CN1345589A - Medicinal preparation containing lactobacillus acidophilus and preparation process thereof - Google Patents

Medicinal preparation containing lactobacillus acidophilus and preparation process thereof Download PDF

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CN1345589A
CN1345589A CN00125469A CN00125469A CN1345589A CN 1345589 A CN1345589 A CN 1345589A CN 00125469 A CN00125469 A CN 00125469A CN 00125469 A CN00125469 A CN 00125469A CN 1345589 A CN1345589 A CN 1345589A
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bacillus acidophilus
preparation
vagina
acid
strain
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CN1234373C (en
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吴刚
刘京
夏凌云
高原
杜育华
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Shanghai Xinyi Pharmaceutical Co. Ltd..
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XINYI PHARMACEUTIC CO Ltd SHANGHAI
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Abstract

Medication preparation containing acidophilic lactobacillus is composed of acidophilic lactobacillus and medicinal excipient. The invented preparations are types of liquid, solid, frost cream, bougie or nebula. It can be used to cure or prevent the diseases of vagina, infection gonorrhea.

Description

The pharmaceutical preparation and the preparation method that contain the bacillus acidophilus
The present invention relates to biological preparation, be specifically related to a kind of bacillus acidophilus's of containing pharmaceutical preparation and preparation method.
Parasitize normal intravaginal micropopulation and mainly contain antibacterial, fungus, protozoon and virus.Secondly their main perch are cervix uteri in the gauffer and the fornix portion of mucosa.Main permanent bacterium has in intravaginal: lactobacillus, staphylococcus epidermidis, escherichia coli, corynebacterium, B family streptococcus, streptococcus faecalis, mycoplasma, Candida albicans, dyspepsiacoccus and bacteroid etc.General record with quantitative methods with qualitative that viable bacteria is 10 in the healthy women vaginal discharge 2CFU/ml-10 9CFU/ml, anaerobe is 5: 1 with the ratio of aerobe, wherein lactobacillus is the permanent bacterium most important in the vagina normal flora, that quantity is maximum.Lactobacillus accounts for more than 95% of all permanent bacterium of intravaginal under the normal condition, is normal flora, contain in the every approximately ml vaginal secretions of its amount≤10 4CFU.Studies show that at present from the healthy women intravaginal is separable and go out 16 kinds of lactobacilluss, separation rate reaches 50-80%, wherein a fairly large number of is bacillus acidophilus's (77.5%), Lactobacillus salivarius and Lactobacillus fermenti, and the strong bacterium of glycogen capacity of decomposition comprises bacillus acidophilus, Lai Shi lactobacillus and Lactobacillus salivarius.Lactobacillus plays an important role in keeping the microecology in vaginas balance.It can decompose the glycogen in the vaginal mucosa epithelial cell, produces a certain amount of lactic acid and then keeps the lower acid-base value of intravaginal, makes vagina keep sour environment, suppresses the undue growth of harmful bacterium, helps the self purification of vagina.It can improve body's immunological function simultaneously, and activating macrophage is kept the partial anti-infection ability of vagina.
Vaginal secretions can not gathered in heapsly when the gynecological specula was checked, polymorphonuclear neutrophil leukocyte is wherein seldom arranged, and the vaginal secretions smear is through Gram, and the epithelial cell sharpness of border has the darker Gram-positive lactobacillus of volume dyeing in the smear.Vaginal wall exists a large amount of lactobacilluss can form a kind of biomembrane, has played the occupancy protective effect.The bacillus acidophilus is one of normal flora bacterial strain of superiority bacteria spp in the healthy women intravaginal micropopulation, and quantity is maximum, accounts for 67% of whole flora.They play balanced action to normal flora, make host's microecology in vaginas system be in dynamic equilibrium, keep the woman vagina normal function, avoid being had the invasion and attack of bacterium or exogenous pathogenic bacterium of dwelling in pathogenic.
The vaginitis morbidity is relevant with the microecology in vaginas environment change: the rising of (1) vagina pH value; (2) variation of gonadal hormone in the menstrual cycle; (3) vagina lavation, the improper vagina part that makes of lavation is injured; Also suppressed the vagina normal flora when (4) pathogenic bacteria has been killed in application such as antibiotic; (5) simultaneously with other infection.Under the systemic disease, vagina local damage, ovarian function or mistakes degree uses such as immunosuppressant, antitumor drug all cause in the vaginal microbial flora some permanent bacterium to be taken advantage of a weak point, cause vaginal infection.The main source that infects be among the vagina microorganism group many potentiality pathogenic microorganism such as escherichia coli, bacteroid, dyspepsiacoccus, staphylococcus aureus, B family streptococcus, Candida albicans, trichomonacide, mycoplasma etc.More than 5 kinds of situations can destroy the microecological balance of vagina, cause the pathogenic bacterium breeding and cause vaginitis.In addition, the anaerobe breeding while, some Ammonia material produces, and the vagina that also further alkalizes comes off vagina epithelium, and vaginal secretions is unusual.And some Ammonia material also can further promote the anaerobe growth to increase the weight of colpitic symptom, causes the microecology in vaginas environmental disruption.Vaginitis is one of common, multiple disease of women, and is relevant with microecological balance in the woman vagina.Nearly more than ten years are because the progress of vagina microorganism fundamental research, for the development and application of this class preparation provides scientific basis.
The preparation that contains lactobacillus, the killed bacterial cell that causes owing to the physical impacts that produces in preparation accounts for certain proportion, and when they when further diffusion and their breeding begin in vaginal canal, the quantity of bacterial cell further reduces, and therefore, needs to handle the suitable kind of lactobacillus, or adjuvant, with the preparation that comprises them, to guarantee preparation good quality is arranged, it is real useful using to patient.In addition, preparation should be able to keep the bacillus acidophilus's that breeds quantity constant, and guarantees bacillus acidophilus's effect unchangeably continuously when the vaginal canal internal diffusion.
The applicant has found to obtain the pharmaceutical preparation for the basis to the very effective bacillus acidophilus of treatment vaginal infection, it provides following feature, can satisfy possible top level: (1) selected strain of lactobacillus acidophilus must have the high-affinity of pair vagina epithelium, promptly adhere to the high-performance of vaginal epithelial cell, this performance can interact antibacterial with vaginal mucosa under physiology and pathological conditions, execution is in the competition effect of epithelium acceptor site and pathogenetic organism, and reconstruction microorganism system and optimum pH condition.(2) high interference performance must be arranged, competition escherichia coli and the adhesion of Candida albicans on human vagina's epithelial cell.(3) high-performance that suppresses escherichia coli, Candida albicans, gonococcus growth must be arranged.
Therefore, the purpose of this invention is to provide and satisfy above feature, reach the preparation of the bacillus acidophilus of possible top level, and be suitable for being used for the treatment of effectively vaginal infection, as vaginitis, gonorrhea etc. for the basis.
The invention provides a kind of bacillus acidophilus's of containing pharmaceutical preparation, said preparation is made up of bacillus acidophilus and pharmaceutical excipient.
