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US20150139969A1 - Use of thiosulfate to potentiate the anti-pathogenic effect of lactobacilli - Google Patents

Use of thiosulfate to potentiate the anti-pathogenic effect of lactobacilli Download PDF

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US20150139969A1
US20150139969A1 US14/413,660 US201314413660A US2015139969A1 US 20150139969 A1 US20150139969 A1 US 20150139969A1 US 201314413660 A US201314413660 A US 201314413660A US 2015139969 A1 US2015139969 A1 US 2015139969A1
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thiosulfate
lactobacilli
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lactobacillus
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Adrien Nivoliez
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K33/00—Medicinal preparations containing inorganic active ingredients
    • A61K33/04—Sulfur, selenium or tellurium; Compounds thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/66—Microorganisms or materials therefrom
    • A61K35/74—Bacteria
    • A61K35/741—Probiotics
    • A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
    • A61K35/747—Lactobacilli, e.g. L. acidophilus or L. brevis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/02—Inorganic compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0034—Urogenital system, e.g. vagina, uterus, cervix, penis, scrotum, urethra, bladder; Personal lubricants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0053—Mouth and digestive tract, i.e. intraoral and peroral administration
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/141—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers
    • A61K9/143—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers with inorganic compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/19—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles lyophilised, i.e. freeze-dried, solutions or dispersions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00—Drugs for disorders of the urinary system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00—Drugs for disorders of the urinary system
    • A61P13/02—Drugs for disorders of the urinary system of urine or of the urinary tract, e.g. urine acidifiers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P15/00—Drugs for genital or sexual disorders; Contraceptives
    • A61P15/02—Drugs for genital or sexual disorders; Contraceptives for disorders of the vagina
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/02—Local antiseptics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/10—Antimycotics

Definitions

  • the present invention relates to compositions including lactobacilli in combination with thiosulfate, the thiosulfate potentiating the anti-pathogenic effect of said lactobacilli.
  • the urogenital flora comprises nearly 50 different species of microorganisms.
  • 95% of the population is composed of various strains of lactobacilli, also called “Doderlein's bacillus ”.
  • lactobacilli help protect against pathogens by various mechanisms, including production of hydrogen peroxide, lactic acid and bacteriocins, inhibition of adhesion and spreading of pathogens.
  • these lactobacilli maintain an acidic pH by producing lactic acid from the glycogen present in the vaginal mucus.
  • the normal vaginal flora is thus principally composed of lactobacilli forming a protective biofilm on the surface of the mucous membrane.
  • lactobacilli most commonly identified in the vagina are notably Lactobacillus crispatus, Lactobacillus jensenii, Lactobacillus vaginalis, Lactobacillus inners and Lactobacillus gasseri.
  • Vulvovaginal candidiasis affects 70-75% of women at least once during their reproductive years, and about 40-50% will have a second episode.
  • the incidence of recurrent vulvovaginal candidiasis (defined as at least 4 episodes per year, including 2 confirmed by mycological examination) has been estimated at 5-8%.
  • This benign ailment has a very negative impact on patients' quality of life and generates significant healthcare expenditures.
  • Such pathology is difficult to treat because of the multifactorial pathogenesis of this affection.
  • the traditional maintenance therapy using an oral or vaginal antifungal must last at least 6 months, but the relapse rate remains high with 60-70% of women having a recurrence in the two months following discontinuation of the treatment.
  • antifungals have frequent side effects and their long-term use can promote an occurrence of bacterial vaginosis.
  • Bacterial vaginosis is due to a qualitative and quantitative imbalance of the normal vaginal flora which can lead to the virtually complete disappearance of lactobacilli, to the benefit of anaerobic flora, and also to the emergence of bacteria such as Gardnerella vaginalis and Atopobium vaginae .
  • Bacterial vaginosis is one of the most frequent vaginal infections with a frequency rate of 10 to 15. This benign pathology in women can be serious during pregnancy as it can cause premature deliveries, low birth-weights and spontaneous abortions.
  • Bacterial vaginosis and other imbalances of the vaginal microflora are commonly treated by antibiotic therapy. This treatment has the typical disadvantages of antibiotic treatments and is proving to be less and less effective. Furthermore, it aims to eliminate the pathogenic flora but also destroys the normal beneficial flora.
  • the preferred lactobacilli are Lactobacillus rhamnosus, Lactobacillus crispatus and Lactobacillus vaginalis .
