阅读说明：本技术 用于稳定***液的酸度和氧化还原状态的基于合适的生化组合物的泌尿生殖器医疗设备制剂 (Urogenital medical device formulation based on suitable biochemical compositions for stabilizing the acidity and redox state of vaginal fluids ) 是由 F.格拉弗 于 2017-11-30 设计创作，主要内容包括：本发明涉及用于阴道或可替代地经口施用的组合物,以及此类组合物用于稳定阴道酸度或可替代地肠道酸度,并且建立用于泌尿生殖器或可替代地胃肠道中的病原体抑制和乳杆菌生长的有利条件的用途。(The present invention relates to compositions for vaginal or alternatively oral administration and the use of such compositions for stabilizing vaginal acidity or alternatively intestinal acidity and establishing favorable conditions for pathogen inhibition and lactobacillus growth in the urogenital or alternatively gastrointestinal tract.)

1. A composition for use in treating or preventing a urogenital infection caused by a pathogenic microorganism in a female subject, wherein the composition comprises an effective amount of each of,

a) a buffering agent, a water-soluble polymer,

b) a preservative that is fully compatible with the vaginal microbiota,

c) pure molecular or ionic biochemical nitrogen sources available to the vaginal microbiota,

d) a prebiotic polysaccharide that is fermentable by the vaginal microbiota,

wherein the treatment or prevention comprises intravaginal administration of the composition to a subject for stabilizing the acidity of vaginal fluid in the subject at a physiological value of about pH4.5 or less.

2. The composition for use according to claim 1, wherein the vaginal formulation is a lubricating gel for personal use.

3. The composition for use according to claim 1 or 2, further comprising a sufficient source of sulfur to be assimilable by the lactobacillus.

4. A composition for use according to claim 3, wherein the sulphur source is at least a sulphur-containing compound.

5. The composition for use according to claim 4, wherein the sulfur-containing compound is at least one selected from the group consisting of sulfate, sulfite, sulfide, N-acetylcysteine, cysteine, methionine, sodium thiosulfate, and a mixture thereof.

6. A composition for use according to any preceding claim comprising a component that modulates the redox potential in vaginal fluid.

7. The composition for use according to claim 6, wherein said component that modulates the redox potential in vaginal fluids is at least one redox system selected from the group consisting of: sulfur-based systems such as sodium thiosulfate/sodium bisulfite, glutathione in the reduced (GSH) and oxidized (GSSG) states, cysteine/cystine pairs, oxygen-based redox systems such as methionine, quinone/hydroquinone, nitrogen-based redox systems such as nitrite/nitrate and selenium-based redox systems (selenite/selenate and selenide), and mixtures thereof.

8. The composition for use according to any of the preceding claims, wherein one or more lactobacillus strains selected from the group consisting of: lactobacillus gasseri, lactobacillus jensenii, lactobacillus crispatus, lactobacillus acidophilus, lactobacillus helveticus, lactobacillus plantarum, lactobacillus fermentum, lactobacillus lactis, lactobacillus johnsonii, and lactobacillus acidophilus KS 400.

9. The composition for use according to any of the preceding claims, wherein one or more lactobacillus strains selected from the group consisting of: lactobacillus gasseri, lactobacillus jensenii, lactobacillus crispatus, lactobacillus acidophilus, lactobacillus helveticus, lactobacillus plantarum, lactobacillus fermentum, lactobacillus lactis, and lactobacillus johnsonii.

10. Composition for use according to any one of the preceding claims, the lactobacillus strain being selected from lactobacillus jensenii KS109, having accession number CNCM I-3482, lactobacillus gasseri KS114.1 deposited at the french national collection of microbial cultures by Medinova AG at 7/22.2005, having accession number CNCM I-3483, lactobacillus jensenii 116.1 deposited at the french national collection of microbial cultures (CNCM) at 22.7/2005 by Medinova AG, having accession number CNCM I-3217, lactobacillus jensenii KS119.1 deposited at french national collection of microbial cultures (CNCM) at 6/4.2004 by Medinova AG, having accession number CNCM I-3484, lactobacillus crispatus deposited at french national collection of microbial cultures (CNCM) at 22.7/2005 by Medinova AG, having accession number CNCM I-3218, Lactobacillus gasseri 120.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus jensenii KS 121.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus jensenii KS122.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus gasseri 123.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 22.7.2005, Lactobacillus gasseri 123.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus gasseri 124.3 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 127.2, Lactobacillus gasseri 126.2 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 7.22.7.9.86, lactobacillus jensenii KS 130.1, having accession number CNCM I-3360, lactobacillus helveticus KS 300 deposited by Medinova AG at french national collection of microbiological cultures (CNCM) at 10/2/2005, and lactobacillus acidophilus KS 400.

11. The composition for use according to claim 10, wherein said lactobacillus strain is selected from lactobacillus jensenii KS119.1 with accession number CNCM I-3217, deposited by Medinova AG at french national collection of microorganisms (CNCM) at 6.4.2004, lactobacillus crispatus 119.4 with accession number CNCM I-3484, deposited by Medinova AG at french national collection of microorganisms (CNCM) at 22.7.2005, lactobacillus gasseri 120.1 with accession number CNCM I-3218, deposited by Medinova AG at french national collection of microorganisms (CNCM) at 4.6.2004, lactobacillus gasseri 124.3 with accession number CNCM I-3220, deposited by Medinova AG at french national collection of microorganisms (CNCM) at 4.6.2004.

12. The composition for use according to any one of the preceding claims, wherein the formulation further comprises an effective amount of a therapeutically effective antimicrobial agent capable of inhibiting or eradicating urogenital pathogens, including parasites, protozoa, bacteria, viruses, and/or fungi.

13. A composition for use according to claim 12, wherein the formulation further comprises a therapeutically effective antifungal agent in an amount effective to inhibit or eradicate fungi in the urogenital area.

14. The composition for use according to claim 13, wherein the therapeutically effective antifungal agent is sorbic acid or a salt thereof.

15. A method for treating or preventing urogenital infections caused by pathogenic microorganisms in a female subject, comprising intravaginally administering to the subject an effective amount of each of:

a) a buffering agent, a water-soluble polymer,

b) a preservative that is fully compatible with the vaginal microbiota,

c) pure molecular or ionic biochemical nitrogen sources available to the vaginal microbiota,

d) a prebiotic polysaccharide that is fermentable by the vaginal microbiota,

wherein the treatment or prevention comprises intravaginal administration of the composition to the subject.

16. A method for stabilizing the acidity of vaginal fluid in a subject at a physiological value of about pH4.5 or less, comprising intravaginal administration to the subject of an effective amount of each of:

a) a buffering agent, a water-soluble polymer,

b) a preservative that is fully compatible with the vaginal microbiota,

c) pure molecular or ionic biochemical nitrogen sources available to the vaginal microbiota,

d) prebiotic polysaccharides fermentable by the vaginal microbiota

Wherein the treatment or prevention comprises intravaginal administration of the composition to the subject.

17. The method according to claim 15 or 16, wherein the buffer a) is an acid or a salt thereof selected from the group comprising or alternatively consisting of: organic acids such as aliphatic acids including acetic acid, propionic acid, butyric acid, dicarboxylic acids including oxalic acid, malonic acid, succinic acid, hydroxy acids such as lactic acid, citric acid, dicarboxylic acids such as malic acid.

18. The method according to any one of claims 15-17, wherein the preservative b) is lactic acid or a derivative and/or salt thereof.

19. The method according to any one of claims 15 to 18, wherein the nitrogen source c) is an ammonium salt, urea, a naturally occurring or synthetic amino acid such as glutamic acid and mixtures thereof.