The bacillus acidophilus that the present invention uses is the viable bacteria of making behind external artificial fermentating breeding.This strain with Tomato juice as culture medium, through external artificial fermentation, cooling, centrifugal, with protective agent after preparationization after the lyophilizing, be used for the treatment of vaginal infection, play the microecology in vaginas therapeutical effect.This strain mycopowder bacterium amount is with 10 8-10 10CFU/g is excellent, and preferred bacterium amount is approximately 10 11CFU/g.This bacillus acidophilus therapeutic dose (bacterium amount) should be greater than 10 4CFU/g, preferred bacterium amount is 10 6-10 7CFU/g.
Another object of the present invention has provided the preparation method of a kind of bacillus acidophilus's of containing pharmaceutical preparation, and this method comprises the following steps:
1, preparation bacillus acidophilus mycopowder:
The bacillus acidophilus SL9404 bacterial strain that secretions in the vagina is separated, obtains behind the purification is rule on the MRS agar slant and is gone down to posterity, and cultivates 48h through 37 ± 1 ℃.Choose full, color and luster is even, medium bacterium colony bigger than normal, once more on the MRS agar slant of streak inoculation in the Fructus Solani melongenae bottle, cultivates 48h through 37 ± 1 ℃.Transferred species inserts beef cysticercosis cellulosae cutis pipe, cultivates 20-50h for 37 ± 1 ℃.Get 1-4 and prop up the cysticercosis cellulosae cutis pipe and change 300ml milk fermentation culture medium or 5-20% Tomato juice fermentation medium or milk+Tomato juice's fermentation medium over to, cultivate the 8-48h fermentation culture for 37 ± 1 ℃, progressively be amplified to bulk fermentation, centrifugal, make mycopowder after the lyophilization;
2, preparation medicament:
Take by weighing bacillus acidophilus and pharmaceutical excipient by recipe quantity, drug formulation process is made various preparations routinely.
The preservation of this strain is excellent with cryodesiccated form.
This strain is preserved in Chinese typical culture collection center on June 12nd, 2000, is numbered CCTCC NO:M200013.
The Pro-Bionate's of the containing of the present invention mechanism of action mainly contains: (1) keeps sour environment.Vagina living preparation of lactobacillus preparation can replenish the most important lactobacillus of patient's intravaginal, it can decompose the glycogen of storing in the vaginal mucosa epithelium and produce acidic materials such as lactic acid, acetic acid, itself also can produce lactic acid simultaneously, this plays an important role but kill multiple pathogenic microorganism to keeping the sour environment of vagina.(2) occupy-place protective effect.Studies have shown that in a large number ectogenic lactobacillus can adhere to the vaginal mucosa epithelium securely, form spatiality occupy-place protective effect.(3) direct antagonism.The inside and outside studies confirm that lactobacillus adds moral sodium Salmonella, infusorian, candidiasis, staphylococcus etc. to vagina direct inhibitory action is arranged.(4) produce multiple antibacterial substance.Lactobacillus can decompose the glycogen of storing in the vaginal mucosa epithelium, produces acidic materials such as lactic acid, acetic acid, and itself also can produce lactic acid simultaneously, keeps the sour environment of vagina, but kills multiple pathogenic microorganism.(5) nutrient competition.Be colonizated in the lactobacillus in the vagina in a large number, nutrition is in the competitive advantage state, is unfavorable for the growth of other bacterium.The lactobacillus therapy is that the living preparation of lactobacillus preparation is put in the vagina, makes living preparation of lactobacillus get back to its natural physiological function of performance in the body, adjusts vaginal microbial flora, and maintaining ecological balance makes vaginal microbial flora reach microecological balance again, reaches the purpose of disease preventing and treating.
Dosage form of the present invention can be liquid form, mastic form or solid form, as tablet, capsule, powder, cream, ointment, suppository or spray.
The described common drug excipient of preparation of the present invention comprises corn starch, pregelatinized Starch, carboxymethyl starch sodium, dextrin, lactose, isolactose, mannitol, glucose, vitamin C, citric acid, sodium dihydrogen citrate, disodium citrate, tartaric acid, fumaric acid, boric acid, acidic amino acid class weak acid such as (as: glutamic acid, aspartic acids), microcrystalline Cellulose, sodium carboxymethyl cellulose (calcium), hypromellose, polyvinylpyrrolidone, gelatin, calcium carbonate, sodium bicarbonate, magnesium stearate, lanoline, Polyethylene Glycol etc.Additive is meant relevant nutrients such as edible defatted milk powder, vitamin C, glutamic acid.
The Pro-Bionate of containing of the present invention can be used for treatment of diseases and preventions such as woman vagina infection aspect, gonorrhea.One, the bacillus acidophilus observes (Nat'l Pharmaceutical ﹠ Biological Products Control Institute's detection) (one) purpose to human vaginal epithelial cell (VEC) adhesive attraction: observe bacillus acidophilus SL9404 to the epithelial adhesive attraction of human vagina.(2) materials and methods: 1, human vagina's epithelial cell (hereinafter to be referred as VEC):
Take from healthy women, the age, the hospital-based outpatient obstetrical clinic health check-up was normal below 40 years old.
Scrape the VEC specimen of getting and be suspended among the 20mlRPMI-1640 vortex mixer concussion washed cell immediately 3 times, 500rpm * 3min, filter is sucking filtration repeatedly, removes the endogenous antibacterial.The blood counting chamber counting is adjusted VEC concentration 10 5/ ml.2, strain: separate from healthy women vagina bacillus acidophilus 9403, SL9404 and one strain of 9405 strain bacterium (2) escherichia coli CMCC44113 strain (3) Candida albicans CMCC 10231 strains (4) Deshi Lactobacilluss (1), separates and make bacterium raw product 3, the preparation of bacterium liquid by oneself:
Lactobacillus is inoculated in MRS culture medium, escherichia coli and Candida albicans is inoculated in ordinary nutrient agar, 37 ℃, cultivated in 18 hours; Scrape before the test and get lawn, 3 times (3000 * 10min), adjust cell concentration is 1 * 10 to RPMI-1640 washing antibacterial 8/ ml.4, method: (1) adherence test: 3 strain bacillus acidophilus adhesive attractions relatively
Get VEC suspension and each 1ml of bacterium liquid, mix, 37 ℃, cultivated in 2 hours; Smear, drying at room temperature, methanol is 10min fixedly, and Gram, mirror are observed antibacterial adhesion situation and counting on VEC down.(2) lactobacillus is divided into following three groups and carries out the influence of escherichia coli, Candida albicans adhesion VEC:
First group: add 1ml Escherichia coli bacteria liquid or 1ml Candida albicans bacterium liquid in the 1ml lactobacillus bacterium liquid, add 1mlVEC suspension mixing again, 37 ℃, 2 hours, observe down and counting by above-mentioned test method mirror.
Second group: 1ml lactobacillus bacterium liquid and 1mlVEC suspension mixing, 37C add 1ml Escherichia coli bacteria liquid or 1ml Candida albicans bacterium liquid after cultivating 30min earlier again, and mixing continues 37 ℃, 2 hours, observe down and counting with above-mentioned test method mirror.5, counting: count 50 VEC under the mirror at random and go up adherent bacterial population, calculate average and standard deviation.(3) result 1, the not comparison of homophyletic bacillus acidophilus adhesive attraction:
Three strain bacillus acidophilus are different to the adhesive attraction of human body VEC, wherein the SL9404 strain has significantly adhesive attraction to VEC, see Table 1, be the bacterial adhesion number (113.8 ± 33.69) of 50 VEC, and 9403 and 9405 strains are through cultivating 37 ℃ with VEC, after 2 hours, a large amount of cell distribution are not seen obvious adhesion on VBC in mixed liquor.The contrast bacterium is promptly also not obvious by deciding the living isolating Deshi Lactobacillus adhesive attraction of bacterium.
Table one bacillus acidophilus SL9404 strain is to the adhesive attraction of VEC
The VEC numbering The adhesion bacterial population (individual/VEC) The VEC numbering The adhesion bacterial population (individual/VEC) The VEC numbering The adhesion bacterial population (individual/VEC)
