CN102586155A - Lactobacillus plantarum N13 and use thereof - Google Patents
Lactobacillus plantarum N13 and use thereof Download PDFInfo
- Publication number
- CN102586155A CN102586155A CN2012100578224A CN201210057822A CN102586155A CN 102586155 A CN102586155 A CN 102586155A CN 2012100578224 A CN2012100578224 A CN 2012100578224A CN 201210057822 A CN201210057822 A CN 201210057822A CN 102586155 A CN102586155 A CN 102586155A
- Authority
- CN
- China
- Prior art keywords
- plant lactobacillus
- temperature
- milk
- lactobacillus
- fermented
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Images
Abstract
The invention relates to a Lactobacillus plantarum N13 and a use thereof. The Lactobacillus plantarum N13 has acid and alkaline resistances, has a very high adhesiveness to intestinal epithelial cell lines HT-29, and can inhibit the EIEC (enteroinvasive Escherichia coli.) growth, and the adhesion and the invasion of the EIEC to the intestinal epithelial cell lines HT-29. Sour soybean milk, fermented oat beverages, fermented vegetables, fermented silages, lactobacillus milk beverages and fermented milk prepared through using the Lactobacillus plantarum N13 have very good application prospects.
Description
[technical field]
The invention belongs to microbial technology field.More specifically, the present invention relates to a kind of plant lactobacillus (Lactobacillus plantarum) N13, also relate to the purposes of said plant lactobacillus N13.
[background technology]
(Enteroinvasive Escherichia coli. EIEC) is one group and can invades big intestinal epithelial cells enteroinvasive E.Coli, host intestinal epithelial cells is inflamed or forms ulcer, causes the coli strain of dysentery appearance symptom.The EIEC virulence of causing a disease is strong, and reproduction speed is fast, as long as there are 10～100 EIEC bacteriums just can be diseases induced, its contaminated food products and water source cause eruption and prevalence, thereby cause extensive property diarrhea disease.The virulence of EIEC depends on the ability of invading cell, and invasion procedure comprises former and spread two processes.EIEC invasion intestinal epithelial cell is divided into four processes: (1) EIEC arrives intestinal mucosa lower floor through the M cell; (2) in scavenger cell, duplicate; (3) invade outside the substrate of colon cell; (4) invade colon epithelial cell, and breeding therein, diffusion causes inflammation and dysentery symptomatic reaction.
The caused intestinal tract disease of treatment EIEC, common treat-ment is to use antibiotic therapy.The antibiotic therapy method has following effect: 1, can effectively control the clinical and subclinical infection that EIEC causes; 2, microbiotic can suppress the hyperplasia of intestinal cell wall, thereby improves the absorption of intestinal cell to nutritive ingredient; 3, playing a part to select the enteric microorganism flora, mainly is through reducing the consumption of harmful microorganism to necessary nutritive substance, increasing a large amount of breedings of beneficial bacteria; 4, influence some physiological metabolism of animal, thereby influence the growth of animal.Antibiotic therapy has instant effect; Advantages such as treatment time is short; But use and abuse of antibiotics along with the mankind are a large amount of, brought a series of problems also for simultaneously environment and society, this mainly comprises: 1, microbiotic can cause alteration of intestinal flora; Endurance strain increases, thereby makes the antibiotic therapy more difficult; 2, the life-time service microbiotic causes immunity of organisms to descend; 3, the easy infection situation increases, and causes the body superinfection; 4, antibiotic etc residual in livestock product and environment threatens human healthy.
In order to overcome antibiotic spinoff, increasing research selects for use probiotic bacterium to treat the infringement of EIEC to body.People such as Resta-Lenert (Resta-Lenert; S.; &Barrett; K.E. (2006) .Live probiotics protect intestinal epithelial cells from the effects of infection with enteroinvasive Escherichia coli (EIEC) .Gut, 52,988-997.) utilize the Streptococcus thermophilus and Lactobacillus acidophilus intervention EIEC that lives; These two kinds of milk-acid bacterias of use are united in discovery can significantly reduce adhesion and the invasion and attack of EIEC to the host, and reduces host's physiological function obstacle that EIEC causes; People such as Medici (Medici, M., Vinderola; C.G.; Weill, R. ， &Perdig ó n; G. (2005) .Effect of fermented milk containing probiotic bacteria in the prevention of an enteroinvasive Escherichia coli infection in mice.Journal of Dairy Research; 72,243-249.) use the fermented milk contain lactobacterium casei, lactobacillus delbruckii to treat the infection that EIEC causes, the infection of EIEC is avoided in the immunoreation of finding the intestinal mucosa that probiotic bacterium can be through strengthening mouse.These researchs show according to the antagonistic action between the bacterium; Main through the competitive inhibition between the different strains; Not only can control the infringement of EIEC to intestinal epithelial cell; The proper distribution that can also be of value to bacterial flora on the intestinal epithelial cell is brought into play the physiological function of enteron aisle more effectively.These researchs are only considered antagonism and the treatment aspect of EIEC from selected lactic bacterium strains, do not investigate the probiotic properties of these milk-acid bacterias.Probiotic bacterium should have following main characteristic: can tolerate GI extreme environment 1.; 2. the body enteron aisle had good adhesive capacity; 3. can regulate the eubiosis of body enteron aisle.The present invention has filtered out through experiment in vitro not only has antagonism EIEC ability, but also the milk-acid bacteria with good probiotic properties, and investigate milk-acid bacteria adhered to and attacked colon epithelial cell to EIEC restraining effect.Compare with the milk-acid bacteria that the treatment EIEC of report infects, plant lactobacillus N13 of the present invention has better bile tolerance ability and the adhesive capacity higher to colon epithelial cell.
In view of probiotic bacterium has significant biological antagonist restraining effect and its unique advantage to enteric pathogenic bacteria, therefore screening has very wide application prospect to the probiotic bacterium that enteroinvasive E.Coli has inhibition and antagonistic action.For this reason, the inventor has carried out lot of experiments work on the basis of summing up prior art, accomplished the present invention.
[summary of the invention]
[technical problem that will solve]
The purpose of this invention is to provide a kind of plant lactobacillus (Lactobacillus plantarum) N13.
Another object of the present invention provides the purposes of said plant lactobacillus N13.
[technical scheme]
The present invention realizes through following technical proposals.
The present invention relates to a kind of plant lactobacillus (Lactobacillus plantarum) N13; In on the November 29th, 2011 of No. 3 China Committee for Culture Collection of Microorganisms of Institute of Microorganism, Academia Sinica common micro-organisms center preservations in the Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, its preserving number is CGMCC No.5495 to this bacteria strain.
Plant lactobacillus (Lactobacillus plantarum) N13 also is preserved in food microorganisms DSMZ of Southern Yangtze University simultaneously, and deposit number is CCFM 233.Therefore in this application, term " CCFM 233 " also refers to plant lactobacillus (Lactobacillus plantarum) N13, and both are same strain bacterial strain.
Described plant lactobacillus N13 has property:
(1) pathogenic bacterium enteroinvasive E.Coli (ATCC 43893 for Enteroinvasive Escherichia coli., EIEC) had significant inhibitory effect;
(2) have acid resistance, under the condition of pH 2.0, can grow;
(3) have the bile tolerance ability, under 1-3 ‰ cholate condition, can grow;
(4) intestinal epithelial cells strain HT-29 cell had very high adhesive capacity;
(5) can obviously suppress the adhesion of EIEC to intestinal epithelial cells strain HT-29 cell;
(6) can obviously reduce the invasion and attack of EIEC to intestinal epithelial cells strain HT-29 cell.
The present invention relates to the purposes of described plant lactobacillus N13 in preparation soy yogurt, oat fermented drink, fermentation silage, fermented vegetables, lactobacillus milk beverage and cultured milk prod.
The described soy yogurt that contains plant lactobacillus N13 is to adopt following method preparation:
Volume ratio 2-4 according to soft water and soybean is following to soft water soaking soybean 1-2h at temperature 75-85 ℃; Remove soybean hulls; Remove soaked in water, again according to the weight ratio 1 of soybean and boiling water: 6-10 adds the boiling water defibrination, and its slurry keeps 10-15min under temperature 80-85 ℃ condition; Filter with 150 eye mesh screens then, obtain a kind of soya-bean milk; In this soya-bean milk, add sucrose earlier in soya-bean milk weight 5%-10%; Re-use clarifixator and under pressure 15-25MPa, carry out homogeneous; Under 95 ℃ of temperature, carry out germicidal treatment 10min subsequently; When its temperature is reduced to 36-38 ℃, insert the plant lactobacillus N13 starter of commercial dry powder leaven lactobicillus bulgaricus, commercial dry powder leaven thermophilus streptococcus and following preparation again according to the inoculum size in soya-bean milk weight 0.04-0.05%, its part by weight is 1: 1: 1; Temperature 35-42 ℃ bottom fermentation 6-8 hour, promptly obtain containing the soy yogurt of plant lactobacillus N13 viable bacteria then 4 ℃ of stored refrigerated;
Plant lactobacillus N13 starter dry powder formulations prepares according to following method: plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 1-3%; Cultivate 20-28h down for 37 ℃ in temperature, make plant lactobacillus N13 viable count reach 10
8More than the cfu/mL; Its nutrient solution centrifugal 8-12min under the condition of rotating speed 3800-4200r/min and temperature 2-6 ℃ then; Its centrifugal sediment washes 2-4 time with the PBS damping fluid of pH 7.2 again, is added to its bacterial sediment in the lyophilized vaccine adjustment cell concn to 10 then
9Cfu/mL carries out vacuum lyophilization after mixing, obtain described plant lactobacillus N13 starter dry powder formulations after the freeze-drying.It is sucrose 1.5%, skimming milk 14%, lactose 5%, glycerine 1%, starch 3% and 75.5% water by weight that described lyophilized vaccine is formed.
