JP6039465B2 - Oral pharmaceutical composition, dry mouth improving agent, dry mouth preventive agent, and aquaporin production promoter - Google Patents

Description

  The present invention relates to an oral pharmaceutical composition, an xerostomia improving agent, an xerostomia preventive agent, and an aquaporin production promoter.

Xerostomia (also known as dry mouth) is a disease in which saliva secretion decreases.
Although there are no accurate statistics on the number of patients with xerostomia, it is said to be about 8 million. Dry mice are more likely to appear in women than men, and 90% of the total are women, and 90% of them are said to be in their 50s and 70s.

  There are various causes of xerostomia such as aging, stress, irregular life, eating habits, living environment, unbalanced diet, smoking, salivary gland disorders, symptoms of systemic diseases such as Sjogren's syndrome, and salivary gland destruction by radiation therapy , HIV infection (AIDS), side effects of drugs and the like.

  Examples of the drug causing the side effect include diuretics such as trichlormethiazide and furosemide, antihypertensives such as reserpine and clonidine hydrochloride, anticholinergic agents such as atropine sulfate, antihistamines such as chlorpheniramine maleate, and various kinds of drugs. Antitussive expectorant, antiparkinsonian, psychotropic, antidepressant, trankinizer, muscle relaxant and the like.

  Radiation therapy is gaining importance for malignant tumors in the field of oral surgery and otolaryngology. However, due to the irradiation range, great damage to the salivary glands due to radiation is unavoidable, and as a result, particularly severe xerostomia is likely to occur. With the spread of radiation therapy, it is predicted that the number of patients with xerostomia will continue to increase.

  In general, as the age increases, the number of regularly used drugs increases. As a side effect due to an increase in the amount of drug, the amount of saliva secretion and the amount of water in the body decrease, and symptoms of xerostomia appear. In addition, the muscle strength of the mouth and jaw decreases with age, and the amount of saliva secretion decreases. As a result, saliva secretion is said to decrease by about 16% for men and about 25% for women at the age of 70 years or older.

In the early stages of xerostomia, a feeling of stickiness in the mouth, plaque, tongue coating, and bad breath occur. However, as the symptoms become severe, burning sensation in the oral cavity, pain, tongue pain, abnormal taste, tongue papillary atrophy, inflammation of the oral mucosa, erosion, ulceration, cracks in the tongue and mouth corners, other chewing, swallowing and vocalization, conversation May cause difficulties, sleep interruptions, eating disorders, and you may have to rely on tube feeding and infusions to supply your body with nutrition.
Xerostomia causes ulcers in the oral mucosa, phagocytosis, periodontal diseases including periodontal disease, increased frequency of oral and respiratory infections, and bad breath. Is the current situation.

  Artificial saliva (eg, thermoreversible composition containing a thermoreversible polymer and a bioadhesive polymer), moisturizer, hyaluronic acid, chondroitin sulfate-containing moisturizing mouthwash as ingredients for the purpose of alleviating xerostomia Oral care compositions (moisturizing creams) containing agents, polyvinylpyrrolidone and anionic mucoadhesive substances, etc., but this only provides temporary oral moistening.

Currently, cevimeline hydrochloride (hereinafter also referred to as “cevimeline”) has received the most attention as a therapeutic agent for xerostomia.
Cevimeline has a continuous promoting effect on salivary secretion. Cevimeline is an agonist of muscarinic acetylcholine receptors, acting on M1 muscarinic receptors in the brain and acting on M3 muscarinic receptors in salivary glands.

  Salivary glands are divided into three major salivary glands (parotid gland, submandibular gland, sublingual gland) and small oral glands. All of their secretory cells are controlled by parasympathetic nerves. doing. The parotid gland and submandibular gland are also controlled by sympathetic nerves and have adrenergic receptors. The parasympathetic center is in the medulla and the sympathetic center is in the spinal cord.

Most of saliva composition is water, and the rest are inorganic components (Na ⁺ , K ⁺ , Ca ²⁺ , Mg ²⁺ chloride, carbonate, phosphate, etc.) and organic components (amylase, mucin, lysozyme, nerves) Growth factors, epidermal growth factors, immunoglobulins, etc.). In the case of a human (adult), 1.0 L to 1.5 L of saliva is secreted per day.
The secretion of saliva is controlled by the excitement of the autonomic nerve (sympathetic nerve, parasympathetic nerve) that controls the salivary glands. The amount of saliva is low when sleeping at night, and the amount is high when waking up in the daytime.

  Aquaporin (AQP), known as a water channel, belongs to the membrane intrinsic protein family and is known to play a role of taking in low-molecular substances such as interstitial water into cells. It has been. In humans, the presence of 13 types of aquaporins (AQP0 to AQP12) is known.

All aquaporins penetrate the membrane six times, and the N-terminal and C-terminal amino acids are located on the cytoplasm side.
Aquaporin is widely distributed throughout the body and is involved in water homeostasis in the body.
For example, AQP1 is present in many organs including the brain, red blood cells, and vascular endothelium, AQP2 is mainly the luminal membrane side of the kidney, AQP3 is the vascular side membrane of the epidermis and kidney, and AQP4 is the glial cell, inner ear, and retina of the brain. AQP5 is distributed in the epidermis, the luminal membrane side of salivary glands and lacrimal glands, and the like.

It is known that cevimeline acts on the M3 muscarinic receptor in the basal side membrane of salivary gland cells to move AQP5 from the cytoplasm into the luminal membrane and increase the amount in the luminal membrane (for example, non Patent Document 1).
That is, it is considered that cevimeline promotes saliva secretion by acting on the M3 muscarinic receptor of salivary gland cells and increasing the amount of AQP5 at the luminal membrane.

  The relationship between saliva secretion and aquaporins other than AQP5 is not known so far, but AQP6 is expressed in salivary gland cells, as is AQP5, and oral mucosal epithelium, and AQP3 and AQP9 are above the spinous cell layer. (See, for example, Non-Patent Document 2).