  • lactobacilli can be administered in lyophilized form or in solution, and optionally in combination with other active agents. Indeed, although lactobacilli have a real and recognized anti-pathogenic effect, it is often insufficient for them to be able to fight an infection. Additional active agents, or agents that potentiate the anti-pathogenic effect of lactobacilli, are actively sought by the scientific community.
  • Sodium thiosulfate is used as an excipient in numerous pharmaceutical and veterinary compositions containing lactobacilli, such as the compositions TROPHIGIL, PROBIOS® and ALLBIO®. Furthermore, the patent application CN 20091145852 describes a drug composed of extracts of Chinese plants and sodium thiosulfate. This drug is proposed notably for the treatment of gonorrhea. In these compositions, sodium thiosulfate is used as an excipient, and not as an active agent.
  • the anti-pathogenic effect of lactobacilli genuinely exists but is not sufficiently strong in most cases to fight infection (see FIG. 2A ), and it is thus often necessary to use in addition other active ingredients such as antibiotics or antifungals.
  • the present invention proposes a means of potentiating the anti-pathogenic effect of lactobacilli.
  • the inventors have indeed discovered that the combination of thiosulfate, notably sodium thiosulfate, and lactobacilli makes it possible to potentiate the anti-pathogenic effect of these lactobacilli, when the thiosulfate is present at a concentration of at least 100 mg/gram of powder, for a powder including from 10 7 to 10 10 CFU/g of lactobacilli.
  • the invention also relates to a pharmaceutical composition including at least 100 mg/g of sodium thiosulfate in combination with Lactobacillus crispatus.
  • FIG. 1 Influence of sodium thiosulfate concentration in the formulation of the strain L. crispatus BLL2005 on inhibition of pathogenic C. albicans .
  • FIG. 2 Compositions including various lactobacilli strains without ( FIG. 2A ) or with ( FIG. 2B ) sodium thiosulfate, for the inhibition of pathogenic C. albicans .
  • FIG. 3 Composition including the strain L. crispatus BLL2005 without ( FIG. 3A ) or with ( FIG. 3B ) sodium thiosulfate, for the inhibition of various pathogens.
  • the results are expressed as the decimal logarithmic (log 10 ) reduction of the number of CFU between the count at T0 and at the various sampling points (T4h, T24h and T28h).
  • FIG. 4 Effect of sodium thiosulfate alone on the growth of pathogenic C. albicans.
  • FIG. 5 Effect of ‘Formulation A’, with or without the presence of L. crispatus , on the growth of pathogenic C. albicans.
  • FIG. 6 Co-culture of the strain Lcr35® ( L. rhamnosus ) formulated with 150 mg of thiosulfate (after lyophilization in the final product) and Candida albicans.
  • the present invention relates to the use of thiosulfate to potentiate the anti-pathogenic effect of Lactobacillus bacteria, the thiosulfate being in a quantity of at least 100 mg/gram of powder, for a powder including from 10 7 to 10 10 CFU/g of lactobacilli.
  • potentiate means to give a greater effectiveness, to increase the beneficial effects of an active ingredient.
  • the potentiating agent here, thiosulfate
  • the potentiating agent does not itself act against pathogens, but acts via lactobacilli whose anti-pathogenic effect it potentiates.
  • anti-pathogenic effect refers to the preventive and curative action of lactobacilli with respect to the various pathogens whose growth can be inhibited by the presence of lactobacilli. This effect is expressed via various mechanisms: lowering of pH, production of hydrogen peroxide (H 2 O 2 ), production of lactic acid and/or acetic acid, production and secretion of bacteriocins, peptides, organic acids and short-chain fatty acids, and inhibition of the adhesion and expansion of pathogens. In particular, this anti-pathogenic effect is measured as the rate of inhibition of pathogen growth, and as the rate of production of bacteriocins and peptide compounds having antimicrobial properties.
  • pathogens refers to all pathogens likely to settle in the urogenital tract, and in particular Gardnerella vaginalis, Prevotella bivia, Neisseria gonorrhoeae, Mycoplasma, Mobiluncus, Candida albicans and Candida glabrata , as well as to pathogens likely to cause enteric infections, such as, for example, Enterobacteriaceae, including Escherichia coli, Salmonella, Staphylococcus, Clostridium difficile and Shigella.
  • lactobacilli refers to all bacteria of the genus Lactobacillus , immotile, facultatively anaerobic, Gram-positive bacteria of variable shapes and sizes. Most lactobacilli convert lactose and other simple sugars into lactic acid. Lactobacilli colonize the vagina and gastrointestinal tract and are an important component of the endogenous intestinal and vaginal flora.
