WO1988002600A1 - Composition bactericide contenant une enzyme et preparation pour le traitement d'infections notamment dentaires comprenant ladite composition - Google Patents

Composition bactericide contenant une enzyme et preparation pour le traitement d'infections notamment dentaires comprenant ladite composition Download PDF

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Publication number
WO1988002600A1
WO1988002600A1 PCT/DK1987/000130 DK8700130W WO8802600A1 WO 1988002600 A1 WO1988002600 A1 WO 1988002600A1 DK 8700130 W DK8700130 W DK 8700130W WO 8802600 A1 WO8802600 A1 WO 8802600A1
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WO
WIPO (PCT)
Prior art keywords
composition
enzyme
wound treatment
peroxidase
glucose
Prior art date
Application number
PCT/DK1987/000130
Other languages
English (en)
Inventor
Otto Melchior Poulsen
Original Assignee
Otto Melchior Poulsen
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Otto Melchior Poulsen filed Critical Otto Melchior Poulsen
Publication of WO1988002600A1 publication Critical patent/WO1988002600A1/fr
Priority to NO882708A priority Critical patent/NO882708L/no
Priority to FI882947A priority patent/FI882947A/fi

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q11/00—Preparations for care of the teeth, of the oral cavity or of dentures; Dentifrices, e.g. toothpastes; Mouth rinses
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/43—Enzymes; Proenzymes; Derivatives thereof
    • A61K38/44—Oxidoreductases (1)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/43—Enzymes; Proenzymes; Derivatives thereof
    • A61K38/44—Oxidoreductases (1)
    • A61K38/443—Oxidoreductases (1) acting on CH-OH groups as donors, e.g. glucose oxidase, lactate dehydrogenase (1.1)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/43—Enzymes; Proenzymes; Derivatives thereof
    • A61K38/46—Hydrolases (3)
    • A61K38/47—Hydrolases (3) acting on glycosyl compounds (3.2), e.g. cellulases, lactases
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/64—Proteins; Peptides; Derivatives or degradation products thereof
    • A61K8/66—Enzymes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders

Definitions

  • This invention relates to a bactericidal composition of the type containing at least one enzyme which in aqueous milieu and in the presence of oxygen and a substrate may form oxygen peroxide together with a peroxidase and a thiocyanate.
  • the bactericidal composition according to the invention is generally applicable, but as the composition is primarily developed and tested with respect to preparing a composition for the prevention and treatment of gingivitis and paradentosis, the invention will be more particularly explainde in the following with a view to obtaining a solution of this problem.
  • dental plaque depositing on the tooth surfaces as bacterial coatings which contain more or less protein and carbon hydrate substances, epithelial cells and food debris or stimulant debris are sharing the responsibility for caries, dental calculus, gingivitis and paradentosis.
  • Certain of the bacteria synthesize some strongly adhesive carbon hydrates (dextrans) from carbon hydrates of the intaken foodstuffs or stimulants and particularly sucrose constitutes the major basis of the formation of the dextrans.
  • the bacteria in plaques develop organic acids, in particular lactic acid and acetic acid due to the fermentative decomposition and reaction of the particularly decomposable mono- and disaccharides occurring in or developed from intaken foodstuffs and stimulants.
  • Dental calculus occurs as a cause of a minirali zation of dental plaque having a tendency to accumulate calcium and phosphate. If dental plaques are not regularly removed, e.g. by tooth brushing the accumulation of calcium and phosphates will reach a stage where a coherent and wear-resisting network of hydroxy apatite embedding the bacteria develops on the surfaces of the teeth.
  • the progressing dental calculus in which hydroxy apatite amounts to 80% of the total material of dental calculus has a tendency to grow, because the bacteria in dental calculus cannot be removed by ordinary tooth brushing.
  • dental calculus develops typically in areas of the teeth that are difficult to clean by tooth brushing. In this context the tooth surfaces facing the spaces between the teeth are particularly important. Dental calculus deposited between the tooth spaces will be spread therefrom and further on to the spaces between the teeth and gingivae, following which the dental calculus under gingivae will be spread from the tooth spaces to other areas of the teeth. In more advanced stages the dental calculus occurs on large areas between gingivae and the neck of the teeth and the organic acids generated by the acids will not only demineralize the teeth but also the bone structures located behind gingivae.
  • Said chemical compounds are said to derange, thereby confining the formation of dextran gels from sucrose caused by the bacteria.
  • the method does not have the intended effect on bacterial coatings in the teeth spaces where plaques are difficult to remove and where adhering food residues most frequently ensure that the bacteria have free access to low-molecular carbon hydrates.
  • the method does not have any effect on already developed dental calculus and, therefore, this method is held to be less suited with respect to the inhibition of gingivitis and paradentosis.
  • dextran gels generated in plaques are according to the accepted German patent application No. 1,955,956 proposed to be broken down by dextranases. According to AT patent No. 318,815 it has, however, been found that dextranase was merely capable of breaking down soluble dextran, while in plaques there is some of the insoluble dextran that cannot be broken down. According to US patent No. 4,154,815 dental plaque is confined by making use of zinc in connection with proteases, carbohydrases and/or Upases. Zinc has been found to be active by interference with the plaque structure and the mineralization of plaques, while the enzymes are entitled to decompose the plaques densifying high-polymeric compositions. A clinical experiment referred to in said patent demonstrates that zinc and protease were capable of reducing the amount of plaques on teeth surfaces, but the experimental report says nothing about the effect in the spaces between the teeth and on dental calculus.
  • DK patent application No. 5504/78 discloses a caries inhibiting dental hygiene agent in tablet form or powder form, said agent containing only lysozyme as an active component. This reference does not specify the type of lysozyme applied.
  • dental hygiene agents regularly used for long periods of time include antibiotics interfering in the intended microflora balance within the mouth, e.g. in connection with developing resistance.
  • German patent application No. 2,027,019 and the parallel US patent No. 4,150,113 deal with a dental hygiene agent containing an addition of oxidoreductase which by oxidative decomposition of a substrate causes hydrogen peroxide to be liberated. This is asserted to provide for obtaining a caries inhibiting effect.
  • the glucose acting as a substrate is said to be obtained from food debris in the mouth containing for example starcj or desaccharides, becouse the centifrice may as well contain carbohydrases generating glucose from the poly- or disaccharides concdrned.
  • EP patent application No. 133,736 relates to dentifrice compositions having an antibacterial activity and are based on the formation of hypothiocyanate (OSCN-) during utilization.
