CN113355375A - Method for synthesizing B vitamins by fermenting lactobacillus paracasei K56 and application - Google Patents

Method for synthesizing B vitamins by fermenting lactobacillus paracasei K56 and application Download PDF

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CN113355375A
CN113355375A CN202011355562.XA CN202011355562A CN113355375A CN 113355375 A CN113355375 A CN 113355375A CN 202011355562 A CN202011355562 A CN 202011355562A CN 113355375 A CN113355375 A CN 113355375A
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vitamin
fermentation
vitamins
lactobacillus paracasei
milk
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CN113355375B (en
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洪维鍊
赵雯
刘伟贤
侯保朝
马国文
吴超
倪丹
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Inner Mongolia Yili Industrial Group Co Ltd
Inner Mongolia Dairy Technology Research Institute Co Ltd
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Inner Mongolia Yili Industrial Group Co Ltd
Inner Mongolia Dairy Technology Research Institute Co Ltd
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
    • C12P17/16—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms containing two or more hetero rings
    • C12P17/167—Heterorings having sulfur atoms as ring heteroatoms, e.g. vitamin B1, thiamine nucleus and open chain analogs
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
    • A23C9/123—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt
    • A23C9/1234—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt characterised by using a Lactobacillus sp. other than Lactobacillus Bulgaricus, including Bificlobacterium sp.
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    • A23L2/00—Non-alcoholic beverages; Dry compositions or concentrates therefor; Their preparation
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    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/135—Bacteria or derivatives thereof, e.g. probiotics
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    • A61K31/00—Medicinal preparations containing organic active ingredients
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    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/4415—Pyridoxine, i.e. Vitamin B6
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/455—Nicotinic acids, e.g. niacin; Derivatives thereof, e.g. esters, amides
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    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/506—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim not condensed and containing further heterocyclic rings
    • A61K31/51—Thiamines, e.g. vitamin B1
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    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
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    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • A61K31/525—Isoalloxazines, e.g. riboflavins, vitamin B2
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/67—Vitamins
    • A61K8/673—Vitamin B group
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/67—Vitamins
    • A61K8/673—Vitamin B group
    • A61K8/675—Vitamin B3 or vitamin B3 active, e.g. nicotinamide, nicotinic acid, nicotinyl aldehyde
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
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    • C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
    • C12P17/10—Nitrogen as only ring hetero atom
    • C12P17/12—Nitrogen as only ring hetero atom containing a six-membered hetero ring
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    • C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
    • C12P17/18—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms containing at least two hetero rings condensed among themselves or condensed with a common carbocyclic ring system, e.g. rifamycin
    • C12P17/182—Heterocyclic compounds containing nitrogen atoms as the only ring heteroatoms in the condensed system
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    • C12P25/00—Preparation of compounds containing alloxazine or isoalloxazine nucleus, e.g. riboflavin
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    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
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    • A61K2800/10—General cosmetic use

Abstract

The invention provides a method for synthesizing B vitamins by fermenting lactobacillus paracasei K56 and application thereof. The invention firstly provides a method for synthesizing B vitamins by fermenting Lactobacillus paracasei (Lactobacillus paracasei subsp. paracasei) K56, which comprises the following steps: the fermentation product containing B vitamins is prepared by fermenting lactobacillus paracasei CGMCC No.15139 or DSM 27447. The research of the invention finds that the lactobacillus paracasei strain can be used for the production and application of foods, daily chemicals, feeds or medicines and the like rich in B vitamins.

Description

Method for synthesizing B vitamins by fermenting lactobacillus paracasei K56 and application
Technical Field
The invention relates to the technical field of microbial fermentation, in particular to a method for synthesizing B vitamins by fermenting Lactobacillus paracasei (Lactobacillus paracasei subsp. paracasei) K56 (with the preservation number of CGMCC No.15139 or DSM27447) and related application.