  • the present invention principally relates to lactobacilli constituting the vaginal flora; it is understood, however, that the invention can be carried out with other types of lactobacilli such as those present in the gastrointestinal tract (in particular Lactobacillus acidophilus, Lactobacillus reuteri, Lactobacillus salivarius, Lactobacillus gasseri, Lactobacillus casei and Lactobacillus rhamnosus ).
  • colony forming unit refers to the unit of measure generally recognized by persons skilled in the art for quantifying bacteria capable of founding a colony.
  • thiosulfate refers to thiosulfate ions (S 2 O 3 2 ⁇ ). This is the basic form of thiosulfuric acid (H 2 S 2 O 3 ), unstable in an aqueous medium.
  • the pathogens whose growth is inhibited by the combination of thiosulfate and lactobacilli belong to the family of Candida albicans, Prevotella bivia, Gardnerella vaginalis, Mycoplasma or Mobiluncus.
  • Candida albicans is the most important and well-known yeast species of the genus Candida . It causes fungal infections (candidiasis or candidosis) mainly of the mucous membranes of the digestive tract and female reproductive system.
  • Prevotella bivia is an anaerobic bacterium, which mainly causes vaginosis and urinary tract infections.
  • Gardnerella vaginalis is a bacterium with a pleomorphic rod or coccobacillus shape. It is frequently found in cases of vaginosis (nonspecific vaginitis), either as the only pathogenic bacterium or in combination with other bacteria. It produces a perforating toxin that only affects human cells. It can also be encountered in the blood, urine and pharynx.
  • Mycoplasma is a family of more than 100 species of bacteria that are insensitive to families of antibiotics such as penicillin or beta-lactams.
  • Mycoplasma genitalium is responsible for genital infections (urethritis, cervicitis, vaginitis, salpingitis) and sterility problems.
  • Mobiluncus is an anaerobic Gram-positive bacterium often found associated with Gardnerella vaginalis in bacterial vaginosis.
  • vaginosis refers to an imbalance of the vagina's microbial flora. It is characterized by the disappearance of lactobacilli and the multiplication of anaerobic bacteria such as Gardnerella vaginalis.
  • Candida albicans the most common species, is part of the usual flora of the oropharynx and digestive track and can also be present in a small quantity in the normal vaginal flora.
  • Vulvovaginal candidiasis is quite common and can be prone to several recurrences.
  • urinary infection refers to colonization of the urine by bacteria, most often expressed as symptoms of urinary infection.
  • the bacteria most commonly concerned are Escherichia coli (75% of cases), Proteus mirabilis and Klebsiella , all three Enterobacteriaceae (Gram-negative bacilli).
  • vaginosis nor candidiasis nor urinary tract infections can be regarded as sexually-transmitted infections such as gonorrhea.
  • the thiosulfate is selected from sodium thiosulfate or potassium thiosulfate.
  • sodium thiosulfate is used.
  • Sodium thiosulfate consists of sodium ions and thiosulfate ions.
  • the quantity used is at least 100 mg/g of powder, preferentially at least 150 mg, and more preferentially at least 200 mg of thiosulfate per gram of powder, this powder including between 10 7 and 10 10 lactobacilli bacteria (CFU).
  • the quantity of sodium thiosulfate in the final product will be approximately 230 mg/gram of powder including 10 7 to 10 10 CFU of lactobacilli.
  • the thiosulfate is used in a quantity of about 230 mg/gram of powder, for a powder including between 10 8 and 10 10 CFU of lactobacilli per gram, and more preferentially between 10 9 and 10 10 CFU/gram.
  • a powder including between 10 8 and 10 10 CFU of lactobacilli per gram, and more preferentially between 10 9 and 10 10 CFU/gram.
  • This final quantity of sodium thiosulfate is obtained by adding 113 g/liter of sodium thiosulfate to the bacterial culture medium, before lyophilization.
  • the examples show, however, that a simple addition of 1 g/liter of thiosulfate in the bacterial culture medium makes it possible to potentiate the effects of lactobacilli in culture.
  • the lactobacilli are Lactobacillus rhamnosus and/or Lactobacillus crispatus .
  • the preferred strains are notably the strain Lcr35® and the strain BLL2005.
  • the invention also relates to a pharmaceutical composition including at least 100 mg of thiosulfate, preferentially at least 150 mg, and more preferentially at least 200 mg of thiosulfate per gram of powder, in combination with a Lactobacillus crispatus strain.
  • the quantity of lactobacilli per gram of composition will be between 10 7 and 10 10 CFU, more preferentially between 10 8 and 10 9 CFU/g.