  • the agents concerned contain an oxidable substrate and an oxidoreductase system specific of said substrate and which in the presence of oxygen in aqueous milieu generates hydrogen peroxide which by reaction wit ⁇ the thiocyanate that is further contained in the agent generates hypothiocyanate under the influence of lactose peroxidase.
  • the agents contain for example glucose as an oxidable substrate, which is oxidized with glucose oxidase under the formation of hydrogen peroxide reacting with thiocyanate (SCN-) added for instance in the form of potassium thiocyanate, and with lactoperoxidase to produce the bacteriostatic hypothiocyanate (OSCN-).
  • SCN- thiocyanate
  • OSCN- lactoperoxidase
  • the agent is proposed to be used in inhibiting caries and for that purpose a bacteriostatic activity may per se provide for obtaining an advantageous effect.
  • a bactericidal activity is, however, not sufficient, since bacterial plaques in the spaces between the teeth and in gingival pockets will continue their growth upon decline of the inhibiting efficacy of the hypothiocyanate.
  • hypothiocyanate-forming system is therefore supposed to afford only a limited effect on the development of gingivitis and paradentosis and a proper inhibition can hardly be obtained thereby.
  • An effective inhibition and treatment of gingivitis and paradentosis can solely be obtained by extinguishing the bacteria localized between the teeth and in the gingival pockets. None of the above mentioned methods of inhibiting the formation of plaques either by preventing dextrans from forming or by attenuating or possibly extinguishing the bacteria involved proves any properly substantiated effect on plaques occurring between the teeth and in gingival pockets and the primary object of the inventions referred to actually also consists in the inhibition of caries.
  • composition according to the invention containing at least one enzyme which in aqueous milieu and in the presence of oxygen and a substrate for the enzyme is capable of generating hydrogen peroxide and a peroxidase and a thiocyanate, the composition according to the invention being characterized in that it further contains lysozyme.
  • the hydrogen peroxide forming enzyme When the bactericidal composition according to the invention, for example in prophylaxis or treatment of gingivitis or paradentosis, is applied in use between the tooth spaces and into the gingival pockets, the hydrogen peroxide forming enzyme will come into contact with food debris and stimulant debris containing substances acting as substrate for the enzyme. This entails oxidation of said substances under the liberation of hydrogen peroxide which by the influence of the peroxidase contained in the composition and naturally available peroxidase is reacted with thiocyanate, thereby generating hypothiocyanate (OSCN-).
  • OSCN- hypothiocyanate
  • composition according to the invention is characterized in that the hydrogen peroxide forming enzyme is glucose oxidase.
  • composition when the composition is to be used in connection with dental hygiene it may be appropriate that it further contains invertase or lactase or both, so that sucrose or lactose present in the oral cavity are decomposed under the formation of glucose serving as substrate for the glucose oxidase.
  • invertase or lactase or both sucrose or lactose present in the oral cavity are decomposed under the formation of glucose serving as substrate for the glucose oxidase.
  • the composition may in itself contain a substrate, also when used for dental hygiene, in which case the formation of hydrogen peroxide is not restricted by the amount of substances naturally present in the oral cavity and which act as substrate for the hydrogen peroxide forming enzyme.
  • a composition according to the invention which contains glucose oxidase is particularly intended to be used for wound treatment, it is expedient that it further includes glucose or glucose forming combinations of materials as substrate for the glucose oxidase and has a water content not exceeding the lowest value at which the glucose oxidase is active.
  • the reactions resulting in the generation of hypothiocyanate attenuating bacteria and causing them to be extinguished by the lysozyme occur only when the composition comes into contact with water, e.g. in the form of blood or lymph and, consequently, the composition is stable until use.
  • the composition may as well contain no glucose, but if so, it is used together with a glucose- containing or forming agent.
  • the alternatively hydrogen peroxide forming enzyme may be an amino acid oxidase, in which case the amino acida present in blood and lymph act as substrate.
  • the peroxidase constituting a component of the composition and ensuring the formation of hypothiocyanate from the thiocyanate and the hydrogen peroxide is lactoperoxidase, myeloperoxidase or horseradish peroxidase, all of which enzymes are classified E.C.1.17.1.7 (IUPAC).
  • the invention further relates to dental hygiene preparations that are particularly suitable to ensure a selectively high mortality of bacteria in tooth spaces and in the gingival pockets without affecting the microflora in the oral cavity.
  • a general affection on the total microflora in the oral cavity is incidentally undesired.
  • the dental hygiene agents according to the invention may be in the form of toothpicks, tooth thread of miniature brushes, in particular for the prevention and treatment of gingivitis and paradentosis and are characterized in that they are impregnated with the above mentioned composition according to the invention.