Cevimeline may cause gastrointestinal symptoms such as nausea, abdominal pain, diarrhea, loss of appetite, salivary gland pain, and enlarged salivary gland as side effects. In addition, palpitation, frequent urination, hyperhidrosis, headache, chest pain, fatigue, etc. have been reported. In addition, the effect diminishes when used in combination with a drug having an anticholinergic action.
Cevimeline is currently not prescribed except for Sjogren's syndrome.
Therefore, an excellent substance that has an effect of improving and preventing xerostomia and has high safety and can be widely used as a composition for oral cavity has not been provided yet. The current situation is strongly demanded.

  In addition, epidermal cells mainly contain AQP3 and AQP5 as the aquaporin. In addition to water, these cells also take in a role of taking in low-molecular compounds such as glycerol and urea involved in water retention. It is thought to carry.

Examples of natural product-derived AQP3 expression promoters include, for example, Nozenhalen plant, gold inversion, himehagi, floret distant, asparasaslinearias, koganebana, licorice, dokudami, choji, maronier, matisus odorachishima, loofah, iridaceae crocus genus saffron In addition, dipotassium glycyrrhizinate is known as a plant extract such as the genus Strawberry of the Rosaceae family, or as a compound having an aquaporin 3 expression promoting action (see, for example, Patent Documents 1 to 6).
As a natural product-derived AQP5 expression promoter, for example, plant extracts such as the genus Strawberry of the Rosaceae family are known (see Patent Document 5).

  The pineapple extract is known to contain glucosylceramide (hydroxy fatty acid derivative) (see, for example, Patent Document 7). Further, the pineapple extract has an EGF production promoting action (see Patent Document 7), FGF-2. Production promoting action (see patent document 7), fibronectin production promoting action (see patent document 7), laminin-5 production promoting action (see patent document 8), epidermal hyaluronic acid production promotion (see patent document 8), and hyaluronidase activity inhibition Although it is known to have an action (see Patent Document 8), it is not known to have an improvement in xerostomia, a preventive effect, or an aquaporin production promoting action.

JP 2004-168732 A JP 2009-46465 A JP 2009-256271 A JP 2005-343882 A JP 2009-298765 A JP 2011-148732 A JP 2012-158573 A JP 2012-097008 A

Ishikawa Y. et al. , FEBS Lett. vol. 477, pp. 253-257 (2004) Takashi Inoue, Dentistry Bulletin, Vol.110, pp.34-49 (2010)

An object of the present invention is to solve various problems in the prior art and achieve the following objects. That is, an object of the present invention is to provide an oral pharmaceutical composition, an oral dryness-improving agent, and an oral dryness-preventing agent, which have excellent dry mouth dryness and preventive effects and are highly safe.
Another object of the present invention is to provide an aquaporin production promoter having an excellent aquaporin production promoting action and high safety.

Means for solving the problems are as follows. That is,
<1> A pharmaceutical composition for oral cavity comprising pineapple ceramide.
<2> An agent for improving xerostomia comprising pineapple ceramide.
<3> The agent for improving xerostomia according to the above <2>, wherein pineapple ceramide is contained as an active ingredient for promoting aquaporin production.
<4> A prophylactic agent for xerostomia comprising pineapple ceramide.
<5> The prophylactic agent for xerostomia according to the above <4>, comprising pineapple ceramide as an active ingredient for promoting production of aquaporin.
<6> An aquaporin production promoter characterized by containing pineapple ceramide.

ADVANTAGE OF THE INVENTION According to this invention, the said various problems in the past can be solved, and it has the outstanding dry mouth improvement and prevention effect, and is highly safe oral pharmaceutical composition, dry mouth improvement agent, and dry mouth A prophylactic agent can be provided.
Moreover, according to this invention, the said various problems in the past can be solved, and the aquaporin production promoter which has the outstanding aquaporin production promotion effect and has high safety | security can be provided.

FIG. 1 is a diagram showing temperature and humidity during an improvement test of oral dryness (Test Examples 1 to 4) in an example of the present invention (September 29, 2012 to October 27, 2012). FIG. 2A is a graph summarizing the evaluation results before and after chewable intake for the placebo group in Test Example 1 (visual observation evaluation in the oral cavity) of the present invention. FIG. 2B is a graph summarizing the evaluation results before and after chewable intake for the pinecer group of Test Example 3 (intraoral macroscopic observation evaluation) of the present invention. FIG. 3A is a graph showing oral moisture measurement values before and after chewable intake of each subject in Test Example 2 (measurement of oral moisture content) of the present invention. FIG. 3B is a graph showing an increase or decrease (relative value with respect to 100% before chewable intake) of each subject before and after chewable intake in Test Example 2 (measurement of oral moisture content) of the present invention. FIG. 3C is a graph showing the results of summarizing the increase and decrease in oral moisture measurement values before and after chewable intake in each group (Pinecera group and placebo group) in Test Example 2 (measurement of oral moisture content) of the present invention. FIG. 4A is a graph summarizing the questionnaire results before and after chewable intake for the placebo group in Test Example 3 (evaluation of subjective symptoms related to dry mouth) of the present invention. FIG. 4B is a graph summarizing the questionnaire results before and after chewable ingestion for the Pinecera group in Test Example 3 (evaluation of subjective symptoms related to dry mouth) of the present invention. FIG. 5A is a graph showing the results of VAS values relating to “moisture in the oral cavity” of Test Example 4 (evaluation by the VAS method) of the present invention in the placebo group and the pinecer group. FIG. 5B is a graph showing the results of summarizing the increase / decrease in VAS values related to “intraoral moisture” in Test Example 4 (evaluation by VAS method) of the present invention in the placebo group and the pinecer group. FIG. 5C is a graph showing the results of summarizing the VAS values for the “feeling of stickiness in the oral cavity when waking up” in Test Example 4 (evaluation by the VAS method) of the present invention in the placebo group and the pinecer group. FIG. 5D is a graph showing the results of summarizing the increase / decrease in VAS values in the placebo group and the pine-cera group related to “feeling of stickiness in the oral cavity when waking up” in Test Example 4 (evaluation by the VAS method) of the present invention. FIG. 5E is a graph showing the results of summarizing the VAS values for “Moisture of the lips” in Test Example 4 (evaluation by the VAS method) of the present invention in the placebo group and the pinecer group. FIG. 5F is a graph showing the results of summarizing the increase and decrease of the VAS value for “Moisture of the lips” in Test Example 4 (evaluation by the VAS method) of the present invention in the placebo group and the pinecer group.

