JPH0517344A - Lipid membranous structure for oral use - Google Patents

Abstract

PURPOSE:To obtain a lipid membranous structure for oral administration capable of efficiently conveying a held medicine to Peyer's patches in digestive tracts after administration and improving the absorbability of medicines which have been poor in absorbability in the oral administration. CONSTITUTION:A lipid membranous structure for oral administration is obtained by adding at least one selected from phosphatidylserine (preferably hydrogenated phosphatidylserine, dipalmitoylphosphatidylserine or distearoylphosphatidylserine), mannose derivatives and mannan derivatives as a membrane ingredient in an amount of >=1%, preferably 5-50% molar fraction based on the total lipid membranous ingredient to a lipid membranous structure such as a liposome, an emulsion or a micelle. This lipid membranous structure has the above-mentioned effects. Furthermore, medicines capable of being held with the aforementioned lipid membranous structure vary with the kind of the lipid membranous structure, but are not especially limited in the case of, e.g. the liposome. Fat-soluble medicines and sparingly water-soluble medicines can be cited as the medicines capable of being held with the lipid membranous structure.

Description

Detailed Description of the Invention

TECHNICAL FIELD The present invention relates to a lipid membrane structure which can be efficiently delivered to Peyer's patches in the digestive tract after oral administration. The lipid membrane structure produced by the present invention can efficiently deliver the retained drug to the Peyer's patches in the digestive tract.

2. Description of the Related Art In recent years, drug delivery aimed at improving drug efficacy, safety and usability.
System (Drug Delivery System)
m, DDS) is actively researched. Academically, this DDS is currently classified into controlled release DDS (first generation DDS) and target-oriented DDS (second generation DDS).
As far as oral drugs are concerned, the former is mainly focused on the controlled release DDS and the latter is targeted D.
It can be said that the study of DS has not been done so much except a part. By the way, the Peyer's patch in the digestive tract according to the present invention is a swelling in the mucosa of the small intestine and is composed of an aggregate of lymph nodules, and in recent years it is a tissue that has been attracting academic attention such as elucidation of its immunological role. Yes, elucidation of the uptake mechanism of latex beads from Peyer's patches (Clinical Experimental Immunology, 7
6, pp. 144, 1989) and the incorporation of a drug encapsulated in a biodegradable polymer from Peyer's patches (European Patent Application No. 87309286.0). However, it can be said that these conventional techniques were not able to selectively incorporate the drug into the Peyer's patches in the digestive tract after oral administration. Studies using liposomes as an oral agent have been made in the past. For example, insulin (Biokimica et biophysica actor, 716, 188, 1982), blood coagulation factor VII (lancet, i, 70, 198).
Year 0), Heparin (Chemical and Pharmaceutical Bulletin, Vol. 30, p. 2245, 1982)
, Etc., high-molecular drugs, vitamin K ₁ (Journal of Pharmacy and Pharmacology, Vol. 36,
527, 1984), Griseofulvin (Journal of Pharmaceutical Sciences,
73, p. 757, 1984) and other poorly water-soluble drugs, as well as liposome-encapsulated streptococcal cell wall antigens (for IgA antibody production) directed to intestinal immunity have been investigated. However, in either case,
In particular, it was not that the Peyer's patches in the digestive tract were made to have selectivity by modifying the liposome membrane with other additives. On the other hand, in the case of liposome as an injection, its membrane is phosphatidylserine (cancer
Research, 40, 4460, 1980), the antitumor effect of activated alveolar macrophages was enhanced after intravenous administration, and the mannose derivative (Biokimica et biophysica. Actor, 632, 562, 1980) or mannan derivatives (pathophysiology, 6, 771, 1985)
There is a report that accumulation by the liver and lungs was confirmed by modification with. In any case, it can be said that none of the conventional techniques can efficiently deliver a drug to the Peyer's patches in the digestive tract after oral administration.

An object of the present invention is to provide a lipid membrane structure capable of efficiently delivering a drug to Peyer's patches in the digestive tract after oral administration.