Background
The B vitamins are also called B vitamins (also called vitamin B, vitamin B group or vitamin B complex group), and are classified as "families" because of 8 types in total. The B vitamins are water-soluble vitamins which are indispensable for maintaining the normal functions and metabolic activities of human bodies, and can be quickly consumed in the human bodies in the tense life and working pressure, improper dietary habits or due to the use of certain specific medicines and the property of dissolving the B vitamins in water. The supplement of vitamin B can relieve emotion. Moreover, according to research reports, the B vitamins can help to maintain the functions of the heart and the nervous system, maintain the health of the digestive system and the skin, participate in energy metabolism, enhance physical strength, tonify and strengthen the body. If vitamin B is absent, cell function is immediately reduced, and metabolic disorder is caused, at which the human body suffers from malaise and inappetence. Furthermore, excessive drinking and the like cause liver damage, which is concurrent with vitamin B deficiency in many cases.
The B vitamins are widely present in rice bran, yeast, animal liver, coarse grain vegetables and other foods. Studies have shown that part of the B vitamins can be synthesized in the human gut.
Lactobacillus paracasei is a gram-positive facultative heterotrophic lactic acid bacterium commonly used in dairy fermentation and probiotics. Lactobacillus paracasei is commonly found in a variety of human habitats, such as the human intestinal tract and oral cavity, as well as silage and naturally fermented foods.
No relevant research report about the B vitamins produced by the fermentation of lactobacillus paracasei is found.
Disclosure of Invention
One object of the present invention is to provide a novel use of lactobacillus paracasei K56.
Lactobacillus paracasei (Lactobacillus paracasei subsp. paracasei) K56 strain has been deposited at the German Collection of Microorganisms and Cell Cultures (German Collection of Microorganisms and Cell Cultures) on day 27 6 month 2013 under accession number DSM 27447; in addition, Lactobacillus paracasei (Lactobacillus paracasei subsp. paracasei) K56 strain has also been stored in China General Microbiological Culture Collection Center (CGMCC) with the Collection number of CGMCC No.15139 in 29.12 months and 29.2017.
The invention discovers that the Lactobacillus paracasei (Lactobacillus paracasei subsp. paracasei) K56 takes feed liquid (milk substrate) mainly containing milk components as a fermentation substrate, and the fermentation product has high-yield B vitamins (containing vitamin B)1Vitamin B2Vitamin B3Vitamin B6Vitamin B9) The ability of the cell to perform.
Thus, in one aspect, the present invention provides a method for the fermentative synthesis of B vitamins from Lactobacillus paracasei (Lactobacillus paracasei subsp. paracasei), comprising: the fermentation product containing B vitamins is prepared by fermenting lactobacillus paracasei CGMCC No.15139 or DSM 27447.
According to a particular embodiment of the invention, the method of the invention further comprises: extracting and purifying B vitamins from the supernatant of the fermentation product.
According to a specific embodiment of the present invention, the B vitamins include vitamin B1Vitamin B2Vitamin B3Vitamin B6And vitamin B9One or more of (a).
According to a specific embodiment of the present invention, the B vitamins include vitamin B1Vitamin B3And/or vitamin B6。
According to a particular embodiment of the invention, the fermentation substrate used in the fermentation may be an emulsion. Preferably, the milk may be raw milk or reconstituted milk, and may be whole milk, low fat milk or skim milk. The emulsion may optionally contain 5% to 10% sucrose.
According to a specific embodiment of the present invention, the fermentation conditions are: fermenting for 1-7 days at 35-45 ℃.
According to a specific embodiment of the invention, the lactobacillus paracasei is inoculated in the fermentation substrate in the form of seed liquid, with an inoculation amount of 2% to 5%. The preparation of the seed liquid may be carried out according to conventional techniques in the field of probiotics.
