CN102166015A - Preparation method of probiotics agent capable of lowering blood fat - Google Patents
Preparation method of probiotics agent capable of lowering blood fat Download PDFInfo
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- 210000004369 blood Anatomy 0.000 title claims abstract description 38
- 239000008280 blood Substances 0.000 title claims abstract description 38
- 239000006041 probiotic Substances 0.000 title claims abstract description 19
- 235000018291 probiotics Nutrition 0.000 title claims abstract description 19
- 238000002360 preparation method Methods 0.000 title claims abstract description 18
- 239000003795 chemical substances by application Substances 0.000 title abstract 3
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims abstract description 40
- 238000000855 fermentation Methods 0.000 claims abstract description 33
- 230000004151 fermentation Effects 0.000 claims abstract description 33
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Abstract
The invention discloses a preparation method of a probiotics agent capable of lowering blood fat. Bacillus licheniformis BLc strains are activated and are subjected to scale-up culture by seed culture; the Bacillus licheniformis BLc strains subjected to scale-up culture are inoculated into a fermentation medium; NaCl, K2HPO4.3H2O and NaH2PO4 in the fermentation medium have actions on accelerating the secretion of various enzymes of the protease, the lipase, the amylase and the like of the BLc strains and on improving activity; the activity of the enzyme can be improved if the pH of the fermentation medium is controlled to 6-7, Enteromorpha is degraded, and a large quantity of Bacillus licheniformis BLc strain sporophyte is contained; after spray drying, the sporophyte temporarily looses activity; after being eaten, the sporophyte revives when entering the stomach and intestine and secretes various enzymes so as to accelerate gastrointestinal foods to decompose and decompose cholesterol, fat and the like; meanwhile, antagonism can be carried out on enteropathogenic bacteria; and the Enteromorpha degradation product has the action on lowering blood fat, resisting lipid peroxidation and preventing fatty liver. The invention provides the preparation method of probiotics agent capable of lowering blood fat, which has the advantages of low raw material cost and simple preparation method and has an action on accelerating the gastrointestinal function.
Description
Technical field
The present invention relates to hypolipemic function food or medicine, be specifically related to a kind of preparation method of reducing blood lipid probiotics.
Background technology
Along with improving constantly of people's living standard, sub-health population is more and more, and especially blood three high phenomenon numbers increase in a large number.Blood fat is the important indicator of cardiovascular pathological changes, the blood fat rising can make the outburst possibility of atherosclerotic and disease of cardiovascular system increase, can cause the cardiovascular various complication that wait internal organs, as concurrent artery sclerosis, concurrent heart disease, problems such as brain blood supply insufficiency appear, the symptom that dysfunction of liver, uremia, pancreatitis due to hyperlipidemia etc. also can concurrent hyperlipidemias appears.Cholesterol all has two parts with triglycerides, and a part is that eat into the outside, and this part is not the major part that body's cholesterol increases, and also having most is self to synthesize endogenous cholesterol, endogenous triglycerides.The medicine or the functional food of relevant reducing blood lipid are varied, and medicine mainly contains Chinese patent drug and Western medicine; Functional food also is formulated with various Chinese medicines and food, as notification number is the CN1099293 patent of invention, disclose by various Chinese medicine compatibilities (Semen Raphani, hawthorn, rice sprout, fig, mung bean, Chinese yam etc.), poach and become water cooking liquid, use saccharomycetes to make fermentation again, obtaining bacterium mud composition becomes blood fat reducing preparation.But because the price of Chinese medicine improves constantly, various is that the reducing blood lipid Chinese patent drug of major ingredient and the cost of health products also improve constantly with the Chinese medicine.Though notification number is the CN101919875 application for a patent for invention, disclose sea grass polysaccharide and mushroom polysaccharide compound and can improve immunity of organisms and effect such as antitumor, relevant Enteromorpha has effect for reducing blood fat and does not appear in the newspapers.
Summary of the invention
Technical problem to be solved by this invention provides that a kind of cost of material is lower, the preparation method is simple, gastrointestinal function is had facilitation, can reduce blood fat the preparation method of reducing blood lipid probiotics.