20. Method according to any one of claims 15-19, wherein the carbon source d) is a non-digestible carbohydrate, such as: fructooligosaccharides, galactooligosaccharides, glucooligosaccharides and mixtures thereof.

21. The method of any one of claims 15-20, wherein the composition further comprises a sufficient source of sulfur that can be assimilated by the lactobacillus.

22. The method of claim 21, wherein the sulfur source is at least a sulfur-containing compound.

23. The method according to claim 22, wherein the sulfur-containing compound is at least one selected from the group consisting of sulfate, sulfite, sulfide, N-acetylcysteine, cysteine, methionine, sodium thiosulfate, and a mixture thereof.

24. The method according to any one of claims 15-23, comprising a component that modulates the redox potential in vaginal fluid.

25. The method according to claim 24, wherein said component that modulates redox potential in vaginal fluid is at least one redox system selected from the group consisting of: sulfur-based systems such as sodium thiosulfate/sodium bisulfite, glutathione in the reduced (GSH) and oxidized (GSSG) states, cysteine/cystine pairs, oxygen-based redox systems such as methionine, quinone/hydroquinone, nitrogen-based redox systems such as nitrite/nitrate and selenium-based redox systems (selenite/selenate and selenide), and mixtures thereof.

26. A formulation having a pH in the range of 4 to 5 suitable for oral administration, wherein the composition comprises an effective amount of each of:

a) a buffering agent, a water-soluble polymer,

b) a preservative that is fully compatible with the vaginal microbiota,

c) pure molecular or ionic biochemical nitrogen sources available to the vaginal microbiota,

d) prebiotic polysaccharides that are fermentable by the vaginal microbiota.

27. The formulation of claim 26, wherein at least one strain of lactobacillus is added to said formulation, said strain being selected from the group consisting of lactobacillus gasseri, lactobacillus jensenii, lactobacillus crispatus, lactobacillus acidophilus, lactobacillus helveticus, lactobacillus plantarum, lactobacillus fermentum, and lactobacillus johnsonii.

28. The formulation of claim 27, wherein at least one strain of lactobacillus selected from lactobacillus jensenii KS109, having accession number CNCM I-3482, lactobacillus gasseri KS114.1 deposited at the french national collection of microbial cultures by Medinova AG at 7.22.2005, lactobacillus crispatus deposited at french national collection of microbial cultures (CNCM) by Medinova AG at 22.7.2005, lactobacillus jensenii 116.1 deposited at french national collection of microbial cultures (CNCM) by Medinova AG at 22.7.2005, having accession number CNCM I-3217, lactobacillus jensenii KS119.1 deposited at french national collection of microbial cultures (CNCM) by Medinova AG at 4.6.2004, having accession number CNCM I-3484, deposited at french central of microbial cultures (CNCM) by Medinova AG at 22.7.2005, having accession number CNCM 119.8-3218, Lactobacillus gasseri 120.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus jensenii KS 121.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus jensenii KS122.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus gasseri 123.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 22.7.2005, Lactobacillus gasseri 123.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus gasseri 124.3 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 127.2, Lactobacillus gasseri 126.2 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 7.22.7.9.86, lactobacillus jensenii KS 130.1, having accession number CNCM I-3360, lactobacillus helveticus KS 300 deposited by Medinova AG at french national collection of microbiological cultures (CNCM) at 10/2/2005, and lactobacillus acidophilus KS 400.

29. The formulation of claim 28, wherein said lactobacillus strain is selected from lactobacillus jensenii KS119.1 having accession number CNCM I-3217, deposited at french national collection of microorganisms (CNCM) by Medinova AG at 6.4.2004, lactobacillus crispatus 119.4 having accession number CNCM I-3484, deposited at french national collection of microorganisms (CNCM) by Medinova AG at 22.7.2005, lactobacillus gasseri 120.1 having accession number CNCM I-3218, deposited at french central collection of microorganisms (CNCM) by Medinova AG at 4.6.2004, lactobacillus gasseri 124.3 having accession number CNCM I-3220, deposited at french collection of microorganisms (CNCM) by Medinova AG at 4.6.2004, and lactobacillus acidophilus KS 400.

30. The formulation according to any one of claims 26-29, wherein said composition further comprises a sufficient source of sulfur to be assimilable by the lactobacillus.

31. The formulation according to claim 30, wherein said sulfur source is at least a sulfur-containing compound.

32. The formulation according to claim 31, wherein said sulfur-containing compound is at least one selected from the group consisting of sulfate, sulfite, sulfide, N-acetylcysteine, cysteine, methionine, sodium thiosulfate, and mixtures thereof.

33. The formulation according to any one of claims 26 to 32, comprising a component that modulates the redox potential in a biological fluid.

34. The formulation according to claim 33, wherein said component that modulates redox potential in vaginal fluid is at least one redox system selected from the group consisting of: sulfur-based systems such as sodium thiosulfate/sodium bisulfite, glutathione in the reduced (GSH) and oxidized (GSSG) states, cysteine/cystine pairs, oxygen-based redox systems such as quinone/hydroquinone, nitrogen-based redox systems such as nitrite/nitrate and selenium-based redox systems (selenite/selenate and selenide), and mixtures thereof.

35. The composition for use according to any one of claims 1 to 14, or the formulation according to any one of claims 26 to 34, wherein the buffer a) is selected from acids or salts thereof comprising or alternatively consisting of: organic acids such as aliphatic acids including acetic acid, propionic acid, butyric acid, dicarboxylic acids including oxalic acid, malonic acid, succinic acid, hydroxy acids such as lactic acid, citric acid, dicarboxylic acids such as malic acid.

36. The composition for use according to any one of claims 1 to 14 or 35, or the formulation according to any one of claims 26 to 35, wherein the preservative b) is lactic acid or a derivative and/or salt thereof.

37. The composition for use according to any one of claims 1 to 14 or 35 to 36, or the formulation according to any one of claims 26 to 36, wherein the nitrogen source c) is an ammonium salt, urea, a naturally occurring or synthetic amino acid such as glutamic acid, and mixtures thereof.

38. The composition for use according to any one of claims 1 to 14 or 35 to 37, or the formulation according to any one of claims 26 to 37, wherein the carbon source d) is a non-digestible carbohydrate, such as: fructooligosaccharides, galactooligosaccharides, glucooligosaccharides and mixtures thereof.

39. The composition for use according to any one of claims 1-14 or 35-38, or the formulation according to any one of claims 26-38, which has a pH of 43 to 4.6 if the solid is brought to a volume of 3ml after dissolution/suspension, and which comprises the following ingredients per single dose:

ca-lactic acid pentahydrate: 30-90, preferably 60 mg

Magnesium citrate dibasic: 15-45, preferably 30mg

Prebiotic fiber: 50-150, preferably 100 mg

Glutamic acid: 20-60, preferably 40 mg

Cysteine/thiosulfate: 10-50, preferably 30mg

Adding 400-1100 mg of excipient: proper amount.

40. A composition for use or formulation according to claim 39, further comprising a lyophilized product of:

lactobacillus gasseri 120.1 having accession number CNCM I-3218, deposited by Medinova AG in French national Collection of cultures of microorganisms (CNCM) on 6/4/2004, and/or

Lactobacillus gasseri 124.3 and/or Lactobacillus crispatus 119.4 with accession number CNCM I-3484, deposited by Medinova AG at 22.7.2005 in the French national Collection of microorganisms (CNCM), and/or

Lactobacillus jensenii KS119.1, deposited by Medinova AG at french national center for microbiological culture collection (CNCM) at 6/4/2004 with accession number CNCM I-3217, amounting to 50 to 200 mg for a total of 20 to 100 hundred million c.f.u.