According to the specific embodiment of the invention, the milk substrate used as the fermentation substrate of lactobacillus paracasei K56 in the invention can be milk (whole milk, low-fat milk or skim milk) with milk content of more than 80%, wherein auxiliary materials such as sweetening agents, stabilizing agents and the like can be selectively added according to requirements, such as fermentation substrates commonly used in the preparation process of fermented milk or milk beverages, the milk substrate fermented by lactobacillus paracasei K56 can achieve the effect of high-yield B vitamins, and the fermentation product is particularly rich in vitamin B1Vitamin B2Vitamin B3Vitamin B6Vitamin B9。
Vitamin B1The content in milk is 0.3 mg/L on average. Vitamin B in milk1The lactobacillus fermented sour milk is not completely from feed, and can be synthesized by microorganisms in rumen of cow, so that vitamin B in sour milk fermented by lactobacillus is more than that in common milk1The content is increased by about 30 percent. Children less than one week in age need vitamin B daily15-10 mg. If vitamin B is deficient1It can cause anorexia, slow growth, damage to nervous system, polyneuritis, milk regurgitation or constipation, dyspepsia, green stool, pale complexion, inappetence, muscle relaxation, frequent crying, and weight loss in children. According to some embodiments of the invention, lactobacillus paracasei K56 ferments vitamin B in the emulsion1The content can reach 30-70 mu g/mL. The yogurt product fermented by K56 is taken 100 mL/day, and can be supplemented with VB for recommended human intake1。
Vitamin B2In human milk, the content of vitamin B in cow milk is not high2The content can reach 1-2 mg/L. The children are recommended to take 21-3 mg vitamin B every day. If vitamin B2 is lacked, it will cause growth retardation, anorexia, inflammation of mouth horn and lip, red and swollen tongue, malnutrition, and asthenia. According to some embodiments of the invention, lactobacillus paracasei K56 ferments vitamin B in the emulsion2The content can reach 2.8-3 mu g/mL.
Vitamin B3(nicotinic acid) is recommended to be taken by adults at 13-19mg per day (dietary guidelines of Chinese residents), 20mg per day is recommended for pregnant women, and 22mg per day is recommended for pregnant women in lactation. Vitamin B3It is mainly used in livestock meat, fish and poultry, and plant food such as mushroom, asparagus, sunflower seed, peanut, bean and grain. Vitamin B in ordinary yoghurt3Is present in an amount of about 0.2mg/100 g. According to some embodiments of the invention, lactobacillus paracasei K56 ferments vitamin B in the emulsion3Is 50-80 mu g/mL and is far higher than the content of the common yoghourt.
Vitamin B6Also known as pyridoxine, which includes pyridoxine, pyridoxal and pyridoxamine, exists in vivo in the form of phosphate esters, and is a water-soluble vitamin. Vitamin B6The functionality involves a number of aspects including: participating in protein synthesis and catabolism and participating in all amino acid metabolism; participating in gluconeogenesis and UFA metabolism, and is related to glycogen metabolism, sphingomyelin metabolism and steroid metabolism; participate in the synthesis of certain nerve mediators (5-hydroxytryptamine, taurine, dopamine, norepinephrine and gamma-aminobutyric acid); involving nucleic acid and DNA synthesis, lacking vitamin B6DNA synthesis is impaired, a process which is important for maintaining proper immune function. Papastoyianidis et al (2006) use commercial ferments YO-1, YO-2 to ferment at 42 ℃ for 3.5h, then store for 1.5h in cold storage, detect vitamin B6The content of (A) is 1.37mg/kg and 1.50mg/kg respectively; vitamin B after fermentation assisted by Bifidobacterium BB-12 and Lactobacillus acidophilus LA-56The contents of (A) were 1.37mg/kg and 1.31mg/kg, respectively. According to some embodiments of the invention, lactobacillus paracasei K56 ferments vitamin B in the emulsion6The content is 8-20 mu g/mL.