The present invention solves the problems of the technologies described above the technical scheme that is adopted: a kind of preparation method of reducing blood lipid probiotics comprises the steps:
A seed culture: the MnSO of the NaCl of the nitrogenous source of the carbon source of percentage part 5～8 %, 2～4 %, 0.3～0.6 %, 0.001～0.003 % by weight
4Surplus is a water, stir after the mixing and be made into seed culture medium, described carbon source is sucrose, maltose, beta-schardinger dextrin-, glucose or soluble starch, described nitrogenous source is casein, yeast extract, soybean protein, gelatin or beef extract, insert bacillus licheniformis BLc strain then in described seed culture medium, the preserving number of this bacillus licheniformis BLc strain is CGMCC No:4324, cultivates down for 37 ℃ and obtains bacillus licheniformis BLc strain bacterium liquid in 8～10 hours; The required clump count of fermentation is satisfied in this bacillus licheniformis BLc strain activation and a large amount of breeding after the seed culture; The depositary institution of this bacillus licheniformis BLc strain is China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) that is positioned at No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, and preservation date is on November 12nd, 2010, and the Latin name is called
Bacillus licheniformisWherein NaCl and MnSO
4It all is chemical pure;
B fermentation medium: the yeast extract of the Enteromorpha dry powder of percentage part 4～10 %, 1～4 % by weight, the NaCl of 0.1～0.5 %, the K of 0.2～0.6 %
2HPO
43H
2The NaH of O, 0.01～0.03 %
2PO
4, surplus is a water, uses Na after the mixing
2CO
3Adjust pH is 6～7, stirs to be made into fermentation medium; Wherein NaCl, K
2HPO
43H
2O and NaH
2PO
4It all is chemical pure; Yeast extract is as the nutrient solution of strain fermentation, NaCl, K
2HPO
43H
2O and NaH
2PO
4Can promote bacillus licheniformis BLc strain secretase and the effect that improves enzymatic activity, pH 6～7 enzymatic activitys are higher, help sea grass polysaccharide, terpene and steroidal etc. in the Enteromorpha are dissociated out;
C inoculation: in described fermentation medium, insert described bacillus licheniformis BLc strain bacterium liquid, described bacillus licheniformis BLc strain
The access weight of bacterium liquid is 8～12 % of described fermentation medium weight, and the viable count of every milliliter of described bacillus licheniformis BLc strain bacterium liquid is 5 * 10
5～5 * 10
7Individual; Need viable count to detect before described bacillus licheniformis BLc strain bacterium liquid inserts, and be diluted to the viable count of inoculation standard;
D fermentation: 37 ℃ of room temperatures, cultivate 36～40h on 120 r/min shaking tables or the fermentation tank, staticly settle, the centrifugal sediment that obtains;
E spray-drying: add the skimmed milk power homogeneous at described sediment, being diluted to solid content with sterilized water is 10～25%(W/V), adding beta-schardinger dextrin-more evenly mixes, at EAT is 100～120 ℃, leaving air temp is 80～100 ℃, and spray-drying under charging rate 8～10mL/min condition obtains the reducing blood lipid probiotics, it is 1～5% of described weight of precipitate that described skimmed milk power adds weight, and it is 5～8% of described weight of precipitate that described beta-schardinger dextrin-adds weight.
Described seed culture medium is by sucrose, the casein of 3 %, the NaCl of 0.5 %, the MnSO of 0.002 % of 6 %
4, surplus is that water is formulated.
Described fermentation medium is by the Enteromorpha dry powder of 8 %, the yeast extract of 3 %, 0.3% NaCl, 0.4 %K
2HPO
43H
2The NaH of O, 0.02 %
2PO
4, surplus is that water is formulated.
The access weight of described bacillus licheniformis BLc strain bacterium liquid is 10 % of described fermentation medium weight.
Culture collection process: the peptone 1% of percentage part, beef extract 0.3% by weight, NaCl 0.05 %, agar powder 2%, surplus is a water, stirs to be made into the culture presevation culture medium, inserts bacillus licheniformis BLc strain in the culture presevation culture medium, preserve down in 4 ℃, but about preservation half a year.Effect with long preservation.