Technical Field

The present invention relates to biochemical compositions of urogenital preparations which stabilize the acidity in the vagina and in which suitable conditions for urogenital pathogen inhibition and lactobacillus growth are established. The present invention provides compositions for vaginal or alternatively oral administration and the use of such compositions to stabilize vaginal or alternatively intestinal acidity and establish favorable conditions for pathogen inhibition and lactobacillus growth in the urogenital or alternatively gastrointestinal tract. These compositions are based on chemically defined ingredients, avoiding the usual natural or ill-defined nutritional components, and may be associated with human vaginal strains having specific antimicrobial and/or anti-inflammatory properties. Furthermore, attention is paid to the appropriate values of the redox potential of such formulations after vaginal administration, in particular with respect to gardnerella vaginalis (b.vaginalis: (b.vaginalis)) (b.G. vaginalis) Inhibition of a type of Bacterial Vaginosis (BV) -associated pathogen.

Finally, formulations containing agents which additionally inhibit opportunistic yeasts such as Candida albicans are presentedCandida albicans) Without impairing the growth of the lactobacilli.

Background

It is well known that useful, well-tolerated non-pathogenic bacteria, the so-called human natural microbiota, dominate in the intestine, vagina and urethra/bladder in a healthy state, whereas overgrowth of pathogenic and opportunistic microorganisms such as bacteria, yeasts, viruses and protozoa often leads to dysbiosis infections in these organs.

The terms "aerobic bacteria" and "anaerobic bacteria" are generally used to describe those microorganisms that require air and those microorganisms that cannot grow in the presence of air, respectively.

Notably, some of the non-pathogenic and pathogenic bacteria that can be found in the vagina can be defined as "non-facultative anaerobes," i.e., they can grow in both the presence and absence of air, and are substantially unaffected by the oxidizing conditions in the growth medium. A non-limiting example of such non-facultative anaerobic bacteria is Escherichia coli (E.coli) (II)E. coli) And Staphylococcus aureus (Staphylococcus aureus) Which is known to cause a vaginal condition commonly indicated as "aerobic vaginitis".

Other microorganisms that may be found in the vagina are "obligate anaerobes," i.e., they do not grow in the presence of free oxygen and can, in fact, be killed in the presence of oxygen or oxidizing conditions.

Anaerobic organisms such as gardnerella vaginalis (an obligate anaerobe) that migrate from the perineum into the vagina, or are obtained through sexual intercourse, can cause dysbiosis with few symptoms, followed by global Bacterial Vaginosis (BV), which is the most common vaginal infection. BV constitutes a serious risk for premature birth in pregnant women and increases the risk for sexually transmitted infections (reference 1). In such cases of vaginal dysbiosis or vaginal infections caused by anaerobic bacteria, the pH of the vaginal environment increases from physiological values of 3.5-4.5 to values of 5-6 or even higher. This increase in pH is a key clinical parameter for diagnosing BV and is one of the major causes of risk associated with BV. The fermentation of metabolites of glycogen and other carbohydrates present in the vagina by the lactobacillus Doderlein microbiota produces mainly lactic acid and to a lesser extent other organic acids and results in a low pH of the vaginal fluid. In addition, certain strains of gram-negative escherichia coli, which originate in the intestinal tract and migrate first to the vagina and then to the urethra/bladder, are the most common cause of urethritis/cystitis in sexually active women. This same E.coli can also cause dysbiosis in the vagina, a condition known as aerobic vaginitis, which is also accompanied by a perturbation of the vaginal microflora, a decrease in vaginal acidity and a corresponding increase in pH (ref.2).

Another pathogen that causes vaginal infections, particularly during pregnancy, is Candida yeast (C.) (R.)Candida) It is expressed opportunistically and often coexists with lactobacilli.

Candida is the only yeast organism that behaves as a facultative anaerobe, i.e., it is essentially unaffected by the presence or absence of air and oxidizing conditions.

These infections are usually cured at least to some extent by anti-infective therapy against pathogens that are expected to be successfully eradicated during the course of such therapy. However, in about up to 70% of cases, patients experience relapse or reinfection within 3 months after primary treatment (reference 3) because the pathogens are not adequately killed or lack protection due to an innate vaginal microbiota through restoration. This microbiota is in fact often compromised by anti-infective treatments. The indigenous vaginal microbiota, historically referred to as the Doderlein microbiota, is sensitive to many anti-infective agents, particularly antibiotics. It is often disturbed or even eradicated by anti-infective therapy and therefore does not inhibit the proliferation of the ever-present opportunistic, often pathogenic organisms. Thus, bacterial vaginosis treated by a narrow spectrum antibiotic specific for anaerobic bacteria, such as clindamycin, can lead to yeast infections, as the vaginal microbiota consumed by lactobacilli can no longer inhibit opportunistic yeast.

In 1892 pioneering work, exogenous lactobacillus was found (Lactobacillus) The physician AlbertDoderlein, i, has proposed the possibility of applying exogenous lactobacillus directly to the vagina to treat vaginal dysbiosis or infection (reference 4). Since then, this concept has been widely developed in the scientific and patent literature, and recently, has been extensively reviewed (reference 5).

WO 2009/123982 a2 relates to methods and compositions for lactobacillus replacement therapy by colonizing the vaginal mucosa with a desired microbial species by contacting the vaginal wall with a dried preparation of viable microbial cells.

WO 2016/020861 a1 relates to film or sponge compositions, and methods for preventing and treating vaginal infections, wherein the film or sponge comprises at least one mucosal polymer (polumer), at least one probiotic and/or prebiotic, and at least one active compound such as an antifungal agent or an antibiotic agent in a pharmacologically effective amount.

In recent years, the present inventors have conducted intensive studies on the properties of previously unknown lactobacillus strains of human vaginal origin that compete with the most common urogenital pathogens. In particular, the present inventors studiedMechanisms involving interactions of lactic acid, hydrogen peroxide and further reagents were found to explain, for example, Lactobacillus gasseri: (A. gasseri.), (B. peroxide)Lactobacillus gasseri) Against pathogenic Gardnerella vaginalis and Prevotella diplodiae (Prevotella bivia) Antibiotic-like activity of particular strength and specificity (reference 6) and Lactobacillus jensenii: (L jensenii) And Lactobacillus crispatus: (L crispatus) Activity against uropathogenic e.coli (reference 7). The method as described in the mentioned papers and patents (reference 8) further has the advantage of using strains isolated from the same area of a fully healthy female for therapeutic applications targeting the urogenital tract. Thereby, possible problems with tolerance of exogenously administered lactobacillus strains may be reduced to a large extent compared to the use of such strains not derived from human origin and not from the urogenital tract of healthy young women. For strains of non-human origin, safety and tolerability issues in therapeutics such as urogenital applications are not explicitly a priori and therefore must be evaluated a posteriori.

In order to provide a suitable composition for a pharmaceutically acceptable carrier, the inventors investigated different formulations used in probiotic products and/or described in the literature. In one disclosed approach (reference 9), the inventors claim that the use of skim milk preparations and derivatives, respectively called LGF (lactobacillus growth factor) or NGF (natural growth factor), favours the growth of the desired lactobacillus by simultaneously inhibiting the growth of undesired uropathogens. According to the same inventors, such specialized growth factors for vaginal applications have also been found in microbial growth media. The most commonly used such growth medium for lactobacillus strains is in fact MRS (DeMan-Rogosa-Sharp) medium (reference 10). In addition to minerals and vitamins, this broth contains peptone, a carbon source and especially nitrogen of undefined composition of chemical composition. Other inventors also recommend the use of complex protein substances such as skim milk and albumin in a preservation matrix for vaginal medications containing lactobacillus strains (reference 11).

