TW201815410A - Transmucosal absorption promoter for drug - Google Patents

Abstract

The present invention addresses the problem of providing a transmucosal absorption promoter for a drug. This problem is solved by using at least one member selected from the group consisting of: (component a) a basic amino acid; (component b) a peptide consisting of 2-5 basic amino acids; and (component c) tryptophan.

Description

Transmucosal absorption enhancer for drugs

The present invention relates to a transmucosal absorption enhancer of a drug and a pharmaceutical composition containing the same.

Entering the 21st century, along with the rapid development of the biological market, it is necessary to introduce innovative Drug Delivery System (DDS) technology to the formulations so that polymers such as peptides, proteins, and nucleic acid medicines can be more efficiently absorbed in the body. In particular, although it is desired to develop an oral preparation that is most convenient for patients, oral preparation of substances with extremely low mucosal absorption and stability such as biopharmaceuticals is the most difficult. At present, 99% The biopharmaceuticals are limited to administration by injection. Therefore, there is a strong demand for the development of a means for promoting transmucosal absorption of biopharmaceuticals and the like.

In the past, it has been reported that, for example, the use of a certain surfactant to cause instability of the membrane structure or the opening of tight junctions between cells can promote transmucosal absorption of biological drugs and the like (non-patent literature) 1 ~ 2). However, due to their principle, these technologies have the potential to adversely affect the living body.

Moreover, in recent years, it has been reported that the use of various cell membrane-penetrating peptides (CPPs) such as TAT peptides and penetratins can promote the introduction of biopharmaceuticals into cells (Non-Patent Document 3) . However, most of these peptides are used in cross-linking with drugs to promote the transmucosal absorption of drugs. For example, a C-terminal side cross-linked cell membrane of leuprolide, which is a therapeutic drug for prostate cancer, has been known to penetrate arginine hexapeptide peptides (Non-Patent Document 4). Therefore, this technology has low general applicability, and there is also a possibility that the drug activity is reduced due to cross-linking.

A report by the inventors of the present case indicates that cell membrane penetrating peptides such as arginine oligopeptides can be used by simply mixing with drugs to promote transmucosal absorption of drugs (Non-Patent Document 5). In the case of this technique, since cross-linking is not required, the possibility of causing the problems described above is low. For arginine oligopeptides, there are currently reported peptides consisting of more than 6 arginine peptides, such as tokamate, octapeptide, and hexapeptide. Among them, although octapeptide and decapeptide show a high transmucosal absorption promotion effect, it is said that the hexapeptide with fewer arginine residues has a lower mucosal absorption (Non-Patent Document 5).

[Habitual technical literature] [non-patent literature] Non-patent literature 1: BF Choonara, EY Choonara, P. Kumar, D. Bijukumar, LC du Toit, V. Pillay, A review of advanced oral drug delivery technologies facilitating the protection and Absorption of protein and peptide molecules, Biotech. Adv. 32 (2014) 1269-1282. Non-patent document 2: S. Gupta, A. Jain, M. Chakraborty, JK Sahni, J. Ali, S. Dang, Oral delivery of therapeutic proteins and peptides: a review on recent developments, Drug Deliv. 20 (2013) 237-246. Non-Patent Literature 3: Copolovici DM, Langel K, Eriste E, Langel U 2014. Cell-penetrating peptides: design, synthesis, and applications. ACS Nano. 8 (3): 1972-1994. Non-Patent Document 4: Kamei N, Morishita M, Ehara J, Takayama K. 2008. Permeation characteristics of oligoarginine through intestinal epithelium and its usefulness for intestinal peptide drug delivery. J Control Release 131: 94-99. Non-Patent Document 5: Morishita M, Kamei N, Ehara J, Isowa K, Takayama K. A novel approach using functional pep tides for efficient intestinal absorption of insulin, J Control Release. 2007 Apr 2; 118 (2): 177-84. Epub 2006 Dec 29. Non-Patent Document 6: Margherita Di Pisa, Ge! rard Chassaing, and Jean-Marie Swiecicki. Translocation Mechanism (s) of Cell-Penetrating Peptides: Biophysical Studies Using Artificial Membrane Bilayers, Biochemistry 2015, 54, 194-207. Non-Patent Document 7: Regulatory Science for Pharmaceutical Medical Devices, PMDRS, 46 (12), 829-836, 2015.