  • the impregnation of for example tooth picks may be effected by humidifying them with solutions of the hydrogen peroxide forming enzyme, peroxidase, thiocyanate and lysozyme and stabilizing agents known per se for enzymes followed by drying, for instance freeze drying, of the humidified tooth picks.
  • the dentifrice may be in the form of a solution or suspension adapted to be introduced into the spaces between the teeth and gingival pockets by means of a brush, tooth thread, toothpicks or the like, said solution containing the hydrogen peroxide generating enzyme, peroxidase, thiocyanate and lysozyme, preferably together with ordinary stabilizing and preservating agents.
  • the bactericidal effect of the composition according to the invention is so high that the comparatively small amount of the components of the composition that may be added by means of impregnated toothpicks will be sufficient to thereby obtain a markable bactericidal effect.
  • the composition by treatment or prophylaxis of gingivitis and paradentosis for example by means of toothpicks is applied locally into gingival pockets and the spaces between the teeth, this composition may advantageously also be used in other dental hygiene agents, such as toothpaste the use of which only affords an inferior localized supply.
  • the composition according to the invention will obvious also be applicable in combination with other agents for enhancing the intended effect.
  • the EDTA will be capable of enhancing the effect of the composition on bacteria having a content of calcium conpounds in the cell walls.
  • the bactericidal composition is to be used for wound treatment a dressing material may be impregnated with it, ot the composition may constitute a component of a vulneary powder.
  • the composition is principally generally applicable else- where, it an effective bactericidal activity is desired which is gentle to the host organism and has a limited duration and which does not involve the same risk of forming resistant strains of micro organisms as when using antibiotics.
  • the composition according to the invention may be prepared by use of such enzymes, all of which are approved for use in foodstuffs.
  • the quantitaive ratios between the components of the bactericidal composition concerned may vary within a wide range.
  • Lactase ⁇ -galactosidase (E.C. 3.2.1.23) is given in the unit L.U. (Lactase unit) defined as the amount of enzyme producing in one minute 1 mg glucose from 6 ml of 5.0% lactose at 20°C and pH 6.5.
  • Invertase ⁇ -fructofuranosidase (E.C. 3.2.1.26) is given in S.U. (Sumner Unit) defined as the amount of enzyme generating in five minutes 1 mg invert sugar from 6 ml of 5 .4% sucrose at 20°C and pH 4.7.
  • Glucose oxidase is classified E.C. 1.1.3.4.
  • Lactoperoxidase is classified E.C. 1.17.1.7. and has the function in the composition concerned to catalyze the reaction H 2 O 2 + SCN ⁇ H 2 O
  • Lysozyme also designated muramidase, is N-acetylmura- mylhydrolase (E.C. 3.2.1.17). This enzyme decomposes N-acethylglucosamine and N-acetylmuramic acid from the cell walls.
  • the activity of lysozyme is measured by lysing of Micrococcus lysodieticus. As a well defined standard of the activity is difficult to determine, the amount of the lysozyme applied is, however, given in absolute values in the following examples.
  • the material as used had a purity of 99% and was extracted from egg white.
  • the bacterial cultures were subsequently incubated for three days at temperatures of 25, 37, 40 or 45°C, during which intervals the cultures were daily inspected. It turned out that in every case the baterial growth ceased even with an addition of only 50 ⁇ l of the solution per 10 ml growth culture.
  • the toothpicks were subsequently freeze-dried. With a view to testing the bactericidal effect of the toothpicks thus impregnated, 10 5 bacteria of different strains of the genera Streptococcus and Lactobacillus were incubated on agar plates.
  • the tooth picks that were impegnated with the composition according to the invention were pressed down into the sucrose- containing agar plates which were then incubated for six days, and the growth was daily observed. Tests were carried out by incubation at 25°C as well as at 40°C, and in particular at 37°C. A clear zone had formed around the toothpicks, completely free of bacterial growth in an area varying between 5 and 15 mm reckoned from the toothpick.
  • toothpicks were impregnated with an aqueous buffer solution of lysozyme and EDTA in the above quoted amounts but without enzymes and NaSCN. Said latter toothpicks could not prove any significant bactericidal actvity by the test carried out.
  • Distilled water ad 100 ml It can be calculated that in use such a solution will be diluted about three times by saliva available in gingival pockets and on the surfaces of the tooth necks. The resulting concentration of the components contained in the composition to about one fourth of the above specified values will be completely sufficient to obtain the desired bacteriostatic activity.
  • Example 5 An aqueous solution as in Example 5 was prepared but without invertase and colour and flavourings. Gauze wads were drenched with the solution and subsequently dried in vacuum. The resulting wound dressing material was suitable to be used in connection with a vulnerable powder containing besides the components ordinary of such a powder, about 2% by weight of glucose.