In our opinion, although preparations containing natural nutrients such as skim milk or peptone may be useful for particular applications, they are not suitable as ingredients in pharmaceutical preparations containing lactobacillus for vaginal applications. This is because, firstly, they are of biological (especially animal) origin, and secondly, these natural components often have an undefined composition of chemical composition and, therefore, as explained further below, they carry an increased risk of causing adverse effects in the patient. Furthermore, from the point of view of selective efficacy, these NGFs may not achieve the desired effect of inhibiting uropathogen growth. After having been contacted and mixed with vaginal fluid, not only lactobacillus strains but also numerous pathogens can grow in such abundant ingredients (with proteins, carbohydrates, vitamins, minerals, etc.) as skim milk. Pathogens such as e.coli sometimes grow faster in milk than in nutrient broths.

With regard to 2 pathogens commonly found in urogenital microbiota, staphylococcus aureus and escherichia coli, both pathogens also grew several logs within about 20 hours when inoculated in a medium specific for lactobacillus, such as MRS (table 1), while similar results were obtained with skim milk (not shown below).

Table 1: growth performance of 2 pathogens in MRS broth.

Even from a relatively high concentration of 10⁸cfu/ml starts, these pathogens can also grow several logs further in MRS, stopping only after about 20 hours by the low pH they generate. That is, a growth medium that is so-called highly specific for lactobacilli may also promote the growth of dangerous pathogens. To inhibit the growth of such pathogens, effective and selective inhibitors must be added to the formulation.

Thus, if a subject has primarily healthy lactobacillus strains present in sufficient numbers in the vagina, administration of LGF may be appropriate as these indigenous lactobacillus bacteria can metabolize them. However, it is not appropriate to administer any kind of LGF or such non-specific growth factors vaginally, which is dysregulated ecologically, already infected or lacking in beneficial lactobacillus, since these may enhance the growth of the pathogens present, rather than the rare lactobacillus strains. Under such treatment with LGF, dysbiosis or infections may even become worse, said risk should be avoided.

In summary: skim milk preparations or components of lactobacillus growth media such as MRS may in some cases sustain the growth of probiotic (eubiotic), indigenous vaginal lactobacilli. However, in general, they are not suitable as therapeutic ingredients for pharmaceutical preparations to be applied in the dysbiosis vagina, since they have low selectivity for lactobacillus and are not well defined in chemical composition. The present invention is intended to overcome these disadvantages.

Due to lactic acid and different fatty acid production by the physiological Doderlein (i.e. vaginal) microbiota, vaginal fluid shows an acidic pH on average between 3.5 and 4.5, which seems to be optimal for vaginal homeostasis, as lactobacillus species can still grow at this pH, thereby inhibiting most pathogens and opportunistic microorganisms. Above pH4.5, pathogens can readily grow, but below pH 3.5, the growth of the lactobacillus Doderlein itself becomes increasingly inhibited. Such acidic vaginal fluids, as produced by fermentation of endogenous carbohydrates such as glycogen metabolites via the Doderlein microbiota, are a very biochemical basis of the defense mechanism of the vagina against infections and are critical for reproductive health in women (reference 12).

In view of these factors, it is not surprising to find references in the literature (e.g., reference 13) to include unspecified pH buffers in vaginal compositions. In another application (reference 14), compositions containing buffering agents (chemical composition is uncertain in the mentioned reference) are recommended in order to stabilize the acidity of the vagina within the range pH 3.5-5.0.

However, the inventors found that in all cases, an acidic pH alone was not sufficient to prevent the growth of pathogens. Experiments conducted in the inventors' laboratory (shown below) indicate that relevant pathogens such as staphylococcus aureus and escherichia coli can grow rapidly even below pH 5. The medium used contained lactose as carbon source and 15% MRS as growth initiation promoter.

In summary: the importance of an acidic vaginal pH is unanimous in the scientific community, and some authors have recognized the usefulness of adding buffers to pharmaceutical formulations designed for vaginal applications. The pH range must therefore be clearly defined in terms of physiology within the range between 3.5 and 4.5. A pH4.5 value at the upper end of the physiological range is intentionally chosen because it is well tolerated by patients. More acidic values of pH about 4 or lower can cause discomfort such as itching and burning, as the vaginal epithelium is more sensitive in the case of dysbiosis and infections.

In addition, the buffer chemistry and dosage must be indicated in the pharmaceutical formulation with respect to, for example, the capsule. Finally, the buffer must be such that its action is exerted shortly after intravaginal administration and at least continues until the next administration, i.e. 12 to 24 hours.

In addition to the acidic buffers and trophic factors set forth in the preceding paragraph, there are other factors that play a role in the design of an effective pre-or pro-or symbiotic formulation for vaginal application. One such factor is the redox potential of the vaginal fluid after application of the device.

As we will show, this is of relevance especially in the case of Bacterial Vaginosis (BV) where the pathogen is obligate anaerobe (gardnerella vaginalis, prevotella diplodi, atropa vaginalis ((BV)Atopobium vaginae) … …). The redox potential can indeed influence the growth of these last-mentioned pathogens.

The effect of redox potential on the growth and survival of microorganisms in vaginal flora is shown in FIG. 3 (according to reference 22, Eschenbach et al 1985)

Finally, another potentially dangerous microorganism is the fungus candida albicans, which often exists as an opportunistic bacterium in the vaginal flora. The authors will propose a method to control fungal growth without inhibiting the growth of lactobacillus vaginalis.

The present invention aims to provide a solution to these objectives.

Summary of The Invention

In one embodiment, the present invention relates to a composition for use in treating or preventing urogenital infections caused by pathogenic microorganisms in a female subject, wherein the composition comprises an effective amount of each of:

a) a buffering agent, a water-soluble polymer,

b) a preservative that is fully compatible with the vaginal microbiota,

c) pure molecular or ionic biochemical nitrogen sources available to the vaginal microbiota,

d) a prebiotic polysaccharide that is fermentable by the vaginal microbiota,

wherein the treatment or prevention comprises intravaginal administration of the composition to a subject for stabilizing the acidity of vaginal fluid in the subject at a physiological value of about pH4.5 or less.

In one embodiment, the present invention provides a method for treating or preventing urogenital infections caused by pathogenic microorganisms in a female subject, comprising intravaginally administering to the subject an effective amount of each of:

a) a buffering agent, a water-soluble polymer,

b) a preservative that is fully compatible with the vaginal microbiota,

c) pure molecular or ionic biochemical nitrogen sources available to the vaginal microbiota,

d) a prebiotic polysaccharide that is fermentable by the vaginal microbiota,

wherein the treatment or prevention comprises intravaginal administration of the composition to the subject.

In another embodiment, the present invention provides a method for stabilizing the acidity of vaginal fluid in a subject at a physiological value of about ph4.5 or less, comprising intravaginally administering to the subject an effective amount of each of:

a) a buffering agent, a water-soluble polymer,

b) a preservative that is fully compatible with the vaginal microbiota,

c) pure molecular or ionic biochemical nitrogen sources available to the vaginal microbiota,

d) a prebiotic polysaccharide that is fermentable by the vaginal microbiota,

wherein the treatment or prevention comprises intravaginal administration of the composition to the subject.

In another embodiment, the present invention relates to a formulation suitable for oral administration having a pH in the range of 4 to 5, wherein the composition comprises an effective amount of each of:

a) a buffering agent, a water-soluble polymer,

b) a preservative that is fully compatible with the vaginal microbiota,

c) pure molecular or ionic biochemical nitrogen sources available to the vaginal microbiota,

d) prebiotic polysaccharides that are fermentable by the vaginal microbiota.