Compared with prior art, the invention has the advantages that a kind of preparation method of reducing blood lipid probiotics, by seed culture bacillus licheniformis BLc strain activation and expansion are cultivated, will enlarge again in the bacillus licheniformis BLc strain access fermentation medium of cultivating and ferment NaCl, K in the fermentation medium
2HPO
43H
2O and NaH
2PO
4Has the protease that promotes the BLc strain, lipase, the secretion of various enzymes such as amylase and the effect that improves enzymatic activity, the pH of culture medium is controlled at 6～7, can improve the activity of enzyme, make the Enteromorpha degraded, the sediment that obtains not only contains Enteromorpha catabolite such as sea grass polysaccharide, terpene, steroidal and short fiber etc., also contain a large amount of bacillus licheniformis BLc strain sporinites, the temporary transient inactivation of spray-drying sporinite, enter stomach and intestine after edible and bring back to life the various enzymes of justacrine, promote stomach and intestine food to decompose, with cholesterol, decomposition such as fat simultaneously can the antagonism pathogenic entero becteria, and the Enteromorpha catabolite has reducing blood lipid, the effect of anti peroxidation of lipid and prevention fatty liver, can reduce serum TC, TG, promote the generation of HDL-C, reduce the deposition of lipid, increase radicals scavenging and increase antioxidant ability of organism at endangium; Therefore a kind of cost of material of the present invention is lower, the preparation method simple, gastrointestinal function is had facilitation, can reduce blood fat the preparation method of reducing blood lipid probiotics.
Above-mentioned bacillus licheniformis BLc strain (
Licheniformis) preservation date be on November 12nd, 2010, depositary institution is China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC), preserving number is CGMCC No:4324.
The specific embodiment
Below in conjunction with embodiment the present invention is described in further detail.
Embodiment 1
A kind of preparation method of reducing blood lipid probiotics at first enlarges bacillus licheniformis BLc strain (preserving number is CGMCC No:4324) and cultivates, and seed culture medium is by 600 gram sucrose, 300 gram caseins, 50 gram NaCl, 2 gram MnSO
4After the mixing, add water to gross weight 10000 grams, stirring is made into seed culture medium, inserts bacillus licheniformis BLc strain, cultivates 8～10 hours down for 37 ℃, and the microscopy viable count is if too much, need be diluted to every milliliter of bacterium liquid viable count is 5 * 10
5～5 * 10
7Individual, obtain bacillus licheniformis BLc strain bacterium liquid and preserve standby; Prepare fermentation medium again: with 8 kilograms of Enteromorpha dry powder, 3 kilograms of yeast extracts, 0.3 kilogram of NaCl, 0.4 kilogram of K
2HPO
43H
2O, 0.02 kilogram of NaH
2PO
4After the mixing, add water to 100 kilograms of gross weights, stirring is made into fermentation medium, uses Na again
2CO
3Transferring pH is 6～7; Insert 10 kilograms of bacillus licheniformis BLc strain bacterium liquid at fermentation medium, under 37 ℃ of room temperatures, on 120 r/min shaking tables, cultivate 36～40h, staticly settle, the centrifugal sediment that obtains; Add the skimmed milk power homogeneous at sediment; The skimmed milk power addition is 3～4% of a weight of precipitate, being diluted to solid content with sterilized water is 15～20%(W/V), adding beta-schardinger dextrin-more evenly mixes, the beta-schardinger dextrin-addition is 6～7% of a weight of precipitate, at EAT is 100～120 ℃, leaving air temp is 80～100 ℃, and spray-drying under charging rate 8～10mL/min condition obtains the reducing blood lipid probiotics.
Embodiment 2
Substantially the same manner as Example 1, different just seed culture mediums by 500 gram maltose, 200 gram yeast extracts, 30 gram NaCl, 1 gram MnSO
4After the mixing, it is formulated to add water to gross weight 10000 grams; Fermentation medium is by 4 kilograms of Enteromorpha dry powder, 1 kilogram of yeast extract, 0.1 kilogram of NaCl, 0.2 kilogram of K
2HPO
43H
2O, 0.01 kilogram of NaH
2PO
4After the mixing, it is formulated to add water to 100 kilograms of gross weights; Insert 8 kilograms of bacillus licheniformis BLc strain bacterium liquid at fermentation medium, the skimmed milk power addition is 1% of a weight of precipitate, and solid content is 10～15%, and the beta-schardinger dextrin-addition is 5% of a weight of precipitate.