[Problems to be Solved by the Invention] The object of the present invention is to provide a transmucosal absorption enhancer of a drug. Furthermore, the subject is to provide a transmucosal absorption enhancer which does not require cross-linking with a drug and has less adverse effects on a living body.

[Means to solve the problem] The inventor of the present case has made careful studies in view of the foregoing problems, and found that basic amino acids such as arginine or the number of residues are smaller than those previously reported (that is, the number of residues is 5 or less) Basic amino acid oligopeptides have a high mucosal absorption promotion effect on drugs. Conventionally, it has been said that if the number of basic amino acid residues is small, the promotion effect on mucosal absorption is low (Non-Patent Document 5), so this discovery of the present invention is surprising. It was further discovered that tryptophan has a high transmucosal absorption promotion effect on the drug. Based on these findings and further research, the present invention has been completed.

That is, the present invention includes the following aspects: Item 1. A drug for promoting transmucosal absorption, comprising at least one member selected from the group consisting of: (a component) a basic amino acid , (B component) 2 to 5 peptides composed of basic amino acids and (C component) tryptophan. Item 2. The transmucosal absorption enhancer according to item 1, wherein the basic amino acid is L-amino acid. Item 3. The transmucosal absorption enhancer according to Item 1 or 2, which contains the aforementioned component a. Item 4. The transmucosal absorption enhancer according to any one of Items 1 to 3, wherein the basic amino acid is at least 1 selected from the group consisting of arginine, lysine, and histidine Species. Item 5. The transmucosal absorption enhancer according to any one of Items 1 to 4, wherein the basic amino acid is arginine. Item 6. The transmucosal absorption enhancer according to any one of Items 1 to 5, wherein the component b is a peptide composed of 2 to 4 basic amino acids. Item 7. The transmucosal absorption enhancer according to any one of Items 1 to 6, which contains the aforementioned component c. Item 8. The transmucosal absorption enhancer according to any one of Items 1 to 7, wherein the mucosa is an intestinal mucosa. Item 9. The transmucosal absorption enhancer according to any one of Items 1 to 8, wherein the isoelectric point of the aforementioned drug is 7 or less. Item 10. The transmucosal absorption enhancer according to any one of Items 1 to 9, wherein the drug is a peptide or a protein. Item 11. A pharmaceutical composition comprising the transmucosal absorption enhancer and a drug according to any one of Items 1 to 10. Item 12. The pharmaceutical composition according to Item 11, which is a solid agent. Item 13. The pharmaceutical composition according to item 11 or 12, which is an oral administration preparation.

[Inventive Effect] According to the present invention, a transmucosal absorption enhancer can be provided, which does not need to be crosslinked with a drug and has less adverse effects on a living body.

Most drugs such as biopharmaceuticals are provided as injections for reasons of low transmucosal absorption. However, injections need to be punctured, which is burdensome for patients. In addition, there is an economic burden or an environmental burden on the disposal of the syringe after use, and furthermore, in developing countries, there may be an infection caused by the repeated use of the syringe. Through the present invention, by improving the transmucosal absorption of a drug, the drug can be provided in a form of a preparation (for example, an oral preparation, etc.) which is less likely to cause the aforementioned problems.

[Forms for Implementing the Invention] Concerning expressions of "contained" and "included" in this specification, the concepts of "contained", "included", "consisting essentially of", and "consisting of only" are included.

The present invention relates to a transmucosal absorption enhancer of a drug (also sometimes referred to as "transmucosal absorption enhancer of the present invention" in this specification), and a pharmaceutical composition containing the transmucosal absorption enhancer and drug of the present invention. (In this specification, it may also be referred to as "the pharmaceutical composition of the present invention.") The transmucosal absorption enhancer of the aforementioned drug contains a component selected from the group consisting of (a component) a basic amino acid and (b component) 2 to 5. At least one of the group consisting of a basic amino acid peptide and (c component) tryptophan. These will be described below.

The basic amino acid is not particularly limited as long as it is an amino acid (preferably an α-amino acid) having a basic functional group in a side chain and an isoelectric point in a basic region.

