JP2006151893A - Yeast-type growth inducing agent of dimorphic fungus - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a yeast-type growth inducing agent of a dimorphic fungus, having high safety and capable of being expected to have an enough effect of improving various symptoms of fungal infections. <P>SOLUTION: This yeast-type growth inducing agent of the dimorphic fungus contains at least one kind selected from a chitosan oligosaccharide, a reduction product of the chitosan oligosaccharide, a chitin oligosaccharide, a reduction product of the chitin oligosaccharide, glucuronic acid, glucosaminitol, lactosylamine, galactosyllactosamine, and salts thereof in an effective amount, so that the agent inhibits mycelium-type growth of the dimorphic fungus and induces yeast-type growth thereof. Thus, the various symptoms of the fungal infections caused by the mycelium-type growth of the dimorphic fungus exhibiting dimorphism are alleviated and improved. It is preferable that the oligosaccharides and the reduction products of the oligosaccharides each have a saccharide polymerization degree of 2 to 8. Further, the agent alleviates and improves the various symptoms of candidiasis, when the dimorphic fungus comprises a Candida fungus, especially Candida albicans, which is a causative fungus of the candidiasis. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

  The present invention provides at least one selected from chitosan oligosaccharide, reduced product of chitosan oligosaccharide, chitin oligosaccharide, reduced product of chitin oligosaccharide, glucuronic acid, glucosaminitol, lactosylamine, galactosyllactosamine and salts thereof. The present invention relates to a yeast-type growth inducer for dimorphic fungi containing a seed as an active ingredient. More specifically, the present invention relates to a dimorphic fungal yeast-type growth inducer that suppresses mycelial growth of a dimorphic fungus that exhibits morphological dimorphism between a yeast form and a mycelium form in growing cells.

  The so-called dimorphic fungus exhibiting morphological dimorphism, both vegetative growth, grows in both yeast-type and oval-spherical yeast-like growths and mycelium-type growths that grow like filamentous mycelia. The growth form can be reversibly converted to proliferate.

  Such reversible transformation of growth forms is induced by specific environmental factors such as pH, nutrients, physical factors, or changes in the environment where the dimorphic fungal cells are placed. It is thought to be a trait that sex fungi acquire and maintain to adapt to the environment during evolution. Therefore, although the unified mechanism of the environmental response phenomenon has not been clarified, it is considered that there is a widely conserved mechanism in these dimorphic fungi.

  In general, fungi are less pathogenic to healthy people, but they cause infection when the body's resistance is weak or the balance of the resident flora in the body is lost. Often becomes.

  In fungal infections, the symptoms are usually caused by the growth and erosion of the fungus attached to the parasitic tissue, such as superficial mycosis occurring on the skin and nails, subcutaneous tissue, It is classified into visceral mycosis (deep mycosis) that invades deep tissues of the body in the visceral organs such as the digestive tract and lungs, or systemic. Visceral mycosis often develops in patients whose resistance is remarkably reduced due to serious underlying diseases or anticancer agents.

  Examples of pathogenic fungi that are pathogenic to humans and cause refractory or serious infections include those belonging to the genus Candida, Aspergillus, Cryptococcus, Trichosporon, etc. It is known that fungi belonging to the genus Aspergillus are the above-mentioned dimorphic fungi.

Candida fungi typified by Candida albicans (Candida albicans) which is caused by bacteria of these candidiasis are normal flora of human parasitizing without causing symptoms with healthy volunteers, the onset of fungal diseases Occasionally, the symptoms are caused by the growth and erosion of fungus parasitic on tissues such as adhesion, erosion site skin, gastrointestinal tract, oral cavity, lungs, intraocular tissues, female vagina, blood and lymph. , Sometimes severe symptoms.

In a dimorphic fungus, it is considered that mechanical invasion to a tissue infested with a fungus is induced during mycelial growth because of the morphological characteristics of the mycelium type. In Candida albicans , pathogenicity to mice is promoted during the growth of mycelia by using a mutant that has been genetically modified so that its growth is induced by drugs. It has been reported that pathogenicity is suppressed by inducing growth (see Non-Patent Document 1 below). Therefore, by inducing the growth state to the yeast type, it is expected that adhesion to a tissue parasitic on bacteria and mechanical invasiveness can be suppressed and pathogenicity of the bacteria can be suppressed.