Described oat fermented drink is to adopt following method preparation:
A) with the oat be raw material, with oat selection, impurity elimination, removal epidermis and clean;
B) the 50-55 ℃ of water that in oat, adds in oat weight 100% mixes, and joins then and breaks into slurries in the hollander;
C) slurries that step B) obtain carry out defibrination with paste roller mill, use 150 purpose strainer filterings again, obtain a kind of slurries;
D) to step C) add in slurry weight 10-15% white sugar and 1000-1200% water in the slurries that obtain, at 121 ℃ of temperature sterilization 20min down, be cooled to 36-38 ℃, obtain the oat base-material that ferments;
E) plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 1-3%; Cultivate 20-24h down for 37 ℃ in temperature, spinning discards fermented supernatant fluid; Remaining bacterial sediment thing is resuspended with sterilized water, makes plant lactobacillus N13 viable bacteria concentration reach 10
8More than the cfu/mL, thereby obtain described plant lactobacillus N13 liquid starter;
F) according to volumeter 6-10% inoculum size with oat fermentation base-material; Lactobicillus bulgaricus liquid starter, thermophilus streptococcus liquid starter and above-mentioned steps E) the plant lactobacillus N13 liquid starter of preparation is inoculated at step D according to volume ratio at 1: 1: 1) in the oat fermentation base-material of preparation; Cultivate 18-22h down at leavening temperature 36-38 ℃, make the concentration of plant lactobacillus N13 reach 10
6More than the cfu/mL, promptly obtain containing the oat fermented drink of plant lactobacillus N13 viable bacteria then temperature 3-4 ℃ of following stored refrigerated.
Described fermented vegetables is to adopt following method preparation:
A) vegetables of selecting for use through clean with drain after, soak with the 0.005-0.01g/kg chlorine bleach liquor and to carry out disinfection in 7-10 minute, with described vegetables remove the peel, cutting, obtain a kind of pre-treatment vegetables;
B) toward steps A) add according to said vegetables weight meter 0.6-1.0% sugar and 4.0-6.0% salt in the pre-treatment vegetables that obtain, stir, pickle 20-28h in advance in room temperature, drain away the water, obtain a kind of preparatory Pickle;
C) take by weighing edible spice, use, is added after the cooling in said edible spice weight 7-9% liquor at temperature 75-85 ℃ of following lixiviate 2-4h in 8-10 times of water of said edible spice weight again, and mixing obtains a kind of spice juice;
D) plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 1-3%; Cultivate 20-28h down for 37 ℃ in temperature, spinning discards fermented supernatant fluid; Remaining throw out is resuspended with sterilized water, makes plant lactobacillus N13 viable bacteria concentration reach 10
8More than the cfu/mL, thereby obtain a kind of bacteria suspension;
E) toward step B) add in said preparatory Pickle weight 12-14% at step C in the preparatory Pickle that obtains) the spice juice that obtains and 1-5% be at step D) bacteria suspension that obtains; Mixing; Tinning, sealing, at room temperature fermentation; Reach 3.6-4.0 up to the pH of its fermented liquid value, obtain described fermented vegetables like this.
Wherein, described vegetables are one or more vegetables that are selected from Caulis et Folium Brassicae capitatae, Radix Dauci Sativae, asparagus lettuce, pimento, onion or celery.
Described spice is one or more spices that are selected from capsicum, Chinese prickly ash, pepper, ginger, garlic, anise, fennel, cassia bark, orange peel or cloves.
Described fermentation silage is to adopt following method preparation:
With the corn straw feedstuff raw material rub cut to 1～4cm for use;
Take by weighing 45-55kg corn straw feedstuff raw material; Be sprayed onto in the described corn straw feedstuff raw material after mixing the plant lactobacillus N13 dry powder formulations of the above-mentioned preparation of 0.5-0.7g and 200-240mL sterilized water; Turn mixed evenly after; 9-12 promptly obtains containing plant lactobacillus N13 after individual month corn straw feedstuff is stored in the compacting sealing in the silage bucket of packing into then under normal temperature condition.
Below the present invention will be described in more detail.
The present invention relates to a kind of plant lactobacillus (Lactobacillus plantarum) N13; In on the November 29th, 2011 of No. 3 China Committee for Culture Collection of Microorganisms of Institute of Microorganism, Academia Sinica common micro-organisms center preservations in the Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, its preserving number is CGMCC No.5495 to this bacteria strain.
Described plant lactobacillus N13 adopts following screening method to obtain.
This screening method step is following:
The screening of A, lactobacterium strain.
Take by weighing 25g traditional fermented food, pickles, butcher's meat, bread dough or infant faeces sample; Be added in the 225mL sterilized water; Cultivate 2h down for 37 ℃ in temperature behind the mixing, get this nutrient solution of 0.5mL then to be added in the 4.5mL sterilized water, the dilution nutrient solution that obtains is cultivated 2h down for 37 ℃ in temperature again; The nutrient solution that obtains like this repeats above-mentioned dilution and cultivates operation, makes this sample concentration be diluted to 10
-4, get extent of dilution and be respectively 10
-1～10
-4Each 50 μ L separate application of bacteria suspension on the MRS solid medium, after 37 ℃ of following anaerobism of temperature are cultivated 46-50h, select typical single bacterium colony, adopt plate streak, obtain pure lactic bacterium strains, particular case is seen table 1.
The test of B, isolated strains antagonism EIEC
That screening is obtained be inoculated in separately in the MRS liquid nutrient medium, under 37 ℃ of temperature, carry out anaerobism and cultivate 24h with lactic bacterium strains other sources (seeing table 1), spinning then, the fermented supernatant fluid that obtains carries out conventional antagonism EIEC test; With indicator intestinal bacteria EIEC (Enteroinvasive Escherichia coli ATCC 43893; Available from American type culture collection ATCC) be inoculated in the LB liquid nutrient medium after the activation; Cultivate 24h down for 37 ℃ in temperature, measure the absorbancy OD of its nutrient solution then at the 600nm place
600Value, and its OD value is adjusted to 0.66-0.70 with aseptic LB liquid nutrient medium, as negative control, the qingfengmeisu qiong of 320 μ g/mL is as positive control with the MRS liquid nutrient medium of pH 3.2, and repeated experiments 3 times is screened the probiotic lactobacillus with antagonism EIEC ability.Its antagonism EIEC test-results is listed in table 2.
Table 1: milk-acid bacteria numbering, strain name and source that the present invention uses
Table 2: the part milk-acid bacteria is to the restraining effect of EIEC
*Has significant difference (P＜0.05) between expression experimental group and the control group.
Table 2 is the result show, plant lactobacillus N13 has the obvious suppression effect to EIEC.Adopt different enzymes, pH and heat treated plant lactobacillus N13 fermented supernatant fluid (seeing accompanying drawing 5) to show that in its fermented supernatant fluid, playing inhibiting material is a kind of protein substance, this material has thermostability, under acidic conditions, has maximum activity.Therefore plant lactobacillus N13 supernatant is the comprehensive action of acid and protein-based antibacterial substance to the restraining effect of EIEC.
C, acid resisting test
The isolated strains (seeing accompanying drawing 1) of freezing preservation is inoculated in respectively in the MRS liquid nutrient medium; Cultivate 24h at 37 ℃; Go down to posterity so repeatedly cultivate 2～3 times after, get the said nutrient solution of 1mL, be inoculated in the MRS liquid nutrient medium of 19mL pH 2.0; Under 37 ℃ of temperature, cultivate 4h respectively, measure initial (0h) with cultivate finish after (4h) at the absorbancy OD at wavelength 600nm place
600Value, with the OD value of every strains of lactic acid bacteria 0h as reference, every strains of lactic acid bacteria OD when investigating cultivation 4h
600Increased value.Through measuring OD
600Variation size, compare with acid proof control strain (lactobacillus helveticus atcc-15019), obtained good acid proof probiotic lactobacillus.
These test-results are listed in accompanying drawing 1.This test-results shows that the increasing amount of plant lactobacillus N13 OD value after cultivating four hours under pH 2.0 conditions is 0.048, is higher than the increasing amount of acid resistance control strain lactobacillus helveticus atcc-15019, and therefore, plant lactobacillus N13 has good acidproof ability.
D, bile tolerance test
With Fel Sus domestica salt the bile salt levels in the MRS liquid nutrient medium is transferred to 1,2,3 ‰ (weight percents) respectively, then at 121 ℃ of following sterilization 15min of temperature.In MRS-biliary salts culture volume 1%, will use liquid MRS culture method activatory isolated strains (bacterial strain is seen accompanying drawing 2,3,4, and the bacterial strain specifying information is seen table 1) nutrient solution to be inoculated in the described MRS-biliary salts substratum, under 37 ℃ of temperature, cultivate.The OD of this probiotic lactobacillus nutrient solution of sampling and measuring when cultivating 0h, 3h and 6h
600Value repeats 3 times.As reference, investigate when cultivating 3h and 6h every strains of lactic acid bacteria OD with the OD value of every strains of lactic acid bacteria 0h
600Increased value, judge the bile tolerance ability of this bacterial strain with this.These test-results are listed in the accompanying drawing 2,3,4.These test-results show, plant lactobacillus N13 cultivates 3h in the MRS liquid nutrient medium that contains 1,2,3 ‰ pig cholate after, and the OD of this probiotic lactobacillus nutrient solution
600The increasing amount of value is respectively 0.56,0.48 and 0.34, this shows, plant lactobacillus N13 has good bile tolerance ability.
The part milk-acid bacteria that table 1 is listed has carried out acid resisting test and bile tolerance test; Its test-results shows; Wherein plant lactobacillus N13 has acid-resistant property and good anti-bile ability; Therefore, can think that plant lactobacillus N13 still has higher survival rate behind low acid environment that passes through human gastric juice and enteron aisle bile extreme environment, have the condition as good probiotic bacterium.