Examples of the basic functional group include a guanidyl group, an amine group, an imidazolyl group, and the like, and preferably a guanidyl group, an amine group, and the like, and a guanidyl group is preferable.

The isoelectric point of the basic amino acid may be, for example, more than 7, and is preferably 7.5 or more, preferably 9 or more, and more preferably 10 or more. The upper limit of the isoelectric point is not particularly limited, and may be, for example, 14.

Specific examples of the basic amino acid include arginine, lysine, histidine, ornithine, citrulline, and the like of natural amino acids. Among these, arginine, lysine, etc. are preferable from the viewpoint of a mucosal absorption promoting effect, and arginine is more preferable.

The stereo configuration of the basic amino acid is not particularly limited, and either the L-form or the D-form may be used. From the viewpoint of a so-called mucosal absorption promotion effect being higher, and / or a so-called dose-dependent enhancement of a mucosal absorption promotion effect, the basic amino acid is preferably L-amino acid.

The so-called peptide composed of a basic amino acid refers to a peptide obtained by binding a plurality of basic amino acids with each other (preferably the amino group and the carboxyl group on the basic amino acid main chain) with a peptide bond. There are no special restrictions.

The number of basic amino acids constituting the peptide is 2-5, and preferably 2-4.

The peptide may be composed of only one basic amino acid, or may be composed of two or more basic amino acids.

The three-dimensional configuration of tryptophan is not particularly limited, and either L-form or D-form may be used. From the viewpoint of a so-called transmucosal absorption promotion effect being higher, and / or a so-called dose-dependent enhancement of the mucosal absorption promotion effect, tryptophan is preferably an L-body.

Basic amino acids, peptides composed of basic amino acids, and tryptophan can be chemically modified as long as they have a mucosal absorption promotion effect. The presence or absence of a mucosal absorption-promoting effect can be determined according to a known method or, for example, a method described in Test Example 1 described later.

The basic amino acid, the peptide composed of the basic amino acid, and the carboxyl group of the tryptophan at the terminal end of the main chain may be a carboxylic acid group (-COO-), amidine (-CONH ₂ ), or an ester (-COOR ).

Here, as the R in the ester, for example, C _1-6 alkyl groups such as methyl, ethyl, n-propyl, isopropyl, and n-butyl groups can be used; for example, C _3-8 cycloalkyl groups such as cyclopentyl and cyclohexyl. ; For example, C _6-12 aryl such as phenyl and α-naphthyl; For example, phenyl-C _1-2 alkyl such as benzyl and phenethyl; and α-naphthyl-C ₁ such as α-naphthylmethyl _-2 alkyl groups such as C _7-14 aralkyl groups; Pivaloyloxymethyl and the like.

Furthermore, basic amino acids, peptides composed of basic amino acids, and tryptophan acids also include protected amino groups at the end of the main chain (for example, C _1-6 alkyl groups such as methylamido and ethylamido). C _1-6 fluorenyl groups such as alkanoyl, etc.), the substituents on the side chain (e.g. amine, imidazolyl, guanidino, etc.) are protected by appropriate protective groups (e.g. formamyl, acetamyl etc. C _1-6 alkyl groups such as C _1-6 alkylfluorenyl and the like) protectors and the like.

The basic amino acid, the peptide composed of the basic amino acid, and tryptophan may be in the form of a pharmaceutically acceptable salt with an acid or a base. The salt is not particularly limited as long as it is a pharmaceutically acceptable salt, and either an acidic salt or a basic salt can be used. Examples of acidic salts include inorganic acid salts such as hydrochloride, hydrobromide, sulfate, nitrate, and phosphate; acetate, propionate, tartrate, fumarate, and maleate Organic acid salts such as diacid, malate, citrate, mesylate, p-toluenesulfonate; amino acid salts such as aspartate, glutamate, etc. Examples of the basic salt include alkali metal salts such as sodium salts and potassium salts; and alkaline earth metal salts such as calcium salts and magnesium salts.

Basic amino acids, peptides composed of basic amino acids, and tryptophan may also be in the form of solvates. The solvent is not particularly limited as long as it is pharmaceutically acceptable, and examples thereof include water, ethanol, glycerol, and acetic acid.