  • the Lactobacillus crispatus strain is in lyophilized form.
  • the strain can be the only lyophilized element of the composition, but preferably the strain is lyophilized in a medium including additional constituents, which will be added before or after the lyophilization step.
  • the sodium thiosulfate can be added before lyophilization, notably in a quantity of 113 g/liter of bacterial culture medium.
  • the sodium thiosulfate can also be added in a quantity of 1 g/liter, 5 g/liter, 10 g/liter, 20 g/liter, 30 g/liter, 50 g/liter or 100 g/liter of bacterial culture medium.
  • the pharmaceutical composition further includes a preservation matrix and/or excipients well-known to persons skilled in the art, and optionally other active ingredients having a complementary action.
  • this composition can include the following active ingredients: hormones (estriol, progesterone, etc.), anti-inflammatory agents and/or bactericidal agents and/or antifungal agents.
  • hormones estriol, progesterone, etc.
  • anti-inflammatory agents and/or bactericidal agents and/or antifungal agents.
  • This composition also includes, according to a specific aspect of the invention, several lactobacilli strains.
  • the invention also relates to a pharmaceutical composition including the culture supernatant of a Lactobacillus strain, preferentially Lactobacillus crispatus , cultivated in the presence of thiosulfate, in particular sodium thiosulfate, in particular at least 100 g/liter of sodium thiosulfate in the culture medium.
  • This culture supernatant contains thiosulfate but does not contain lactobacilli. It has a strong anti-pathogenic activity.
  • the presence of thiosulfate potentiates the secretion of bacteriocins, peptide compounds having antimicrobial properties. It is thus likely that the “culture supernatant” no longer containing lactobacilli contains large amounts of bacteriocins, secreted beforehand by the lactobacilli.
  • compositions are formulated for vaginal administration or oral administration.
  • dosage forms used will be capsules, tablets, creams, liquid or oily suspensions, or any other suitable medical device.
  • the invention also relates to a pharmaceutical composition including at least 100 mg, preferentially at least 150 mg, and more preferentially at least 200 mg of thiosulfate per gram of powder, in combination with a Lactobacillus crispatus strain, for its use in the treatment of urogenital infections such as vaginosis, candidosis and urinary tract infections.
  • this composition will include about 230 mg of sodium thiosulfate in combination with 10 8 to 10 9 CFU/g of a Lactobacillus crispatus strain.
  • the optimal modes of administration, dosing schedules and dosage forms of the compounds and compositions according to the invention can be determined according to the criteria generally taken into account in the establishment of a pharmaceutical treatment suited to a patient, such as, for example, the patient's age or weight, the gravity of the patient's general state, the tolerance for the treatment and the side effects observed.
  • Urogenital infections will preferably be treated by the administration of one or two 350 mg capsules per day.
  • the present invention also relates to a pharmaceutical composition including at least 100 mg, preferentially at least 150 mg, and more preferentially at least 200 mg of thiosulfate per gram of powder, in combination with 10 7 to 10 10 CFU of a Lactobacillus strain, for its use in the treatment of urogenital infections such as vaginosis, candidosis and urinary tract infections, by potentiation of the anti-pathogenic effect of lactobacilli.
  • the invention also relates to a novel use of thiosulfate, and in particular sodium thiosulfate, for the treatment of urogenital infections such as vaginosis, candidiasis and urinary tract infections, by potentiation of the anti-pathogenic effect of lactobacilli.
  • the thiosulfate can be used in combination with lactobacilli, as described above, or administered alone for the purpose of potentiating the anti-pathogenic effect of lactobacilli present endogenously in the vaginal cavity.
  • Formulation A includes the following constituents:
  • Culture medium 116 g/l milk, 15 g/l dextrose, 10 g/l autolyzed yeast, 2 ml/l Tween
  • Lyophilization medium 110 g/l milk, 101.5 g/l FOS, 9.5 ml/I glutamate, 5.25 g/l ascorbic acid, 113 g/l sodium thiosulfate
  • MRS (titer near 1 ⁇ 10 8 CFU/ml) is placed with 5 ml of non-inoculated Sabouraud broth.
  • results are presented in FIG. 1 and in Table 2 below.
  • the results are expressed as the decimal logarithmic (log 10 ) reduction of the number of CFU between the count at T0 and at the various sampling points (T4h, T24h and T28h).