E, isolated strains are to the adherence test of intestinal epithelial cells strain HT-29 cell
Adopt the intestinal epithelial cells of 0.25% trypsin solution digestion by weight strain HT-29 cell, the RPMI-1640 cell culture fluid that uses U.S. Gibco company to produce dilutes, and its cell concn is adjusted to 2 * 10
5Individual/mL, in six orifice plates, in every hole, add earlier the deckglass of 18 * 18mm, add the above-mentioned cell culture fluid that contains the HT-29 cell of 2mL in each hole again, concussion makes cell evenly be tiled on the deckglass.Place 5% CO2gas incubator to cultivate 24h down for 37 ℃ then, make this cell be attached to the orifice plate bottom in temperature.Pipette and adopt liquid MRS culture method activatory three generations milk-acid bacteria (stationary phase) (bacterial strain is seen table 2, and the bacterial strain specifying information is seen table 1), clean 2 times, adjust its milk-acid bacteria concentration and reach 1～2 * 10 with PBS damping fluid (pH7.2)
8Cfu/mL.Each hole of six orifice plates is with the PBS buffer solution for cleaning of pH 7.2 3 times, and each hole adds said lactic acid bacterial liquid of 1mL and 1mLRPMI-1640 cell culture fluid (not containing two resisting and calf serums), in 5% CO2gas incubator, under 37 ℃ of temperature, hatches altogether two hours; The PBS buffer solution for cleaning of each Kong Zaiyong pH 7.2 5 times; To remove not adherent milk-acid bacteria, each hole is with the fixing 30min of anhydrous methanol, after the cleaning then; Take out deckglass and carry out gramstaining; Under oily mirror, choose 20 visuals field and observe, calculate adherent milk-acid bacteria number on 100 HT-29 cells, with lactobacillus rhamnosus (L.rhamnosus GG; ATCC533103) as contrast, filter out the high milk-acid bacteria of intestinal epithelial cells strain HT-29 cell adhesion ability.Its test-results is listed in table 3.
Table 3 part milk-acid bacteria is to the adhesion rate of HT-29 cell
*The difference (P＜0.01) that has highly significant between expression experimental group and the contrast.
The result of table 3 shows that plant lactobacillus N13 is the strongest to the adhesive capacity of intestinal epithelial cells strain HT-29 cell.
The inhibition test that F, milk-acid bacteria adhere to and attack EIEC
According to above-mentioned E) mode that step is identical carries out; Join probiotic lactobacillus (bacterial strain is seen accompanying drawing 6 and 7) in six orifice plates that are paved with intestinal epithelial cells strain HT-29 cell; Add 1mL probiotic lactobacillus bacterium liquid and 1mL RPMI-1640 cell culture fluid (not containing two resisting and calf serums) in each hole, in 37 ℃, 5% CO2gas incubator, hatched altogether two hours, then the PBS buffer solution for cleaning 5 times of each Kong Zaiyong pH 7.2; To remove not adherent milk-acid bacteria, in every hole, add concentration 1～2 * 10 again
8The EIEC of cfu/mL is hatched 4h altogether under 37 ℃ of temperature in 5% CO2gas incubator, with the PBS buffer solution for cleaning of pH 7.2 3 times, remove not adherent EIEC.With 0.5%Triton X-100 (polyoxyethylene octyl phenyl ether) lysing cell, the PBS damping fluid with pH 7.2 carries out the gradient series dilution again, and the dilution gradient is respectively 10
-2, 10
-3With 10
-4, and carry out plate count, calculating adheres on the cell and invades intracellular EIEC cell number.Likewise, hatch 1h altogether with 100 μ g/mL qingfengmeisu qiongs and this cell, to kill the EIEC cell that sticks on this cell, with 0.5%Triton X-100 lysing cell, the PBS damping fluid with pH 7.2 carries out the gradient series dilution again, and the dilution gradient is respectively 10
-1, 10
-2With 10
-3, and carry out plate count, calculate the cell number that invades EIEC in the cell.
Choose probiotic lactobacillus CCFM 235, N13, CCFM 232, CCFM 236, CCFM 231 with good acid resistance, bile tolerance property and high-adhesiveness; And be reference with plant lactobacillus LP-onlly, studied them EIEC is adhered to (seeing accompanying drawing 6) and attacks the effect of (seeing accompanying drawing 7) HT-29 cell inhibiting.
Can find out by accompanying drawing 6; High adherent plant lactobacillus N13 can obviously reduce the adhesion of EIEC to intestinal epithelial cells strain HT-29; Compare with control group (not adding the milk-acid bacteria intervention group); The EIEC number that adheres on the cell has reduced by 46%, explains that high adherent plant lactobacillus N13 can occupy EIEC and stick on the intestinal epithelial cells strain HT-29 cell in the attachment sites on the intestinal epithelial cells strain HT-29 cell or through steric restriction effect inhibition EIEC.
Can find out by accompanying drawing 7; The plant lactobacillus N13 of high-adhesiveness ability reduces the invasion and attack of EIEC to intestinal epithelial cells strain HT-29 cell equally significantly; EIEC reduces the reduction of the invasion and attack rate that has directly caused EIEC to the adhesion rate of HT-29, and N13 has the significant protection effect to intestinal epithelial cell HT-29 cell.
Through above-mentioned these evidences, plant lactobacillus N13 of the present invention (CCFM233) has property:
(1) pathogenic bacterium enteroinvasive E.Coli (ATCC 43893 for Enteroinvasive Escherichia coli., EIEC) had significant inhibitory effect;
(2) have acid resistance, under the condition of pH 2.0, still can grow;
(3) have the bile tolerance ability, under 1-3 ‰ cholate condition, can grow;
(4) intestinal epithelial cells strain HT-29 cell had good adhesive capacity;
(5) can obviously suppress the adhesion of EIEC on intestinal epithelial cells strain HT-29 cell;
(6) can obviously reduce the invasion and attack of EIEC to intestinal epithelial cells strain HT-29 cell.
The biological characteristics of described plant lactobacillus N13 is following:
(1) thalli morphology is shaft-like, does not move, and does not form gemma, Gram-positive;
(2) can obligate decompose sugar, main metabolites is a lactic acid;
(3) the catalase reaction is negative.
The spawn culture method of described plant lactobacillus N13 is following:
(1) the dull and stereotyped purifies and separates of MRS: choose plant lactobacillus N13 with transfering loop under the aseptic condition, on the MRS flat board, rule, flat board places incubator to cultivate 72h for 37 ℃, selects single bacterium colony and carries out microscopy, realizes the purebred separation of bacterial strain;
(2) bacterial strain activation: from-80 ℃, take out the cold storage bacterial classification, normal temperature melts down, and gets 100 microlitre fungus preserving liquids and (contain 30% glycerine; The lactic bacterium strains suspension of 70% sterilized water) is inoculated in the MRS liquid nutrient medium of sterilization; After 37 ℃ of temperature were grown 18-24h down, line was cultivated 72h down for 37 ℃ in temperature in solid MRS; Select the MRS liquid nutrient medium of single colony inoculation, experimentize behind the activation three generations in sterilization;
(3) composition of MRS substratum: 10g peptone, 10g Carnis Bovis seu Bubali cream, 5g yeast extract, 20g glucose, 2g potassium hydrogenphosphate, 5g sodium acetate, 2g trisodium citrate, 1g tween 80,200mg sal epsom, 54mg manganous sulfate and 1000mL zero(ppm) water; PH 6.4, this substratum 20min that under 115 ℃ of temperature, sterilizes;
(4) culture condition: under 37 ℃ of anaerobic condition and temperature, leave standstill cultivation;
(5) detection method of bacterium quantity: adopt ten times of serial dilution methods; Choose suitable extent of dilution, join diluent in the flat board, adopt the Standard Plate Count method; Pour 20mL solid MRS (temperature is lower than 50 ℃) in each flat board into; Mixing is waited to solidify the back and under 37 ℃ of temperature and anaerobic condition, is cultivated 72h, and the bacterium colony that forms is counted.
The invention still further relates to the purposes of described plant lactobacillus N13 in preparation soy yogurt, oat fermented drink, fermentation silage, fermented vegetables, lactobacillus milk beverage and cultured milk prod.
Described soy yogurt is to adopt following method preparation:
Volume ratio 2-4 according to soft water and soybean is following to soft water soaking soybean 1-2h at temperature 75-85 ℃; Remove soybean hulls; Remove soaked in water, again according to the weight ratio 1 of soybean and boiling water: 6-10 adds the boiling water defibrination, and its slurry keeps 10-15min under temperature 80-85 ℃ condition; Filter with 150 eye mesh screens then, obtain a kind of soya-bean milk; In this soya-bean milk, add sucrose earlier in soya-bean milk weight 5%-10%; Re-use clarifixator and under pressure 15-25MPa, carry out homogeneous; Under 95 ℃ of temperature, carry out germicidal treatment 10min subsequently; When its temperature is reduced to 36-38 ℃, insert the plant lactobacillus N13 starter dry powder formulations of commercial dry powder leaven lactobicillus bulgaricus, commercial dry powder leaven thermophilus streptococcus and following preparation again according to the inoculum size in soya-bean milk weight 0.04-0.05%, its part by weight is 1: 1: 1; Temperature 35-42 ℃ bottom fermentation 6-8 hour, promptly obtain containing the soy yogurt of plant lactobacillus N13 viable bacteria then 4 ℃ of stored refrigerated.
Described soft water should be appreciated that it is the water that does not contain or contain a small amount of solubility calcium, magnesium compound.
Water hardnessMainly by wherein calcium (Ca
2+), magnesium (Mg
2+) the ion formation.When the former water that contains hardness passed through the resin layer of interchanger, the calcium in the water, mg ion were discharged sodium ion simultaneously by resin absorption, and effusive water is exactly the softening water that has removed hardness ions in the interchanger like this.
In the present invention, described strainer is that Haining City joins the 150 purpose stainless steel duplex deep bed filter that many straining installations Science and Technology Ltd. sells.