Brief Description of Drawings

FIG. 1: graphical representation of data obtained in growth experiments with Lactobacillus gasseri KS 120.1 CNCM I-3218.

FIG. 2: graphical representation of data obtained in growth experiments with multiply resistant staphylococcus aureus CCOS 461.

FIG. 3: schematic two-dimensional representation of the redox potential Eh and acidic pH in the vagina according to the composition of the vaginal microbiota (flora).

(according to reference Eschenbach et al 1985)

The present invention provides a composition for stabilizing the pH of vaginal fluids at physiological values of about 4.5, and optionally for a sufficient duration of time for the redox potential of the vaginal fluid to be higher than 150 mV, and selectively favouring the growth of lactobacillus species, according to the appended claims. Against the growth of pathogens, particularly anaerobic bacteria, as a means of preventing, alleviating or treating vaginal and genitourinary infections such as BV caused by pathogenic microorganisms.

The composition according to the invention may optionally comprise natural, chemically defined substances in concentrations that inhibit candida species, but not lactobacillus species bacteria.

Acidity and nutrients: formulations against harmful anaerobic pathogens

The method uses a well-defined pharmaceutical formulation composition of specific, biochemical components to be applied vaginally.

The compositions in the context of the present invention contain the following biochemical components:

-a biochemical buffer to maintain the pH of the composition (in vitro and in vivo) in the desired range of 4.0 to 5.0, preferably 4.5, after administration and up to 12-24 hours thereafter;

anti-pathogenic preservatives having biochemical properties otherwise fully compatible with the bacterial components of the Doderlein (vaginal) microbiota;

-selecting a biological, chemically-defined carbon source for selectively enhancing the growth of lactobacillus species;

a bioavailable, otherwise chemically pure, nitrogen source that supports adequate growth and proliferation of lactobacillus species (already present or administered); and optionally

-a sufficient, chemically defined, source of sulphur assimilable by the lactobacilli;

redox regulators (oxidizers combined with antioxidants in appropriate ratios);

chemical compounds (organic and/or inorganic) selected for their anti-infective activity specific and targeted against different kinds of pathogens, such as parasites and/or protozoa and/or bacteria and/or fungi and/or viruses, as well as selected minerals, vitamins and other optional components, which may be associated with the growth of selected lactobacillus species or the inhibition of undesired pathogens. A non-limiting example of an antifungal agent, particularly against candida, is sorbate.

The ingredients may be present in the compositions of the invention alone or in combination with: as living cells or lyophilized products thereof, free ofProbiotic Lactobacillus strains of cell culture supernatant or its tyndallisate are selected for their specific properties, including but not limited to antibiotic-like activity against urogenital pathogens, adhesion to vaginal/urethral epithelial cells, H₂O₂Lactic acid and related bacteriocins and enzyme production.

The formulation itself is in a suitable galenical form in a pharmaceutically acceptable carrier or delivery system. The actual composition may be instilled in the form of a freeze-dried formulation, cream, paste, gel liquid or suppository, capsule or tablet/stylus for enteral, urethral or vaginal application. Alternatively, the formulation may be a capsule or tablet or suppository, cream or gel (lubricant) for vaginal delivery.

The probiotic lactobacillus species mentioned above should be taken as being generally at 10 in the practice of the method⁵To 10¹⁰The amount of cfu of viable cells is administered or as a cell-free supernatant of the corresponding culture in liquid, semi-solid or lyophilized form. The Lactobacillus species is preferably selected from Lactobacillus crispatus, Lactobacillus jensenii, Lactobacillus gasseri, Lactobacillus acidophilus: (A)L. acidophilus) Inert lactobacillus (I), (II)L. iners) Lactobacillus plantarum (II)L. plantarum) Lactobacillus fermentum (I)L. fermentum) Lactobacillus lactis (I), (II)L. lactis) And Lactobacillus johnsonii: (L johnsonii）。

In contrast, the method according to the invention does not involve the use of culture media such as natural dairy nutrients or components of animal origin as previously proposed elsewhere. Apart from the fact that in clinical studies further confirmation of the efficacy of said growth factors is required, such nutrients and components are not suitable for pharmaceutical intravaginal or intraurethral applications for the following medical reasons. They may contain known and unknown substances with proteinaceous properties, having allergenic or irritating potential against sensitive epithelium of the vagina and urethra and/or bacterial spores, viruses, prions or their components/metabolites that may have been sterilized and become a potential health threat to the patient.

The same considerations detailed above with respect to the optimal composition of the probiotic formulation apply analogously to the Gastrointestinal (GI) environment. Although the GI tract is rich in nutrients for the resident microbiota and the pathogens resident therein, compared to the urogenital tract, the probiotic formulation should help maintain optimal acidity and redox status in the GI tract, while at the same time the concept of promoting the growth of healthy probiotics while inhibiting the growth of pathogens and opportunistic species remains effective.

However, when the focus shifts from the urogenital to the GI tract, which is the ecosystem, two major differences need to be considered.

First, acidity in the GI tract is variable due to its nature and length (reference 17). The following pH values are typically measured in normal humans:

a. stomach: 1.0 to 2.5

b. Proximal small intestine: 6.6

c. Terminal of ileum: 7.5

d. Cecum: 6.4

e. Rectum: 7.0

The pH in the intestine varies between 6 and 7 compared to values ranging from 3.5 to 5 in a healthy vagina.

Another difference is that the major acidic fermentation product of the GI microbiota is not the predominant lactic acid, but a full range of Short Chain Fatty Acids (SCFAs), including acetate or butyrate, which constitute the major carbon flux from the diet through the GI microbiota to the host and a significant contribution to the metabolism of the host (reference 18).

Thus, when applied to the intestinal tract, the prebiotic fibre included in the composition according to the invention to improve the metabolism of strains of the Doderlein microbiota will benefit not only lactobacillus species but also e.g. bifidobacteria and other resident species. Advantageously, the composition according to the invention comprises at least two different fibres that act in different levels of the intestinal tract, since the GI microbiota in the upper and lower intestinal tract, respectively, are to some extent different.

The method according to the invention for urogenital applications can be converted without major changes to oral application. In fact, the optimal pH of an oral formulation containing probiotics is that of yoghurt or curd, which is in the region of about 4.5 (reference 19), the same pH found in a healthy vagina.

For this reason, the considerations set out above and the experiments carried out accordingly retain their effectiveness when considering suitable oral formulations containing probiotics. As indicated above, the main difference consists of the choice of prebiotic fiber, since it is preferred for the intestine to comprise at least two fibers.

As used hereinafter, the term "composition" refers to compositions for intravaginal administration as defined in the appended claims as well as formulations for oral use.

The composition according to the invention may contain the following biochemical components:

a. a biochemical buffer to maintain the pH of the composition (in vitro and once dissolved in vivo) within the desired range of 4 to 5, preferably 4.5;

b. an anti-pathogenic preservative having biochemical properties fully compatible with the bacterial composition of the intestinal lactic acid microbiota;

c. a bioavailable, otherwise chemically pure, nitrogen source that supports adequate growth and proliferation of lactobacillus species (already present or administered);

d. selecting a biologically, chemically defined carbon source for selectively enhancing the growth of lactobacillus species and fermenting in lactic acid and SCFA; preferably in combination with

e. A bioavailable, otherwise chemically pure, source of sulfur that supports adequate growth and proliferation of lactobacillus species (already present or administered);

f. a component for modulating the redox potential in vaginal fluids, which may be included in the composition in the presence or absence of a source of sulfur,

g. selected minerals, and/or vitamins, oxidants/antioxidants and other components which may be associated with growth or pathogen inhibition of selected lactobacillus species, and

h. as living cells or lyophilised products thereof, cell-free culture supernatants thereof or intermittently sterilised products thereofProbiotic Lactobacillus strains selected for their specific properties, including but not limited to antibiotic-like activity against urogenital and/or intestinal pathogens, adhesion to intestinal and vaginal/urethral epithelial cells, H₂O₂Lactic acid and related bacteriocins and enzyme production.