  • Candida albicans vs E289-2005 (0 g/l) 0 1 2
  • Candida albicans vs E290-2005 (10 g/l) 0 3 3
  • Candida albicans vs E291-2005 25 g/l) 0 3 5
  • Candida albicans vs E292-2005 50 g/l) 0 3 3
  • Candida albicans vs E293-2005 75 g/l) 0 3 7
  • Complete inhibition Candida albicans vs E115-2005 0 6 7 (113 g/l) Complete inhibition
  • counts of the strain BLL2005 remained constant during co-incubation.
  • the protocol is equivalent to that described in Example 1.
  • the count of the lactobacilli strains remains constant during co-incubation.
  • the pathogen and the probiotic are counted at T0, T4h, T24h and T28h.
  • the count of the strain BLL2005 remains constant during co-incubation.
  • results are presented in FIG. 3 .
  • the results are expressed as the decimal logarithmic (log 10 ) reduction of the number of CFU between the count at T0 and at the various sampling points (T4h, T24h and T28h).
  • Formulation A of the strain L. crispatus BLL2005 makes it possible to obtain after 28 hours of co-incubation an inhibition greater than or equal to 3 log against all Candida strains tested.
  • the strain BLL2005 is cultivated in the following culture medium:
  • fermentation is maintained at a temperature of 37° C. by a suitable thermostatically controlled device for 48 to 72 hours.
  • the culture having reached a count greater than or equal to 10 8 CFU/ml, is poured into a mixing tank.
  • the culture is shaken rapidly to break the curd and the following lyophilization adjuvants are added: 110 g/l milk, 101.5 g/l FOS, 9.5 ml/I sodium glutamate, 5.25 g/l ascorbic acid, 113 g/l sodium thiosulfate.
  • Lyophilization is then carried out according to the standard conditions well-known to persons skilled in the art: the bacteria are distributed aseptically on stainless steel trays, frozen quickly at ⁇ 40° C., and then subjected to a sublimation operation at ⁇ 22° C. Incubation is then carried out at 37° C. The product is then crushed and filtered under controlled atmosphere.
  • the powder obtained can advantageously be mixed with 1% of magnesium stearate in order to facilitate the filling of capsules.
  • the powder thus obtained is distributed in “0” size capsules in an amount of 350 mg of powder per capsule.
  • Candida strain was tested: PBp0001— Candida albicans ATCC 10231.
  • results are expressed in decimal logarithmic (log 10 ) reduction of the number of CFU between the count at T0 and at the various sampling points (T24h, T48h and T72h).
  • test culture medium is a homogeneous mixture of the pathogen's culture medium and MRS medium containing sodium thiosulfate.
  • Candida control 5 ml of inoculated Sabouraud broth is placed with the pathogenic strain (titer near 1.10 8 CFU/ml) and 5 ml of non-inoculated MRS broth. Samples are taken and titers determined at T0, T4h, T24h and T28h.
  • FIG. 5 shows several culture types:
  • the strain L. rhamnosus Lcr35® is able to inhibit the growth of pathogens. Moreover, the strain formulated with thiosulfate is more effective than the strain alone. In order to understand the mechanisms that differ between these two conditions, genes involved in bacteriocin production were sought. In Lcr35®, 7 genes coding for bacteriocins were found.

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CN113564075A (zh) * 2015-06-29 2021-10-29 辉凌制药公司 用于治疗例如细菌性阴道病的鼠李糖乳杆菌细菌
WO2018100533A1 (fr) * 2016-11-30 2018-06-07 Probioswiss Ag Formulation de dispositif médical urogénital à base de compositions biochimiques appropriées pour la stabilisation de l'acidité et de l'état redox des sécrétions vaginales
CN110799198A (zh) * 2016-11-30 2020-02-14 普罗生物瑞士股份公司 用于稳定阴道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
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CA2877666C (fr) 2020-07-14
ES2604880T3 (es) 2017-03-09
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US11202799B2 (en) 2021-12-21
CN112999240A (zh) 2021-06-22
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EP2869825B1 (fr) 2016-08-24
BR112014033013A2 (pt) 2017-06-27
US20220062330A1 (en) 2022-03-03
CN104519892A (zh) 2015-04-15
WO2014009349A1 (fr) 2014-01-16
PL2869825T3 (pl) 2017-04-28
FR2992861B1 (fr) 2014-10-17
US12390489B2 (en) 2025-08-19
MX360411B (es) 2018-10-30
AU2013289241A1 (en) 2015-01-29
RU2667122C2 (ru) 2018-09-14
TW201414481A (zh) 2014-04-16
FR2992861A1 (fr) 2014-01-10

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