Embodiment 3
Substantially the same manner as Example 1, different just seed culture mediums by 800 gram soluble starches, 400 gram gelatin, 60 gram NaCl, 3 gram MnSO
4After the mixing, it is formulated to add water to gross weight 10000 grams; Fermentation medium is by 10 kilograms of Enteromorpha dry powder, 4 kilograms of yeast extracts, 0.5 kilogram of NaCl, 0.6 kilogram of K
2HPO
43H
2O, 0.03 kilogram of NaH
2PO
4After the mixing, it is formulated to add water to 100 kilograms of gross weights; Insert 12 kilograms of bacillus licheniformis BLc strain bacterium liquid at fermentation medium, the skimmed milk power addition is 5% of a weight of precipitate, and solid content is 20～25%, and the beta-schardinger dextrin-addition is 8% of a weight of precipitate.
Embodiment 4
Substantially the same manner as Example 1, in the different just seed culture mediums 600 gram sucrose substitute by 550 gram glucose, 300 gram caseins are substituted by 250 gram soybean proteins.
Embodiment 5
Basic identical with the real example 1 of putting, in the different just seed culture medium 600 gram sucrose substitute by 700 gram beta-schardinger dextrin-s, 300 gram caseins are substituted by 350 gram beef extracts, shaking table is cultivated by fermentation tank culture and is substituted.
Application examples
Get 60 of ICR mouse, male and female are divided into 5 groups at random than 1:2, and 12 every group, claim respectively to organize body weight, that is: the blank group (give, hyperlipidemia model group, low dose group, high dose group, positive controls by filling.Except that blank group (feeding basal feed), what other each group was all fed is high lipid food (20 % lards, 2 % cholesterol, 0.2 % cholate, 0.1 % propylthiouracil).Low dose group is irritated stomach with the reducing blood lipid fermentate of embodiment 1 once a day by 0.1 mL/10g dosage of mouse body weight, high dose group is than the low dose group consumption that doubles, positive controls is irritated stomach with commercially available spirulina once a day by 0.1 mL/10g dosage of mouse body weight, blank group and hyperlipidemia model group are irritated stomach with clear water once a day by 0.1 mL/10g dosage of mouse body weight, the filling stomach that regularly feeds intake every day, and freely drink water; Irritate 4 weeks of stomach, claim respectively to organize body weight, after water 12 h are can't help in fasting, eyeball is got blood, centrifugal behind 37 ℃ of waters 30 min (3000 r/min, 15 min) separation of serum is surveyed cholesterol in serum (TC), triglycerides (TG), low-density lipoprotein (LDL-C), HDL (HDL-C); Dissect internal organ simultaneously, win liver, abdomen fat and claim that each group is heavy.Obtain result as shown in Table 1 and Table 2 respectively.
Table 1: each organizes the water-glass of the blood fat of mouse
| Group | TC(mmol/L) | TG (mmol/L) | LDL-C(mmol/L) | HDL-C(mmol/L) |
| The blank group | 2.42±0.63 | 0.90±0.47 | 0.46±0.03 | 1.88±0.47 |
| The hyperlipidemia model group | 3.56±0.59 | 1.89±0.32 | 1.15±0.10 | 1.15±0.46 |
| Low dose group | 2.92±0.41 | 1.09±0.42 | 0.48±0.18 | 1.98±0.60 |
| High dose group | 2.86±0.34 | 1.10±0.41 | 0.40±0.26 | 2.00±0.37 |
| Positive controls | 2.61±0.74 | 1.35±0.45 | 0.58±0.25 | 2.05±0.57 |
The all higher fat model group of each blood lipid level of low dose group, high dose group, positive controls is low as can be seen from Table 1, and the same effect with reduction blood fat with spirulina of reducing blood lipid fermentate of the present invention is described.