????1 ????132 ????18 ????92 ????35 ????85
????2 ????113 ????19 ????120 ????36 ????123
????3 ????74 ????20 ????89 ????37 ????144
????4 ????149 ????21 ????65 ????38 ????131
????5 ????186 ????22 ????78 ????39 ????156
????6 ????118 ????23 ????158 ????40 ????98
????7 ????111 ????24 ????75 ????41 ????143
????8 ????63 ????25 ????126 ????42 ????125
????9 ????84 ????26 ????118 ????43 ????95
????10 ????89 ????27 ????109 ????44 ????194
????11 ????94 ????28 ????138 ????45 ????88
????12 ????118 ????29 ????80 ????46 ????155
????13 ????119 ????30 ????128 ????47 ????178
????14 ????125 ????31 ????74 ????48 ????122
????15 ????90 ????32 ????77 ????49 ????167
????16 ????120 ????33 ????50 ????50 ????133
????17 ????129 ????34 ????62 ????X±S ??113.8±33.69
2, bacillus acidophilus SL9404 strain adheres to influence (1) escherichia coli 44113 strains and the Candida albicans 10231, white 157.14 ± 54.06 and 31.84 ± 11.65 (seeing Table 2) of VEC to escherichia coli, Candida albicans.Table 2, first group of result of the test
Bacterial strain Escherichia coli 44113 adhesion number/VEC (X ± S) Candida albicans 10231 adhesion number/VEC (X ± S) Statistical disposition
Escherichia coli 44113 Lactobacillus lactis SL9404+Escherichia coli 44113 Lactobacillus delbrueckiis+Escherichia coli 44113 Candida albicans 10321 Lactobacillus acidophilus S9404+Candida albicans 10231 Lactobacillus delbrueckiis+Candida albicans 10231 ?157.14±54.06 ?118.13±49.24 ? ?147.78±45.12 ? ? ? ? ? ? ?31.84±11.65 ?9.76±5.94 ?15.66±8.45 ? ?P＜0.01 ? ?P＞0.05 ? ? ?P＜0.01 ?P＜0.01
(2) first groups of result of the tests (table 2) are indicated, after bacillus acidophilus SL9404 strain and escherichia coli 44113 strains or Candida albicans 10231 strains are incubated at 37 ℃ simultaneously, the SL9404 strain can suppress the adhesive attraction of other two strains bacterium, statistical procedures has notable difference, and comparison is more obvious according to the inhibitory action of Deshi Lactobacillus.(3) second groups of result of the tests show that after bacillus acidophilus's 9405 strains were at that time cultivated with VEC earlier, when adding escherichia coli 44113 or candida albicans 10231 again (table 3), then bacillus acidophilus SL9404 strain meeting obviously suppressed the effect that back two strain bacterium adhere to VEC.Table 3, the 2nd group of result of the test
Bacterial strain Escherichia coli 44113 adhesion number/VEC (X ± S) Candida albicans 10231 adhesion number/VEC (X ± S) Statistical disposition
Escherichia coli 44113 ????267.22±97.80 ????P＜0.01
Bacillus acidophilus SL9404+escherichia coli 44113 ????125.26±51.64
Candida albicans 10231 ????29.24±8.45 ????P＜0.01
Bacillus acidophilus's SL9404+Candida albicans 10231 ????19.54±10.24
(4) the 3rd groups of result of the tests show, after escherichia coli 44113, Candida albicans 10231 are cultivated with VEC earlier, when adding bacillus acidophilus SL9404 strain again, the SL9404 strain adheres to VEC to preceding two strain bacterium also has obvious suppression effect (seeing Table 4) table 4, bacillus acidophilus SL9404 strain that colibacillus 44113 or Candida albicans 10231 are influenced at the VEC adhesive attraction
Bacterial strain Escherichia coli 44113 adhesion number/VEC (X ± S) Candida albicans 10231 adhesion number/VEC (X ± S) Statistical disposition
Escherichia coli 44113 ????180.7±252.20 ??P＜0.01
Escherichia coli 44113+bacillus acidophilus SL9404 ????134.18±40.63
Candida albicans 10231 ????29.24±8.85 ??0.01＜P＜0.05
Candida albicans 10231+bacillus acidophilus SL9404 ????24.40±11.36
(4) conclusion: 1, bacillus acidophilus SL9404 strain has tangible adhesive attraction to this vaginal epithelial cell of people.2, bacillus acidophilus SL9404 strain can suppress escherichia coli 44113 strains and Candida albicans 10231 strains at the epithelial adhesive attraction of human vagina.3, bacillus acidophilus SL9404 strain is more obvious than separating the Deshi Lactobacillus of making the living preparation of bacterium by oneself to human vagina's epithelial cell adhesive attraction.Two, the research of 3 kinds of pathogenic bacterium of the external antagonism of bacillus acidophilus (Chinese biological goods calibrating institute measure) (one) purpose: observation bacillus acidophilus SL9404 external to three kinds of common vaginal pathogenic, i.e. the antagonism of Escherichia coli, gonococcus and Candida albicans.(2) materials and methods: (1) strain: 1. bacillus acidophilus SL9404 strain: provide by friendship Pharma Inc. of last Hisense.2. three kinds of pathogenic bacterium provide by Chinese medicine antibacterial preservation administrative center: Escherichia coli CMCC44l13 gonococcus (hereinafter to be referred as gonorrhea) CMCC29403 Candida albicans CMCC10231 (2) culture medium: 1. contain 2% calf serum TPY culture fluid: be used for liquid culture.2. lactobacillus selective medium: adopt the MRS agar culture medium.3. escherichia coli selective medium: adopt the MRS agar culture medium.4. Candida albicans selective medium: adopt brave red agar culture medium.5. gonorrhea selective medium: adopt the NG agar culture medium.(3) bacterium liquid preparation:
With each bacterial strain fresh cultured thing, use standard antibacterial opacity tube than turbid respectively, and adjust bacterial concentration with 2% calf serum TPY culture fluid, the bacillus acidophilus is: 1 * 10 7/ ml, other three kinds of pathogenic bacterium are 1 * 10 5/ ml.(4) test grouping: bacillus acidophilus bacterium liquid mixes the formation matched group respectively with three kinds of pathogenic bacterium bacterium liquid mixed in equal amounts (9ml+9ml) in triangular flask.
Test group and matched group all place 37 ℃ of aqueous solutions to be cultured to 72 hours or 96 hours.(5) bacterium quantitative determination:
Above-mentioned each test group, matched group are being cultivated initial (), each sampling in 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, bacillus acidophilus in the determination test group triangular flask and the antibacterial production concentration of three kinds of pathogenic bacterium respectively at 0 o'clock.Also measure simultaneously the bacterium amount of each matched group.The bacterium quantity measuring method carries out with reference to the count plate method of nineteen ninety-five version " Chinese biological goods rules " regulation, and adds up into table with the logarithm value of bacterium amount concentration.(3) result and analysis (1) table 5 shows the antagonism of bacillus acidophilus SL8404 to three kinds of common vaginal pathogenic, show when the SL9404 strain respectively with escherichia coli, gonococcus and 72-96 hour well-grown of Candida albicans Mixed culture; Every ml viable bacteria concentration is all 1 * 10 8More than the individual bacterium: the number of viable of SL9404 strain and matched group are approximate in each test group.(2) table 5 shows that three kinds of pathogenic bacterium are cultured to 12 hours in test group or the matched group, and viable bacteria concentration obviously increases, and two groups of bacterium amounts are close.(3) after table 5 showed that gonorrhea bacterium in the test group was cultured to 12 hours, growth obviously suppressed, and to 24 hours, every ml bacterium liquid viable bacteria concentration was less than 10, and this moment matched group the bacterium amount still be consistent with 12 hours bacterium amount.(4) table 5 shows matched group escherichia coli cultivation 72 hours, and the still every ml of bacterial concentration is greater than 10 8Escherichia coli are cultured to 24 hours in the test group, and viable bacteria concentration is also 10 8More than/the ml, similar to matched group, but growth obviously suppresses after 24 hours.The viable bacteria amount is lower than 10 in 48 hours every ml bacterium liquid 5(5) table 5 has shown that also the growth of Candida albicans also has certain inhibition in the test group, and from cultivating 12 hours to 96 hours, its viable bacteria concentration is low than matched group.
Table five bacillus acidophilus SL9404 strain antagonistic effect result
Group Viable count (CFU/ml) the Log value of different incubation times
0 hour 12 hours 24 hours 48 hours 72 hours 96 hours
Test group SL9404+gonococcus 29403 ??5.9165 ? ??3.3979 ????8.3502 ? ????3.8692 ????8.8209 ? ????＜1 ??8.4983 ? ??＜1 ??8.0253 ? ??＜1