(Oral pharmaceutical composition, dry mouth improving agent, dry mouth preventive agent, aquaporin production promoter)
The oral pharmaceutical composition, dry mouth ameliorating agent, dry mouth prophylactic agent, and aquaporin production promoter of the present invention contain pineapple ceramide, and further contain other components as necessary.
The content of the pineapple ceramide in the oral pharmaceutical composition, the xerostomia ameliorating agent, the xerostomia prevention agent, and the aquaporin production promoter is not particularly limited and is appropriately selected depending on the purpose. be able to.

<Pineapple ceramide>
The pineapple ceramide contains a hydroxy fatty acid derivative represented by the following structural formula (1) as a main component as glucosylceramide (hydroxy fatty acid derivative) extracted from pineapple, and further extracted from pineapple as necessary. And other hydroxy fatty acid derivatives. The pineapple ceramide preferably further contains at least one of hydroxy fatty acid derivatives represented by the following structural formulas (2) to (5).
As said pineapple ceramide, what was obtained by extracting a pineapple edible part with a solvent is preferable, and what was obtained by extracting the residue after pressing of a pineapple edible part with a solvent is more preferable.

<< Hydroxy fatty acid derivative >>
The hydroxy fatty acid derivative represented by the structural formula (1) includes a glucosyl group, a linear α-hydroxy acid having 20 carbon atoms as a fatty acid, and sphingosine (2-amino-4-octadecene-1,3 as a sphingoid base. -Diol) is a hydroxy fatty acid derivative of the chemical formula C ₄₄ H ₈₃ NO ₉ consisting of 2-amino-4,8-octadecidiene-1,3-diol having a double bond at the 8-position.
The hydroxy fatty acid derivative represented by the structural formula (2) includes a glucosyl group, a linear α-hydroxy acid having 18 carbon atoms as a fatty acid, and 2-amino-8-octadecidiene-1,3-diol as a sphingoid base. A hydroxy fatty acid derivative of the chemical formula C ₄₄ H ₇₉ NO ₉ consisting of
The hydroxy fatty acid derivative represented by the structural formula (3) includes a glucosyl group, a linear α-hydroxy acid having 24 carbon atoms as a fatty acid, and 2-amino-4-octadecene-1,3,4 as a sphingoid base. - it consists of a triol, a hydroxy fatty acid derivative of the formula _C 49 _H 95 _{NO 10.}
The hydroxy fatty acid derivative represented by the structural formula (4) includes a glucosyl group, a linear α-hydroxy acid having 25 carbon atoms as a fatty acid, and 2-amino-4-octadecene-1,3,4 as a sphingoid base. - it consists of a triol, a hydroxy fatty acid derivative of the formula _C 50 _H 97 _{NO 10.}
The hydroxy fatty acid derivative represented by the structural formula (5) includes a glucosyl group, a straight chain α-hydroxy acid having 26 carbon atoms as a fatty acid, and 2-amino-4-octadecene-1,3,4 as a sphingoid base. - it consists of a triol, a hydroxy fatty acid derivative of the formula _C 51 _H 99 _{NO 10.}

-Identification method of hydroxy fatty acid derivatives-
There is no restriction | limiting in particular as an identification method of the said hydroxy fatty acid derivative, It can carry out by a conventional method. For example, it is confirmed by TCL analysis that it has a molecular skeleton of monohexosylceramide (CMH), which is a glycolipid with monosaccharide, and then the molecular weight measured by mass spectrometry such as MALDI-TOFMS analysis, The molecular structure is estimated from the molecular skeleton information. Subsequently, by hydrolyzing the hydroxy fatty acid derivative, it is decomposed into a glucosyl group, a fatty acid, and a sphingoid base as structural units, and a structure analysis is performed on the fatty acid part and the sphingoid base part, respectively, and the estimated molecule Together with the structural information, the molecular structure of the hydroxy fatty acid derivative can be determined.
As a method for identifying a fatty acid moiety and a sphingoid base moiety, it can be performed by mass spectrometry such as GC-MS analysis.

  There is no restriction | limiting in particular as content of the said hydroxy fatty acid derivative in the said pineapple ceramide, Although it can select suitably according to the objective, It is preferable that it is at least 10 mass%, and it is at least 20 mass% more. preferable.

<< Pineapple >>
Pineapple is a perennial plant belonging to the genus Ananas of the pineapple family, and has a scientific name: Ananas comosus (L.) Merr. It is Ananas sativus Schult and is also called pear in China. The fruit is large, succulent, yellow, ripe and fragrant, and is used for food. The pineapples are mainly produced in the United States, the Philippines, Malaysia, Brazil, Australia, and the like. However, in obtaining the pineapple ceramide used in the present invention, the type and the production area are not particularly limited.

The extraction material of the pineapple ceramide is not particularly limited as long as it is a pineapple edible part such as a pulp and a fruit core part (core), and can be appropriately selected according to the purpose, but after squeezing the pineapple edible part The residue (ie, pineapple pulp) remaining after collecting the pineapple juice is particularly preferred.
The extraction raw material is preferably used after being collected, washed, dried and pulverized. The drying may be performed in the sun or using a commonly used dryer.

  The pineapple ceramide can be easily obtained by putting the pineapple edible portion into the solvent and extracting the pineapple ceramide using an arbitrary apparatus at a temperature between room temperature and the boiling point of the solvent.