In the context of the present invention, the terms "chemically pure" and "chemically defined" indicate synthetic media or substances in which the exact chemical composition is known (cf. textbook "Basic microbiology" 8 th edition, W. A: Volk, J.C, Brown, 1997, pages 39-40).

A complex (undefined) medium is one in which the exact chemical composition of the medium is unknown. It is established that the culture medium is usually composed of pure biochemicals ready for use; complex media usually contain complex materials of biological origin, such as blood or milk or yeast extract or beef extract, the exact chemical composition of which is clearly uncertain. The defined medium is a minimal medium. The formulation itself may be a suitable galenic formulation in a pharmaceutically acceptable carrier or delivery system. The actual composition is in liquid (yoghurt drink), semi-solid (yoghurt, cheese) or solid form, the latter including tablets, capsules, sachets and sticks.

The following are preferred embodiments of the present invention.

a. Buffering agent

The purpose of such ingredients is to help maintain the pH of the vaginal fluid after application in the physiological range of pH4 to 5. Acids and their salts having such desired properties in solution are, for example, organic acids such as aliphatic acids: acetic acid, propionic acid, butyric acid, etc. (a.s.o.), dicarboxylic acids such as oxalic acid, malonic acid, succinic acid, etc., hydroxy acids such as lactic acid, citric acid, dicarboxylic acids such as malic acid, etc. In addition, other acids found in human organisms are also contemplated for this use, such as phosphoric acid.

b. Anti-pathogen preservatives compatible with the Doderlein microbiota

As the name itself indicates, the specificity of lactic acid bacteria metabolism is the production of lactic acid. This is the main reason why lactic acid fermentation is used in traditional and industrial food preparation to kill potential pathogens present in the original nutrients and to preserve them at the end of the fermentation process by an effective amount of lactic acid (e.g. sauerkraut). Since lactobacillus species, especially vaginal species, are well-tolerated in the acidic environment containing lactic acid, the latter compound is in fact a Doderlein compatible preservative. Lactic acid, and individually selected salts thereof (calcium, magnesium, potassium, sodium, ammonium, etc.) are included in the composition to achieve such effects.

c. Biochemically utilizable nitrogen source for supporting growth of lactobacillus species

Much attention is now given to the properties and uses of so-called prebiotics, polysaccharides specifically metabolized by probiotics such as lactobacilli and bifidobacteria (see next). Prebiotics are widely used in combination with probiotics to produce so-called symbiotics (symbiotics) which, when taken orally, ensure better proliferation of the probiotic bacteria in the GI tract. However, in the vaginal environment, the availability of suitable nutrients to probiotic bacteria already present or administered exogenously is more limited, particularly with respect to the necessary nitrogen source (N source). The addition of a suitable organic and/or inorganic, chemically defined and pure source of N to a pharmaceutically suitable carrier appears to be an essential factor allowing the growth and proliferation of lactobacillus species in the vagina.

Such chemically defined N sources are, for example, ammonium salts (e.g. ammonium chloride, reference 15), urea, amino acids such as glutamic acid and the like (arginine etc.) capable of enhancing the growth of lactobacilli and bifidobacteria.

d. Biological, chemically defined carbon source for supporting the growth of lactobacilli

Obviously, a carbon source (C source) is also necessary for the growth of Lactobacillus vaginalis. Preferably, one such source is a nutrient specific for lactobacillus species rather than opportunistic or pathogenic bacteria. Examples thereof are non-digestible carbohydrates such as: fructooligosaccharides, galactooligosaccharides, glucooligosaccharides (glyco-oligosaccharides), and others. The presence of these so-called prebiotics confers a further differential competitive advantage over the growth of lactobacillus species in the vagina and gut with respect to the presence of pathogens or opportunistic bacteria.

Optionally, other components are present in the composition according to the invention, such as sulphur sources, minerals, redox (redox) regulators and vitamins.

Preferably, the composition according to the invention comprises a bioavailable, otherwise chemically pure source of sulphur that supports sufficient growth and proliferation of lactobacillus species (already present or administered), which may be, but is not limited to, organic/inorganic compounds such as sulphates, sulphites, sulphides, thiosulphates, sulphur-containing amino acids such as cysteine and methionine, cysteine (the oxidized dimer of cysteine), and the like.

Basic, chemically defined media for the growth of different lactobacillus species have been discovered and published (reference 18 a). In addition to the components listed above, in cases where vaginal nutrient availability is poor, it may be helpful to consider also adding the following to the preferred formulation: metal ions: for example Mg²⁺And Mn²⁺Antioxidant, antioxidant: for example, vitamin C, N-acetylcysteine, thiosulfate, vitamin: such as riboflavin and niacin; alone or in combination with one another.

Preferred ingredients are also probiotics as live cells, lyophilised products, tyndallized products, dried or liquid supernatants of species and strains with demonstrated activity against infectious or inflammatory conditions of vaginal and preferably human urogenital origin. Such species can be found and isolated in the Doderlein microbiota of healthy young women and are represented worldwide by Lactobacillus gasseri, Lactobacillus jensenii, Lactobacillus crispatus, Lactobacillus acidophilus, Lactobacillus fermentum, Lactobacillus plantarum and Lactobacillus inerticus.

Preferred ingredients are also compounds with activity against pathogens (bacteria, viruses, fungi, etc.), which for the purpose and understanding of the present invention are those that are generally capable of inhibiting the growth of pathogens without interfering with the metabolism and growth of probiotic bacteria. Examples of such suitable antifungal compounds of organic or inorganic origin are cysteine, sorbate or thiosulfate, respectively.

Preferably, the composition according to the invention comprises a bioavailable redox mediator, i.e. a component that modulates the redox potential in a biological fluid. In the context of the present invention, the term "redox-modulator" indicates a substance or at least a pair of substances capable of reducing or enhancing the redox potential of a certain biological medium. A method for determining vaginal redox potential in bacterial vaginosis is indicated in reference 22.

The redox modulators may be included in the compositions of the present invention in the presence or absence of a sulfur source as defined above.

When redox modulators are present, the compositions according to the invention are particularly suitable for the treatment or prevention of infections due to obligate anaerobes, for example Bacterial Vaginosis (BV).