SL9404+escherichia coli 44113 ??5.9165 ? ??3.7782 ????8.2577 ? ???8.8727 ????8.8395 ? ????8.7404 ??8.9004 ? ??＜5 ??8.7308 ? ??＜1
SL9404+Candida albicans 10231 ??5.9165 ??3.7782 ????8.3201 ????7.6821 ????8.5775 ????7.8420 ??8.7745 ??7.7474 ??8.8028 ??7.8921 ??8.7559 ??7.9731
Matched group Gonococcus 29403 ??3.3979 ????3.8865 ????3.6990 ??＜1 ??＜1
Escherichia coli 44113 ??3.7782 ????8.8882 ????8.8176 ??8.8987 ??8.4409
Candida albicans 10231 ??3.7782 ????7.8481 ????8.1614 ??8.1903 ??8.4314 ??8.2041
Bacillus acidophilus SL9404 ??5.9165 ????8.4100 ????8.7752 ??8.2455 ??7.8722 ??7.1303
(4) conclusion: 1. bacillus acidophilus SL9404 strain can comparatively fast suppress in external growth to gonococcus.2. bacillus acidophilus SL9404 strain is grown at the external Escherichia coli that suppresses.3. bacillus acidophilus SL9404 strain has certain effect external to Candida albicans.Three, vagina with the lactobacillus effervescent tablet to rat vagina irritant test (Chinese Academy of Sciences's Shanghai medicine is measured)
24 of female sd inbred rats are divided into 3 groups, administration group, vehicle group and untreated fish group, 8 every group.Continuous 7 days vagina administrations are whenever in observation behavior and external genital situation.Getting rat vagina in 24 hours after the last administration organizes macroscopy and does the pathology histological examination.Experimental result shows, the perusal tangible abnormal response of pudendum of not regarding sb. as an outsider, and histopathologic examination does not have obvious pathological changes.1, experiment purpose:
Observe vagina with continuous 7 days rat vagina administrations of lactobacillus effervescent tablet after irritant reaction.2, be subjected to the reagent thing: (1) title: vagina lactobacillus effervescent tablet, white olive-shaped tablet.(2) specification: 1.7 * 10 7CFU/ sheet (0.25g/ sheet) (3) unit of providing: lot number: the DM-032573 of Shanghai Xinyi Pharmaceutical Co., Ltd (4), tester: (1) excipient: the 0.25g/ sheet, Shanghai Xinyi Pharmaceutical Co., Ltd provides.(2) lot number: DM032504, animal: give processing before (1) source, kind, the strain test: 24 of female sd inbred rats are provided by Chinese Academy of Sciences's Shanghai Experimental Animal Center.The quality certification: No. the 005th, the moving pipe of middle section.Single cage is raised, and animal freely ingests, drinking-water.Raise a week at institute of materia medica, Chinese Academy of Sciences Shanghai Animal House before the experiment.(2) body weight: 242 ± 24g (3) sex: female (4) every treated animal thing: be divided into 3 groups at random by body weight, 8 every group, promptly tried thing group, excipient and untreated fish group.(5) Animal House temperature: 23 ± 2 ℃ 5, dosage (1) dosage setting: clinical human vagina lactobacillus effervescent tablet 10 6CFU/ sheet/people.The rat dosage is decided to be 1.7 * 10 7The CFU/ sheet/only.Local application's amount is about 10 times of human dosage.(2) every animals administer capacity: the 0.25g/ sheet/only.6, route of administration
Rat vagina administration 7, method
To be tried thing during test and be inserted gently in the rat vagina, be guaranteed that medicine contacts until dissolving with vaginal mucosa looking on to examine.Once a day, each a slice continuous 7 days, is observed rat behavior and external genital situation every day.Dissected in 24 hours after the last administration and carry out macroscopy and draw materials and make histology's microscopy.8, observation index (1) perusal: observe rat behavior reaction, vaginal orifice situation every day.After the last administration, put to death rat in 24 hours, take out vagina tissue, observe stimulation phenomenons such as having or not hyperemia, redness.(2) histological examination: every rat vagina sample of tissue is done the pathology histological examination.(3) result judges: result of the test and untreated fish group compare judgement.9, result: (1) perusal: after the rat administration in continuous 7 days behavior normal, perusal external genital Non Apparent Abnormality reaction.After the last administration 24 hours, take out vagina tissue, do not see that hyperemia, redness etc. stimulate phenomenon.
(2) histological examination: histopathology is observed vagina epithelium and is stratified squamous epithelium, and structure is normal, there is no ANOMALOUS VARIATIONS.
10, conclusion:
Vagina lactobacillus effervescent tablet 1.7 * 10 7CFU/ sheet/only continuous 7 days, perusal rat vagina Non Apparent Abnormality reaction after the administration, histopathologic examination also no abnormality seen changes.The vagina lactobacillus effervescent tablet reaction non-stimulated to rat vagina.
Four, rat vagina is with lactobacillus effervescent tablet mtd test (being measured by Chinese Academy of Sciences's medicine)
24 of female sd inbred rats are divided into 4 groups.Test vagina and give the acute toxicity of lactobacillus effervescent tablet.Dosage is 1.7 * 10 7, 6.9 * 10 9The CFU/ sheet/only, other establishes vehicle group and untreated fish group, totally 4 dosage groups.Observed behavior in continuous 14 days reaction and record body weight gain situation.Activities in rats is normal after the administration, and Non Apparent Abnormality changes, no animal dead.Dissected all animals on the 15th day, the macroscopy internal organs are not seen obvious pathological changes.Rat is to lactobacillus effervescent tablet maximum tolerated dose〉6.9 * 10 9DFU/ only.Calculate by absolute magnitude, be people's consumption 10 6More than 1000 times of CFU/ sheet/people.
1, experiment purpose:
Observe rat vagina and once contact the influence that is tried the acute toxic reaction, death condition and the body weight that are produced behind the thing.
2, be subjected to the reagent thing:
(1) title: vagina lactobacillus effervescent tablet, white olive-shaped tablet.
(2) specification: 1.7 * 10 7, 6.9 * 10 9CFU/ sheet (0.25g/ sheet)
(3) unit of providing: Shanghai Xinyi Pharmaceutical Co., Ltd
(4) lot number: DM-03257; DM-03255
3, tester:
(1) excipient: the 0.25g/ sheet, Shanghai Xinyi Pharmaceutical Co., Ltd provides.
(2) lot number: DM03250
4, animal:
Source, kind, the quality certification: 4 of female sd inbred rats are provided by Chinese Academy of Sciences's Shanghai Experimental Animal Center.The quality certification: No. the 005th, the moving pipe of middle section.Test adaptability the last week is raised.Feedstuff must triumphant laboratory animal company limited available from west, the joint Shanghai of China and Britain pul one.Freely fetch water, ingest, raising temperature is 23 ± 2 ℃.
5, dosage (1) dosage setting: clinical human vagina lactobacillus effervescent tablet 10 6CFU/ sheet/people (1g/ sheet).The effective dose of animal is 10 6The CFU/ sheet/only.Be observation toxic reaction big as far as possible dosage and the technical relation of considering film-making.If 1.7 * 10 7The CFU/ sheet/only with 6.9 * 10 9The CFU/ sheet/only.Calculate respectively quite clinical consumption 10 and 1000 times by local application amount, and establish excipient and untreated fish group.
(2) every animals received capacity: the l sheet/only.
6, route of administration;
The rat vagina administration.
7, method
Earlier use the physiologic dose brine washing vagina during test, will be tried thing then and insert in the rat vagina gently.Examine 6 hours and guarantee that tablet contacts with vaginal mucosa looking on.Promptly observe the reaction of animals situation after the administration,, check internal organs if dead animal is promptly dissected.
8, observation index:
(1) observation period: 14 days
(2) toxic reaction: situations such as observation and inspection rat outward appearance, behavior, feed, feces, body weight gain, dead animal becomes celestial.When experiment finished in the 15th day, survival rats was dissected, and checked organ diseases such as the heart, lung, liver,spleen,kidney.
9, result:
After rat vagina awarded the lactobacillus effervescent tablet, situations such as outward appearance behavior, feed and feces there is no ANOMALOUS VARIATIONS, no animal dead, and it is similar to untreated fish group with vehicle group that rat body weight increases the administration group.The body weight gain situation is listed following table in:
The influence of lactobacillus effervescent tablet of table six, vagina to rat body weight
Time (my god) Untreated fish group body weight (g) Excipient body weight (g) O.25 x 107CFU/g/ body weight (g) O.25 X 109CFU/g/ body weight (g)
??DO ??D2 ??D4 ??D6 ??D8 ??D10 ??D12 ??D14 ??219.7±11.1 ??221.8±13.7 ??224.9±14.1 ??230.4±12.9 ??231.2±13.O ??233.2±9.96 ??235.24±9.66 ??238.7±10.1 ??220.2±8.2 ??223.9±7.1 ??229.2±6.2 ??237.3±8.6 ??240.4±11.1 ??246.7±14.4 ??250.6±14.6 ??253.0±14.5 ??220.0±8.9 ??228.8±5.3 ??236.3±7.3 ??247.9±8.7 ??250.9±7.8 ??259.1±8.7 ??259.5±10.7 ??260.9±10.8 ????219.3±7.7 ????223.4±8.9 ????232.8±11.7 ????244.5±12.8 ????250.5±14.2 ????255.5±14.2 ????251.O±16.3 ????254.3±15.5
10, evaluation of result:
It is clinical consumption 10 or 1000 times that effervescent tablet, contained antibacterial are used in experiment.Much smaller because of rat volume ratio people, dwindle so the size of effervescent tablet is also corresponding, contain the nectar degree comparatively speaking and want high than the human effervescent tablet.Reached capacity from tabletting technology.Rat vagina awards lactobacillus effervescent tablet 1.7 * 10 7, 6.9 * 10 9CFU/ sheet/only, observed continuously 14 days, activities in rats is normal after the administration, and Non Apparent Abnormality changes, no animal dead.The administration group increases with the vehicle group rat body weight and shows that processed group is similar.Rat vagina uses the maximum tolerated dose of lactobacillus effervescent tablet greater than 6.9 * 10 9CFU/ only.Five, bacillus acidophilus SL9404 strain stability test (measuring) 1, purpose: observe the stability after bacillus acidophilus SL9404 involves continuous 30 generations of going down to posterity in a criminal case by Nat'l Pharmaceutical ﹠ Biological Products Control Institute.2, materials and methods: (1) bacterial strain: bacillus acidophilus SL9404 strain is provided by friendship Pharma Inc. of last Hisense.(2) culture medium: 1. MRS fluid medium-go down to posterity with 2. MRS agar culture medium-flag check with 3. sugared biochemical reaction culture medium-observations biochemical reaction with 4. PYG fluid medium-survey metabolite with (3) method:
Strain carries out former generation strain and the strain of 30 generations the comparison of phenotypic characteristic biology (comprising lactic acid content mensuration in growth characteristics, colonial morphology, dyeing microscopic examination, biochemical reaction, drug sensitive test, toxicity test and the metabolite) and hereditism's feature (G+C Mol% mensuration) by after 30 generations of MRS fluid medium continuous passage.3, result:
It is 7 listed that former generation strain and 30 generations strain characteristic check see Table.
Table 7 bacillus acidophilus 9404 former generation strains and 30 stable comparisons of feature in generation
Inspection item 9404 strains (former generation) 9404 strains (30 generation)
Growth characteristics (1) all can grow in the aerobic or anaerobic environment (2) on MRS agar, be grown to light yellow, the about 0.5-1mm of the circular colony diameter of central protrusion. (1) all can grow in the aerobic or anaerobic environment (2) on MRS agar, be grown to light yellow, the about 0.5-1mm of the circular colony diameter of central protrusion.
Dyeing microscopic examination The Gram-positive thalline is quarter butt shape, is dispersed in or in pairs, becomes short chain to arrange.The about 0.5-0.8 μ m of thalline size * 1.5-3 μ m The Gram-positive thalline is quarter butt shape, is dispersed in or in pairs, becomes short chain to arrange.The about 0.5-0.8 μ m of thalline size * 1-2.5 μ m
Biochemical reaction The sweet fruit half sugarcane Mai Lali grape in A Mu Mus nuclear Portugal reveal newborn bud uncle sugar sugar sugar sugar sugar sugar sugar sugar sugar sugar---+++++++ The sweet fruit half sugarcane Mai Lali grape in A Mu Mus nuclear Portugal reveal newborn bud uncle sugar sugar sugar sugar sugar sugar sugar sugar sugar sugar---+++++++
The cotton pine of fine newborn gill fungus honey can form sediment the pure glycosides sugar of sweet mountain 7 degree of freedom two sons, three molten dew Folium pyri disaccharide sugar sugar powder sugar sugared+++--+-+++ The cotton pine of fine newborn gill fungus honey can form sediment the pure glycosides sugar of sweet mountain 7 degree of freedom two sons, three molten dew Folium pyri disaccharide sugar sugar powder sugar sugared+++--+-+++
The bright nitre of water also cattle touch the plain liquid salt of poplar glue yoghourt former coagulate change solid enzyme+--+- The bright nitre of water also cattle touch the plain liquid salt of poplar glue yoghourt former coagulate change solid enzyme+--+-
Drug sensitive test Amikacin R tetracycline S gentamicin R erythromycin S streptomysin R PB R neomycin R chloramphenicol S ampicillin S furazolidone R closes spore azoles quinoline S SMZco R cefuroxime S orfloxacin R Amikacin R tetracycline S gentamicin R erythromycin S streptomysin R PB R neomycin R chloramphenicol S ampicillin S furazolidone R closes spore azoles quinoline S SMZco R cefuroxime S orfloxacin R
Toxicity test Mouse peritoneal injection 1.0 * 10 9Behind the CFU viable bacteria, mice is strong deposits weight increase.
The metabolite organic acid is measured Lactic acid 0.5331% acetic acid 0.0796% Lactic acid 0.5482% acetic acid 0.0600%
G+CMol% 39.76 39.76
Conclusion: in 30 generations of 9404 strain continuous passages, biology and genetics characteristic are stable.
9403,9405 strain bios are learned, hereditism's feature compares
Inspection item 9403 9405
Growth characteristics (1) all can grow in the aerobic or anaerobic environment (2) on MRS agar, be grown to ecru, circle, projection smooth colony 1-2mm size. (1) all can grow in the aerobic or anaerobic environment (2) on MRS agar, be grown to light yellow, the about 0.5-1mm of the circular bacterium colony of central protrusion size.
Dyeing microscopic examination The Gram-positive thalline is quarter butt shape, about 0.3-0.5 μ m * 1-1.5 μ m. The Gram-positive thalline is quarter butt shape, about 0.5-0.8 μ m * 1-2.5 μ m
Biochemical reaction The sweet fruit half sugarcane Mai Lali grape in A Mu Mus nuclear Portugal reveal newborn bud uncle sugar sugar sugar sugar sugar sugar sugar sugar sugar sugar--±+++++++ The sweet fruit half sugarcane Mai Lali grape in A Mu Mus nuclear Portugal reveal newborn bud uncle sugar sugar sugar sugar sugar sugar sugar sugar sugar sugar+-++++++++
The cotton pine of fine newborn gill fungus honey can form sediment the pure glycosides sugar of sweet mountain 7 degree of freedom two sons, three molten dew Folium pyri disaccharide sugar sugar powder sugar sugared+++--+-+++ The cotton pine of fine newborn gill fungus honey can form sediment the pure glycosides sugar of sweet mountain 7 degree of freedom two sons, three molten dew Folium pyri disaccharide sugar sugar powder sugar sugared+-++-+-+++
The bright nitre of water also stone touch the plain liquid salt of poplar glue acid stamen aurochsization milk enzyme+--+- The bright nitre of water also cattle touch the plain liquid salt of poplar glue yoghourt former coagulate change solid enzyme+--+-
Drug sensitive test Amikacin R tetracycline S gentamicin S erythromycin S streptomysin S PB R neomycin R chloramphenicol S ampicillin S furazolidone R closes spore azoles quinoline S SMZco R cefuroxime S orfloxacin R Amikacin R tetracycline S gentamicin R erythromycin S streptomysin R PB R neomycin R chloramphenicol S ampicillin S furazolidone R closes spore azoles quinoline S SMZco S cefuroxime S orfloxacin R
The metabolite organic acid is measured Lactic acid 0.9106% acetic acid 0.0560% Lactic acid 0.6884% acetic acid 0.1115%
G+CMol% 41.84 42.88
Conclusion: (1) two strain bacteria growing feature, colonial morphology and dyeing microscopic examination meet the lactobacillus feature.(2) two strain bacterium all can produce lactic acid.(3) 9403 strain ribose, melezitose, mannitol, sorbitol biochemical reaction do not meet bacillus acidophilus's feature of uncle Jie Shi handbook regulation.
9405 strain arabinose, ribose, melezitose, mannitol biochemical reaction do not meet bacillus acidophilus's feature of uncle Jie Shi handbook regulation.The G+C Mol% measured value of the DNA of (4) two strain bacterium is higher than the G+C Mol% content of the heavy breathing Lactobacillus lactis of uncle Jie Shi handbook regulation.