<< solvent >>
The solvent is not particularly limited and may be appropriately selected depending on the intended purpose. Examples thereof include lower aliphatic alcohols having 1 to 5 carbon atoms such as methanol, ethanol, propanol and isopropanol; lower fats such as acetone and methyl ethyl ketone. Group ketones; C2-C5 polyhydric alcohols such as 1,3-butylene glycol, propylene glycol and glycerin; hexane; hydrophilic organic solvents such as lower aliphatic alcohols, lower aliphatic ketones and polyhydric alcohols, and water Or a mixed solvent thereof. Among these, a mixed solvent of a hydrophilic organic solvent and water is preferable. The hydrophilic organic solvent is not particularly limited and may be appropriately selected depending on the intended purpose. However, methanol, ethanol, propanol, 1,3-butylene glycol, glycerin and propylene glycol are preferable, and ethanol is more preferable.
In a mixed solvent of water and a hydrophilic organic solvent, when a lower aliphatic alcohol is used as the hydrophilic organic solvent, the content of the lower aliphatic alcohol in the mixed solvent is preferably 10% by volume to 100% by volume. 70 volume% to 100 volume% is more preferable, and 90 volume% is particularly preferable. When a lower aliphatic ketone is used as the hydrophilic organic solvent, the content of the lower aliphatic ketone in the mixed solvent is preferably 10% by volume to 80% by volume. When polyhydric alcohol is used as the hydrophilic organic solvent, the content of the polyhydric alcohol in the mixed solvent is preferably 10% by volume to 90% by volume.

The extraction method of the pineapple ceramide is not particularly limited as long as the fat-soluble component contained in the pineapple edible portion can be eluted in the solvent, and can be performed according to a conventional method. In the extraction process, it is not necessary to employ a special extraction method, and any apparatus can be used at room temperature or under reflux heating.
Specifically, as an extraction method of the pineapple ceramide, for example, the extraction raw material such as a residue (pineapple pulp) after squeezing the pineapple edible portion is put into a treatment tank filled with the solvent such as an ethanol aqueous solution. The mixture is heated and extracted with a reflux extractor at 80 ° C. for 2 hours with appropriate stirring as necessary, filtered while hot to elute the fat-soluble component, and then concentrated under reduced pressure using an evaporator. And a method for obtaining the desired pineapple ceramide.
At this time, the extraction conditions can be appropriately adjusted according to the extraction raw material and the like, but the amount of the extraction solvent is 5 to 20 times (mass ratio) with respect to the pineapple edible portion as the extraction raw material. The extraction time is preferably 1 hour to 3 hours, and the extraction temperature is preferably 20 ° C to 95 ° C.

The obtained pineapple ceramide is subjected to treatments such as dilution, concentration, drying and purification according to a conventional method in order to obtain a diluted, concentrated, dried, roughly purified or purified product of the pineapple ceramide. May be.
The obtained pineapple ceramide can be used as it is as the oral pharmaceutical composition, the xerostomia ameliorating agent, the xerostomia prophylactic agent and the aquaporin production promoter, but is easy to use. The said concentrate and the said dried material are preferable at a point. In obtaining the dried product, a carrier such as dextrin or cyclodextrin may be added to improve hygroscopicity.

<Other ingredients>
The other components are not particularly limited as long as they do not impair the effects of the present invention, and can be appropriately selected depending on the purpose. For example, excipients, lubricants, binders, fluids Examples include agents, sweeteners, disintegrants, stabilizers, colorants, fragrances, coating agents, flavoring agents, and flavoring agents.

-Excipient-
The excipient is not particularly limited and may be appropriately selected depending on the intended purpose. For example, D-mannitol, sucrose (including purified sucrose), sodium bicarbonate, corn starch, potato starch, wheat starch, rice Examples include starch, partially pregelatinized starch, crystalline cellulose, light anhydrous silicic acid, anhydrous calcium hydrogen phosphate, precipitated calcium carbonate, calcium silicate and the like.

-Lubricant-
The lubricant is not particularly limited and may be appropriately selected depending on the intended purpose. Examples thereof include magnesium stearate, calcium stearate, and sodium stearyl fumarate.

-Binder-
The binder is not particularly limited and may be appropriately selected depending on the intended purpose. For example, povidone, dextrin, hydroxypropylcellulose, hydroxypropylmethylcellulose, methylcellulose, polyvinyl alcohol, sodium carboxymethylcellulose, pregelatinized starch, alginic acid Examples include sodium, pullulan and gum arabic powder.

-Fluidizer-
The fluidizing agent is not particularly limited and may be appropriately selected depending on the intended purpose. Examples thereof include hydrous silicon dioxide, light anhydrous silicic acid, and talc.

-Sweetener-
The sweetener is not particularly limited and may be appropriately selected depending on the intended purpose. Examples thereof include aspartame, sodium saccharin, dipotassium glycyrrhizin, stevia, thaumatin, sucrose, mannitol, acesulfame potassium, and sucralose.

-Disintegrant-
There is no restriction | limiting in particular as said disintegrating agent, According to the objective, it can select suitably, For example, low substituted hydroxypropyl cellulose, carmellose, carboxymethyl starch sodium, crospovidone etc. are mentioned.

-Stabilizer-
There is no restriction | limiting in particular as said stabilizer, According to the objective, it can select suitably, For example, a citric acid, dibutylhydroxytoluene, talc, dextran, magnesium hydroxide etc. are mentioned.

-Colorant-
There is no restriction | limiting in particular as said coloring agent, According to the objective, it can select suitably, For example, ferric sesquioxide, yellow ferric oxide, titanium oxide etc. are mentioned.

-Fragrance-
There is no restriction | limiting in particular as said fragrance | flavor, According to the objective, it can select suitably, For example, menthol, bran oil, lemon oil, orange oil etc. are mentioned.

-Coating agent-
There is no restriction | limiting in particular as said coating agent, According to the objective, it can select suitably, For example, a hydroxypropyl cellulose, shellac, zein, a yeast origin substance, etc. are mentioned.

-Flavoring agent, flavoring agent-
There is no restriction | limiting in particular as said flavoring agent and said flavoring agent, According to the objective, it can select suitably, For example, sucrose, orange peel, a citric acid, tartaric acid etc. are mentioned.