The basic amino acid (component a) may be used alone or in combination of any two or more of them.

The peptide (component b) composed of a basic amino acid may be used alone or in combination of any two or more thereof.

Tryptophan (component c) may be used alone or in combination of any two or more.

The transmucosal absorption enhancer of the present invention is not particularly limited as long as it contains at least one selected from the group consisting of component a, component b, and component c in terms of effective ingredients. From the viewpoint that the so-called transmucosal absorption promotion effect is higher, and / or that the living body has been routinely vaccinated and has higher safety, the transmucosal absorption promoter of the present invention preferably contains an active ingredient selected from the group consisting of It is preferable that at least one of the group consisting of a component and c component contains only at least one selected from the group consisting of a component and c component in terms of active ingredients, so that only Containing c component is even better.

The a component, the b component, and the c component can be used as they are, or they can be produced according to a well-known synthesis method.

Although it is not intended to limit the explanation, when the a component and the b component are used, the mechanism of the present invention can be considered as follows. The well-known cell membrane penetrating peptides, such as arginine octapeptide, which contain many basic amino acids, can be considered to be due to their positive charge forming a complex with the drug and penetrating the cell membrane. Transmucosal absorption is improved (Non-Patent Documents 6 and 7). Therefore, the a component and the b component of the active ingredient of the present invention also have a positive charge, and it is considered that the effect is exerted based on the same mechanism.

Therefore, the drug targeted for the transmucosal absorption enhancer of the present invention, especially when the component a and the component b are used, as long as they are biologically active and can interact with the component a and / or b by electrical interaction The drug forming the complex is not particularly limited. Examples include drugs having an isoelectric point (pI) from a neutral region to an acidic region, and preferably drugs having a pI: 7 (or less than 7), and pI: 7 to 0.01. The medicine is preferably, the medicine with pI: 6 ~ 0.1 is more preferable, and the medicine with pI: 6 ~ 1 is particularly preferable.

The molecular weight of the drug is not particularly limited. For example, a drug having a molecular weight of 100 to 1,000,000, and preferably a drug of 300 to 200,000, a drug of 800 to 100,000 is preferable, a drug of 1,500 to 50,000 is more preferable, and a drug of 2000 to 10,000 is particularly preferable.

Specific examples of the drug include peptides, proteins, sugars, polysaccharides, nucleic acids, and low-molecular compounds. More specific examples include insulin, gastrin, Exendin-4, GLP1, leuprolide, calcitonin, interferon beta, PEGylated protein, dextran, Nano particles and so on.

The target mucosa of the transmucosal absorption enhancer of the present invention is not particularly limited. Mucosa can be, for example, intestinal mucosa, gastric mucosa, nasal mucosa, oral mucosa, lung mucosa, etc., and is preferably intestinal mucosa.

The pharmaceutical composition of the present invention is not particularly limited as long as it contains the above-mentioned transmucosal absorption enhancer and drug of the present invention, and may contain other ingredients as needed. The other ingredients are not particularly limited as long as they are pharmaceutically acceptable. Examples include bases, carriers, solvents, dispersants, emulsifiers, buffers, stabilizers, excipients, binders, disintegrating agents, and lubricants. , Tackifiers, humectants, colorants, perfumes, chelating agents, etc.

The dosage form of the pharmaceutical composition of the present invention is not particularly limited as long as it is prepared for the purpose of absorption from the mucosa, and examples thereof include lozenges, capsules, granules, powders, fine granules, syrups, and enteric solvents. Oral agents such as slow-release capsules, chewable tablets, drops, pills, internal liquids, snack tablets, slow-release agents, slow-release granules, etc .; nasal drops, inhalants, anal suppositories, inserts, External preparations such as raccoon, gels, etc. In addition, the pharmaceutical composition of the present invention may be any of a solid agent, a semi-solid agent, and a liquid agent, but is preferably a solid agent, a semi-solid agent, and a solid agent is preferred.

The content of the drug in the pharmaceutical composition of the present invention will vary without limitation depending on the type of drug, the object to be administered, the route of administration, the dosage form, the state of the patient, and the judgment of the physician, etc., and it can be set to 0.0001 to 95% by weight, for example It is set to 0.001 to 50% by weight.