Table 2: each organizes the water-glass of the body weight gain rate liver exponential sum fat coefficient of mouse
| Group | Body weight gain rate (%) | Liver coefficient (%) | Fat coefficient (%) |
| The blank group | 31.71±6.40 | 5.87±0.66 | 1.58±0.41 |
| The hyperlipidemia model group | 36.76±3.62 | 4.90±0.24 | 3.26±0.52 |
| Low dose group | 32.67±3.93 | 5.24±0.27 | 2.57±0.34 |
| High dose group | 31.09±4.03 | 5.31±0.39 | 2.38±0.70 |
| Positive controls | 33.31±3.85 | 5.23±0.46 | 2.70±0.51 |
Wherein the liver coefficient is the ratio of mouse liver weight and body weight, and fat coefficient is the interior abdomen fat of mouse body and the ratio of body weight; Take low dose group of the present invention and high dose group as can be seen from Table 2 and blank group body weight is approaching, be starkly lower than the hyperlipidemia model group, also be lower than positive controls, illustrate to show fat; Liver coefficient and the fat coefficient of taking low dose group of the present invention and high dose group and positive controls are approaching, and the liver coefficient is obviously greater than the hyperlipidemia model group, fat coefficient is significantly less than the hyperlipidemia model group, and the same effect with reduction blood fat and fat with spirulina of the present invention is described, can lose weight.
We also find in clinical research, take reducing blood lipid probiotics of the present invention, and TC, TG have remarkable reduction, and HDL-C significantly raises, LDL-C significantly descends (P＜0.01), and have tangible dose-effect relationship.Main component is viable bacteria and the Enteromorpha product after strains for degrading in the fermentation postprecipitation thing, and viable bacteria exists with the endospore form in the reducing blood lipid probiotics.Take the back spore and enter enteron aisle, can bring back to life the very strong protease of justacrine activity, lipase, amylase rapidly in upper intestines, help the complicated carbohydrate of degrading, generation has the multiple material of antagonism pathogenic entero becteria, the oxygen that consumes in the enteron aisle by biological oxidation causes anaerobic environment simultaneously, enteron aisle ancestral home dominant bacteria belongs to anaerobic bacteria mostly, and harmful bacterium and foreign bacteria mostly are aerobic bacteria, thereby helps keeping intestinal eubiosis.Not only contain the essential various nutriments of animal in the Enteromorpha catabolite, balanced in nutrition, but also contain multiple bioactivators such as polysaccharide, terpene and steroidal, have that reducing blood lipid is anti-, an effect of oxidation and prevention fatty liver.
Claims (4)
1. the preparation method of a reducing blood lipid probiotics is characterized in that comprising the steps:
A, seed culture: by weight the NaCl, 0.001 of the nitrogenous source of the carbon source of percentage part 5～8 %, 2～4 %, 0.3～0.6 %～
0.003 the MnSO of %
4Surplus is a water, stir after the mixing and be made into seed culture medium, described carbon source is sucrose, maltose, beta-schardinger dextrin-, glucose or soluble starch, described nitrogenous source is casein, yeast extract, soybean protein, gelatin or beef extract, insert bacillus licheniformis BLc strain then in described seed culture medium, the preserving number of this bacillus licheniformis BLc strain is CGMCC No:4324, cultivates down for 37 ℃ and obtains bacillus licheniformis BLc strain bacterium liquid in 8～10 hours;
B, fermentation medium: the yeast extract of the Enteromorpha dry powder of percentage part 4～10 %, 1～4 % by weight, the NaCl of 0.1～0.5 %, the K of 0.2～0.6 %
2HPO
43H
2The NaH of O, 0.01～0.03 %
2PO
4, surplus is a water, uses Na after the mixing
2CO
3Adjust pH is 6～7, stirs to be made into fermentation medium;
C, inoculation: in described fermentation medium, insert described bacillus licheniformis BLc strain bacterium liquid, described bacillus licheniformis BLc
The access weight of strain bacterium liquid is 8～12 % of described fermentation medium weight, and the viable count of every milliliter of described bacillus licheniformis BLc strain bacterium liquid is 5 * 10
5～5 * 10
7Individual;
D, fermentation: 37 ℃ of room temperatures, cultivate 36～40h on 120 r/min shaking tables or the fermentation tank, staticly settle, the centrifugal sediment that obtains;
E, spray-drying: add the skimmed milk power homogeneous at described sediment, being diluted to solid content with sterilized water is 10～25%(W/V), adding beta-schardinger dextrin-more evenly mixes, at EAT is 100～120 ℃, leaving air temp is 80～100 ℃, and spray-drying under charging rate 8～10mL/min condition obtains the reducing blood lipid probiotics, it is 1～5% of described weight of precipitate that described skimmed milk power adds weight, and it is 5～8% of described weight of precipitate that described beta-schardinger dextrin-adds weight.