It is the common disease of women that woman vagina infects, and is that microecological balance imbalance institute causes disease in the woman vagina, and main and sex hormone level, sexual life are excessively, peace intrauterine device and antibacterial therapy upset vagina normal flora etc. are relevant.The ratio that the woman vagina incidence of infection accounts in the female diseases is higher, by replenishing the vagina probiotic bacteria, make probiotic bacteria field planting in vagina, thereby stop the procreation of antibacterial in the vagina, play colpitic effect of treatment and effect, alleviate and free women's sufferings.The present invention has great economic efficient latent and high exploitation is worth.
Example one bacillus acidophilus is to the obvious adhesion of human vagina's epithelial cell (VEC)
By this test, tested bacillus acidophilus SL9404 to the epithelial adhesive capacity of human vagina.
Get each 1ml of human vagina's epithelial cell suspension and bacteria suspension, mix, 37 ℃, cultivated in 2 hours; Smear, drying at room temperature, methanol is 10min fixedly, and Gram, mirror are observed antibacterial adhesion situation and counting on human vagina's epithelial cell down.
The result who obtains represents in Table 1, shows that bacillus acidophilus SL9404 has very strong adhesive capacity to human vagina's epithelial cell, is suitable for preparing preparation of the present invention.
Table one bacillus acidophilus SL9404 strain is to the adhesive attraction of VEC
The VEC numbering The adhesion bacterial population (individual/VEC) The VEC numbering The adhesion bacterial population (individual/VEC) The VEC numbering The adhesion bacterial population (individual/VEC)
????1 ????132 ????18 ????92 ????35 ????85
????2 ????113 ????19 ????120 ????36 ????123
????3 ????74 ????20 ????89 ????37 ????144
????4 ????149 ????21 ????65 ????38 ????131
????5 ????186 ????22 ????78 ????39 ????156
????6 ????118 ????23 ????158 ????40 ????98
????7 ????111 ????24 ????75 ????41 ????143
????8 ????63 ????25 ????126 ????42 ????125
????9 ????84 ????26 ????118 ????43 ????95
????10 ????89 ????27 ????109 ????44 ????194
????11 ????94 ????28 ????138 ????45 ????88
????12 ????118 ????29 ????80 ????46 ????155
????13 ????119 ????30 ????128 ????47 ????178
????14 ????125 ????31 ????74 ????48 ????122
????15 ????90 ????32 ????77 ????49 ????167
????16 ????120 ????33 ????50 ????50 ????133
????17 ????129 ????34 ????62 ????X±S ??113.8±33.69
Example two bacillus acidophilus' Chinese People's Anti-Japanese Military and Political College enterobacteria and Candida albicans effect
Bacillus acidophilus SL9404 disturbs escherichia coli and Candida albicans to obtain check in the epithelial adherent ability of human vagina with competitive way.
Get 1ml Escherichia coli bacteria liquid or 1ml Candida albicans bacterium liquid and 1ml VEC suspension mixing earlier, behind 37 ℃ of pre-cultivation 30min, add 1ml lactobacillus bacterium liquid again, mixing continues 37 ℃, 2 hours, observes down and counting by above-mentioned example one test method mirror.
The result who obtains represents in table two, shows that bacillus acidophilus SL9404 has reduced escherichia coli and Candida albicans in the epithelial adhesion of human vagina.
Table two bacillus acidophilus SL9404 strain influences at the VEC adhesive attraction escherichia coli 44113 or Candida albicans 10231
Bacterial strain Escherichia coli 44113 adhesion number/VEC (X ± S) Candida albicans 10231 adhesion number/VEC (X ± S) Statistical disposition
Escherichia coli 44113 ????180.7±252.20 ????P＜0.01
Escherichia coli 44113+bacillus acidophilus SL9404 ????134.18±40.63
Candida albicans 10231 ????29.24±8.85 ??0.01＜P＜0.05
Candida albicans 10231+bacillus acidophilus SL9404 ????24.40±11.36
Example three bacillus acidophilus are in external inhibition to gonococcus, escherichia coli, albicans growth
Bacillus acidophilus bacterium liquid constitutes test group with three kinds of pathogenic bacterium bacterium liquid mixed in equal amounts in triangular flask (9ml+9ml) respectively.Every kind of bacterium liquid mixes the formation matched group with equivalent 2% calf serum TPY culture fluid (9ml+9ml) simultaneously.Test group and matched group all place 37 ℃ of water-baths to be cultured to 72 hours or 96 hours.Each test group, matched group are being cultivated initial (0 hour), each sampling in 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, bacillus acidophilus in the determination test group triangular flask and the bacterial growth concentration of three kinds of pathogenic bacterium respectively.Also measure simultaneously the bacterium amount of each matched group.
The result who obtains represents in table three, shows that bacillus acidophilus SL9404 can comparatively fast suppress in external growth to gonococcus, also can suppress the growth of Escherichia coli, and Candida albicans is also had certain effect.
Table three bacillus acidophilus SL9404 strain antagonistic effect result
Group Viable count (CFU/ml) the Log value of different incubation times
0 hour 12 hours 24 hours 48 hours 72 hours 96 hours
Test group SL9404+gonococcus 29403 ??5.9165 ? ??3.3979 ????8.3502 ? ????3.8692 ??8.8209 ? ??＜1 ??8.4983 ? ??＜1 ??8.0253 ? ??＜1
SL9404+escherichia coli 44113 ???5.9165 ???3.7782 ????8.2577 ????8.8727 ??8.8395 ??8.7404 ??8.9004 ??＜5 ??8.7308 ??＜1
SL9404+Candida albicans 10231 ????5.9165 ????3.7782 ????8.3201 ????7.6821 ??8.5775 ??7.8420 ??8.7745 ??7.7474 ??8.8028 ??7.8921 ???8.7559 ???7.9731
Matched group Gonococcus 29403 ????3.3979 ????3.8865 ??3.6990 ??＜1 ??＜1
Escherichia coli 44113 ????3.7782 ????8.8882 ??8.8176 ??8.8987 ??8.4409
Candida albicans 10231 ????3.7782 ????7.8481 ??8.1614 ??8.1903 ??8.4314 ???8.2041
Bacillus acidophilus SL9404 ????5.9165 ????8.4100 ??8.7752 ??8.2455 ??7.8722 ???7.1303
Example four freeze-dried Lactobacillus acidophilus powders warp and excipient composition, preparation forms the prescription of tablet.
The lyophilizing mycopowder 100mg
Microcrystalline Cellulose 500mg
Lactose 320mg
Carboxymethyl starch sodium 50mg
Magnesium stearate 20mg
Colloidization silicon dioxide 10mg
Above-mentioned material needed 40 mesh sieves before the tabletting, and after fully mixing, in rotary tablet machine, was pressed into tablet with oval punch die, and sheet heavily is the tablet,vaginal of 1000mg.This film can soften within several minutes.Example five freeze-dried Lactobacillus acidophilus powders warp and weak acid and carbonate formulation, preparation becomes the prescription of vagina effervescence.
The lyophilizing mycopowder 100mg
Tartaric acid 400mg
Sodium bicarbonate 80mg
Calcium carbonate 60mg
Microcrystalline Cellulose 200mg
Lactose 120mg
Colloidization silicon dioxide 10mg
Magnesium stearate 20mg
Hypromellose 10mg
When preparing above-mentioned tablet, sodium bicarbonate and calcium carbonate need be made granule in advance, in rotary tablet machine, are pressed into tablet,vaginal with oval punch die, and sheet heavily is 1000mg.Through the gas release inspection, the result in 2ml water 20 minutes gas releases greater than 6ml, less than 10ml.Example six
The lyophilizing mycopowder 500mg (in two aluminium foil small package bags)
Physiological saline solution 10ml (in little plastic bottle)
The lyophilizing mycopowder is loaded in the aluminium foil pouch, in addition physiological saline solution is loaded in the plastic jar.During use the lyophilizing mycopowder is poured in the above-mentioned plastic jar into use-case seven vagina ointment (5g pipe) behind the mixing
The lyophilizing mycopowder 500mg
Lanoline 1g
Vaseline 1g
Colloidization silicon dioxide 2.5g
Lanoline, vaseline are preheating to 35-42 ℃, make whole thawings, add colloidization silicon dioxide and lyophilizing mycopowder again and stir, make evenly, in the flexible pipe of packing into.The preparation of example eight lactobacilluss sprays mould dose (10g)
The lyophilizing mycopowder 100mg
PVP 500mg
Physiological saline solution 10
Carbon dioxide In right amount
When preparing above-mentioned spray, earlier with the lyophilizing mycopowder, PVP and physiological saline solution are mixed with suspension, pour in the spray special-purpose bottle, with spray nozzle specially used airtight after, pour into carbon dioxide.