The total content of component a and component b in the pharmaceutical composition of the present invention varies without limitation depending on the type of drug, the object to be administered, the route of administration, the dosage form, the state of the patient, and the judgment of the physician, and can be set to, for example, 0.0001 to 95% by weight, and preferably 0.001 to 50% by weight.

The a component and the b component exhibit their effects even when they are small. From such a point of view, the pharmaceutical composition of the present invention, for example, should be used in an amount of 1 μg to 200 mg, and preferably 10 μg to 150 mg. 100 μg to 100 mg is more preferred, and 500 μg to 50 mg is particularly preferred.

[Examples] The present invention will be described in detail below based on examples, but the present invention is not limited by these examples.

Test Example 1: Evaluation of the mucosal absorption promotion effect of arginine or arginine peptides <Test Example 1-1: Preparation of administration solution> Various amounts of recombinant human insulin (27.5 IU / mg) (Wako Pure Chemical Industries, Ltd.) (Made by the company) After dissolving in 50 μL of 0.1N hydrochloric acid aqueous solution in a polypropylene tube, it was diluted with 1.4 mL of a PBS solution (pH 6.0) containing methylcellulose at a concentration of 0.001%, and then 0.1 μN in 50 μL The sodium hydroxide aqueous solution was standardized to obtain an insulin solution. On the other hand, in terms of the test substance, various amounts of basic amino acid (L-spermine acid) (manufactured by Wako Pure Chemical Industries, Ltd.) or various amounts of basic amino acid oligopeptide (2 mer, 4 mer, or 8 mer) into a polypropylene tube, and then dissolved in a PBS solution (pH 6.0) containing methyl cellulose at a concentration of 0.001% to obtain a test substance solution. The insulin solution obtained as described above is directly mixed, or the insulin solution obtained as described above and the test substance solution are each mixed in equal amounts to prepare a dosing solution and used in the following administration test.

<Test Example 1-2: Administration Test> A administration test was performed using male SD (Sprague Dawley) rats (body weight 180 to 220 g) (manufactured by SLC). The anesthetized rat fixed on the plate was cut open in the middle to expose the ileum. A catheter was inserted at the proximal end of the ileocecal section, and to prevent fluid loss, it was returned to its original position in the abdominal cavity after ligation. Using the infusion pump, PBS (37 ° C) was passed through the catheter and circulated (5.0 mL / min, 4 minutes). Remove the tube from the catheter and close its part. In order to return the blood glucose level caused by the surgery to the original state, the rats were directly left for 30 minutes. Thereafter, 0.5 mL of the administration solution obtained in the aforementioned Test Example 1-1 was directly administered into the ileal loop. Table 1 shows the amounts of insulin administered, and the types and concentrations of test substances in the administration solution. The concentrations of the test substances in Examples 1 to 3 and Comparative Example 2 were adjusted to the same concentration (8 mM) when converted to arginine monomer.

[Table 1]

<Experimental Example 1-3: Evaluation of the promotion effect for transmucosal absorption> In Experimental Example 1-2, blood was collected before administration and after 5, 10, 15, 30, 60, 120, and 180 minutes after administration. The obtained blood was centrifuged (13,400 g, 1 minute) to obtain plasma. The concentration of human insulin (that is, insulin absorbed through the intestinal mucosa after administration) in plasma was measured using a human insulin ELISA kit (manufactured by Mercodia). The measurement results are shown in FIG. 1. As shown in FIG. 1, insulin absorption can be promoted by administering arginine or arginine oligopeptide together with insulin. The order of the intensity of the absorption-promoting effect is arginine (Example 1: LR)> arginine dipeptide (Example 2: L-R2)> arginine tetrapeptide (Example 3: L-R4) )> Arginine octapeptide (Comparative Example 2: L-R8).

Test example 2: Evaluation of the effect of the amount-dependent dependence of arginine on the promotion of transmucosal absorption Except that the type and concentration of the test substance in the administration solution are shown in Table 2, the test was performed in the same manner as in Test Example 1. The results are shown in Figure 2. As shown in Figure 2, arginine promotes insulin absorption in a dose-dependent manner.