  As an attempt to induce the growth state of a dimorphic fungus into a yeast type, Patent Document 1 listed below discloses that farnezole is a Candida genus when the dimorphic fungus that reversibly converts to a yeast cell and a mycelial cell. , A form-transformation control active agent for Candida, which is a bimorphic fungus, characterized in that it contains, in its components, farzoneate, farnesyl acetate, farnesene, or nerolidol.

  The above-mentioned farnesol is a component of an aromatic essential oil contained in plants and the like, and is a compound having a dodecatriene or dodecatetraene structure having 15 carbon atoms having a linear unsaturated hydrocarbon in the main chain.

  On the other hand, chitin, which is a natural material that can be prepared from shells of crustaceans such as crabs and shrimps, is a natural polymer having a linear polymer structure with N-acetyl-D-glucosamine as a constituent sugar as its basic structure. It is a polysaccharide. Chitosan is a polysaccharide obtained by hydrolyzing and removing an N-acetyl residue that is constitutively contained in chitin, and a linear chain having D-glucosamine (2-amino-D-glucose) as a constituent sugar. It is a natural polysaccharide having a polymer structure as its basic structure.

  Chitin oligosaccharide or chitosan oligosaccharide, which is an oligosaccharide obtained by hydrolyzing the β-1,4 glucoside bond of chitin or chitosan, is a general term for oligosaccharides having a degree of polymerization of constituent sugars of about 2 to several tens. Various uses and applications as natural materials are expected.

  For example, with respect to the antibacterial activity of chitosan oligosaccharide, the following Patent Document 2 and Patent Document 3 disclose caries preventive / suppressive agents containing chitosan oligosaccharide as an active ingredient. Patent Document 4 listed below discloses a liver dysfunction prevention / amelioration agent characterized by containing at least one selected from chitin oligosaccharides, chitosan oligosaccharides and salts thereof as an active ingredient by the present applicant. Has been.

  Patent Document 5 listed below also proposes a reduced chitosan oligosaccharide (chitosan oligosaccharide alcohol) that prevents coloring and loses its characteristics in an aqueous solution state without losing the characteristics of chitosan oligosaccharide. . In Patent Document 6 below, impurities such as partially acetylated chitooligosaccharides derived from raw materials can be efficiently removed, and coloring of chitosan oligosaccharides in the production process can be prevented, and high-purity chitosan oligos are obtained. A method capable of efficiently producing sugar and chitosan oligosaccharide alcohols is disclosed.

Further, Patent Document 7 below discloses a novel food material containing N-acetylglucosaminitol, which is an alcohol form of N-acetylglucosamine, and / or a reduced product of chitooligosaccharide, particularly non-cariogenic and resistant to digestion. A novel food material having the following characteristics is disclosed.
Stephan P. Saville et al. Eukaryotic Cell, 2003 Vol.2 (5) p.1053-1060 Japanese Patent No. 3056730 JP 2004-91413 A JP 2004-256557 A Japanese Patent Laid-Open No. 10-287572 Japanese Patent Publication No. 4-74358 JP 2003-212889 A Japanese Patent No. 2529166

  Since fungi are eukaryotes and their basic mechanisms in cell physiology are similar to those of higher organisms, there is a problem that compounds having antifungal activity also act on human cells. Therefore, it is generally difficult to obtain an antifungal agent that selectively acts on fungi and has few side effects compared to compounds against bacteria (so-called antibiotics) whose basic physiological mechanism is different from that of higher organisms. Accompanying sex.

  Conventional antifungal agents are compounds having antifungal activity that suppresses fungal cell growth or kills fungal cells, and although their activity is high, they have problems of side effects and resistance due to the above reasons. There was a problem that it was not suitable for long-term medication in terms of bacteria.

  The present invention has been made in view of the above-mentioned problems of the prior art, and the object thereof is to provide a yeast-type growth inducer for dimorphic fungi that is highly safe and can be expected to sufficiently improve various symptoms of mycosis. There is to do.

  As a result of diligent research to achieve the above object, the present inventors have determined that mycelium-type growth of a dimorphic fungus exhibiting a morphological dimorphism of yeast form and mycelium in growing cells is expressed as chitosan oligosaccharide, The present inventors have found that it can be suppressed by a specific saccharide such as a chitosan oligosaccharide reduced product, and have completed the present invention.

