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Patent Issued for Method for Producing Coenzyme Q10 by Fermentation Using Stock Culture from Solid Phase Fermentation

July 15, 2014



By a News Reporter-Staff News Editor at Life Science Weekly -- From Alexandria, Virginia, NewsRx journalists report that a patent by the inventors Chen, Jinqing (Xiamen, CN); Chen, Dajun (Xiamen, CN); Chen, Junhuang (Xiamen, CN); Wu, Meiqiong (Xiamen, CN); Su, Songgang (Xiamen, CN); Huang, Jianzhong (Xiamen, CN); Zheng, Yi (Xiamen, CN), filed on April 26, 2011, was published online on July 1, 2014 (see also Xiamen Kingdomway Group Company).

The patent's assignee for patent number 8765421 is Xiamen Kingdomway Group Company (Xiamen, Fujian, CN).

News editors obtained the following quote from the background information supplied by the inventors: "Coenzyme Q10 (2,3-dimethoxy-5-methyl-6-decaprenyl-1,4-benzoquinone, CoQ10), also named as ubiquinone or ubidecarenone, exists widely in animals, plants and microorganisms, and is an essential hydrogen carrier in respiratory chain of biological cells. Coenzyme Q10 is an important biochemical medicament, and recently has been widely applied in the treatment of various diseases such as cardiovascular diseases, diabetes, cancers, acute and chronic hepatitis, and Parkinson's Disease. In addition, coenzyme Q10 also exhibits a significant effect on the treatment of scurvy, duodenal ulcer and necrotic periodontitis, as well as on the enhancement of the function and secretion of pancreas. Meanwhile, coenzyme Q10 also has an anti-aging effect, and is widely applied in the fields of cosmetics and health products.

"There are mainly three kinds of methods for producing coenzyme Q10, namely, i) extraction from animal or plant tissues, ii) chemical synthesis, and iii) fermentation using microorganisms. In the method of extraction from animal or plant tissues, due to low content of animal coenzyme Q10, complicated chemical components, and the limitation of raw materials and sources, the product cost and price are high, and the industrial production is limited to some extent. Chemical synthesis methods are relatively mature technically, but the products are mixtures of cis-isomers and trans-isomers and have low biological activity, and the production cost is high. Therefore, chemical synthesis methods are limited in industrial production. Fermentation methods using microorganisms have become the most prospective methods for producing coenzyme Q10 for the following reasons: raw materials are cheap and abundant; processes for isolation of products are relatively simple; the products are natural products, without the problems generated by isomers; the products have good biological activity and are capable of being absorbed easily in human bodies; and the production in an industrial scale can be accomplished by using fermentation tanks.

"During fermentation using microorganisms, seed incubation is the first step for fermentation production, and the quality of seed liquid plays a key role in fermentation production. Seeds with good quality may shorten the fermentation period, stabilize yield and quality, and enhance utilization of devices. Therefore, seed incubation is essential for the production of coenzyme Q10 by fermentation. Currently, the general processes for producing coenzyme Q10 by fermentation using stock culture are as follows: culture in a slant medium.fwdarw.preparation of stock culture.fwdarw.primary enlargement culture.fwdarw.secondary enlargement culture.fwdarw.tertiary enlargement culture (if necessary).fwdarw.fermentation. In most of the processes, stock culture are cultured by liquid phase fermentation.

"Chinese Patent No. CN101519681, which is incorporated by reference, discloses a method for producing coenzyme Q10 by fermentation, wherein the quality of coenzyme Q10 produced thereby is maintained. In this method, the initial metabolic reaction is modulated in a feedback manner, depending on the synthetic rate and the content of the main side-product (i.e., 5-demethoxy coenzyme Q10) during metabolism of coenzyme Q10-producing bacteria, and has a rate-limiting effect on synthetic metabolism, so as to modulate the accumulation of the product of interest (i.e., coenzyme Q10) in cells in a feedback manner. By optimization of the concentration of the main substrates in the fermentation formulation and by optimized modulation of stirring rate during fermentation, the fermentation level of coenzyme Q10 is greatly enhanced, and the fermentation level of coenzyme Q10 can be enhanced to 3000 u/ml or more; and the determination of the optimized timing of discharging into the fermentation tanks improves time availability for producing coenzyme Q10 in cells, reduces fermentation cost, and greatly reduces the consumption of raw materials for fermentation."

As a supplement to the background information on this patent, NewsRx correspondents also obtained the inventors' summary information for this patent: "To overcome the disadvantages of fermentation production of coenzyme Q10 using stock culture in the prior art, an aspect of the invention provides a method for producing coenzyme Q10 by fermentation using stock culture from solid phase fermentation, which can improve productivity.

"The bacterial strain used in the invention is Rhodobacter sphaeroides strain JDW-610, which was deposited in China General Microbiological Culture Collection Center (CGMCC) on Dec. 21, 2010, with an accession number of CGMCC No. 4497.

"The method of the invention comprises the following steps:

"1) passaging the strain by culturing in a slant medium;

"2) preparing a solid medium, wherein the solid medium is steam-cooked and dried in air, subpackaged and sterilized, wherein said solid medium comprises solid components and liquid components; and

"3) culturing stock culture from solid phase fermentation, wherein a bacterial suspension is prepared by adding the culture of fresh Rhodobacter sphaeroides in the slant medium into sterile water, the suspension is added into the solid medium, and then cultured and used as stock culture for primary fermentation.