[Table 2]

Test Example 3: Evaluation of the promotion effect of mucosal absorption of basic amino acid The test was performed in the same manner as in Test Example 1 except that the types and concentrations of the test substance in the administration solution were as shown in Table 3. The results are shown in Figure 3. As shown in FIG. 3, even basic amino acids other than arginine promote insulin absorption. The order of the strength of the absorption-promoting effect is arginine (Example 1: L-R)> lysine (Example 6: L-K)> histidine (Example 7: L-H).

[table 3]

Test Example 4: Evaluation 1, Effect on intestinal epithelial tissue integrity () After Test Example 1, the ileum was treated with 20 mL of PBS (37 ° C), and then air was passed through. 0.5 mL of the solution was directly administered into the ileal loop. The solutions for administration are the following 4 types: Solution 1: PBS Solution 2: PBS containing insulin (insulin amount: 50 IU / body weight (kg)) Solution 3: PBS solution containing L-arginine at a concentration of 40 mM 4: PBS containing sodium taurodeoxycholate at a concentration of 5% (w / v).

Three hours after the administration, the ileum loop was washed with 5.0 mL of PBS, and the intestinal fluid was recovered. The LDH (lactate dehydrogenase) concentration in the intestinal fluid was measured using the LDH kit. The measurement results are shown in FIG. 4. As shown in FIG. 4, when sodium taurodeoxycholate was administered in a positive control group, the LDH concentration in the intestinal fluid was extremely high compared with the case where PBS was administered. This phenomenon shows that, because sodium taurodeoxycholate degrades the integrity of the intestinal epithelium, LDH, which is originally present in blood, leaks through the intestinal epithelium. On the other hand, when arginine is administered, the concentration of LDH in the intestinal fluid is almost unchanged from the case of PBS. Therefore, it has been shown that arginine has little effect on the integrity of intestinal epithelial tissue. And is a highly safe ingredient.

Test Example 5: Evaluation 1. The effect on the tight junction structure of epithelial tissue was evaluated based on the resistance of the cultured cell layer (TEER: transepithelial electrical resistance) after the test substance such as arginine was applied. The effect of the tight connection structure of the test substance. Specifically, it proceeds as follows.

Human colon cancer-derived cells (Caco-2 cells) were seeded into 12-well transwell plates (1.0 × 10 ⁵ cells / cm ² ) and cultured in DMEM medium for 21 days (until a certain amount of TEER (> 500 Ωcm ² ) (showing that a tight connection is formed in a single layer.)). Add 500 μL of transport buffer 1 (HBs buffered with 10 mM MES) to the upper side of the transwell, and 1.5 mL of transport buffer 2 (to 10 at the bottom) mM HEPES (pH 7.4) buffered HBSS) and pre-cubed for 30 minutes. After the pre-culture, TEER was measured using a voltage resistance meter (Millicell ERS-2, manufactured by Millipore), and the obtained 値 was the TEER (T ₀ ) at the start of the test. 100 μL of the upper transfer buffer 1 was replaced with 100 μL of the test solution containing the test substance, and cultured for 2 hours. After replacing the test solution, the types and concentrations of the test substances in the upper chamber are as follows: Test solution 1: Insulin 15 μM Test solution 2: Insulin 15 μM, L-arginine 480 μM Test solution 3: Insulin 15 μM, L-arginine 960 μM test solution 4: insulin 15 μM, D-arginine 480 μM test solution 5: insulin 15 μM, D-arginine 960 μM test solution 6: insulin 15 μM, D-arginine octapeptide 60 μM.

After incubation, TEER was measured using a voltage resistance meter (Millicell ERS-2, manufactured by Millipore), and the obtained erbium was set to TEER after the test (T ₁₂₀ ). The TEER (T ₁₂₀ ) after the test is divided by the TEER (T ₀ ) at the start of the test and then multiplied by 100. 値 is shown in FIG. 5. As shown in Fig. 5, even if arginine is allowed to act at a very high concentration, TEER is hardly changed. This phenomenon shows that arginine has little effect on the tight junction structure of intestinal epithelial tissue, and is a highly safe component.