  That is, the yeast-type growth inducer for dimorphic fungi of the present invention is a yeast-type growth inducer for dimorphic fungi that exhibits morphological dimorphism between yeast and mycelium in growing cells, and is a chitosan oligosaccharide. An active ingredient containing at least one selected from reduced products of chitosan oligosaccharide, chitin oligosaccharide, reduced product of chitin oligosaccharide, glucuronic acid, glucosaminitol, lactosylamine, galactosyllactosamine and salts thereof It is characterized by that.

  According to the present invention, an effective amount of the above-mentioned specific saccharide is mixed into a nutrient source necessary for the growth of fungal cells to suppress the mycelium-type growth of dimorphic fungi and yeast-type growth. Can be induced.

  In the present invention, the chitosan oligosaccharide, the reduced product of chitosan oligosaccharide, the chitin oligosaccharide, and the reduced product of chitin oligosaccharide preferably have a sugar polymerization degree of 2-8. According to this, since low-activity chitosan oligosaccharide and chitin oligosaccharide are removed, it can be made higher in specific activity.

  If the dimorphic fungus is a fungus belonging to the genus Candida, the fungal yeast-type growth inducer of the present invention can be used effectively.

Moreover, if the said dimorphic fungus is Candida albicans , the yeast type growth inducer of the dimorphic fungus of this invention can be used more effectively.

That is, according to these, a fungus belonging to the genus Candida, the symptoms of fungal infections caused by particularly mycelial growth of Candida albicans (Candida albicans), reduced, can be improved.

  By using the yeast growth inducer of dimorphic fungi of the present invention, it is possible to suppress the growth of dimorphic fungi in a mycelium-type growth form, and to suppress the pathogenicity of these dimorphic fungi However, infectious diseases caused by these dimorphic fungi, for example, if the dimorphic fungus is a Candida fungus, an effect of reducing and improving various symptoms of candidiasis can be expected. In addition, since it is a highly safe saccharide, it can be added to pharmaceuticals, foods and drinks, gargles, creams for skin application, cosmetics, feeds, feeds, agricultural chemicals, etc. and used regularly.

  In the present invention, chitosan oligosaccharide, a reduced product of chitosan oligosaccharide, a chitin oligosaccharide, a reduced product of chitin oligosaccharide is chitin prepared from a shellfish shell such as crab, shrimp, etc. It can be obtained by biochemical treatment. In that case, according to the method disclosed in Patent Document 4 or Patent Document 6 by the present applicant, it can be more suitably manufactured.

  In other words, chitosan oligosaccharide removes acid after heating chitosan partial hydrolyzate obtained by partial hydrolysis of chitosan obtained by heat-concentrating alkaline treatment with chitin together with acids such as hydrochloric acid, acetic acid and formic acid. Or by neutralizing and desalting and powdering by crystallization or the like, or after dissolving in dilute acid, a chitosan-degrading enzyme such as chitosanase or D-glucosaminidase is allowed to act. The chitin oligosaccharide can be obtained by partially hydrolyzing chitin with an acid or an enzyme.

As a method of reducing chitosan oligosaccharide and chitin oligosaccharide into a reduced product (oligosaccharide alcohol form) in which the terminal constituent sugar is a sugar alcohol form, a known method can be adopted. For example, Campbell, BJ et al: J Chromatogr., 622, 137-146 (1993), using sodium borohydride (NaBH ₄ ), P. Karrer and J. Meyer: Helvetica, 20, 626 (1937), or Yoshio Matsushita: Bull. Chem. Soc. Jpn., 24, 144-147 (1950) can be used.

  Chitosan oligosaccharide, chitosan oligosaccharide reduced product, chitin oligosaccharide, and chitin oligosaccharide reduced product obtained as described above are fractionated and purified by methods such as column chromatography and solvent fractionation as necessary. By doing this, a composition containing a desired average molecular weight in a high concentration can be obtained.

  The chitosan oligosaccharide used in the present invention, the reduced product of chitosan oligosaccharide, the chitin oligosaccharide, and the reduced product of chitin oligosaccharide may be any oligosaccharide having a reduced molecular weight, but preferably have an average molecular weight of 200 to 4, 000 are used, and more preferably those having an average molecular weight of 600 to 1,400 are used.

  For example, “COS-YS” (trade name, manufactured by Yaizu Suisan Kagaku Kogyo Co., Ltd.) is obtained by the method described above, and is a mixture having a degree of polymerization of chitosan oligosaccharide of about 2 to 8, that is, chitobiose, chitotriose, chitotetraose. , Chitopentaose, chitohexaose, chitoheptaose, chitooctaose and the like, and can be preferably used. Moreover, it is more preferable if it is a mixture with a polymerization degree of 2-6.