In the present invention, the clarifixator that uses is the GJB type clarifixator (pressure is 10-80MPa) that Changzhou homogeneous machinery ltd produces.The employed equipment of sterilizing for example is the SANYO board Autoclave of being produced by Sanyo Electric company.
Plant lactobacillus N13 starter dry powder formulations prepares according to following method: plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 1-3%; Cultivate 20-28h down for 37 ℃ in temperature, make plant lactobacillus N13 viable count reach 10
8More than the cfu/mL; Its nutrient solution centrifugal 8-12min under the condition of rotating speed 3800-4200r/min and temperature 2-6 ℃ then; Its centrifugal sediment washes 2-4 time with the PBS damping fluid of pH 7.2 again, is added to its bacterial sediment in the lyophilized vaccine adjustment cell concn to 10 then
9Cfu/mL carries out vacuum lyophilization after mixing, obtain described plant lactobacillus N13 starter dry powder formulations after the freeze-drying.It is sucrose 1.5%, skimming milk 14%, lactose 5%, glycerine 1%, starch 3% and 75.5% water by weight that described lyophilized vaccine is formed.
In the present invention, the measuring method of plant lactobacillus N13 concentration is to adopt ten times of above-mentioned serial dilution methods, uses the MRS flat board to carry out Standard Plate Count method counting.
Described oat fermented drink is to adopt following method preparation:
A) with the oat be raw material, with oat selection, impurity elimination, removal epidermis and clean;
B) the 50-55 ℃ of water that in oat, adds in oat weight 100% mixes, and joins then and breaks into slurries in the hollander;
C) slurries that step B) obtain carry out defibrination with paste roller mill, use 150 purpose strainer filterings again, obtain a kind of slurries;
D) to step C) add in slurry weight 10-15% white sugar and 1000-1200% water in the slurries that obtain, at 121 ℃ of temperature sterilization 20min down, be cooled to 36-38 ℃, obtain the oat base-material that ferments;
E) plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 1-3%; Cultivate 20-24h down for 37 ℃ in temperature, spinning discards fermented supernatant fluid; Remaining bacterial sediment thing is resuspended with sterilized water, makes plant lactobacillus N13 viable count reach 10
8More than the cfu/mL, thereby obtain plant lactobacillus N13 liquid starter;
F) according to inoculum size with the volumeter 6-10% of oat fermentation base-material; Lactobicillus bulgaricus liquid starter, thermophilus streptococcus liquid starter and above-mentioned steps E) the plant lactobacillus N13 liquid starter of preparation is inoculated at step D according to volume ratio at 1: 1: 1) in the oat fermentation base-material of preparation; Cultivate 18-22h down at leavening temperature 36-38 ℃, make the concentration of plant lactobacillus N13 reach 10
6More than the cfu/mL, promptly obtain containing the oat fermented drink of plant lactobacillus N13 viable bacteria then temperature 3-4 ℃ of following stored refrigerated.
Described fermented vegetables is to adopt following method preparation:
A) vegetables of selecting for use through clean with drain after, soak with the 0.005-0.01g/kg chlorine bleach liquor and to carry out disinfection in 7-10 minute, with described vegetables remove the peel, cutting, obtain a kind of pre-treatment vegetables;
B) toward steps A) add according to said vegetables weight meter 0.6-1.0% sugar and 4.0-6.0% salt in the pre-treatment vegetables that obtain, stir, pickle 20-28h in advance in room temperature, drain away the water, obtain a kind of preparatory Pickle;
C) take by weighing edible spice, use, is added after the cooling in said edible spice weight 7-9% liquor at temperature 75-85 ℃ of following lixiviate 2-4h in 8-10 times of water of said edible spice weight again, obtains a kind of spice juice;
D) plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 1-3%; Cultivate 20-28h down for 37 ℃ in temperature, spinning discards fermented supernatant fluid; Remaining throw out is resuspended with sterilized water, makes plant lactobacillus N13 viable bacteria concentration reach 10
8More than the cfu/mL, thereby obtain a kind of bacteria suspension;
E) toward step B) add in said preparatory Pickle weight 12-14% at step C in the preparatory Pickle that obtains) the spice juice that obtains and 1-5% be at step D) bacteria suspension that obtains; Mixing; Tinning, sealing, at room temperature fermentation; Reach 3.6-4.0 up to the pH of its fermented liquid value, obtain described fermented vegetables like this.
According to the present invention, described vegetables are one or more vegetables that are selected from Caulis et Folium Brassicae capitatae, Radix Dauci Sativae, asparagus lettuce, pimento, onion or celery.
These vegetables need be removed the peel, and go processing such as mashed leaf, Caulis et Folium Brassicae capitatae, the Radix Dauci Sativae fibrous root of need pruning for example, and asparagus lettuce, onion need remove tertia, and pimento need be removed base of a fruit handle, and celery need be removed random leaf.
These vegetables are pickled with sugar and salt.Salt can make vegetable tissue softening, to preserve soluble cell content local flavor, can also prevent the spoilage organism breeding, reaches the purpose of preserving vegetables.The sugar that adds can be sucrose, glucose, fructose etc., and it can make taste soft, promotes appetite.
According to the present invention, described spice is that some have pungency fragrance, gives the food particular flavor, improves a poor appetite the plant seed that helps digest and absorb, bud, base of leaf, root piece and extract thereof.Spice all contains compositions such as volatile oil (essential oil), pungency component and organic acid, fiber, starch, colloid usually.
According to the present invention, described spice is one or more spices that are selected from capsicum, Chinese prickly ash, pepper, ginger, garlic, anise, fennel, cassia bark, orange peel or cloves.
The MRS liquid nutrient medium that uses among the present invention is the substratum that the technician in present technique field knows; It is formed as follows: 10g peptone, 10g Carnis Bovis seu Bubali cream, 5g yeast soak powder, 20g glucose, 2g potassium hydrogenphosphate, 5g sodium acetate, 2g trisodium citrate, 1g tween 80,200mg sal epsom, 54mg manganous sulfate and 1000mL zero(ppm) water; PH 6.4, and this substratum is sterilized under 115 ℃ of temperature and can be used behind the 20min.
Described fermented feed is to adopt following method preparation:
With the corn straw feedstuff raw material rub cut to 1～4cm for use;
Take by weighing 45-55kg corn straw feedstuff raw material; Be sprayed onto in the described corn straw feedstuff raw material after mixing the plant lactobacillus N13 starter dry powder formulations of the above-mentioned preparation of 0.5-0.7g and 200-240mL sterilized water; Turn mixed evenly after; 9-12 promptly obtains containing plant lactobacillus N13 after individual month corn straw feedstuff is stored in the compacting sealing in the silage bucket of packing into then under normal temperature condition.
Outside the corn straw as feedstuff raw material, can also use stalk and various green fodders such as paddy rice, corn, Chinese sorghum, wheat.Green fodder mainly comprises native grass, tame forage grass, weeds in field, dish leaf class, waterplant, spray leaf etc.
The PBS damping fluid that uses among the present invention (pH 7.2) is formed as follows: NaCl 137mmol/L, KCl 2.7mmol/L, Na
2HPO
410mmol/L, KH
2PO
42mmol/L.
Use plant lactobacillus N13 to prepare fermented-milk, its preparation method is following:
At first, preparation plant lactobacillus N13 working stock culture.
The skimming milk of above-mentioned preparation is subsequent use at 121 ℃ of following autoclaving 7min postcooling of temperature.With plant lactobacillus N13 bacterial classification according to being inoculated in the skimming milk with skimming milk volumeter 8-10%; Leave standstill under 37 ℃ of temperature that to cultivate 8-12h good to curdled milk; The skimming milk culture concussion of curdled milk is even; Again plant lactobacillus N13 bacterial classification is inoculated in the described curdled milk skimming milk according to the inoculum size with skimming milk volumeter 1-3%, leaves standstill under 37 ℃ of temperature that to cultivate 8-12h good to curdled milk, obtain plant lactobacillus N13 working stock culture.
Then, the preparation fermented-milk, its step is following:
A) select the fine raw dairy, filter, remove impurity with 200 eye mesh screens.Adopt ordinary method to carry out stdn then, make that lipid content is 8.0-11.0%, skim solid content is 22.5-24.5%.
B) in raw dairy, add sucrose, stirring and dissolving in raw dairy weight 8-12%.
C) emulsion stabilizer dissolving back being joined in the cow's milk by the 0.2-0.4% amount, is that 18-22MPa carries out homogeneous at pressure.
D) homogenized milk is heated to 90 ℃, keeps 10min to carry out pasteurize, cooling.
E) inoculate the plant lactobacillus N13 working stock culture (its volume ratio is 1: 1: 1) of lactobicillus bulgaricus, thermophilus streptococcus and above-mentioned preparation to the Ruzhong according to newborn volumeter 3-5%.
F) postvaccinal cow's milk ferments under 37 ℃ of temperature, and fermentation time is 6-8 hour, to curdled milk, then the yogurt after the curdled milk is placed 4 ℃ of refrigeration 22-26h after-ripening, promptly obtains described fermented-milk.
Described breast is cow's milk, horse breast, sheep is newborn or their mixture.
Use plant lactobacillus N13 to prepare lactobacillus milk beverage, its preparation method is following:
Take by weighing skim-milk and add the water stirring and dissolving, be prepared into the skimming milk of 10-12% by weight.According to inoculate in said skimming milk in skimming milk weight 3-5% inoculum size lactobicillus bulgaricus liquid starter, thermophilus streptococcus liquid starter and above the plant lactobacillus N13 starter (its volume ratio is 1: 1: 1) for preparing when the preparation fermented-milk ferment.Under the condition of 37 ℃ of temperature, cultivate 6-8h;, the pH of nutrient solution stops fermentation when reaching 4.3-4.4; Use clarifixator under the condition of pressure 18-22MPa, to carry out homogeneous 4-6min then, be cooled to 4 ℃ of temperature again, allocate toward wherein adding white sugar, compound stabilizer, Hydrocerol A, composite sweetener, essence, water then; Stirring and dissolving makes that the total acidity of fermentation system is 0.4-0.6%, pH 3.9-4.0.Use clarifixator under the condition of pressure 18-22MPa, to carry out the homogeneous 4-6min second time then, then under 95 ℃ of temperature, carry out sterilization 10min, carry out can after being cooled to below 10 ℃, promptly obtain described lactobacillus milk beverage.