Vitamin B9The content of the (folic acid) in the cow milk is generally 40 +/-10.00 mu g/L, and the content of the (folic acid) in the common yoghurt is 80 +/-20.00 mu g/L. Vitamin B9The main effects on organisms are as follows: are involved in the metabolism of genetic material and proteins; influence animal reproductive performance; affecting secretion of the animal pancreas; promoting the growth of animals; improving immunity of organism. Vitamin B9As one of the essential vitamins for the growth and reproduction of body cells, the deficiency can affect the normal physiological activities of human bodies. Many reports in the literature of vitamin B deficiency9Has direct relation with diseases such as neural tube deformity, megaloblastic anemia, cleft lip and palate, depression, tumor and the like. In general, normal persons require vitamin B9The amount of the compound is 100-200 mu g/d, and the recommended amount of the world health organization is as follows: adult 200 ug/d, pregnant woman and lactating mother 400 ug/d. The latest folic acid consumption standard of the FDA is as follows: between 25 and 50 years old (male) 240 μ g/d; 190 μ g/d between 25 and 50 years old (female); the concentration of the extract is 400 mug/d for the nannie and the pregnant woman; the infant is 200-400 mu g/d. According to some embodiments of the invention, lactobacillus paracasei K56 ferments vitamin B in the emulsion9The content of (b) is 1.6-1.8 mu g/mL.
Thus, in another aspect, the invention also provides the use of lactobacillus paracasei for the fermentative preparation of a composition rich in B vitamins, wherein the fermentation product rich in B vitamins is prepared by fermentation using lactobacillus paracasei CGMCC No.15139 or DSM 27447.
According to a particular embodiment of the invention, the lactobacillus paracasei of the invention is used in the fermentative preparation of a composition enriched in B vitamins, which may be a food composition, a daily chemical composition or a pharmaceutical composition. Specifically, the fermentation product obtained by fermenting the milk substrate with lactobacillus paracasei K56 of the present invention can be used directly as food (e.g., fermented milk, milk drink, milk powder, solid drink, etc.), daily chemical (cosmetics, etc.), feed (including pet food or probiotic products for livestock), or medicine, etc., or further used for preparing such food, daily chemical, feed, or medicine. The process for preparing food, daily chemical, feed or pharmaceutical products from the fermentation product can be carried out with reference to the prior art in the field.
In a particular embodiment of the invention, the B vitamin containing fermentation product of the invention is used as or further for the preparation of a food product, preferably a fermented milk or fermented milk beverage. The emulsion used as a fermentation substrate may further comprise other materials commonly used in fermented milk or fermented milk beverages, such as appropriate amounts of sweeteners, stabilizers, nutritional additives, and the like.
According to a particular embodiment of the invention, the fermented milk matrix of lactobacillus paracasei of the invention results in a fermentation product containing B vitamins which is also enriched in conjugated linoleic acid and/or gamma-aminobutyric acid. Therefore, the invention also provides the application of the lactobacillus paracasei K56 in the preparation of a composition rich in conjugated linoleic acid and/or gamma-aminobutyric acid through fermentation. In the composition, conjugated linoleic acid and/or gamma-aminobutyric acid are mainly produced by fermentation of the lactobacillus paracasei.
In conclusion, the invention provides the new application of the lactobacillus paracasei K56, and the invention discovers that the single-strain fermented milk substrate of the strain has good vitamin B inclusion synthesized by vitamin B1Vitamin B2Vitamin B3Vitamin B6Vitamin B9The ability of the cell to perform. The lactobacillus paracasei K56 strain can be used for production and application of foods, daily chemicals, feeds or medicines rich in B vitamins.
Detailed Description
For a more clear understanding of the technical features, objects and advantages of the present invention, reference is now made to the following detailed description taken in conjunction with the accompanying specific embodiments, and the technical solutions of the present invention are described, it being understood that these examples are intended to illustrate the present invention and are not intended to limit the scope of the present invention. In the examples, each raw reagent material is commercially available, and the experimental method not specifying the specific conditions is a conventional method and a conventional condition well known in the art, or a condition recommended by an instrument manufacturer.
Example 1: production of B vitamins from Lactobacillus paracasei fermented milk
1. Experimental strains
The lactobacillus paracasei K56(CGMCC No.15139) used in the technical proposal of the invention;
control strain 1: lactobacillus paracasei strain l.casei 431;
control strain 2: the commercial strain Lactobacillus paracasei LPC-37.
