from fresh yak heart and the screening of coenzyme Q10 high-yield strain with heavy ion irradiation,in order to obtain the basic theoretical data to support China's large-scale production of coenzyme Q10.Methods:(1)The coenzyme Q10 was extracted from the heart of fresh yak by alkali saponification:The yak heart cells were sieved by acid heat.The single factor experiment was designed by saponification p H,saponification temperature,saponification liquid ratio and saponification time to determine the best level of single factor.The L₉(3⁴)orthogonal test was designed on the basis of single factor experiment.The optimal combination of coenzyme Q10 extracted from the best results and the optimal combination of coenzyme Q10 extracted from the orthogonal test results was used to verify whether the optimal combination obtained in the orthogonal experiment results was consistent with the actual result.(2)Breeding of Coenzyme Q10 High-yield Bacteria Using Heavy Ion Irradiation Rhodobacter sphaeroides:Using the heavy ion accelerator of Lanzhou Institute of Modern Physics,Chinese Academy of Sciences,⁵⁶Fe¹⁷⁺heavy ion beam irradiation was carried out on the prepared bacterium powder at 2Gy,4Gy,6Gy,8Gy and 16Gy respectively.After the irradiation,the initial screening.Then,the mutant strains were screened by reasoning on the resistant plates with the lowest inhibitory concentrations of vitamin K3,kanamycin,roxithromycin,p-hydroxybenzoic acid and sodium azide.Then,the strains screened by screening and screening factor were re-screened by shake flask fermentation,and the positive mutant strains of high coenzyme Q10 was selected.Finally,the high yield strain was subcultured and its genetic stability was determined.Results:(1)Extraction of coenzyme Q10 from fresh yak heart by alkali saponification method:The optimum saponification conditions determined in the saponification single factor experiment were:saponification p H 11,saponification ratio 1:15,saponification temperature 90?,saponification time 30 min.In the orthogonal experiment with the degree of freedom of 3,with the saponification p H,feed/liquid ratio,time and temperature of 4 factors,the optimal combination of the results was A₂B₁C₂D₃,the yield was 24.046 ug/g,the purity was 73.8%.The combination of orthogonal experiments is relatively high,indicating that the results of orthogonal experiments and actual match.(2)Breeding of Coenzyme Q10 High-yield Bacteria Using Heavy Ion Irradiation Rhodobacter sphaeroides:The lethal rate of ⁵⁶ Fe¹⁷⁺heavy ion irradiation was very high,and the lethality rate was proportional to the irradiation dose,and the lethality rate at 16Gy was 99.7%.⁵⁶ Fe¹⁷⁺heavy ion beam irradiation mutagenesis,combined with five screening factor screening,the highest yield mutant strains coenzyme Q10yield of 127.86 mg/L,which was 69.73%higher than that of the original strain.The mutant strain was cultured for 5 passages to verify that the genetic traits were stable.Conclusions:(1)It is feasible to extract coenzyme Q10 from fresh yak heart by alkali saponification method.The cost of raw materials used in the process is relatively low.For local enterprises with sufficient raw material source and low cost,this method can be used for small-scale production.(2)It is proved that the method of breeding the coenzyme Q10 high-yield strain by using heavy ion beam irradiation is possible.The technology can be used to screen the excellent strains which can be used for large-scale commercial production of coenzyme Q10.