  The oral pharmaceutical composition, the xerostomia ameliorating agent, the xerostomia prophylactic agent, and the aquaporin production promoter of the present invention are at least one of xerostomia amelioration, preventive action, and aquaporin production promoting action as necessary. It can be used by blending with other natural extracts having

<Oral pharmaceutical composition, xerostomia improving agent, xerostomia preventive agent, aquaporin production promoter>
The dosage form of the oral pharmaceutical composition, dry mouth improving agent, dry mouth prevention agent, and aquaporin production promoter of the present invention is not particularly limited and can be appropriately selected according to the purpose. Examples include tablets, pills, troches, chewables, powders, fine granules, granules, liquids, gummi, jelly and other foods, and pharmaceutical dosage forms.

<Method for producing oral pharmaceutical composition, xerostomia improving agent, xerostomia preventive agent, aquaporin production promoter>
The method for producing the oral pharmaceutical composition, the xerostomia improving agent, the xerostomia preventive agent, and the aquaporin production promoter of the present invention is not particularly limited and can be appropriately selected according to the purpose. Well-known methods for producing chewable agents can be mentioned.

The oral pharmaceutical composition, the xerostomia preventive agent, and the xerostomia ameliorating agent exert an improvement and preventive effect on xerostomia by the action of the pineapple ceramide contained as an active ingredient.
The aquaporin production promoter exhibits an aquaporin production promoting action by the action of the pineapple ceramide contained as an active ingredient.

  The oral pharmaceutical composition, the xerostomia ameliorating agent, and the xerostomia prophylactic agent of the present invention can be used for all applications that are meaningful in exhibiting excellent xerostomia improvement and preventive effects.

  The aquaporin production promoter of the present invention maintains the moisture retention function and the barrier function of the oral cavity, skin, etc. through the excellent aquaporin production promoting action, thereby preventing and improving dry mouth, drying the oral cavity, lips, and skin. , As well as all uses for preventing and improving lips and rough skin. However, the aquaporin production promoter of the present invention can be used for all uses other than these uses that are meaningful for exerting the aquaporin production promoting action.

  The xerostomia ameliorating agent, xerostomia prevention agent, and aquaporin production promoter of the present invention are excellent in action, and the components thereof are glucosylceramide (pineapple ceramide) obtained from edible pineapple. Since it is excellent in safety, it is suitable for blending into skin cosmetics, pharmaceuticals, and foods and drinks.

  Moreover, since the aquaporin production promoter of this invention has the outstanding effect | action, it can utilize suitably as a reagent for research of aquaporin and the function thru | or disease related to aquaporin.

  The oral pharmaceutical composition, xerostomia ameliorating agent, xerostomia prophylactic agent, and aquaporin production promoter of the present invention are preferably applied to humans, but each has its effects. It can also be applied to animals other than humans as long as possible.

  The amount of the oral pharmaceutical composition, the dry mouth improving agent, the dry mouth preventive agent, and the aquaporin production promoter of the present invention added to a pharmaceutical product differs depending on the pharmaceutical product to be added and cannot be specified unconditionally. However, in the case of a tablet, a capsule, etc., 1 mass%-90 mass% are preferable, and 0.001 mass%-50 mass% are preferable in another pharmaceutical. In addition, taking into account the general intake of the drug to be added, intake of any one of the oral pharmaceutical composition, the dry mouth ameliorating agent, the dry mouth prevention agent and the aquaporin production promoter per adult day It is preferable to adjust the amount to be about 1 mg to 1,000 mg.

  EXAMPLES Hereinafter, the present invention will be described more specifically with reference to examples, but the present invention is not limited thereto.

(Production Example 1: Production of pineapple ceramide)
10 kg of the residue (pineapple pulp) after squeezing the pineapple edible portion was added to 1,000 mL of 90% by volume ethanol, heated and extracted at 80 ° C. for 2 hours with a reflux extractor, and filtered while hot. Then, it concentrated under reduced pressure using the evaporator and performed the same filtration process further. The obtained residue was washed with 500 mL of water to obtain 15 g of a paste-like pineapple ceramide. The yield of the extract was 0.15 (mass%).

About the obtained pineapple ceramide, the component analysis was performed as follows.
-Measurement of hydroxy fatty acid derivatives-
A solution obtained by dissolving 100 mg of dried pineapple ceramide in 1 mL of ethanol was used as a test sample, and a commercially available glycosphingolipid standard solution ethanol solution (0.25 mg / mL, 0.5 mg / mL, 1, 2 mg / mL). 5 mg / mL) and applied to a silica gel thin layer chromatography plate, and developed with a chloroform: methanol mixed solution (9: 1, volume ratio). After the development, sulfuric acid was sprayed and heated, and a spot having the same Rf value as the standard glycosphingolipid product was used as a glycosphingolipid spot. The color intensity of thin-layer chromatography is measured with a densitometer (CS-9300PC, manufactured by Shimadzu Corporation), and a calibration curve is created based on the color intensity of the standard product obtained. Lipid content was determined. As a result of the measurement, the pineapple ceramide was found to contain 20% by mass of a hydroxy fatty acid derivative.

-Identification of hydroxy fatty acid derivatives-
The hydroxy fatty acid derivative contained in the obtained pineapple ceramide was identified by the following procedure.

<1. Estimation of molecular skeleton by TLC analysis>
As a result of developing the test sample together with the following standard sample under the following TLC analysis conditions, it was estimated that the test sample contained monohexosylceramide (CMH), which is a glycolipid having a monosaccharide.
[TLC analysis conditions]
Plate: HPTLC silica gel 60 (Merck)
Immediately before use, activation is performed at 120 ° C. for 30 minutes. Developing solvent: chloroform: methanol: water = 65: 25: 4 (volume ratio)
Color development: Orcinol sulfate reagent Standard sample: Monohexosylceramide (CMH) and steryl glycoside

<2. Estimation of molecular structure by MALDI-TOFMS analysis>
Matrix-assisted laser desorption / ionization-time-of-flight mass spectrometry (Matrix Assisted Laser Desorption / Ionization-Time of Flight Mass Spectrometry; MALDI-TOFMS), the following procedure for hydroxy derivatives of pineapple ceramides contained in fatty acid derivatives The molecular structure was estimated.
A solution prepared by adjusting 2,5-dihydroxybenzoic acid (DHB) as a matrix (sample molecule ionization aid) to a concentration of 10 mg / mL with a 10% by volume ethanol aqueous solution was used as the matrix solution. Next, a test sample is dissolved in a chloroform: methanol = 1: 1 (volume ratio) solution to a concentration of 1 mg / mL to prepare a glycolipid solution, and 0.2 μL of the glycolipid solution and 1.0 μL of the matrix solution are prepared. Were mixed on a sample plate and then air-dried to crystallize. This sample plate was set in Voyager DE-STR (manufactured by Applied Biosystems) which is a MALDI-TOFMS analyzer, and mass spectrometry was performed.