2. Activation of strains and liquid culture of seeds
(1) Strain activation: activating lactobacillus paracasei by using MRS culture medium, taking the strain growing to logarithmic phase, and preserving glycerol tubes for later use. Wherein, MRS culture medium (g/L): peptone 10.0 g; 5.0g of yeast extract; 20.0g of glucose; 5.0g of anhydrous sodium acetate; 2.0g of dihydroamine citrate; tween-801.0 mL; dipotassium phosphate 2.0 g; magnesium sulfate heptahydrate 0.2 g; 0.05g of manganese sulfate; 10.0g of beef extract; 15g to 20g of agar.
(2) Determination of growth dynamics of strains
Dissolving 12% whole milk powder and 6% white sugar in water at normal temperature, sterilizing at 95 deg.C for 5min, cooling to 36 + -1 deg.C, inoculating strain as milk matrix, fermenting and culturing for 120h, sampling at different fermentation time periods, and measuring viable count and pH value.
And (3) viable count detection: after mixing 25ml of the sample with 75ml of physiological saline uniformly, counting by a plate dilution method.
And (3) acidity detection: the sample pH was measured using a pH meter and recorded. Taking 10g of the fermented dairy product, adding 2 drops of phenolphthalein solution, mixing uniformly, and titrating 0.1M standard NaOH solution to pink. The calculation formula of the titration acidity is as follows:
°T＝C×V×100/(m×0.1)。
the results are shown in tables 1 and 2, respectively.
TABLE 1 colony count of Lactobacillus paracasei in milk matrix for different fermentation times
Figure BDA0002802513100000051
Figure BDA0002802513100000061
From the analysis of the test data, the pH value of K56 fermentation is rapidly increased at 28h, and then is reduced to reach a stable value. The viable count of the control strain 1 and the control strain 2 is increased rapidly after 36 hours, and then the viable count is reduced to be stable. K56 was not significantly different from control strain 1, control strain 2 by p > 0.05; control strain 1 and control strain 2 analyzed p >0.05 with no significant difference. Wherein the blank group is an emulsion matrix without added bacteria liquid.
TABLE 2 pH determination of Lactobacillus paracasei in milk substrates for different fermentation times
Figure BDA0002802513100000062
Figure BDA0002802513100000071
From the analysis of the above test data, K56, control strain 1 and control strain 2 gradually decreased within 12 to 36 hours. K56 was very significantly different from control strain 2 by p < 0.01; k56 was no significantly different from control strain 1 for p > 0.05; control strain 1 and control strain 2 analyzed p >0.05 with no significant difference. The presence of non-killed micro-organisms in the blank resulted in milk spoilage and a decrease in pH due to temperature. Wherein the blank group is an emulsion matrix without added bacteria liquid.
(3) Preparation of strain seed liquid: dissolving 12% whole milk powder and 6% white sugar in water at normal temperature, sterilizing at 95 deg.C for 5min, cooling to 36 + -1 deg.C, inoculating strain, fermenting at 36 + -1 deg.C for 48 + -2 h, and prolonging to 72 + -2 h to obtain seed liquid.
3. Synthesis of B vitamins from lactobacillus paracasei fermented milk
Preparing fermented milk: adding sucrose accounting for 6.5% of the total weight of raw milk into raw milk, stirring, heating to 95 ℃, sterilizing for 5min, cooling the sterilized emulsion to 37 ℃, respectively adding various bacteria seed liquids (the inoculation amount of lactobacillus paracasei K56 is 3%, the inoculation amount of control bacteria is equivalent to K56 according to the number of viable bacteria), respectively fermenting each sample at 37 ℃ for 24, 48, 72, 96 and 120h, centrifuging the fermented emulsion at 8000r/min for 15min, collecting fermented supernatant, filtering with a 0.22 mu m filter membrane, and measuring the content of B vitamins in the filtrate.
The content of B vitamins is determined by high performance liquid chromatography, and the specific operation conditions refer to HPLC detection of water-soluble vitamins (Liyan, food research and development, 2018) in Collybia nigricans.