Preferably, the redox-modulator is selected from:

a sulfur-containing redox system, such as sodium thiosulfate/bisulfite and glutathione in reduced (GSH) and oxidized (GSSG),

oxygen-based redox systems, such as quinone/hydroquinone,

nitrogen-based redox systems, such as nitrite/nitrate,

selenium-based redox systems (selenite/selenate and selenide). For example, the redox potential may be enhanced so as to become positive: (>150 mV, preferably 200 mV) and inhibiting anaerobic bacteria such as Gardnerella (G.sub.), (B.sub.))Gardnerella）。

Preferably, in the context of the present invention, at least one strain of lactobacillus selected from lactobacillus gasseri, lactobacillus jensenii, lactobacillus crispatus, lactobacillus acidophilus, lactobacillus helveticus: (a)L. helveticus) Lactobacillus plantarum, Lactobacillus fermentum and Lactobacillus johnsonii, more preferably said strain is selected from the group having accession number CNCM I-3482, deposited at 22.7.2005 by Medinova AG in the French national Collection of microorganisms cultures (Collection national a)Lactobacillus gasseri KS114.1 of le de Cultures de Microorganises) (CNCM), Lactobacillus crispatus 116.1 with accession number CNCM I-3483, deposited by Medinova AG at 22.7.2005 in the center for French national collections of microorganisms (CNCM), Lactobacillus jensenii KS119.1 with accession number CNCM I-3217, deposited by Medinova AG at 4.6.2004 in the center for French national collections of microorganisms (CNCM), Lactobacillus crispatus 119.4 with accession number CNCM I-3484, deposited by Medinova AG at 22.7.2005 in the center for French collections of microorganisms (CNCM), Lactobacillus crispatus 120.1 with accession number CNI-3218, deposited by Medinova AG at 2004 in the center for French collections of microorganisms (CNCM) 120.1, deposited by Medinova AG 9.3216.9 in the center for 2004 in French collections of microorganisms (CNCM) and cultured by Medinova AG 121.121, lactobacillus gasseri 123.1 with accession number CNCM I-3485, deposited at french national collection of microbiological cultures (CNCM) by Medinova AG at 22.7.2005, lactobacillus gasseri 126.2, lactobacillus crispatus 127.1 with accession number CNCM I-3486, deposited at french national collection of microbiological cultures (CNCM) by Medinova AG at 22.7.2005, and lactobacillus acidophilus KS 400, most preferably the lactobacillus strains are selected from lactobacillus jensenii KS119.1 CNCM I-3217, lactobacillus crispatus 119.4 CNCM I-3484, lactobacillus gasseri 120.1 CNCM I-3218 and lactobacillus gasseri 124.3CNCM I-3220.

The above bacterial strains have been deposited under the conditions of the Budapest treaty and are publicly available.

Preferably, in the context of the present invention, the buffer a) is selected from acids or salts thereof comprising or alternatively consisting of: organic acids such as aliphatic acids including acetic acid, propionic acid, butyric acid, dicarboxylic acids including oxalic acid, malonic acid, succinic acid, hydroxy acids such as lactic acid, citric acid, dicarboxylic acids such as malic acid.

Preferably, in the context of the present invention, the preservative b) is lactic acid or a derivative and/or salt thereof.

Preferably, in the context of the present invention, the nitrogen source c) is an ammonium salt, urea, a naturally occurring or synthetic amino acid such as glutamic acid and mixtures thereof.

Preferably, in the context of the present invention, the carbon source d) is a non-digestible carbohydrate, such as: fructooligosaccharides, galactooligosaccharides, glucooligosaccharides and mixtures thereof. Preferably, the composition according to the invention comprises the following ingredients, for example, based on a solution/suspension of 1 ml of ingredients. The actual composition of e.g. a tablet or capsule is obtained by multiplying the number of ingredients by 3:

。

the present invention includes the following embodiment (E).

E1. A composition for use in treating or preventing urogenital infections caused by pathogenic microorganisms in a female subject, wherein the composition comprises an effective amount of each of:

a) a buffering agent, a water-soluble polymer,

b) a preservative that is fully compatible with the vaginal microbiota,

c) pure molecular or ionic biochemical nitrogen sources available to the vaginal microbiota,

d) a prebiotic polysaccharide that is fermentable by the vaginal microbiota, and optionally,

e) an antioxidant component in a mixture of at least one antioxidant component,

wherein the treatment or prevention comprises intravaginal administration of the composition to a subject for stabilizing the acidity of vaginal fluid in the subject at a physiological value of about pH4.5 or less.

E2. The composition for use according to E1, wherein the vaginal formulation is a lubricating gel for personal use.

E3. Composition for use according to E1 or E2, wherein the antioxidant is selected from sulphur containing compounds, such as N-acetylcysteine, cysteine, methionine, sodium thiosulphate.

E4. Composition for use according to any one of E1-E3, wherein one or more lactobacillus strains selected from the group consisting of: lactobacillus gasseri, lactobacillus jensenii, lactobacillus crispatus, lactobacillus acidophilus, lactobacillus helveticus, lactobacillus plantarum, lactobacillus fermentum, lactobacillus lactis, lactobacillus johnsonii, and lactobacillus acidophilus KS 400.

E5. Composition for use according to any one of E1-E4, wherein one or more lactobacillus strains selected from the group consisting of: lactobacillus gasseri, lactobacillus jensenii, lactobacillus crispatus, lactobacillus acidophilus, lactobacillus helveticus, lactobacillus plantarum, lactobacillus fermentum, lactobacillus lactis, and lactobacillus johnsonii.

E6. A composition for use according to any one of E1-E5, the lactobacillus strain is selected from lactobacillus jensenii KS109, having accession number CNCM I-3482, lactobacillus gasseri KS114.1 deposited at the french national collection of microbial cultures by Medinova AG at 7/22.2005, lactobacillus jensenii KS116.1 deposited at the french national collection of microbial cultures (CNCM) by Medinova AG at 22.7/2005, lactobacillus jensenii KS119.1 having accession number CNCM I-3217, lactobacillus jensenii KS119.1 deposited at the french national collection of microbial cultures (CNCM) by Medinova AG at 4.6/2004, lactobacillus jensenii KS 119.84 having accession number cni-3484, deposited at the french collection of national collection of microbial cultures (CNCM) by Medinova AG at 22.7/2005, and lactobacillus crispus 3218-3218, Lactobacillus gasseri 120.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus jensenii KS 121.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus jensenii KS122.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus gasseri 123.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 22.7.2005, Lactobacillus gasseri 123.1 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus gasseri 124.3 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 127.2, Lactobacillus gasseri 126.2 deposited by Medinova AG on French national center for microbiological culture Collection (CNCM) at 7.22.7.9.86, lactobacillus jensenii KS 130.1, having accession number CNCM I-3360, lactobacillus helveticus KS 300 deposited by Medinova AG at french national collection of microbiological cultures (CNCM) at 10/2/2005, and lactobacillus acidophilus KS 400.

E7. Composition for use according to any one of E1-E6, wherein the lactobacillus strain is selected from lactobacillus jensenii KS119.1 with accession number CNCM I-3217, deposited at french national collection of microbial cultures (CNCM) by Medinova AG at 6.4.2004, lactobacillus crispatus 119.4 with accession number CNCM I-3484, lactobacillus gasseri 119.4 deposited at french national collection of microbial cultures (CNCM) by Medinova AG at 22.7.2005, lactobacillus gasseri 120.1 with accession number CNCM I-3218, deposited at french national collection of microbial cultures (CNCM) by Medinova AG at 4.6.2004, lactobacillus gasseri 124.3 with accession number CNCM I-3220, deposited at french collection of national collection of microbial cultures (CNCM) by Medinova AG at 4.6.2004.

E8. A composition for use according to any one of E1-E7, wherein the formulation further comprises an effective amount of a therapeutically effective antimicrobial agent capable of inhibiting or eradicating urogenital pathogens.

E9. A method for treating or preventing urogenital infections caused by pathogenic microorganisms in a female subject, comprising intravaginally administering to the subject an effective amount of each of:

a) a buffering agent, a water-soluble polymer,

b) a preservative that is fully compatible with the vaginal microbiota,

c) pure molecular or ionic biochemical nitrogen sources available to the vaginal microbiota,

d) a prebiotic polysaccharide that is fermentable by the vaginal microbiota, and optionally,

e) an antioxidant component.