"In step 1), the slant medium may be comprised of: glucose 10 g/L, yeast extract paste 5 g/L, peptone 5 g/L, sodium chloride 5 g/L, ammonium sulfate 0.5 g/L, vitamin B.sub.1 1 .mu.g/L, vitamin K 1 .mu.g/L, vitamin A 1.5 .mu.g/L, Na.sub.2MoO.sub.4.2H.sub.2O 0.8 .mu.g/L, ZnSO.sub.4.7H.sub.2O 1.2 .mu.g/L, KNO.sub.3 0.33 .mu.g/L, NaBr 0.44 .mu.g/L, agar 20 g/L, pH 7.2; the culture condition may be: sterilization temperature of 121.degree. C., sterilization time of 25 min, culture at 30.degree. C. in dark for 24 h, stored at 4.degree. C. for further use, activated prior to use.

"In step 2), the formulation of the solid components may be comprised of: bran, rice, and millet in a ratio of 25:25:50 by mass; the formulation of the liquid components may be comprised of: glucose 10 g/L, yeast extract paste 5 g/L, peptone 5 g/L, sodium chloride 5 g/L, calcium chloride 2 g/L, ammonium sulfate 0.5 g/L, vitamin B.sub.1 1 .mu.g/L, vitamin K 1 .mu.g/L, vitamin A 1.5 .mu.g/L, CuSO.sub.2.5H.sub.2O 0.6 .mu.g/L, Na.sub.2MoO.sub.4.2H.sub.2O 0.8 .mu.g/L, ZnSO.sub.4.7H.sub.2O 1.2 .mu.g/L, KNO3 0.33 .mu.g/L, NaBr 0.44 .mu.g/L, pH 7.2; the ratio of the solid components and the liquid components may be 10:7 by mass; the temperature for steam cooking may be 80.degree. C., and the time for steam cooking may be 40 min. The solid medium may be subpackaged into 1000 mL kjeldahl flasks with 200 g per flask, and the sterilization may be carried out at a temperature of 121.degree. C. for 30 min.

"In step 3), the culturing may be carried out at 30.degree. C. in dark for 12 h, and continued for another 12 h after shaking homogeneously.

"An aspect of the present invention, a process of producing coenzyme Q10 using stock culture from solid phase fermentation is established according to the physiological characteristics of the coenzyme Q10-producing bacteria, with the original fermentation quality of coenzyme Q10 maintained. The process aims at simplifying seed process and enhancing productivity of coenzyme Q10, by carrying out a large number of experiments to optimize the seed process. By optimization of the conditions for seed culture, the invention provides a method for producing coenzyme Q10 by fermentation using stock culture from solid phase fermentation, which finally affects the fermentation and enhances productivity.

"The invention employs, for the first time, a new fermentation method of culturing stock culture from solid phase fermentation in the seed incubation process of coenzyme Q10 fermentation, wherein solid materials such as bran, rice and millet are used in place of agar powder to prepare a solid medium. As solid supporters, bran, rice, and millet provide a larger surface area than agar plate, and thus they can provide more seeds in a limited volume. In addition, these solid raw materials comprise a variety of growth factors. Particularly, bran and millet are more abundant in trace nutrients. Therefore, these solid materials are very suitable for seed culture. Meanwhile, stock culture from solid phase fermentation are more convenient for storage and are long-lasting, which is convenient for scheduling fermentation production.

"It can be seen that, as compared to liquid phase fermentation, the invention has the following advantages: {circle around (1)} bacterial cells have strong growth activity and good synchronism, can grow quickly after being transferred to a fermentation tank with a short lag phase; {circle around (2)} bacterial cells are in a stable physiological state and retain stable productivity; and {circle around (3)} the seed incubation process is simplified and the fermentation process is more readily scheduled.

"The invention establishes a method for producing coenzyme Q10 by fermentation using stock culture from solid phase fermentation for the first time. The method is applicable in middle-scale and large-scale fermentation, and achieves desirable fermentation effects."

For additional information on this patent, see: Chen, Jinqing; Chen, Dajun; Chen, Junhuang; Wu, Meiqiong; Su, Songgang; Huang, Jianzhong; Zheng, Yi. Method for Producing Coenzyme Q10 by Fermentation Using Stock Culture from Solid Phase Fermentation. U.S. Patent Number 8765421, filed April 26, 2011, and published online on July 1, 2014. Patent URL: http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO1&Sect2=HITOFF&d=PALL&p=1&u=%2Fnetahtml%2FPTO%2Fsrchnum.htm&r=1&f=G&l=50&s1=8765421.PN.&OS=PN/8765421RS=PN/8765421

Keywords for this news article include: Chemicals, Chemistry, Sterilization, Sodium Chloride, Ammonium Sulfate, Rhodobacteraceae, Infection Control, Gram-Negative Bacteria, Rhodobacter sphaeroides, Xiamen Kingdomway Group Company.

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Source: Life Science Weekly


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