  In addition, for example, “NA-COS-Y” (trade name, manufactured by Yaizu Suisan Chemical Co., Ltd.) is obtained by the above method, and N-acetylglucosamine and chitin oligosaccharide having a degree of polymerization of about 2 to 8, That is, N-acetylchitobiose, N-acetylchitotriose, N-acetylchitotetraose, N-acetylchitopentaose, N-acetylchitohexaose, N-acetylchitoheptaose, N-acetylchitooctaose, etc. And can be preferably used. Moreover, it is more preferable if the degree of polymerization is a mixture of 2 to 5.

  In the present invention, as described above, oligosaccharides or reduced oligosaccharides can also be used in the form of a mixture. However, the fraction can be further fractionated to a desired degree of polymerization by a method such as column chromatography or solvent fractionation. It may be purified. In that case, according to the method disclosed in Patent Document 6 by the present applicant, it can be more suitably manufactured.

  The chitosan oligosaccharide, chitosan oligosaccharide reduced product, chitin oligosaccharide, and chitin oligosaccharide reduced product prepared by the above method are preferably desalted when they contain a large amount of salts after preparation. A known method can be adopted as the desalting method, but it is preferable to use an RO membrane having an NaCl rejection of 55.0 to 99.5%. Moreover, when the solution contained in each fraction fractionated and refined | purified by ion exchange chromatography exhibits salt or alkali, it is preferable to desalinate after neutralizing.

  On the other hand, glucosaminitol, which is a sugar alcohol form of glucosamine, can be obtained by reducing glucosamine by the above-described reduction method. The origin of glucosamine is not particularly limited, and can be obtained by, for example, complete hydrolysis of chitin with an acid or fermentation using chlorovirus disclosed in JP-A No. 2004-283144.

  Glucuronic acid (GlcA) is a monosaccharide in which the carbon at the 6-position of glucose constitutes a carboxyl group, and is widely distributed in the animal and plant world as constituent sugars such as hyaluronic acid, chondroitin, fucoidan, and the like. Although it can be obtained by extraction from natural products, decomposition, synthesis, fermentation, or the like, its origin and production method are not particularly limited. Glucuronic acid, glucuronic acid amide, sodium glucuronic acid, and the like are commercially available as pharmaceutical raw materials and may be used.

Lactosylamine (Lac-NH ₂ ) is a disaccharide having an amino group at the 1-position carbon of glucose constituting lactose. For example, lactose can be used in water or a solvent such as DMSO (dimethyl sulfoxide). It can be obtained by dissolving and adding an excess amount of ammonium bicarbonate.

  Furthermore, galactosyllactosamine (Galb1-4Galb1-4GlcN) is an oligosaccharide having a degree of polymerization of 3 in which two galactose and one glucose are linearly formed by β-1,4 glucoside bonds, A trisaccharide having an amino group at the 2nd carbon of glucose, for example, obtained by adding β-galactosidase derived from Bacillus circulans to a mixed aqueous solution of lactose and glucosamine.

  In the yeast-type growth inducer for dimorphic fungi of the present invention, chitosan oligosaccharide, chitosan oligosaccharide reduced product, chitin oligosaccharide, chitin oligosaccharide reduced product, glucuronic acid, glucosaminitol obtained by the above means , Lactosylamine and galactosyllactosamine can be used as active ingredients.

  The active ingredient may be a free form or may form a salt as long as it does not affect its activity. Examples of the salt include various inorganic salts and organic acid salts, such as chitosan oligosaccharide, reduced product of chitosan oligosaccharide, glucosaminitol, lactosylamine, galactosyllactosamine, hydrochloride, lactate, Examples include acetate, sulfate, citrate, ascorbate, malate, succinate, adipate, gluconate, tartrate, etc. If it is glucuronic acid, potassium salt, sodium salt, calcium Salts, magnesium salts, ammonium salts and the like can be mentioned, but the salt form of the final product is not particularly limited.

  When the yeast-type growth inducer of the dimorphic fungus of the present invention is used for the prevention and improvement of fungal infection, the active ingredients are pharmaceuticals, foods and drinks, gargles, creams for skin application, cosmetics, feeds, feeds, agricultural chemicals. It can be used by adding and blending them. Moreover, it can also be taken or administered substantially directly as the active ingredient.