As a result, the pineapple ceramide is mainly composed of a hydroxy fatty acid derivative represented by the following structural formula (1) having a molecular formula of C ₄₄ H ₈₃ NO ₉ , and the following structural formula (2) in which the fatty acid portion and the sphingoid base portion are different. It was estimated that it was a mixture containing the hydroxy fatty acid derivative represented by (5) to (5).

<3. Structure identification of fatty acid moiety by GC-MS analysis>
The structure of the fatty acid moiety was identified by the following procedure using a mass spectrometer (Gas Chromatography-Mass Spectrometer; GC-MS) directly connected to a gas chromatograph.
0.3 mL of 2.5% by volume anhydrous hydrochloric acid methanol per 100 μg to 200 μg of glycolipid in the test sample was added and hydrolyzed at 80 ° C. for 12 hours (methanolysis). An equal amount of hexane was added to the reaction solution, and the resulting fatty acid methyl ester was extracted with hexane. Hexane extraction was repeated three times, and the obtained hexane layer was once dried under a nitrogen stream, and then the residue was trimethylsilyl (TMS) reagent (pyridine: 1,1,1,3,3,3-hexamethyldisilazane). (HMDS): trimethylchlorosilane (TMCS) = 1: 1.3: 0.8, volume ratio) 200 μL was added and heated at 60 ° C. for 10 minutes. The reaction solution was centrifuged, and 0.2 μL of the obtained supernatant was analyzed by GC-MS. The column for GC-MS analysis was J & W Scientific DB-5M (0.25 mm × 30 m), and the column temperature was kept at 60 ° C. for the first minute after sample injection, and then at 8 ° C. per minute. The temperature was raised to 300 ° C. and kept at 300 ° C. for 9 minutes.

  As a result of the fatty acid partial analysis by GC-MS, it was possible to identify that the fatty acid moiety constituting the main component hydroxy fatty acid derivative was a linear α-hydroxy acid having 20 carbon atoms. Moreover, as the fatty acid moiety derived from the test sample, linear α-hydroxy acids having carbon numbers of 18, 19, 20, 21, 22, 23, 24, 25, and 26, respectively, could be identified.

<4. Structural identification of sphingoid base moiety by GC-MS analysis>
Aqueous methanolic hydrochloric acid (prepared by mixing 8.6 mL of concentrated hydrochloric acid, 0.4 mL of water, and 41.0 mL of methanol) was added to 200 μg of glycolipid in the test sample, and hydrolysis was performed at 75 ° C. for 16 hours. An equal amount of hexane was added to the reaction solution, and the fatty acid methyl ester was extracted and removed with hexane. After the acidic methanol layer was dried under a nitrogen stream, 0.6 mL of 0.1N sodium hydroxide solution and 1.0 mL of methanol were added, and then 2.0 mL of chloroform was added and mixed, followed by centrifugation to remove the upper layer. . The lower chloroform layer was washed twice with the upper layer of Folch (chloroform: methanol: water = 1: 50: 49, volume ratio). After the obtained chloroform layer was dried under a nitrogen stream, 100 μL of a TMS reagent (pyridine: HMDS: TMCS = 1: 1.3: 0.8, volume ratio) was added to the residue and heated at 60 ° C. for 10 minutes. did. The reaction solution was centrifuged, and 0.2 μL of the obtained supernatant was analyzed by GC-MS. GC-MS analysis was performed under the same conditions as fatty acid analysis.

  As a result of the sphingoid base moiety analysis by GC-MS, it was possible to identify that the sphingoid base moiety constituting the main component glucosylceramide was 2-amino-4,8-octadecidiene-1,3-diol. Moreover, 2-amino-4,8-octadecidiene-1,3-diol and 2-amino-4-octadecene-1,3,4-triol could be identified as sphingoid base moieties derived from the test sample.

<5. Identification of hydroxy fatty acid derivatives>
From the above analysis results, as estimated by the MALDI-TOFMS analysis, the main components of the hydroxy fatty acid derivative contained in the pineapple ceramide are a glucosyl group, a linear α-hydroxy acid having 20 carbon atoms as a fatty acid, and a sphingoid. It was confirmed that it was a hydroxy fatty acid derivative represented by the above structural formula (1) of the chemical formula C ₄₄ H ₈₃ NO ₉ , consisting of 2-amino-4,8-octadecidiene-1,3-diol as a base. Further, the pineapple ceramide is represented by the structural formulas (2) to (5) in which the hydroxy fatty acid derivative represented by the structural formula (1) is a main component and the number of carbon atoms and sphingoid bases of the fatty acid portion are different. It was confirmed that this was a mixture containing a hydroxy fatty acid derivative.

(Example 1: Production of pineapple ceramide-containing chewable)
A pineapple ceramide-blended chewable having the composition shown in Table 1 below was produced.
6 g of pineceramide containing 20% by mass of glucosylceramide of Production Example 1, 250 g of starch (F-melt (registered trademark) F-1, manufactured by Fuji Chemical Industry Co., Ltd.), 121.5 g of dextrin (made by Matsutani Chemical Industry Co., Ltd.), cellulose (stock) A pineapple ceramide-containing chewable was manufactured using 100 g of Fushimi Factory), 7.5 g of fine silicon oxide (Fuji Syria Chemical Co., Ltd.), and 15 g of sucrose fatty acid ester (Mitsubishi Chemical Foods Co., Ltd.). As the chewable manufacturing method, a known method was used.
Table 1 shows the composition (content of each component) of the obtained pineapple ceramide-containing chewable.