[beneficial effect]
The invention has the beneficial effects as follows: plant lactobacillus N13 of the present invention (CCFM 233) has acid resistance, bile tolerance ability; Intestinal epithelial cells strain HT-29 cell had very high adhesivity; Has the EIEC of inhibition growth; Reduce the ability of EIEC, so N13 is the plant lactobacillus that a strain has good probiotic properties to the adhesion and the invasion and attack of colon epithelial cell.Soy yogurt, oat fermented drink, fermented vegetables, lactobacillus milk beverage, fermented-milk and fermentation silage prodn with plant lactobacillus N13 preparation of the present invention have extraordinary application prospect.
Plant lactobacillus (Lactobacillus plantarum) N13 bacterial strain on November 29th, 2011 in the preservation of common micro-organisms DSMZ of China Committee for Culture Collection of Microorganisms, its preserving number is CGMCC 5495.
[description of drawings]
Fig. 1 representes the growing state of different lactic bacterium strains after cultivating 4h under the condition of pH 2;
Fig. 2 is illustrated in the growing state of 1 ‰ pig gallbladder salinity condition lactic acid will bacterium;
Fig. 3 is illustrated in the growing state of 2 ‰ pig gallbladder salinity condition lactic acid will bacterium;
Fig. 4 is illustrated in the growing state of 3 ‰ pig gallbladder salinity condition lactic acid will bacterium;
Fig. 5 is illustrated in the CCFM233 supernatant of handling under the different condition it is suppressed the influence of EIEC;
The 1-untreated fish group; 2-trypsin treatment group; 3-Proteinase K treatment group; 4-katalase treatment group; 5-thermal treatment group; 6-fermented supernatant fluid pH is adjusted to neutral treatment group;
Fig. 6 representes that the milk-acid bacteria pre-treatment adheres to the influence of HT-29 cell to EIEC;
Fig. 7 representes that milk-acid bacteria and EIEC cultivate the influence to EIEC invasion and attack HT-29 cell altogether.
[embodiment]
Embodiment 1: utilize plant lactobacillus (Lactobacillus plantarum) N13 (CCFM 233) to make soy yogurt
The preparation method is following:
Prepare plant lactobacillus N13 starter dry powder formulations of the present invention: plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 3%, cultivates 24h down for 37 ℃ in temperature, makes plant lactobacillus N13 viable count reach 10
8More than the cfu/mL; Its nutrient solution centrifugal 10min under the condition of 4 ℃ of rotating speed 4000r/min and temperature then; Its centrifugal sediment washes 3 times with the PBS damping fluid of pH 7.2 again, then be added in the lyophilized vaccine adjustment cell concn to 10 through the centrifugal bacterial sediment that obtains of washing
9Cfu/mL carries out vacuum lyophilization after mixing, just obtain a kind of plant lactobacillus N13 starter dry powder formulations after the freeze-drying.The weight percent of described lyophilized vaccine consists of: sucrose 1.5%, skimming milk 14%, lactose 5%, glycerine 1%, starch 3% and 75.5% water.
Embodiment 2: utilize plant lactobacillus N13 to make fermentation pickled vegetable
The preparation method is following:
A) vegetables of selecting for use through clean with drain after, soak with the 0.008g/kg chlorine bleach liquor and to carry out disinfection in 8 minutes, with described vegetables remove the peel, cutting, obtain a kind of pre-treatment vegetables;
B) toward steps A) add according to said vegetables weight meter 0.8% sugar and 5.0% salt in the pre-treatment vegetables that obtain, stir, pickle 24h in advance in room temperature, drain away the water, obtain a kind of preparatory Pickle;
C) take by weighing edible spice, use, is added after the cooling in said edible spice weight 8% liquor at 80 ℃ of following lixiviate 3h of temperature in 8 times of water of said edible spice weight again, obtains a kind of spice juice;
D) plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 2%, cultivates 24h down for 37 ℃ in temperature, and spinning discards fermented supernatant fluid, and remaining throw out is resuspended with sterilized water, makes plant lactobacillus N13 viable bacteria concentration reach 10
8More than the cfu/mL, thereby obtain a kind of bacteria suspension;
E) toward step B) add in said preparatory Pickle weight 12% at step C in the preparatory Pickle that obtains) the spice juice and 3% that obtains is at step D) bacteria suspension that obtains; Mixing; Tinning, sealing, at room temperature fermentation; Reach 3.8 up to the pH of its fermented liquid value, obtain described fermented vegetables like this.
Embodiment 3: make the oat fermented drink that contains plant lactobacillus N13
The preparation method is following:
A) with the oat be raw material, with oat selection, impurity elimination, removal epidermis and clean;
B) 52 ℃ of water that in oat, add in oat weight 100% mix, and join then and break into slurries in the hollander;
C) slurries that step B) obtain carry out defibrination with paste roller mill, use 150 purpose strainer filterings again, obtain a kind of slurries;
D) to step C) add in slurry weight 12% white sugar and 1000% water in the slurries that obtain, at 121 ℃ of temperature sterilization 20min down, be cooled to 36-38 ℃, obtain oat fermentation base-material;
E) plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 2%, cultivates 24h down for 37 ℃ in temperature, and spinning discards fermented supernatant fluid, and remaining bacterial sediment thing is resuspended with sterilized water, makes plant lactobacillus N13 viable count reach 10
8More than the cfu/mL, thereby obtain plant lactobacillus N13 liquid starter.
F) with the plant lactobacillus CCFM233 liquid starter of above-mentioned preparation, commercial lactobicillus bulgaricus liquid starter and commercial thermophilus streptococcus liquid starter (three's volume ratio is 1: 1: 1); Inoculum size according to the volume 8% of oat fermentation base-material is linked at step D) in the oat fermentation base-material of preparation; At 37 ℃ of bottom fermentation 20h of leavening temperature, make plant lactobacillus CCFM233 concentration reach 10
6More than the cfu/mL, promptly obtain containing the oat fermented drink of plant lactobacillus N13 viable bacteria then temperature 3-4 ℃ of following stored refrigerated.
Embodiment 4: make the cornstalk silage that contains plant lactobacillus N13
The preparation method is following:
With the corn straw feedstuff raw material rub cut to 1～4cm for use;
Plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 1-3%, cultivates 24h down for 37 ℃ in temperature, makes plant lactobacillus N13 viable bacteria concentration reach 10
8More than the cfu/mL; Its nutrient solution centrifugal 10min under the condition of 4 ℃ of rotating speed 4000r/min and temperature then; Its centrifugal sediment washes 3 times with the PBS damping fluid of pH 7.2 again, then be added in the lyophilized vaccine adjustment cell concn to 10 through the centrifugal bacterial sediment that obtains of washing
9Cfu/mL carries out vacuum lyophilization after mixing, obtain a kind of plant lactobacillus N13 dry powder formulations after the freeze-drying; Said lyophilized vaccine is by 1.5% sucrose, 14% skimming milk, 5% lactose, 1% glycerine, 3% starch and 75.5% water are formed by weight;
Then; Take by weighing 50kg corn straw feedstuff raw material; Mix 0.6g described plant lactobacillus N13 dry powder formulations and 220mL sterilized water, sprinkling joins in the described corn straw feedstuff raw material, turns evenly mixed; The compacting sealing in the silage bucket of packing into then, storage promptly obtains containing the corn straw feedstuff of plant lactobacillus N13 after 10 months under normal temperature condition then.
Embodiment 5: preparation plant lactobacillus N13 starter, and preparation fermented-milk
The starter preparation method of plant lactobacillus N13 is following:
The skimming milk of this specification sheets embodiment 5 preparations is subsequent use at 121 ℃ of following autoclaving 7min postcooling of temperature.Plant lactobacillus N13 bacterial classification is inoculated in the skimming milk according to volumeter 10%; Leave standstill under 37 ℃ of temperature that to cultivate 10h good to curdled milk; The skimming milk culture concussion of curdled milk is even; Be inoculated in the described skimming milk according to inoculum size again and leave standstill under 37 ℃ of temperature that to cultivate 10h good to curdled milk, obtain plant lactobacillus N13 working stock culture with skimming milk volumeter 2%.
The method for preparing fermented-milk with above-mentioned starter is following:
Raw material: the plant lactobacillus N13 working stock culture of fresh cow milk, above-mentioned preparation, lactobicillus bulgaricus, thermophilus streptococcus, sucrose, emulsion stabilizer (Guangzhou is made a concerted effort to tie up ltd and produced all newborn plain CL 6121).
Concrete preparation process is following:
A) select fine raw material cow's milk, filter, remove impurity with 200 eye mesh screens.Carry out stdn then, make that lipid content is 10.0%, skim solid content is 23.5%.
B) in raw dairy, add sucrose, stirring and dissolving in raw dairy weight 10%.
C) emulsion stabilizer dissolving back being joined in the cow's milk by 0.3% amount, is that 20MPa carries out homogeneous at pressure.
D) cow's milk with homogeneous is heated to 90 ℃, keeps 10min to carry out pasteurize, cooling.
E) in cow's milk, inoculate the plant lactobacillus N13 starter of lactobicillus bulgaricus liquid starter, thermophilus streptococcus liquid starter and above-mentioned preparation according to cow's milk volumeter 4%, its volume ratio is 1: 1: 1.