  For example, when used as a medicine for humans, an effective amount of the above-mentioned active ingredient is adopted by employing any of various administration methods such as oral, inhalation to the lungs, application on the skin, and application to the external mucosa. Can be mixed with nutrients from outside the cells necessary for fungal cells to proliferate, so that mycelium-type growth can be suppressed and yeast-type growth of dimorphic fungi can be induced.

  In that case, using pharmaceutically acceptable carriers, excipients and other additives, sprays, creams for skin application, sustained release formulations, powders, tablets, fine granules, capsules, rounds Preparations such as pills and granules. It may also contain auxiliary additives such as stabilizers, thickeners, extenders, colorants and fragrances.

  Daily administration, ingestion, and application amount of the above-mentioned active ingredient vary depending on various conditions, but per kg of body weight, 0.1 to 300 mg for oral, 0.1 to 300 mg for inhalation into the lung, onto the skin 10 to 1,500 mg for application to the skin, 10 to 1,500 mg for application to the outer mucous membrane is preferred, 1 to 30 mg for oral administration, 1 to 30 mg for inhalation to the lung, and application to the skin 10 to 150 mg, and 10 to 150 mg is more preferable when applied to the outer mucosa. In the case of a mouthwash, the active ingredient may be used so as to be immersed in the oral cavity for a certain period of time with a solution containing preferably 0.1 to 200 mg / ml, more preferably 1 to 20 mg / ml.

  The safety of chitosan oligosaccharides, reduced products of chitosan oligosaccharides, chitin oligosaccharides, and reduced products of chitin oligosaccharides has already been confirmed. In both cases, LD50> 5 g / kg or more in both cases, the safety of which has been confirmed.

  In addition, glucuronic acid (GlcA), glucosaminitol, lactosylamine (Lac-NH2), and galactosyllactosamine (Galb1-4Galb1-4GlcN) are low in toxicity. It may be administered and ingested in view of its effectiveness.

  On the other hand, the above-mentioned active ingredient can be used by blending in foods and drinks, feeds, feeds, cosmetics and the like. In that case, it is preferable to mix | blend so that it may not influence with respect to each main objectives, such as food-drinks, feed, a feed, and cosmetics. Specifically, the blending amount per solid content thereof is preferably 0.01 to 80% by weight for foods and drinks, feeds and feeds, and 0.01 to 40% by weight for cosmetics. In addition, since all the above active ingredients are easily dissolved in water, addition and blending are easy.

  The dimorphic fungus to which the yeast growth-inducing agent of the dimorphic fungus of the present invention is applied may be any dimorphic fungus that exhibits a morphological dimorphism between the yeast form and the mycelium form in the growing cells. As an example of such a thing, the Aspergillus genus and the Sporotrix genus can be illustrated besides the fungi which belong to Candida genus.

  EXAMPLES Hereinafter, the present invention will be specifically described with reference to examples, but these examples do not limit the scope of the present invention.

<Test Example 1>
(Effects of chitosan oligosaccharide on Candida albicans growth morphology)
Each chitosan oligosaccharide having a predetermined degree of polymerization was compared for induction of yeast type growth. As a control, D-glucosamine (2-amino-D-glucose; GlcN) was compared.

  For each chitosan oligosaccharide, “COS-YS” (trade name, manufactured by Yaizu Suisan Kagaku Kogyo Co., Ltd.), which is a mixture of chitosan oligosaccharides having the above-mentioned low sugar polymerization degree, is converted into one having a predetermined polymerization degree by ion exchange column chromatography. Fractionated and purified chitobiose (COS2), chitotriose (COS3), chitotetraose (COS4), chitopentaose (COS5) and chitohexaose (COS6) hydrochlorides were used. In addition, the purity was 97.5% or more, and each contained chitosan oligosaccharides other than a predetermined degree of polymerization at a concentration that did not affect the activity.