2. the preparation method of a kind of reducing blood lipid probiotics as claimed in claim 1 is characterized in that sucrose, the casein of 3 %s, the NaCl of 0.5 %, the MnSO of 0.002 % of described seed culture medium by 6 %
4, surplus is that water is formulated.
3. the preparation method of a kind of reducing blood lipid probiotics as claimed in claim 1 is characterized in that described fermentation medium is by the Enteromorpha dry powder of 8 %, the yeast extract of 3 %, 0.3% NaCl, the K of 0.4 %
2HPO
43H
2The NaH of O, 0.02 %
2PO
4, surplus is that water is formulated.
4. the preparation method of a kind of reducing blood lipid probiotics as claimed in claim 1 is characterized in that the access weight of described bacillus licheniformis BLc strain bacterium liquid is 10 % of described fermentation medium weight.
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| CN105541033A (en) * | 2016-01-12 | 2016-05-04 | 沈勤 | Livestock and poultry aquaculture wastewater ecological treatment system |
| TWI622398B (en) * | 2014-06-20 | 2018-05-01 | Use of mountain flour round fermentation product and preparation method thereof | |
| CN110237101A (en) * | 2019-06-25 | 2019-09-17 | 东北制药集团沈阳第一制药有限公司 | A kind of bacillus licheniformis and the composition of xylo-oligosaccharide and application thereof |
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| CN101669580A (en) * | 2009-09-21 | 2010-03-17 | 宁波大学 | Method for preparing feed additive by fermenting enteromorpha |
| CN101869269A (en) * | 2010-07-13 | 2010-10-27 | 福建农林大学 | Method for preparing gelatinous enteromorpha polysaccharide |
| CN101919875A (en) * | 2010-08-24 | 2010-12-22 | 温州大学 | Enteromorpha mushroom polysaccharide compound, preparation method and application as health-care medicament thereof |
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| CN101669580A (en) * | 2009-09-21 | 2010-03-17 | 宁波大学 | Method for preparing feed additive by fermenting enteromorpha |
| CN101869269A (en) * | 2010-07-13 | 2010-10-27 | 福建农林大学 | Method for preparing gelatinous enteromorpha polysaccharide |
| CN101919875A (en) * | 2010-08-24 | 2010-12-22 | 温州大学 | Enteromorpha mushroom polysaccharide compound, preparation method and application as health-care medicament thereof |
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| 《中国公共卫生》 20090515 林文庭等 浒苔多糖降血脂及抗脂质过氧化作用 第567－568页 1－4 第25卷, 第05期 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| TWI622398B (en) * | 2014-06-20 | 2018-05-01 | Use of mountain flour round fermentation product and preparation method thereof | |
| CN105541033A (en) * | 2016-01-12 | 2016-05-04 | 沈勤 | Livestock and poultry aquaculture wastewater ecological treatment system |
| CN110237101A (en) * | 2019-06-25 | 2019-09-17 | 东北制药集团沈阳第一制药有限公司 | A kind of bacillus licheniformis and the composition of xylo-oligosaccharide and application thereof |
| CN110237101B (en) * | 2019-06-25 | 2021-04-27 | 东北制药集团沈阳第一制药有限公司 | Composition of bacillus licheniformis and xylo-oligosaccharide and application thereof |
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