Test Example 6: Evaluation of the promotion effect of mucosal absorption through the combined use of basic amino acid and tryptophan. Except that the type and concentration of the test substance in the administration solution are as shown in Table 4, the rest are the same as those in the test example. 1 to test. The results are shown in Fig. 6. As shown in FIG. 6, the combination of basic amino acid and tryptophan can promote insulin absorption.

[Table 4]

Test Example 7: Evaluation of the promotion effect of L-tryptophan alone through mucosal absorption was performed in the same manner as in Test Example 1 except that the type and concentration of the test substance in the administration solution were as shown in Table 5. test. The results are shown in Fig. 7. As shown in Fig. 7, the absorption of insulin can be further promoted by L-tryptophan.

[table 5]

Test Example 8: Evaluation of the promotion effect of D-tryptophan alone through mucosal absorption was performed in the same manner as in Test Example 1 except that the type and concentration of the test substance in the administration solution were as shown in Table 6. test. The results are shown in Figure 8. As shown in FIG. 8, D-tryptophan can further promote insulin absorption.

[TABLE 6]

Test Example 9: Evaluation 2, the effect on the integrity of intestinal epithelial tissue was tested in the same manner as in Test Example 4 except that the following 7 solutions were used in the administration of the solution: Solution A: Contained at a concentration of 0.001% Methylcellulose in PBS solution B: PBS containing insulin (insulin amount: 50 IU / body weight (kg)) Solution C: PBS containing L-arginine at a concentration of 40 mM Solution D: L-containing at 16 mM Tryptophan PBS solution E: PBS solution containing L-tryptophan at a concentration of 32 mM F: PBS solution containing L-arginine at a concentration of 40 mM and L-tryptophan at a concentration of 50 mM G: % (w / v) concentration of PBS containing sodium taurodeoxycholate.

The measurement results are shown in FIG. 9. As shown in FIG. 9, in the case where sodium taurodeoxycholate was administered to the positive control group, the LDH concentration in the intestinal fluid was extremely high compared to the case where PBS was administered. This phenomenon shows that, because sodium taurodeoxycholate collapses the integrity of the intestinal epithelium, LDH, which is originally present in cells, leaks through the intestinal epithelium. In contrast, when tryptophan is administered, the concentration of LDH in the intestinal fluid is almost unchanged from the case of PBS administration. Therefore, it has been shown that tryptophan has little effect on the integrity of intestinal epithelium. It is a highly safe ingredient.

Test Example 10: Oral administration test in vivo For mice 24 hours after hunger strike, oral sonde (i..d. 0.9 × length 50 mm) (Natsume Seisakusho Co., Ltd., Tokyo, Japan), orally administered insulin (50 IU / kg), L-arginine (200 mM), or a mixed solution of insulin and L-arginine (40 or 200 mM) (100 μL). Blood was collected from the tail vein (one drop) before administration, and 15, 30, 60, 120, 180, 240, 300, and 360 minutes after administration, and one Touch Ultra View (Johnson & Johnson KK, Tokyo, Japan) was used. ) Measure blood glucose over time.

The measurement results are shown in FIG. 10. As shown in FIG. 10, oral administration of insulin or L-arginine alone does not make blood glucose levels. However, oral administration of insulin and L-arginine together can suppress blood glucose levels.