These chitosan oligosaccharides (COS2-6) or, as a control, D-glucosamine (GlcN) was mixed with RPMI1640 (containing 0.2% glucose) (manufactured by Nissui Pharmaceutical Co., Ltd.) which is a liquid nutrient medium of Candida albicans. 1 × 10 ⁵ (cells /) of Candida albicans previously adjusted to grow into a yeast type by shaking culture at 27 ° C. using Sabouraud medium (1% peptone, 4% glucose, pH 5.6 to 5.8). Inoculated at a cell concentration of 5 ml) and cultured overnight at 37 ° C. under aerobic conditions of 5% CO ₂ , and the form of the bacteria was observed under a microscope to determine the number of yeast-type growing bacteria. For the determination of morphology, monocyte or ellipsoidal cells are yeast type, filamentous cells are mycelium type, and for cells with Germ tube which is a transformation process cell, from the diameter of yeast type which is mother cell If the length is too long, the mycelium type was used, and the others were yeast type. In addition, it was 7.2-7.5 of the culture solution which mix | blended each chitosan oligosaccharide, and was pH near neutrality.

  (A) Final concentration 20 mg / ml, or its molar concentration, with the weight being constant, the concentration of chitosan oligosaccharide (COS2-6) or D-glucosamine (GlcN) blended in RPMI 1640 which is a liquid nutrient medium (B), the final concentration was 10 mM, and the ratio of the number of yeast-type growing bacteria to the total number of bacteria was compared as the yeast-type growth-inducing activity.

The results are shown in Table 1.

  As shown in Table 1, when nothing is added, the proportion of bacteria exhibiting yeast-type growth is about 0.1%, and Candida albicans is proliferating by mycelium-type growth under this culture condition. I understand. In addition, even when D-glucosamine (GlcN) as a control was added, the proportion of bacteria exhibiting yeast-type growth was 8.0% under the condition that the blending concentration condition (A) had a constant weight concentration, and (B) It was 0.4% under a constant molar concentration, and almost no change was observed in its growth form.

  On the other hand, when chitosan oligosaccharide (COS2-6) was added, the proportion of bacteria exhibiting yeast-type growth was 100% under the condition of constant weight concentration of the blending concentration condition (A), and the mole of (B). It was 43.5 to 100% under a constant concentration condition, and a yeast-type growth inducing activity significant for chitosan oligosaccharide was observed. In particular, the proportion of bacteria showing yeast-type growth was high in COS3 and COS4, and yeast-type growth inducing activity depending on the degree of polymerization of the oligosaccharide was observed.

<Test Example 2>
(Effects of chitin oligosaccharides on the growth of Candida albicans)
For each chitin oligosaccharide having a predetermined degree of polymerization, the yeast type growth induction was compared. In addition, N-acetyl-D-glucosamine (GlcNAc) was compared as a control.

  For each chitin oligosaccharide, “NACOS-Y” (trade name, manufactured by Yaizu Suisan Kagaku Kogyo Co., Ltd.), which is a chitin oligosaccharide mixture having a low sugar polymerization degree, is separated into those having a predetermined polymerization degree by activated carbon column chromatography. And purified N-acetylchitobiose (NACOS2), N-acetylchitotriose (NACOS3), N-acetylchitotetraose (NACOS4) and N-acetylchitopentaose (NACOS5) were used. In addition, the purity was 97.5% or more, and each contained chitin oligosaccharides other than a predetermined degree of polymerization at a concentration in a range not affecting the activity.

  Using these chitin oligosaccharides (NACOS2-5) or N-acetyl-D-glucosamine (GlcNAc) as a control, the same test as in Test Example 1 was performed. In addition, pH of the culture solution which mix | blended each chitin oligosaccharide was 7.2-7.3, and was pH near neutrality. Table 2 shows the result of comparison of the yeast type growth induction activity.

  As shown in Table 2, when N-acetyl-D-glucosamine (GlcNAc), which is a control, was added, the proportion of bacteria exhibiting yeast-type growth was 7 under the condition of constant weight concentration in the blending concentration condition (A). It was 6.4% under the conditions where the molar concentration of 1% and (B) was constant, and almost no change was observed in its growth form. In addition, even when NACOS2 or NACOS3 having a polymerization degree of 2 or 3 was added, the proportion of bacteria exhibiting yeast-type growth was low.

  On the other hand, when chitin oligosaccharides (NACOS4, 5) were added, the proportion of bacteria exhibiting yeast-type growth was 25.8%, 26.8%, respectively, under the constant weight concentration conditions of (A), (B) 24.4% and 21.9% respectively under the condition where the molar concentration is constant, and chitin oligosaccharides having a polymerization degree of 4 and 5 are weaker than the activity of chitosan oligosaccharide in Test Example 1, Significant yeast-type growth-inducing activity was observed.