F) postvaccinal cow's milk fermented 8 hours under 37 ℃ of temperature, to curdled milk, then the cultured milk after the curdled milk was placed 4 ℃ of refrigeration 24h after-ripening, promptly obtained described fermented-milk.
Embodiment 6: use plant lactobacillus N13 to prepare lactobacillus milk beverage:
The preparation method is following:
Raw material: skim-milk (inspiring confidence in the nz NZMP skim-milk that company sells among the Tian Ze of Tianjin), lactobacillus inoculation (plant lactobacillus N13), stablizer (Guangzhou is made a concerted effort to tie up ltd and produced all newborn plain CL 6121), white sugar.Batching is formed: the good sweeting agent ltd that adds in compound stabilizer, 0.4% Hydrocerol A, 0.2% Shenzhen of selling with composite peanut milk soup stablizer trade(brand)name in skim-milk weight 35% white sugar, 0.3% Xiamen Kang Hui biotechnology Science and Technology Ltd. is a skim-milk with the composite sweetener of the sweet trade(brand)name sale of knob, essence, the surplus that 0.06% Shanghai Ke Dao essence and flavoring agent company sells with prairiedir milk flavour (58509) trade(brand)name.
Preparation process: take by weighing skim-milk and add the water stirring and dissolving, be prepared into 10% skimming milk by weight.Plant lactobacillus N13 working stock culture (its volume ratio is 1: 1: 1) according in said skimming milk, inoculate lactobicillus bulgaricus liquid-liquid starter, thermophilus streptococcus liquid-liquid starter and embodiment 5 preparations in skimming milk weight 4% inoculum size ferments.Under the condition of 37 ℃ of temperature, cultivate 8h;, the pH of nutrient solution stops fermentation when reaching 4.3-4.4; Use clarifixator under the condition of pressure 20MPa, to carry out homogeneous 5min then, be cooled to 4 ℃ of temperature again, allocate toward wherein adding white sugar, compound stabilizer, Hydrocerol A, composite sweetener, essence, water then; Stirring and dissolving makes that the total acidity of fermentation system is 0.5%, pH 3.9-4.0.Use clarifixator under the condition of pressure 20MPa, to carry out the homogeneous 5min second time then, then under 95 ℃ of temperature, carry out sterilization 10min, carry out can after being cooled to below 10 ℃, promptly obtain described lactobacillus milk beverage.
Claims (9)
1. a plant lactobacillus (Lactobacillus plantarum) N13, this bacteria strain on November 29th, 2011 in the preservation of common micro-organisms DSMZ of China Committee for Culture Collection of Microorganisms, its preserving number is CGMCC 5495.
2. plant lactobacillus N13 according to claim 1 is characterized in that it has property:
(1) pathogenic bacterium enteroinvasive E.Coli (ATCC 43893 for Enteroinvasive Escherichia coli., EIEC) had significant inhibitory effect;
(2) have acid resistance, under the condition of pH 2.0, can grow;
(3) have the bile tolerance ability, under 1-3 ‰ cholate condition, can grow;
(4) intestinal epithelial cells strain HT-29 cell had very high adhesive capacity;
(5) can obviously suppress the adhesion of EIEC to intestinal epithelial cells strain HT-29 cell;
(6) can obviously reduce the invasion and attack of EIEC to intestinal epithelial cells strain HT-29 cell.
3. the purposes of plant lactobacillus N13 according to claim 1 in preparation soy yogurt, oat fermented drink, fermentation silage, fermented vegetables, lactobacillus milk beverage and cultured milk prod.
4. purposes according to claim 3 is characterized in that the described soy yogurt that contains plant lactobacillus N13 is to adopt following method preparation:
Volume ratio 2-4 according to soft water and soybean is following to soft water soaking soybean 1-2h at temperature 75-85 ℃; Remove soybean hulls; Remove soaked in water, again according to the weight ratio 1 of soybean and boiling water: 6-10 adds the boiling water defibrination, and its slurry keeps 10-15min under temperature 80-85 ℃ condition; Filter with 150 eye mesh screens then, obtain a kind of soya-bean milk; In this soya-bean milk, add sucrose earlier in soya-bean milk weight 5%-10%; Re-use clarifixator and under pressure 15-25MPa, carry out homogeneous; Under 95 ℃ of temperature, carry out germicidal treatment 10min subsequently; When its temperature is reduced to 36-38 ℃, insert the plant lactobacillus N13 starter dry powder formulations of commercial dry powder leaven lactobicillus bulgaricus, commercial dry powder leaven thermophilus streptococcus and following preparation again according to the inoculum size in soya-bean milk weight 0.04-0.05%, its part by weight is 1: 1: 1; Temperature 35-42 ℃ bottom fermentation 6-8 hour, promptly obtain containing the soy yogurt of plant lactobacillus N13 viable bacteria then 4 ℃ of stored refrigerated;
Plant lactobacillus N13 starter dry powder formulations prepares according to following method: plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 1-3%; Cultivate 20-28h down for 37 ℃ in temperature, make plant lactobacillus N13 viable count reach 10
8More than the cfu/mL; Its nutrient solution centrifugal 8-12min under the condition of rotating speed 3800-4200r/min and temperature 2-6 ℃ then; Its centrifugal sediment washes 2-4 time with the PBS damping fluid of pH 7.2 again, is added to its bacterial sediment in the lyophilized vaccine adjustment cell concn to 10 then
9Cfu/mL carries out vacuum lyophilization after mixing, obtain described plant lactobacillus N13 starter dry powder formulations after the freeze-drying; It is sucrose 1.5%, skimming milk 14%, lactose 5%, glycerine 1%, starch 3% and 75.5% water by weight that described lyophilized vaccine is formed.
5. purposes according to claim 3 is characterized in that described oat fermented drink is to adopt following method preparation:
A) with the oat be raw material, with oat selection, impurity elimination, removal epidermis and clean;
B) the 50-55 ℃ of water that in oat, adds in oat weight 100% mixes, and joins then and breaks into slurries in the hollander;
C) slurries that step B) obtain carry out defibrination with paste roller mill, use 150 purpose strainer filterings again, obtain a kind of slurries;
D) to step C) add in slurry weight 10-15% white sugar and 1000-1200% water in the slurries that obtain, at 121 ℃ of temperature sterilization 20min down, be cooled to 36-38 ℃, obtain the oat base-material that ferments;
E) plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 1-3%; Cultivate 20-24h down for 37 ℃ in temperature, spinning discards fermented supernatant fluid; Remaining bacterial sediment thing is resuspended with sterilized water, makes plant lactobacillus N13 viable count reach 10
8More than the cfu/mL, thereby obtain plant lactobacillus N13 liquid starter;
F) according to volumeter 6-10% inoculum size with oat fermentation base-material; Lactobicillus bulgaricus liquid starter, thermophilus streptococcus liquid starter and above-mentioned steps E) the plant lactobacillus N13 liquid starter of preparation is inoculated at step D according to volume ratio at 1: 1: 1) in the oat fermentation base-material of preparation; Cultivate 18-22h down at leavening temperature 36-38 ℃, make the concentration of plant lactobacillus N13 reach 10
6More than the cfu/mL, promptly obtain containing the oat fermented drink of plant lactobacillus N13 viable bacteria then temperature 3-4 ℃ of following stored refrigerated.
6. purposes according to claim 3 is characterized in that described fermented vegetables is to adopt following method preparation:
A) vegetables of selecting for use through clean with drain after, soak with the 0.005-0.01g/kg chlorine bleach liquor and to carry out disinfection in 7-10 minute, with described vegetables remove the peel, cutting, obtain a kind of pre-treatment vegetables;
B) toward steps A) add according to said vegetables weight meter 0.6-1.0% sugar and 4.0-6.0% salt in the pre-treatment vegetables that obtain, stir, pickle 20-28h in advance in room temperature, drain away the water, obtain a kind of preparatory Pickle;
C) take by weighing edible spice, use, is added after the cooling in said edible spice weight 7-9% liquor at temperature 75-85 ℃ of following lixiviate 2-4h in 8-10 times of water of said edible spice weight again, and mixing obtains a kind of spice juice;
D) plant lactobacillus N13 is inoculated in the MRS liquid nutrient medium with MRS liquid nutrient medium volumeter 1-3%; Cultivate 20-28h down for 37 ℃ in temperature, spinning discards fermented supernatant fluid; Remaining throw out is resuspended with sterilized water, makes plant lactobacillus N13 viable bacteria concentration reach 10
8More than the cfu/mL, thereby obtain a kind of bacteria suspension;
E) toward step B) add in said preparatory Pickle weight 12-14% at step C in the preparatory Pickle that obtains) the spice juice that obtains and 1-5% be at step D) bacteria suspension that obtains; Mixing; Tinning, sealing, at room temperature fermentation; Reach 3.6-4.0 up to the pH of its fermented liquid value, obtain described fermented vegetables like this.
7. according to claim 3 or 6 described purposes, it is characterized in that described vegetables are one or more vegetables that are selected from Caulis et Folium Brassicae capitatae, Radix Dauci Sativae, asparagus lettuce, pimento, onion or celery.
8. purposes according to claim 6 is characterized in that described spice is one or more spices that are selected from capsicum, Chinese prickly ash, pepper, ginger, garlic, anise, fennel, cassia bark, orange peel or cloves.