Figure 1 shows the results of Test Example 1. The vertical axis shows the human insulin concentration in plasma, the horizontal axis shows the elapsed time after administration, and each data shows the mean ± SEM (n = 3-5). "Insulin" indicates Comparative Example 1, "LR" indicates Example 1, "L-R2" indicates Example 2, "L-R4" indicates Example 3, and "L-R8" indicates Comparative Example 2 (see Table 1) . The results of Test Example 2 are shown in FIG. 2. The vertical axis shows the human insulin concentration in plasma, the horizontal axis shows the elapsed time after administration, and each data shows the mean ± SEM (n = 4). "0 mM" indicates Comparative Example 1, "8 mM" indicates Example 1, "16 mM" indicates Example 4, and "40 mM" indicates Example 5 (see Table 2). The results of Test Example 3 are shown in FIG. 3. The vertical axis shows the human insulin concentration in plasma, the horizontal axis shows the elapsed time after administration, and each data shows the mean ± SEM (n = 3-4). "Insulin" indicates Comparative Example 1, "LR" indicates Example 1, "LK" indicates Example 6, and "LH" indicates Example 7 (see Table 3). Fig. 4 shows the results of Test Example 4. The horizontal axis shows the concentration of LDH (lactate dehydrogenase) in the intestinal juice, and each data shows the mean ± SEM (n = 4). "PBS" means solution 1, "insulin" means solution 2, "LR" means solution 3, and "Sodium taurodeoxycholate" means solution 4. The results of Test Example 5 are shown in FIG. 5. The horizontal axis shows a value obtained by dividing the TEER (T ₁₂₀ ) after the test by the TEER (T ₀ ) at the start of the test and multiplying by 100. Each data is an average ± SEM (n = 4). The use of test solution 1, test solution 2, test solution 3, test solution 4, test solution 5, and test solution 6 is sequentially shown from the top. The results of Test Example 6 are shown in FIG. 6. The vertical axis shows the human insulin concentration in plasma, the horizontal axis shows the elapsed time after administration, and each data shows the mean ± SEM (n = 2-4). "Insulin" indicates Comparative Example 1, "L-R1" indicates Example 5, and "L-R1 + L-W1" indicates Example 8 (see Table 4). The results of Test Example 7 are shown in FIG. 7. The vertical axis shows the human insulin concentration in plasma, the horizontal axis shows the elapsed time after administration, and each data shows the mean ± SEM (n = 2-6). "Insulin" indicates Comparative Example 1, "16 mM" indicates Example 9, and "32 mM" indicates Example 10 (see Table 5). Figure 8 shows the results of Test Example 8. The vertical axis shows the human insulin concentration in plasma, the horizontal axis shows the elapsed time after administration, and each data shows the mean ± SEM (n = 2-6). "Insulin" indicates Comparative Example 1, "16 mM" indicates Example 11, and "32 mM" indicates Example 12 (see Table 6). The results of Test Example 9 are shown in FIG. 9. The vertical axis shows the LDH (lactate dehydrogenase) concentration in the intestinal fluid, and each data shows the mean ± SEM (n = 3). "PBS" means solution A, "insulin" means solution B, "＋ L-R1 (40 mM)" means solution C, "＋ L-W1 (16 mM)" means solution D, and "＋ L-W1 (32 mM)" means Solution E, "+ L-R1 (40 mM) + L-W1 (50 mM)" means solution F, and "sodium taurodeoxycholate" means solution G. FIG. 10 shows the results of Test Example 10. The vertical axis shows the relative value of the blood glucose level (when the value before the administration is 100%), the horizontal axis shows the elapsed time after the administration, and each data is an average ± SEM (n = 4-6). "L-R1" means L-arginine.

Claims (13)

Transmucosal absorption enhancer for a drug, comprising at least one selected from the group consisting of: (a component) a basic amino acid, (b component) 2 to 5 basic amino acids The constituent peptide and (c) tryptophan. The transmucosal absorption enhancer according to claim 1, wherein the basic amino acid is L-amino acid. The transmucosal absorption enhancer according to claim 1 or 2, which contains the aforementioned component a. The transmucosal absorption enhancer according to any one of claims 1 to 3, wherein the basic amino acid is at least one selected from the group consisting of arginine, lysine, and histidine. The transmucosal absorption enhancer according to any one of claims 1 to 4, wherein the basic amino acid is arginine. The transmucosal absorption enhancer according to any one of claims 1 to 5, wherein the aforementioned component b is a peptide composed of 2 to 4 basic amino acids. The transmucosal absorption enhancer according to any one of claims 1 to 6, which contains the aforementioned component c. The transmucosal absorption enhancer according to any one of claims 1 to 7, wherein the aforementioned mucosa is an intestinal mucosa. The transmucosal absorption enhancer according to any one of claims 1 to 8, wherein the isoelectric point of the aforementioned drug is 7 or less. The transmucosal absorption enhancer according to any one of claims 1 to 9, wherein the aforementioned drug is a peptide or a protein. A medicinal composition containing the transmucosal absorption enhancer according to any one of claims 1 to 10 and a drug. The pharmaceutical composition according to claim 11, which is a solid agent. The pharmaceutical composition according to claim 11 or 12, which is an oral agent.