Abstract

La composition bactéricide contient une enzyme formant du peroxyde d'hydrogène, en particulier la glucose oxydase et la glucose peroxydase et notamment la lactoperoxydase, un thiocyanate et une lysozyme. Durant l'utilisation, du peroxyde d'hydrogène est produit, lequel sous l'influence de la peroxydase transforme le thiocyanate en hypothiocyanate (OSCN-) qui a le pouvoir d'atténuer les bactéries d'une manière telle qu'elles sont lysées par la lysozyme. Ladite composition est particulièrement appropriée dans la prophylaxie et le traitement de gingivites et de paradontoses, où elle est utilisée localement, par exemple appliquée comme agent d'imprégnation sur des cure-dents.
PCT/DK1987/000130 1986-10-20 1987-10-19 Composition bactericide contenant une enzyme et preparation pour le traitement d'infections notamment dentaires comprenant ladite composition WO1988002600A1 (fr)

Priority Applications (2)

Application Number Priority Date Filing Date Title
NO882708A NO882708L (no) 1986-10-20 1988-06-17 Enzymholdig baktericid blanding og tann- og saarbehandlingspreparater som omfatter blandingen.
FI882947A FI882947A (fi) 1986-10-20 1988-06-20 Enzyminnehaollande bakteriedoedande komposition och vaordpreparat foer taender och saor innehaollande denna komposition.

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
DK5016/86 1986-10-20
DK501686A DK501686A (da) 1986-10-20 1986-10-20 Enzymholdigt, baktericidt middel samt tandplejemidler og saarbehandlingsmidler, som indeholder det

Publications (1)

Publication Number Publication Date
WO1988002600A1 true WO1988002600A1 (fr) 1988-04-21

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PCT/DK1987/000130 WO1988002600A1 (fr) 1986-10-20 1987-10-19 Composition bactericide contenant une enzyme et preparation pour le traitement d'infections notamment dentaires comprenant ladite composition

Country Status (6)

Country Link
EP (1) EP0293407A1 (fr)
JP (1) JPH01501000A (fr)
AU (1) AU8175087A (fr)
DK (1) DK501686A (fr)
FI (1) FI882947A (fr)
WO (1) WO1988002600A1 (fr)

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EP0461530A2 (fr) * 1990-06-13 1991-12-18 HAARMANN & REIMER CORP. Compositions antimicrobiennes synergiques
WO1992010165A1 (fr) * 1990-12-05 1992-06-25 Warner-Lambert Company Solution nettoyante pour dentiers, contenant des enzymes
EP0500387A2 (fr) * 1991-02-21 1992-08-26 Exoxemis, Inc. Méthodes et compositions pour le traitement des infections et le contrôle de la flore utilisant l'haloperoxydase
FR2685202A1 (fr) * 1991-12-24 1993-06-25 Sederma Sa Nouvelle methode de traitement pharmaceutique et cosmetique pour la regulation de la seborrhee, de l'acne et de la flore cutanee.
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FR2695034A1 (fr) * 1992-09-01 1994-03-04 Oreal Composition cosmétique ou pharmaceutique comprenant en association une peroxydase et un agent anti-oxygène singulet.
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Cited By (75)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
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FI882947A0 (fi) 1988-06-20
JPH01501000A (ja) 1989-04-06
DK501686A (da) 1988-04-21
EP0293407A1 (fr) 1988-12-07
FI882947A (fi) 1988-06-20
DK501686D0 (da) 1986-10-20
AU8175087A (en) 1988-05-06

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