9. purposes according to claim 3 is characterized in that described fermentation silage is to adopt following method preparation:
With the corn straw feedstuff raw material rub cut to 1～4cm for use;
Take by weighing 45-55kg corn straw feedstuff raw material; Be sprayed onto in the described corn straw feedstuff raw material after mixing 0.5-0.7g plant lactobacillus N13 according to claim 4 dry powder formulations and 200-240mL sterilized water; Turn mixed evenly after; 9-12 promptly obtains containing plant lactobacillus N13 after individual month corn straw feedstuff is stored in the compacting sealing in the silage bucket of packing into then under normal temperature condition.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012100578224A CN102586155B (en) | 2012-03-07 | 2012-03-07 | Lactobacillus plantarum N13 and use thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012100578224A CN102586155B (en) | 2012-03-07 | 2012-03-07 | Lactobacillus plantarum N13 and use thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN102586155A true CN102586155A (en) | 2012-07-18 |
| CN102586155B CN102586155B (en) | 2013-04-03 |
Family
ID=46475394
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2012100578224A Active CN102586155B (en) | 2012-03-07 | 2012-03-07 | Lactobacillus plantarum N13 and use thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102586155B (en) |
Cited By (34)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102960620A (en) * | 2012-12-12 | 2013-03-13 | 江南大学 | Direct-throwing type sour dough fermenting agent as well as preparation method and application thereof |
| CN103060228A (en) * | 2012-12-12 | 2013-04-24 | 广州立达尔生物科技股份有限公司 | Lactobacillus plantarum and micro-ecological preparation for silage |
| CN103205374A (en) * | 2013-03-15 | 2013-07-17 | 厦门和美科盛生物技术有限公司 | Lactic acid bacterium capable of reducing aflatoxin B1 in fermented feed and application of lactic acid bacterium |
| CN103300333A (en) * | 2013-06-27 | 2013-09-18 | 四川省旺平食品有限责任公司 | Method for producing pickled vegetables with lactic acid bacteria |
| CN103584251A (en) * | 2013-11-07 | 2014-02-19 | 南昌大学 | Ginger preservative or bacteriostatic agent |
| CN103637132A (en) * | 2013-11-05 | 2014-03-19 | 浙江省农业科学院 | Rapid processing method of low-salt mustard leaf by inverted compaction |
| CN103750225A (en) * | 2013-11-07 | 2014-04-30 | 南昌大学 | Method for improving organoleptic quality of ginger |
| CN104207115A (en) * | 2014-08-26 | 2014-12-17 | 吉林农业大学 | Lactobacillus plantaru fermented shiitake mushroom and preparation method thereof |
| CN104293689A (en) * | 2014-01-29 | 2015-01-21 | 浙江农林大学 | Lactobacillus strain with high adhesive power, and application thereof |
| CN104489084A (en) * | 2014-12-29 | 2015-04-08 | 江南大学 | Lactobacillus plantarum for producing high-ester fermented bactrian camel milk |
| CN105062918A (en) * | 2015-07-31 | 2015-11-18 | 吉林省命之元生物科技有限公司 | Lactobacillus plantarum and applications thereof |
| CN105238727A (en) * | 2015-11-13 | 2016-01-13 | 江南大学 | Lactobacillus plantarum capable of antagonizing campylobacter jejuni and inhibiting expression of flaA gene of campylobacter jejuni |
| CN105255752A (en) * | 2015-04-27 | 2016-01-20 | 南昌大学 | Lactobacillus plantarum ZDY2013 with high acid resistance |
| CN105420152A (en) * | 2015-12-16 | 2016-03-23 | 石家庄君乐宝乳业有限公司 | High-activity lactobacillus plantarum N3117 as well as screening method and application thereof |
| CN105505814A (en) * | 2015-11-30 | 2016-04-20 | 南昌大学 | Lactobacillus plantarum capable of delaying aging |
| CN105494629A (en) * | 2015-11-12 | 2016-04-20 | 南昌大学 | Preparation method of yoghourt |
| CN105969681A (en) * | 2016-03-28 | 2016-09-28 | 四川农业大学 | Highly oxidation-resistant, cholate-tolerant and acid-resistant lactobacillus plantarum JR4 and application thereof |
| CN106616042A (en) * | 2016-12-23 | 2017-05-10 | 江南大学 | Lactobacillus plantarum feed and application thereof |
| CN107312726A (en) * | 2017-05-27 | 2017-11-03 | 武汉科缘生物发展有限责任公司 | One lactobacillus plantarum ZN 3 and application |
| CN107354104A (en) * | 2017-05-03 | 2017-11-17 | 华中农业大学 | A kind of branch bacillus for Pickle |
| CN107988094A (en) * | 2017-11-14 | 2018-05-04 | 北京好实沃生物技术有限公司 | One plant of lactobacillus fermenti HEW-A846 and its application |
| CN108102983A (en) * | 2018-02-09 | 2018-06-01 | 上海交通大学 | A kind of lactobacillus plantarum of high yield amylase and its application |
| CN109619457A (en) * | 2019-01-11 | 2019-04-16 | 大连工业大学 | The method of lactobacillus plantarum and its mixed fermentation acid Preserved-fish fish |
| CN109619195A (en) * | 2018-12-29 | 2019-04-16 | 光明乳业股份有限公司 | A kind of oat base normal temperature acid drink and preparation method thereof |
| JP2019187251A (en) * | 2018-04-19 | 2019-10-31 | 国立研究開発法人農業・食品産業技術総合研究機構 | Method for producing fermentation product and fermentation product |
| CN111011685A (en) * | 2019-12-30 | 2020-04-17 | 江南大学 | Fermented milk rich in probiotics and preparation method thereof |
| CN111269849A (en) * | 2019-12-30 | 2020-06-12 | 北京博锦元生物科技有限公司 | Lactobacillus plantarum and application thereof |
| CN111436203A (en) * | 2018-11-12 | 2020-07-21 | 中粮营养健康研究院有限公司 | Fermented lactobacillus plantarum and application thereof |
| CN111705011A (en) * | 2020-03-30 | 2020-09-25 | 四川农业大学 | Lactobacillus plantarum C20 for vegetable tailed vegetable silage fermentation and application thereof |
| CN111705086A (en) * | 2020-07-03 | 2020-09-25 | 兰州大学 | Serous acellular culture solution for resisting digestive system tumors |
| CN113186139A (en) * | 2021-06-21 | 2021-07-30 | 西北民族大学 | Lactobacillus plantarum LR002 and application thereof |
| CN114437997A (en) * | 2022-04-07 | 2022-05-06 | 山东向日葵生物工程有限公司 | Lactobacillus plantarum SF-L38 and application thereof in preparation of blood sugar control product |
| CN114940956A (en) * | 2022-05-10 | 2022-08-26 | 中国农业科学院农产品加工研究所 | Lactobacillus plantarum WL-100 and application thereof |
| CN115058372A (en) * | 2022-08-11 | 2022-09-16 | 中国肉类食品综合研究中心 | Compound leavening agent and application thereof in low-sugar bubble type fermented soybean milk |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1844363A (en) * | 2006-04-21 | 2006-10-11 | 江南大学 | Lactobacillus plantarum CW006 with antihypertensive function |
| CN102100252A (en) * | 2009-12-18 | 2011-06-22 | 光明乳业股份有限公司 | Fermented dairy product and preparation method and raw material composition thereof, lactobacillus plantarum application |
-
2012
- 2012-03-07 CN CN2012100578224A patent/CN102586155B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1844363A (en) * | 2006-04-21 | 2006-10-11 | 江南大学 | Lactobacillus plantarum CW006 with antihypertensive function |
| CN102100252A (en) * | 2009-12-18 | 2011-06-22 | 光明乳业股份有限公司 | Fermented dairy product and preparation method and raw material composition thereof, lactobacillus plantarum application |
Non-Patent Citations (1)
| Title |
|---|
| M.VESCOVO ET AL.: "application of antimicrobial-producing lactic acid bacteria to control pathogens in ready-to-use vegetables", 《JOURNAL OF APPLIED MICROBIOLOGY》, vol. 81, no. 2, 31 August 1996 (1996-08-31) * |
Cited By (50)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103060228B (en) * | 2012-12-12 | 2014-06-25 | 广州立达尔生物科技股份有限公司 | Lactobacillus plantarum and micro-ecological preparation for silage |
| CN103060228A (en) * | 2012-12-12 | 2013-04-24 | 广州立达尔生物科技股份有限公司 | Lactobacillus plantarum and micro-ecological preparation for silage |
| CN102960620A (en) * | 2012-12-12 | 2013-03-13 | 江南大学 | Direct-throwing type sour dough fermenting agent as well as preparation method and application thereof |
| CN103205374A (en) * | 2013-03-15 | 2013-07-17 | 厦门和美科盛生物技术有限公司 | Lactic acid bacterium capable of reducing aflatoxin B1 in fermented feed and application of lactic acid bacterium |
| CN103205374B (en) * | 2013-03-15 | 2015-05-13 | 厦门和美科盛生物技术有限公司 | Lactic acid bacterium capable of reducing aflatoxin B1 in fermented feed and application of lactic acid bacterium |
| CN103300333A (en) * | 2013-06-27 | 2013-09-18 | 四川省旺平食品有限责任公司 | Method for producing pickled vegetables with lactic acid bacteria |
| CN103637132A (en) * | 2013-11-05 | 2014-03-19 | 浙江省农业科学院 | Rapid processing method of low-salt mustard leaf by inverted compaction |
| CN103750225A (en) * | 2013-11-07 | 2014-04-30 | 南昌大学 | Method for improving organoleptic quality of ginger |
| CN103584251A (en) * | 2013-11-07 | 2014-02-19 | 南昌大学 | Ginger preservative or bacteriostatic agent |
| CN104293689A (en) * | 2014-01-29 | 2015-01-21 | 浙江农林大学 | Lactobacillus strain with high adhesive power, and application thereof |
| CN104207115A (en) * | 2014-08-26 | 2014-12-17 | 吉林农业大学 | Lactobacillus plantaru fermented shiitake mushroom and preparation method thereof |
| CN104489084A (en) * | 2014-12-29 | 2015-04-08 | 江南大学 | Lactobacillus plantarum for producing high-ester fermented bactrian camel milk |