Claims (4)

1, a kind of pharmaceutical preparation that contains the bacillus acidophilus is characterized in that said preparation is made up of bacillus acidophilus and pharmaceutical excipient.
2, a kind of a kind of preparation method that contains bacillus acidophilus's pharmaceutical preparation as claimed in claim 1 is characterized in that this method comprises the following steps: (1) preparation bacillus acidophilus mycopowder:
The bacillus acidophilus SL9404 bacterial strain that secretions in the vagina is separated, obtains behind the purification is rule on the MRS agar slant and is gone down to posterity, and cultivates 48h through 37 ± 1 ℃.Choose full, color and luster is even, medium bacterium colony bigger than normal, once more on the MRS agar slant of streak inoculation in the Fructus Solani melongenae bottle, cultivates 48h through 37 ± 1 ℃.Transferred species inserts beef cysticercosis cellulosae cutis pipe, cultivates 20-50h for 37 ± 1 ℃.Get 1-4 and prop up the cysticercosis cellulosae cutis pipe and change 300ml milk fermentation culture medium or 5-20% Tomato juice fermentation medium or milk+Tomato juice's fermentation medium over to, cultivate the 8-48h fermentation culture for 37 ± 1 ℃, progressively be amplified to bulk fermentation, centrifugal, make mycopowder after the lyophilization; (2) preparation medicament:
Take by weighing bacillus acidophilus and pharmaceutical excipient by recipe quantity, drug formulation process is made various preparations routinely.
3, a kind of pharmaceutical preparation that contains bacillus acidophilus or its flora according to claim 1 is characterized in that wherein said medicinal excipient is corn starch, pregelatinized Starch, carboxymethyl starch sodium, dextrin, lactose, isolactose, mannitol, glucose, vitamin C, citric acid, sodium dihydrogen citrate, disodium citrate, tartaric acid, fumaric acid, boric acid or acidic amino acid class: glutamic acid, aspartic acid weak acid; Microcrystalline Cellulose, sodium carboxymethyl cellulose (calcium), hypromellose, polyvinylpyrrolidone, gelatin, calcium carbonate, sodium bicarbonate, magnesium stearate, lanoline, Polyethylene Glycol; Additive is meant edible defatted milk powder, vitamin C or glutamic acid.
4, a kind of pharmaceutical preparation that contains the bacillus acidophilus according to claim 1 is characterized in that said preparation is liquid form, mastic form or solid form, tablet, capsule, powder, cream, ointment, suppository or spray.
CNB001254693A 2000-09-28 2000-09-28 Medicinal preparation containing lactobacillus acidophilus and preparation process thereof Expired - Fee Related CN1234373C (en)