Blank milk base (without added probiotics) was used as a control, and blank control supernatant could not be obtained by centrifugation due to the inability to curd during the same fermentation time. Therefore, the following data are the vitamin contents directly measured in the supernatant of the fermentation broth, without deducting the vitamin contents in the blank milk matrix.
(1) Vitamin B in fermentation samples1In an amount of
Vitamin B in probiotic fermentation supernatant with different fermentation times1See table 3 for the contents of (d).
TABLE 3 fermentation supernatant vitamin B1In an amount of
Figure BDA0002802513100000081
From the analysis of the above test data, it can be seen that vitamin B is present in the fermentation supernatant of each strain1The content showed a clear rising trend with time. Differences were found, but not significant, between K56, control strain 1, and control strain 2.
(2) Fermentation sample vitamin B2In an amount of
Vitamin B in probiotic fermentation supernatant with different fermentation times2See table 4 for the contents of (d).
TABLE 4 fermentation supernatant vitamin B2In an amount of
Figure BDA0002802513100000082
From the analysis of the above test data, it can be seen that vitamin B in the supernatant of the fermentation of each strain was observed at different times2The contents are all close and the difference is not obvious. Wherein vitamin B in the supernatant in the first 72h2The content shows a clear ascending trend, and the content is basically unchanged after 48 h.
(3) Fermentation sample vitamin B3In an amount of
Vitamin B in probiotic fermentation supernatant with different fermentation times3See table 5 for the contents of (d).
TABLE 5 vitamin B in fermentation supernatants3In an amount of
Figure BDA0002802513100000083
From the analysis of the above test data, it can be seen that vitamin B is present in the K56120 h fermentation supernatant3The content was about 1.4 times that of the control strain 2. Vitamin B in fermentation supernatant of each strain3The content showed a clear rising trend with time, the rising trend of the control strain 2 was relatively slow. Vitamin B in supernatant from fermentation of K56 and control Strain 1 for 120h3The content difference is obvious (p is less than 0.05). Vitamin B in fermentation supernatants of K56 and control Strain 23The content difference is very obvious (p is less than 0.01).
(4) Fermentation sample vitamin B6In an amount of
Vitamin B in probiotic fermentation supernatant with different fermentation times6See table 6 for the contents of (d).
TABLE 6 vitamin B in fermentation supernatants6In an amount of
Figure BDA0002802513100000091
From the analysis of the above test data, it can be seen that vitamin B is present in the K56120 h fermentation supernatant6The content was about 1.4 times that of the control strain 2. Vitamin B in fermentation supernatant of each strain6The content showed a clear rising trend with time, the rising trend of the control strain 2 was relatively slow. Vitamin B in supernatant after 72h fermentation of K56 and control Strain 16The content difference is obvious (p is less than 0.05). Vitamin B in supernatant after 72h fermentation of K56 and control Strain 26The content difference is obvious (p is less than 0.05).
(5) Fermentation sample vitamin B9In an amount of
Vitamin B in probiotic fermentation supernatant with different fermentation times9See table 7 for the contents of (d).
TABLE 7 vitamin B9In an amount of
Figure BDA0002802513100000092
From the analysis of the above test data, it can be seen that vitamin B is present in the fermentation supernatant of each strain9The content of vitamin B is substantially constant with time9The contents were all at a lower level.
According to the experimental result of the viable count of each strain, the lactobacillus paracasei reaches the maximum value of the viable count within about 48 hours, namely the stable period, and the viable counts are kept unchanged after 48 hours. After 24h fermentation, fermentation supernatant can not be obtained to detect the content of folic acid, but folic acid is supposed to be metabolized and generated in the process of strain growth. There was no significant difference in the folate content over time during the subsequent fermentation time, a possible reason: the folic acid content reaches the maximum value in the stable growth period (48h) of the strain, and as the strain is continuously metabolized, the thalli age and the folic acid synthesis amount is reduced; the influence of environmental factors such as light, heat, temperature and the like on the folic acid content in the experimental process.