(Comparative Example 1: Production of chewable without pineapple ceramide)
Instead of the pineapple ceramide of Example 1, the same procedure as in Example 1 was carried out except that 4 g of a pigment (gardenia pigment, powdered Sun Yellow (registered trademark) No. 2FU, manufactured by Saneigen FFI Co., Ltd.) was used. A pineapple ceramide-free chewable (placebo) was produced. Table 2 shows the composition (content of each component) of the resulting pineapple-ceramide-free chewable.

Intraoral dry improvement test subjects were treated with either the pineapple ceramide-containing chewable of Example 1 or the pineapple-ceramide-free chewable (placebo) of Example 1 within the oral cavity for 4 weeks every day within 1 hour after lunch. For more than a minute, they were ingested in the form of holding with the sensation of licking their heels. At that time, each subject was not told whether he was taking pineapple ceramide-containing chewable or pineapple-ceramide-free chewable (placebo).
There were a total of 12 subjects (5 men and 7 women). Hereinafter, subjects who took the pineapple ceramide-containing chewable for 4 weeks are referred to as the pinecer group, and subjects who took the pineapple-ceramide-free chewable (placebo) for 4 weeks are referred to as the placebo group.
(Pinecera group)
Number of people: 6 Age: 47.5 years on average (placebo group)
Number of people: 6 Age: Average: 44.2 years old The following was evaluated for the effect of improving dry mouth.
FIG. 1 shows the temperature and humidity during the test period (September 29, 2012 to October 27, 2012).

(Test Example 1: Evaluation by visual observation in the oral cavity)
The oral state of each subject was observed with the naked eye at each time before ingestion and 4 weeks after ingestion, and evaluated by the following five-step evaluation.
A: Dry tongue mucosa and buccal mucosa are observed B: Saliva mass and bubbles in saliva are observed C: Saliva consistency is increased D: Palate and tongue back are moistened with saliva E : Saliva on the palate and tongue back is fully visible Table 3 shows the evaluation results.

2A (placebo group) and FIG. 2B (pinecer group) are graphs of the evaluation results by visual observation shown in Table 3 above.
In the Pinecera group, improvement in oral condition was observed 4 weeks after ingestion.

(Test Example 2: Measurement of oral moisture content)
The oral moisture content of each subject was measured three times using an oral moisture meter (Mucus (registered trademark), manufactured by Life Co., Ltd.) at each time before ingestion and 4 weeks after ingestion. It was adopted. The results are shown in FIGS. 3A to 3C.
FIG. 3A shows oral moisture measurements before and after chewable intake for each subject.
FIG. 3B shows the amount of increase or decrease in oral moisture measurement values before and after chewable intake for each subject (relative value with 100% before chewable intake).
FIG. 3C shows the results of summarizing the amount of increase or decrease in oral moisture measured before and after chewable intake in each group. The measured value by an oral moisture meter was compared using t-test which is Parametric-test.
As a result of measuring the moisture content in the oral cavity with an oral moisture meter, the oral water content decreased before and after ingestion in the placebo group, whereas there was no significant difference in salivation before and after ingestion in the pinecera group. As a result, a significant difference was confirmed.

(Test Example 3: Evaluation of subjective symptoms related to dry mouth)
Each subject directly entered and answered the questionnaire regarding subjective symptoms of dry mouth (selected from the following five-step evaluation) at each time before ingestion and 4 weeks after ingestion.
A: Very dry B: Dry C: Slightly dry D: Not very dry E: Not dried at all Table 4 shows the evaluation results.

FIG. 4A (placebo group) and FIG. 4B (pinecera group) are graphs showing the results of the questionnaire shown in Table 4.

(Test Example 4: Evaluation by VAS method)
The VAS (visual analog scale) method was used to quantify the subjective dry mouth symptoms, which is a subjective item. Three items of dry symptoms (“moisture in the mouth”, “feeling of stickiness in the mouth when waking up”, “moisturization of the lips”) were evaluated by the VAS method. The scale was set to 10.0 cm, and subjects were asked to freely mark their subjectivity between two points, and the length from the 0.0 cm point was measured.
Regarding “intraoral moisture”, the point of 0.0 cm was “not felt at all”, and the point of 10.0 cm was “most moistened”.
Regarding the “feeling of stickiness in the oral cavity when waking up”, the point of 0.0 cm was “not felt at all” and the point of 10.0 cm was “most sticky”.
Regarding “moisturization of the lips”, a point of 0.0 cm was “not felt at all”, and a point of 10.0 cm was “most moistened”.
FIG. 5A shows the results of summarizing the VAS values for “moisture in the oral cavity” in the placebo group and the pinecer group.
FIG. 5B shows the results of summarizing the increase and decrease of the VAS value regarding “moisture in the oral cavity” in the placebo group and the pinecer group.
FIG. 5C shows the results of summarizing VAS values for “feeling sticky in the oral cavity when waking up” in the placebo group and the pine-cera group.
FIG. 5D shows the result of summarizing the increase and decrease of the VAS value regarding “feeling of stickiness in the oral cavity when waking up” in the placebo group and the pine-cera group.
FIG. 5E shows the results of summarizing the VAS values related to “moisturization of the lips” in the placebo group and the pinecer group.
FIG. 5F shows the result of summarizing the increase and decrease of the VAS value regarding “moisturization of the lips” in the placebo group and the pinecer group.
Compared to the placebo group, an improvement effect was confirmed in the Pinecera group for “feeling sticky in the mouth when waking up” and “moisturization of the lips”.

(Example 2: Aquaporin 3 mRNA expression promoting test)
Using the pineapple ceramide containing 20% by mass of glucosylceramide of Production Example 1 as a test sample, the aquaporin 3 (AQP3) production promoting action was tested by the following test method.