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US10286017B2 (en) 2011-05-09 2019-05-14 Probiotical S.P.A. Probiotic bacterial strains and symbiotic composition containing the same intended for infant food
US10384847B2 (en) 2011-09-23 2019-08-20 Probiotical North America Inc. Material impermeable to humidity and oxygen for packaging dietary products, cosmetics and medicinal specialities
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US11110136B2 (en) 2013-05-14 2021-09-07 Probiotical S.P.A. Composition comprising lactic acid bacteria for use in the preventive and/or curative treatment of recurrent cystitis
CN114591875A (en) * 2022-05-07 2022-06-07 微康益生菌(苏州)股份有限公司 Lactobacillus acidophilus LA88 for improving vaginal environment and culture method and application thereof

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US9492377B2 (en) 2011-01-28 2016-11-15 Probiotical S.P.A. Effervescent composition in solid form for use in vaginal applications for the treatment of vaginal infections
JP2014503581A (en) * 2011-01-28 2014-02-13 プロバイオティカル・ソシエタ・ペル・アチオニ Foam composition in solid form for use in vaginal applications to treat vaginal infections
KR101913797B1 (en) * 2011-01-28 2018-11-01 프로바이오티컬 에스.피.에이. Effervescent composition in solid form for use in vaginal applications for the treatment of vaginal infections
US10286017B2 (en) 2011-05-09 2019-05-14 Probiotical S.P.A. Probiotic bacterial strains and symbiotic composition containing the same intended for infant food
US9925224B2 (en) 2011-05-09 2018-03-27 Probiotical S.P.A. Bacterial strains belonging to the genus bifidobacterium for use in the treatment of hypercholesterolaemia
US10982184B2 (en) 2011-05-09 2021-04-20 Probiotical S.P.A. Bacterial strains capable of metabolizing oxalates
US10028982B2 (en) 2011-09-09 2018-07-24 Probiotical North America Inc. Composition comprising N-acetylcysteine and/or microencapsulated gastroprotected lysozyme in association with probiotic bacteria capable of restoring the stomach's own barrier effect which is lost during the pharmacological treatment of gastric hyperacidity
US10384847B2 (en) 2011-09-23 2019-08-20 Probiotical North America Inc. Material impermeable to humidity and oxygen for packaging dietary products, cosmetics and medicinal specialities
US11110136B2 (en) 2013-05-14 2021-09-07 Probiotical S.P.A. Composition comprising lactic acid bacteria for use in the preventive and/or curative treatment of recurrent cystitis
CN103695442B (en) * 2013-12-13 2016-01-20 上海市农业科学院 A kind of NFSAI gene coming from Lactobacterium acidophilum and its preparation method and application
CN103695442A (en) * 2013-12-13 2014-04-02 上海市农业科学院 NFSAI gene from lactobacillus acidophilus and preparation method and application thereof
CN107714672A (en) * 2017-09-11 2018-02-23 广州市微生物研究所 A kind of woman is with vagina active probiotic gel packing and preparation method thereof
CN107714672B (en) * 2017-09-11 2019-08-06 广州市微生物研究所 A kind of woman's vagina active probiotic gel packing and preparation method thereof
CN114591875A (en) * 2022-05-07 2022-06-07 微康益生菌(苏州)股份有限公司 Lactobacillus acidophilus LA88 for improving vaginal environment and culture method and application thereof

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Patentee before: Sine Pharmaceutical Factory Co., Ltd.

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Address after: 201206 Pudong New Area new Jinqiao Road, Shanghai, No. 905

Patentee after: Shanghai Xinyi Pharmaceutical Co. Ltd..

Address before: 201206 Pudong New Area new Jinqiao Road, Shanghai, No. 905

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Granted publication date: 20060104

Termination date: 20190928

CF01 Termination of patent right due to non-payment of annual fee