(6) Content of Conjugated Linoleic Acid (CLA) and gamma-aminobutyric acid in fermentation sample
In the invention, the beneficial substances such as Conjugated Linoleic Acid (CLA) and gamma-aminobutyric acid are generated in the fermented milk of K56. See tables 8 and 9 for relevant test results. Wherein, the content determination method of the conjugated linoleic acid is based on the gas chromatography NY/T1671-2008 for the content determination of the Conjugated Linoleic Acid (CLA) in milk and dairy products; the method for measuring the content of the gamma-aminobutyric acid is based on QB/T4587-2013 gamma-aminobutyric acid.
TABLE 8 content of conjugated linoleic acid in fermentation supernatant of K56
Figure BDA0002802513100000101
TABLE 9 content of gamma-aminobutyric acid in fermentation supernatant of K56
Figure BDA0002802513100000102
Example 2 fermentation of Lactobacillus paracasei K56 milk beverage enriched in vitamin B group
Heating fresh milk to more than 50 ℃, adding 6% of white granulated sugar, stirring until the white granulated sugar is completely dissolved, preheating to 60-65 ℃, homogenizing under 20Mpa pressure, carrying out heat treatment for 5-8 minutes at about 90 ℃, cooling to 37 ℃, and inoculating lactobacillus paracasei K56 seed liquid, wherein the inoculation amount is 2-3%. Fermenting at 37 deg.C for 72-96 h. Diluting the fermented product with sterilized and cooled mixture of water, syrup, emulsion stabilizer, etc. by 2-4 times, and making into directly drinkable lactobacillus beverage. The lactobacillus beverage has sour and sweet taste, good flavor, and high vitamin B content1Vitamin B3Vitamin B6Conjugated linoleic acid and gamma-aminobutyric acid content.

Claims (10)

1. A method for synthesizing B vitamins by fermenting Lactobacillus paracasei (Lactobacillus paracasei subsp. paracasei), comprising: the fermentation product containing B vitamins is prepared by fermenting lactobacillus paracasei CGMCC No.15139 or DSM 27447.
2. The method of claim 1, further comprising: extracting and purifying B vitamins from the supernatant of the fermentation product.
3. The method according to claim 1 or 2, wherein the B vitamins comprise vitamin B1Vitamin B2Vitamin B3Vitamin B6And vitamin B9One or more of (a).
4. The method of claim 3 wherein the B vitamins comprise vitamin B1Vitamin B3And/or vitamin B6。
5. The method according to claim 1, wherein the fermentation substrate used in the fermentation is an emulsion; preferably, the emulsion is fresh milk or reconstituted milk, and optionally contains 5-10% of sucrose.
6. The method of claim 1 or 5, wherein the fermentation conditions are: fermenting for 1-7 days at 35-45 ℃.
7. The method according to claim 1 or 5, wherein the Lactobacillus paracasei is inoculated in the fermentation substrate in the form of seed liquid, in an amount of 2% to 5%.
8. The application of lactobacillus paracasei in preparing the composition rich in B vitamins by fermentation, wherein the fermentation product rich in B vitamins is prepared by fermenting lactobacillus paracasei with CGMCC No.15139 or DSM 27447; the obtained fermentation product containing the B vitamins is used as or further used for preparing food, daily chemicals, feed or medicaments.
9. Use according to claim 8, wherein the food product is a fermented milk, a fermented milk beverage, a milk powder or a solid beverage.
10. Use according to claim 8 or 9, wherein the composition is also enriched in conjugated linoleic acid and/or gamma-aminobutyric acid, produced by fermentation of said lactobacillus paracasei.