| CN107593908A (en) * | 2014-12-29 | 2018-01-19 | 江南大学 | A kind of Lactobacillus plantarum for being used to produce high ester fermentation bactrian camel milk |
| CN105255752A (en) * | 2015-04-27 | 2016-01-20 | 南昌大学 | Lactobacillus plantarum ZDY2013 with high acid resistance |
| CN105255752B (en) * | 2015-04-27 | 2018-08-17 | 南昌大学 | High acid plant lactobacillus ZDY2013 |
| CN105062918A (en) * | 2015-07-31 | 2015-11-18 | 吉林省命之元生物科技有限公司 | Lactobacillus plantarum and applications thereof |
| CN105062918B (en) * | 2015-07-31 | 2019-01-08 | 吉林省命之元生物科技有限公司 | One lactobacillus plantarum and its application |
| CN105494629A (en) * | 2015-11-12 | 2016-04-20 | 南昌大学 | Preparation method of yoghourt |
| CN105238727A (en) * | 2015-11-13 | 2016-01-13 | 江南大学 | Lactobacillus plantarum capable of antagonizing campylobacter jejuni and inhibiting expression of flaA gene of campylobacter jejuni |
| CN105238727B (en) * | 2015-11-13 | 2019-01-11 | 江南大学 | A kind of antagonism campylobacter jejuni and the lactobacillus plantarum for inhibiting its flaA gene expression |
| CN105505814B (en) * | 2015-11-30 | 2020-01-17 | 南昌大学 | Lactobacillus plantarum for delaying senescence |
| CN105505814A (en) * | 2015-11-30 | 2016-04-20 | 南昌大学 | Lactobacillus plantarum capable of delaying aging |
| CN105420152A (en) * | 2015-12-16 | 2016-03-23 | 石家庄君乐宝乳业有限公司 | High-activity lactobacillus plantarum N3117 as well as screening method and application thereof |
| CN105420152B (en) * | 2015-12-16 | 2019-04-16 | 石家庄君乐宝乳业有限公司 | Lactobacillus plantarum N3117, its screening technique and application |
| CN105969681A (en) * | 2016-03-28 | 2016-09-28 | 四川农业大学 | Highly oxidation-resistant, cholate-tolerant and acid-resistant lactobacillus plantarum JR4 and application thereof |
| CN106616042A (en) * | 2016-12-23 | 2017-05-10 | 江南大学 | Lactobacillus plantarum feed and application thereof |
| CN107354104A (en) * | 2017-05-03 | 2017-11-17 | 华中农业大学 | A kind of branch bacillus for Pickle |
| CN107354104B (en) * | 2017-05-03 | 2019-09-20 | 华中农业大学 | A kind of branch bacillus for Pickle |
| CN107312726A (en) * | 2017-05-27 | 2017-11-03 | 武汉科缘生物发展有限责任公司 | One lactobacillus plantarum ZN 3 and application |
| CN107988094A (en) * | 2017-11-14 | 2018-05-04 | 北京好实沃生物技术有限公司 | One plant of lactobacillus fermenti HEW-A846 and its application |
| CN108102983B (en) * | 2018-02-09 | 2021-02-09 | 上海交通大学 | Lactobacillus plantarum for high yield of amylase and application thereof |
| CN108102983A (en) * | 2018-02-09 | 2018-06-01 | 上海交通大学 | A kind of lactobacillus plantarum of high yield amylase and its application |
| JP2019187251A (en) * | 2018-04-19 | 2019-10-31 | 国立研究開発法人農業・食品産業技術総合研究機構 | Method for producing fermentation product and fermentation product |
| CN111436203B (en) * | 2018-11-12 | 2022-06-28 | 中粮营养健康研究院有限公司 | Fermented lactobacillus plantarum and application thereof |
| CN111436203A (en) * | 2018-11-12 | 2020-07-21 | 中粮营养健康研究院有限公司 | Fermented lactobacillus plantarum and application thereof |
| CN109619195A (en) * | 2018-12-29 | 2019-04-16 | 光明乳业股份有限公司 | A kind of oat base normal temperature acid drink and preparation method thereof |
| CN109619457A (en) * | 2019-01-11 | 2019-04-16 | 大连工业大学 | The method of lactobacillus plantarum and its mixed fermentation acid Preserved-fish fish |
| CN111269849A (en) * | 2019-12-30 | 2020-06-12 | 北京博锦元生物科技有限公司 | Lactobacillus plantarum and application thereof |
| CN111011685A (en) * | 2019-12-30 | 2020-04-17 | 江南大学 | Fermented milk rich in probiotics and preparation method thereof |
| CN111705011A (en) * | 2020-03-30 | 2020-09-25 | 四川农业大学 | Lactobacillus plantarum C20 for vegetable tailed vegetable silage fermentation and application thereof |
| CN111705011B (en) * | 2020-03-30 | 2021-09-14 | 四川农业大学 | Lactobacillus plantarum C20 for vegetable tailed vegetable silage fermentation and application thereof |
| CN111705086A (en) * | 2020-07-03 | 2020-09-25 | 兰州大学 | Serous acellular culture solution for resisting digestive system tumors |
| CN111705086B (en) * | 2020-07-03 | 2023-04-07 | 兰州大学 | Serous acellular culture solution for resisting digestive system tumors |
| CN113186139A (en) * | 2021-06-21 | 2021-07-30 | 西北民族大学 | Lactobacillus plantarum LR002 and application thereof |
| CN113186139B (en) * | 2021-06-21 | 2022-06-07 | 西北民族大学 | Lactobacillus plantarum LR002 and application thereof |
| CN114437997A (en) * | 2022-04-07 | 2022-05-06 | 山东向日葵生物工程有限公司 | Lactobacillus plantarum SF-L38 and application thereof in preparation of blood sugar control product |
| CN114437997B (en) * | 2022-04-07 | 2022-06-10 | 山东向日葵生物工程有限公司 | Lactobacillus plantarum SF-L38 and application thereof in preparation of blood sugar control product |
| CN114940956A (en) * | 2022-05-10 | 2022-08-26 | 中国农业科学院农产品加工研究所 | Lactobacillus plantarum WL-100 and application thereof |
| CN115058372A (en) * | 2022-08-11 | 2022-09-16 | 中国肉类食品综合研究中心 | Compound leavening agent and application thereof in low-sugar bubble type fermented soybean milk |
| CN115058372B (en) * | 2022-08-11 | 2022-11-08 | 中国肉类食品综合研究中心 | Compound leavening agent and application thereof in low-sugar bubble type fermented soybean milk |
Also Published As
| Publication number | Publication date |
|---|---|
| CN102586155B (en) | 2013-04-03 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102586155B (en) | Lactobacillus plantarum N13 and use thereof | |
| CN104830731B (en) | One strain has plant lactobacillus AB-1 and the application thereof of broad-spectrum antibacterial characteristic | |
| CN103725633B (en) | A kind of pickle fermentation bacteria agent and preparation method and application | |
| CN102174450B (en) | Lactobacillus plantarum for resisting helicobacter pylori infection and application thereof | |
| JP3789303B2 (en) | New species of microorganisms and their use | |
| CN105779350B (en) | A kind of lactobacillus plantarum and application thereof of anti-Bacterium enteritidis infection | |
| CN1911118B (en) | Kefir mushroom freeze-dried powder, production method and use thereof | |
| CN105707648A (en) | Anti-fungus biological protection agent and application thereof | |
| CN103651742A (en) | Biological preservative with food preservation effect and application thereof | |
| CN103070285A (en) | Microbe feed additive and preparation method thereof | |
| JP2006325576A (en) | Fermented product of unpolished rice, method for producing the same, beverage and food containing the same, method for producing fermented and germinated unpolished rice and the fermented and germinated unpolished rice | |
| CN102986872A (en) | Fermented egg yoghurt containing chickpeas and method for preparing egg yoghurt | |
| CN102559563B (en) | Pediococcus acidilactici CCFM7902 and application thereof | |
| CN100589710C (en) | An instant pickled vegetables containing live probiotics and preparation method thereof | |
| CN104611256B (en) | A kind of microorganism lyophilized formulations and preparation method thereof | |
| KR20090086796A (en) | Lactobacillus plantarum bls41 originated from kimchi and use thereof | |
| CN102329763A (en) | Lactobacillus helveticus H9 and application of Lactobacillus helveticus H9 to preparation of fermented milk | |
| CN103952336A (en) | Bacillus licheniformis-Bacillus subtilis-Lactobacillus casei preparation and preparation method thereof | |
| JP2008048683A (en) | Viable bacteria agent for animal, feed composition, and method for using the agent and the composition | |
| CN103667093A (en) | Lactobacillus plantarum 929-2 strain having food preservative and fresh-keeping effect and application thereof | |
| CN105670959A (en) | Probiotic strain and application thereof | |
| CN101437414B (en) | Antimicrobial preparations | |
| CN109393390B (en) | Fermented hawthorn pulp and preparation method thereof | |
| KR100951273B1 (en) | Feed additive containing an extract of bamboo and microorganism cultivate,for prevention of Porcinediarrhea caused by enterotoxigenic enterobacteria | |
| CN103952345B (en) | Probiotic preparation containing Lactobacillus casei and Bacillus licheniformis and preparation method thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| TR01 | Transfer of patent right | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20181123 Address after: No. 1033 Longqiao Road, Wujiang District, Suzhou City, Jiangsu Province, 215000 Patentee after: JIANGSU WECARE BIOTECHNOLOGY CO., LTD. Address before: 1800 No. 214122 Jiangsu city of Wuxi Province Li Lake Avenue Patentee before: Jiangnan University |
|
| CP03 | Change of name, title or address | ||
| CP03 | Change of name, title or address |
Address after: 215000 No. 1033, long Qiao Road, Wujiang economic and Technological Development Zone, Suzhou, Jiangsu Patentee after: Weikang probiotics (Suzhou) Co.,Ltd. Address before: No. 1033 Longqiao Road, Wujiang District, Suzhou City, Jiangsu Province, 215000 Patentee before: JIANGSU WECARE BIOTECHNOLOGY Co.,Ltd. |