Normal human newborn foreskin epidermal keratinocytes (NHEK) in an 80 cm ³ flask using a serum-free liquid medium for epithelial keratinocyte proliferation (Epilife-KG2, manufactured by Kurashiki Boseki Co., Ltd.) at 37 ° C., 5% CO _2. The cells were precultured under the above conditions and cells were collected by trypsin treatment.
Using Epilife-KG2 as a medium, the collected cells were seeded in a 35 mm petri dish (manufactured by FALCON) at 40 × 10 ⁴ cells / 2 mL each and cultured at 37 ° C. under 5% CO ₂ for 24 hours. After 24 hours, the culture solution was discarded, and 2 mL of a test sample (sample concentration: 50 μg / mL) dissolved in Epilife-KG2 was added to each dish, and the condition was maintained at 37 ° C. and 5% CO ₂ for 24 hours. Cultured. After culturing, the culture solution is discarded, and total RNA is extracted with ISOGEN (Cat. No. 311-02501, manufactured by Nippon Gene Co., Ltd.), and the amount of each RNA is measured with a spectrophotometer (manufactured by JASCO Corporation). Total RNA was prepared to 200 μg / mL.

Using this total RNA as a template, the expression level of AQP3 and the internal standard GAPDH mRNA was measured. Detection is performed using a real-time PCR device SmartCycler (registered trademark) (manufactured by Cepheid) using a TaKaRa SYBER PrimeScript (registered trademark) RT-PCR kit (Perfect Real Time, code No. RR063A, manufactured by Takara Bio Inc.). The RT-PCR reaction was performed.
The AQP3 mRNA expression promotion rate (%) was calculated based on the total RNA preparation prepared from the cells cultured with and without the test sample added, and the correction value was obtained from the GAPDH value. The correction value of the test sample addition was calculated when the correction value of 100 was taken as 100. The results are shown in Table 5.
The calculation method of the expression rate (%) of AQP3 mRNA is as follows.
AQP3 mRNA expression promotion rate (%) = A / B × 100
However, in the said formula, A represents the correction value at the time of test sample addition, B represents the correction value at the time of a test sample non-addition (control).

The pineapple ceramide containing 20% by mass of glucosylceramide was found to have an AQP3 mRNA expression promoting effect about 2.2 times that of the control at an addition concentration of 50 μg / mL.
Incidentally, the AQP3 mRNA expression promotion rate in the positive control experiment in which retinoic acid was added to a concentration of 10 μM was 518.1%.

(Example 3: Aquaporin 5 mRNA expression promoting effect test)
Using the pineapple ceramide containing 20% by mass of glucosylceramide of Production Example 1 as a test sample, the aquaporin 5 (AQP5) production promoting effect was tested by the following test method.

A mouse alveolar epithelial cell line (MLE-12) was cultured in D-MEM containing 4% by mass FBS in an 80 cm ³ flask at 37 ° C. under 5% CO ₂ , and cells were collected by trypsin treatment.
Using the D-MEM containing 4% by mass FBS in the medium, the collected cells were seeded in a 35 mm petri dish (manufactured by FALCON) at 90 × 10 ⁴ cells / 2 mL each, under conditions of 37 ° C. and 5% CO ₂ for 24 hours. Cultured. After 24 hours, the culture solution was discarded, and 2 mL of a test sample (sample concentration: 0.2 μg / mL or 12.5 μg / mL) dissolved in D-MEM containing 4% by mass FBS was added to each dish, and 37 ° C., and cultured for 24 hours in 5% CO ₂ conditions. After culturing, the culture solution is discarded, and total RNA is extracted with ISOGEN (Cat. No. 311-02501, manufactured by Nippon Gene Co., Ltd.), and the amount of each RNA is measured with a spectrophotometer (manufactured by JASCO Corporation). Total RNA was prepared to 200 μg / mL.

Using this total RNA as a template, the expression levels of mRNA of AQP5 and GAPDH as an internal standard were measured. Detection is performed using a real-time PCR device SmartCycler (registered trademark) (manufactured by Cepheid) using a TaKaRa SYBER PrimeScript (registered trademark) RT-PCR kit (Perfect Real Time, code No. RR063A, manufactured by Takara Bio Inc.). The RT-PCR reaction was performed.
The AQP5 mRNA expression promotion rate (%) was calculated based on the total RNA preparation prepared from the cells cultured without the test sample and with the test sample, respectively, and a correction value was obtained from the GAPDH value. The correction value of the test sample addition was calculated when the correction value of 100 was taken as 100. The results are shown in Table 6.
The method for calculating the acceleration rate (%) of AQP5 mRNA expression is as follows.
AQP5 mRNA expression promotion rate (%) = A / B × 100
However, in the said formula, A represents the correction value at the time of test sample addition, B represents the correction value at the time of a test sample non-addition (control).

The pineapple ceramide containing 20% by mass of glucosylceramide showed AQP5 mRNA expression promoting action.
Incidentally, the AQP5 mRNA expression promotion rate in the positive control experiment in which retinoic acid was added to a concentration of 10 μM was 241.2%.

  The oral pharmaceutical composition, oral dryness-improving agent, oral dryness-preventing agent, and aquaporin production promoter of the present invention are excellent in safety and can be ingested on a daily basis. Since it has a dryness improvement, preventive effect, and aquaporin production promoting action, it can be suitably used as a component for oral preparations, dermatological preparations, cosmetic foods and drinks, and a research reagent.

Claims (5)

An oral pharmaceutical composition comprising a hydroxy fatty acid derivative represented by the following structural formula (1) (except for use as an antibacterial agent against Candida) .
An agent for improving xerostomia comprising a hydroxy fatty acid derivative represented by the following structural formula (1) ( excluding use as an agent for improving xerostomia against Candida).
An agent for preventing xerostomia comprising a hydroxy fatty acid derivative represented by the following structural formula (1) (excluding use as an agent for preventing xerostomia against Candida).
The aquaporin production promoter characterized by containing the hydroxy fatty acid derivative represented by following Structural formula (1).
The oral cavity pharmaceutical composition according to claim 1, further comprising at least one of hydroxy fatty acid derivatives represented by the following structural formulas (2) to (5), the xerostomia improving agent according to claim 2, Item 5. The agent for preventing xerostomia according to Item 3 or the aquaporin production promoter according to Item 4.