E10. A method for stabilizing the acidity of vaginal fluid in a subject at a physiological value of about pH4.5 or less, comprising intravaginal administration to the subject of an effective amount of each of:

a) a buffering agent, a water-soluble polymer,

b) a preservative that is fully compatible with the vaginal microbiota,

c) pure molecular or ionic biochemical nitrogen sources available to the vaginal microbiota,

d) a prebiotic polysaccharide that is fermentable by the vaginal microbiota, and optionally,

e) an antioxidant component.

E11. The method according to claim 9 or 10, wherein the buffer a) is an acid or a salt thereof selected from the group comprising or alternatively consisting of: organic acids such as aliphatic acids including acetic acid, propionic acid, butyric acid, dicarboxylic acids including oxalic acid, malonic acid, succinic acid, hydroxy acids such as lactic acid, citric acid, dicarboxylic acids such as malic acid.

E12. The method according to any one of E9-E11, wherein the preservative b) is lactic acid or a derivative and/or salt thereof.

E13. The method according to any one of E9-E12, wherein the nitrogen source c) is an ammonium salt, urea, a naturally occurring or synthetic amino acid such as glutamic acid and mixtures thereof.

E14. The method according to any one of claims E9-E13, wherein the carbon source d) is a non-digestible carbohydrate, such as: fructooligosaccharides, galactooligosaccharides, glucooligosaccharides and mixtures thereof.

E15. A formulation having a pH in the range of 4 to 5 suitable for oral administration, wherein the composition comprises an effective amount of each of:

a) a buffering agent, a water-soluble polymer,

b) a preservative that is fully compatible with the vaginal microbiota,

c) pure molecular or ionic biochemical nitrogen sources available to the vaginal microbiota,

d) a prebiotic polysaccharide that is fermentable by the vaginal microbiota, and optionally,

e) an antioxidant component.

E16. The formulation of claim E15, wherein the antioxidant E) is selected from the group consisting of sulfur-containing compounds, such as N-acetylcysteine, cysteine, methionine, sodium thiosulfate.

E17. The formulation of claim E15 or E16, wherein at least one strain of lactobacillus is added to the formulation, said strain being selected from the group consisting of lactobacillus gasseri, lactobacillus jensenii, lactobacillus crispatus, lactobacillus acidophilus, lactobacillus helveticus, lactobacillus plantarum, lactobacillus fermentum, and lactobacillus johnsonii.

E18. E15 to E17, wherein at least one strain of Lactobacillus is added to the formulation, said strain being selected from the group consisting of Lactobacillus jensenii KS109, Lactobacillus gasseri KS114.1 having accession number CNCM I-3482, Lactobacillus gasseri KS114.1 deposited at the French national collection of microbial cultures by Medinova AG at 7.22.2005, Lactobacillus crispatus 116.1 having accession number CNCM I-3483, Lactobacillus jensenii KS119.1 deposited at the French national collection of microbial cultures (CNCM) by Medinova AG at 22.7.2005, Lactobacillus crispatus having accession number CNCM I-3217, Lactobacillus crispatus deposited at the French national collection of microbial cultures (CNCM) by Medinova AG at 4.6.2004, CNCM I-3484, Lactobacillus crispatus deposited at the French collection of microbial cultures (national collection of microorganisms) by Medinova AG at 22.7.2005, lactobacillus gasseri 120.1 having accession number CNCM I-3218, deposited by Medinova AG on the French national center for microbiological culture Collection (CNCM) at 6.4.2004, Lactobacillus jensenii KS 121.1 having accession number CNCM I-3219, deposited by Medinova AG on the French national center for microbiological culture Collection (CNCM) at 6.4.2004, Lactobacillus jensenii KS122.1, having accession number CNCM I-3485, Lactobacillus gasseri 123.1 deposited by Medinova AG on the French national center for microbiological culture Collection (CNCM) at 22.7.2005, Lactobacillus gasseri 124.3 having accession number CNCM I-3220, deposited by Medinova AG on the French national center for microbiological culture Collection (CNCM) at 4.6.2004, Lactobacillus gasseri 126.2, deposited by Medinova AG on the French national center for microbiological culture Collection (CNCM) at 127.127.86, lactobacillus jensenii KS 130.1, having accession number CNCM I-3360, lactobacillus helveticus KS 300 deposited by Medinova AG at french national collection of microbiological cultures (CNCM) at 10/2/2005, and lactobacillus acidophilus KS 400.

E19. Formulation according to any one of E15 to E18, wherein the lactobacillus strain is selected from lactobacillus jensenii KS119.1 with accession number CNCM I-3217, deposited at french national collection of microorganisms (CNCM) by Medinova AG at 6.4.2004, lactobacillus crispatus 119.4 with accession number CNCM I-3484, lactobacillus crispatus 119.4 deposited at french national collection of microorganisms (CNCM) by Medinova AG at 22.7.2005, lactobacillus gasseri 120.1 with accession number CNCM I-3218, deposited at french national collection of microorganisms (CNCM) by Medinova AG at 4.6.2004, lactobacillus gasseri 124.3 with accession number CNCM I-3220, deposited at french national collection of microorganisms (CNCM) by Medinova AG at 4.6.2004, and lactobacillus acidophilus deposit KS 400.

E20. A composition for use according to any one of E1-E8, or a formulation according to any one of E15-E19, wherein the buffer a) is selected from an acid or a salt thereof comprising or alternatively consisting of: organic acids such as aliphatic acids including acetic acid, propionic acid, butyric acid, dicarboxylic acids including oxalic acid, malonic acid, succinic acid, hydroxy acids such as lactic acid, citric acid, dicarboxylic acids such as malic acid.

E21. The composition for use according to any one of E1-E8 or E20, or the formulation according to any one of claims 15-20, wherein the preservative b) is lactic acid or a derivative and/or salt thereof.

E22. The composition for use according to any one of E1-E8 or E20-E21, or the formulation according to any one of claims 15-21, wherein the nitrogen source c) is an ammonium salt, urea, a naturally occurring or synthetic amino acid such as glutamic acid, and mixtures thereof.

E23. Composition for use according to any one of E1-E8 or E20-E22, or a formulation according to any one of E15-E22, wherein the carbon source d) is a non-digestible carbohydrate, such as: fructooligosaccharides, galactooligosaccharides, glucooligosaccharides and mixtures thereof.

E24. A composition for use according to any one of E1-E8 or E20-E23, or a formulation according to any one of claims 15-23, which has a pH of 43 to 4.6 if the solid is to a volume of 3ml after dissolution/suspension, and which comprises the following ingredients per single dose:

ca-lactic acid pentahydrate: 30-90, preferably 60 mg

Magnesium citrate dibasic: 15-45, preferably 30mg

Prebiotic fiber: 50-150, preferably 100 mg

Glutamic acid: 20-60, preferably 40 mg

Cysteine/thiosulfate: 10-50, preferably 30mg

Adding 400-1100 mg of excipient: proper amount of

E25. A composition for use or formulation according to E24, further comprising a lyophilized product of:

lactobacillus gasseri 120.1 having accession number CNCM I-3218, deposited by Medinova AG in French national Collection of cultures of microorganisms (CNCM) on 6/4/2004, and/or

Lactobacillus gasseri 124.3 and/or Lactobacillus crispatus 119.4 with accession number CNCM I-3484, deposited by Medinova AG at 22.7.2005 in the French national Collection of microorganisms (CNCM), and/or

Lactobacillus jensenii KS119.1, deposited by Medinova AG at french national center for microbiological culture collection (CNCM) at 6/4/2004 with accession number CNCM I-3217, amounting to 50 to 200 mg for a total of 20 to 100 hundred million c.f.u.

Example (b):