<Test Example 3>
For COS3 showing high specific activity in Test Example 1 above, the compounding concentrations added to the liquid nutrient medium were 3, 6, and 12 mg / ml, and the same test as in Test Example 1 was conducted. The dependence on the blending concentration was examined. Moreover, the hydrochloride of chitotriol, which is a reduced product of COS3 that has lost the intramolecular hemiacetal property of D-glucosamine, which is a constituent sugar of COS3, was also examined. The results are shown in Table 3.

  As shown in Table 3, it was clarified that the proportion of bacteria exhibiting yeast-type growth can be increased by increasing the amount of COS3 added. Moreover, although it was weaker than its activity, yeast type growth inducing activity was also observed for chitotriol.

<Test Example 4>
Various sugar-related compounds were tested in the same manner as in Test Example 1 to examine the yeast-type growth-inducing activity and evaluated relative to a control to which no activity was added.

As shown in Table 4, galactosyllactosamine (Galb1-4Galb1-4GlcN), glucuronic acid (GlcA), and reduced product of glucosamine sugar having an amino group at the 2-position of glucose of the terminal constituent sugar of an oligosaccharide having a polymerization degree of 3 glucosaminide Torr (CH1H), or sugar polymerization degree reduction of chitosan oligosaccharide 2-5 is (CH2H-CH5H), in lactosyl amine (Lac-NH _2), observed significant yeast-type growth inducing activity It was. Here, the reduced products (CH2H to CH5H) of each chitosan oligosaccharide having a sugar polymerization degree of 2 to 5 have a sugar polymerization degree of about 2-5, similar to the chitosan oligosaccharides COS2-6 and chitin oligosaccharides NACOS2-5. The prepared reduced product of chitosan oligosaccharide is further prepared by molecular weight fractionation by chromatography.

  In the abbreviated structure of oligosaccharides shown in Table 4, the oligosaccharide is glucose (Glc), galactose (Gal), glucosamine (GlcN), acetylglucosamine (GlcNAc) or glucosamine hydrochloride (GlcNHCl) as a constituent sugar. It is shown that they are composed of β-1,4 glucoside bonds (β1-4) or α-1,6 glucoside bonds (α1-6). GlcAN (6COOH-GlcNHCl) is glucosamic acid.

<Test Example 5>
Glucuronic acid (GlcA), reduction of chitosan oligosaccharide (CH4H and CH5H), the lactosyl amine (Lac-NH _2), the formulation concentration to be added to the liquid nutrient medium, and 0.1,1,10mg / ml, The same test as in Test Example 1 was conducted to examine the yeast type growth inducing activity.

As shown in Table 5, in the reduced products of lactosylamine (Lac-NH ₂ ) and chitosan oligosaccharide (CH 4 H and CH 5 H), concentration-dependent yeast type growth induction activity was observed up to 10 mg / ml. On the other hand, glucuronic acid having no amino group (GlcA) shows activity at 10 mg / ml, but no activity at a concentration of 1 mg / ml, and its specific activity is higher than that of other sugars having amino groups. It became clear that it was weak.

ADVANTAGE OF THE INVENTION According to this invention, the yeast type growth inducer of the dimorphic fungus which can be added and used for a pharmaceutical, food-drinks, a mouthwash, cream for skin application, cosmetics, feed, a feed, an agricultural chemical, etc. is provided. be able to.

Claims (4)

  Yeast type growth inducer of dimorphic fungi that exhibits morphological dimorphism between yeast and mycelium types in growing cells, chitosan oligosaccharide, chitosan oligosaccharide reduct, chitin oligosaccharide, chitin oligosaccharide reduction A yeast-type growth inducer for a dimorphic fungus, comprising at least one selected from the group consisting of a product, glucuronic acid, glucosaminitol, lactosylamine, galactosyllactosamine and salts thereof.   2. The yeast-type growth inducer for dimorphic fungi according to claim 1, wherein the chitosan oligosaccharide, the reduced product of chitosan oligosaccharide, the chitin oligosaccharide, and the reduced product of chitin oligosaccharide have a sugar polymerization degree of 2-8.   The yeast-type growth inducer for a dimorphic fungus according to claim 1 or 2, wherein the dimorphic fungus is a fungus belonging to the genus Candida. The dimorphic fungi, yeast-type growth inducers of dimorphic fungi of claims 1 to 3 wherein the Candida albicans (Candida albicans).