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Citations (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN105076402A (en) * 2015-09-25 2015-11-25 齐齐哈尔医学院 Method for preparing health yogurt rich in natural folic acid
KR20170088668A (en) * 2016-01-25 2017-08-02 충남대학교산학협력단 Preparation Method for Kimchi with High GABA Content
CN107916236A (en) * 2016-10-05 2018-04-17 益福生医股份有限公司 Lactobacillus paracasei subspecies paracasei K56
CN108348559A (en) * 2015-07-07 2018-07-31 阿尔法西格玛有限公司 Lactobacillus paracasei, the nutritional preparation containing it and pharmaceutical preparation and application thereof for producing conjugated linoleic acid
CN110301480A (en) * 2018-03-20 2019-10-08 内蒙古蒙牛乳业（集团）股份有限公司 Milk-contained drink and preparation method thereof
CN110882280A (en) * 2019-06-27 2020-03-17 内蒙古伊利实业集团股份有限公司 New application of lactobacillus paracasei K56
WO2020063515A1 (en) * 2018-09-30 2020-04-02 内蒙古伊利实业集团股份有限公司 Novel application of lactobacillus paracasei k56 capable of regulating balance of gastrointestinal flora
CN111733105A (en) * 2020-07-03 2020-10-02 上海商学院 Fermentation agent for improving content of gamma-aminobutyric acid in fermented milk and preparation method of fermented milk rich in gamma-aminobutyric acid

Patent Citations (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108348559A (en) * 2015-07-07 2018-07-31 阿尔法西格玛有限公司 Lactobacillus paracasei, the nutritional preparation containing it and pharmaceutical preparation and application thereof for producing conjugated linoleic acid
CN105076402A (en) * 2015-09-25 2015-11-25 齐齐哈尔医学院 Method for preparing health yogurt rich in natural folic acid
KR20170088668A (en) * 2016-01-25 2017-08-02 충남대학교산학협력단 Preparation Method for Kimchi with High GABA Content
CN107916236A (en) * 2016-10-05 2018-04-17 益福生医股份有限公司 Lactobacillus paracasei subspecies paracasei K56
CN110301480A (en) * 2018-03-20 2019-10-08 内蒙古蒙牛乳业（集团）股份有限公司 Milk-contained drink and preparation method thereof
WO2020063515A1 (en) * 2018-09-30 2020-04-02 内蒙古伊利实业集团股份有限公司 Novel application of lactobacillus paracasei k56 capable of regulating balance of gastrointestinal flora
CN110882280A (en) * 2019-06-27 2020-03-17 内蒙古伊利实业集团股份有限公司 New application of lactobacillus paracasei K56
CN111733105A (en) * 2020-07-03 2020-10-02 上海商学院 Fermentation agent for improving content of gamma-aminobutyric acid in fermented milk and preparation method of fermented milk rich in gamma-aminobutyric acid

Non-Patent Citations (10)

* Cited by examiner, † Cited by third party
Title
HONGYUN LU等: "Safety Evaluation of Bifidobacterium lactis BL-99 and Lacticaseibacillus paracasei K56 and ET-22 in vitro and in vivo", 《FRONT. MICROBIOL.》 *
JEAN GUYLEBLANC等: "Bacteria as vitamin suppliers to their host: a gut microbiota perspective", 《CURRENT OPINION IN BIOTECHNOLOGY》 *
M.JUAREZ DEL VALLE等: "Riboflavin producing lactic acid bacteria as a biotechnological strategy to obtain bio-enriched soymilk", 《FOOD RESEARCH INTERNATIONAL》 *
刘艺茹 等: "分离自人体肠道的副干酪乳杆菌K56和ET-22的菌株鉴定", 《食品与发酵工业》 *
常若毅: "益生菌发酵果蔬产维生素的研究及其产品的研制", 《中国优秀硕士论文全文数据库（电子期刊）工程科技I辑》 *
张丽莉 等: "产共轭亚油酸干酪乳杆菌的诱变选育", 《食品研究与开发》 *
张俊 等: "副干酪乳杆菌产γ-氨基丁酸的发酵培养基优化", 《西华大学学报（自然科学版）》 *
耿文超 等: "副干酪乳杆菌的功能特性及其应用研究进展", 《生物加工过程》 *
芦红云 等: "三株酸乳发酵菌株的发酵特性评估及风味分析", 《第十五届益生菌与健康国际研讨会摘要集》 *
邵群 等: "乳酸菌发酵产生共轭亚油酸条件的研究", 《山东师大学报（自然科学版）》 *

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