The present invention is based on the finding that the above problems can be solved by using at least one selected from phosphatidylserine, mannose derivatives and mannan derivatives as a membrane component of a lipid membrane structure. It was. That is, the lipid membrane structure according to the present invention means a particle having a membrane structure in which the polar groups of the amphipathic lipid are arranged toward the aqueous phase side of the interface, and specifically, liposome, emulsion or water-soluble Micelle etc. are illustrated. The lipid membrane structure of the present invention is characterized in that phosphatidylserine and / or mannose derivative and / or mannan derivative is added as a membrane component for efficient delivery to the Peyer's patches in the digestive tract after oral administration. . Examples of the phosphatidylserine according to the present invention include naturally-occurring soybean or egg yolk phosphatidylserine,
Hydrogenated phosphatidylserine obtained by hydrogenating these,
Alternatively, semisynthetic dimyristoylphosphatidylserine, dipalmitoylphosphatidylserine, distearoylphosphatidylserine and the like can be mentioned, but hydrogenated phosphatidylserine, dipalmitoylphosphatidylserine and distearoylphosphatidylserine are preferable. Examples of the mannose derivative according to the present invention include a derivative of phosphatidylethanolamine having a mannose residue (Biokimica et Biophysica Actor, 632, 562, 1980) and a cholesterol derivative (Proceeding of National. Academic Sciences USA 77, 4430, 1980, Dimannosyl diglyceride (Biochemical and Biophysical Research Communications, 110, 14)
0, 1983), mannobiose fatty acid esters and amides (JP-A-1-104088), phosphatidyl mannose and the like. Further, as the mannan derivative of the present invention, a derivative in which a fatty acid or cholesterol is partially substituted in the mannan which is a polysaccharide (pathophysiology, 6
Vol. 771, 1985) and the like. Additives as these film components can be used individually or as a mixture.
It may be added to the lipid membrane structure mainly composed of other membrane components such as lecithin, but these additives may be used alone to form the lipid membrane structure. That is, the phosphatidylserines can form liposomes and emulsions by themselves, and the mannose derivatives such as mannobiose fatty acid esters and amides can form emulsions and micelles by themselves. Therefore, in the present invention, the addition ratio of phosphatidylserine, mannose derivative and mannan derivative used as a membrane component in the present lipid membrane structure is not limited at all in the upper limit. The lower limit is preferably 1% or more, preferably 5 to 50%, in terms of mole fraction based on the total lipid membrane components. The drug that the lipid membrane structure of the present invention can hold depends on the type of the lipid membrane structure. For example, what the liposome can hold is not particularly limited, and examples thereof include high-molecular compounds such as insulin, hormones and polysaccharides, water-soluble drugs such as immunostimulants and immunosuppressants, and fat-soluble drugs. In the case of an emulsion, a fat-soluble drug can be mentioned, and in the case of a micelle, a poorly water-soluble drug can be mentioned. These can be retained in the lipid membrane structure produced by an ordinary method in the method for producing a lipid membrane structure described below. Next, a method for producing a lipid membrane structure to which the phosphatidylserine and / or mannose derivative and / or mannan derivative of the present invention is added will be described. a) Method for producing liposome Using a membrane component substance such as lecithin and sphingomyelin and a substance added with a phosphatidylserine and / or a mannose derivative and / or a mannan derivative, a known method (Annual Review of Biophysics
And Bioengineering, Vol. 9, pp. 467, 1
980) to prepare an aqueous dispersion of liposomes. Such liposomes may contain, as stabilizers, sterols such as cholesterol, dialkyl phosphates, charged lipids such as diacylphosphatidic acid and stearylamine, and antioxidants such as α-tocopherol. b) Emulsion production method Known using an amphipathic substance such as lecithin and polyoxyethylene sorbitan fatty acid ester (Tween), an additive substance of a phosphatidylserine and / or a mannose derivative and / or a mannan derivative, and an oil and fat such as soybean oil. An emulsion can be prepared according to the emulsion preparation method of. c) Method for producing micelle Polyoxyethylene sorbitan fatty acid ester (Twe
en), sodium fatty acid and polyoxyethylene hydrogenated castor oil and other micelle-forming surface-active substances (added to the aqueous solvent at a concentration above the micelle-forming critical concentration) and phosphatidylserine and / or mannose and / or mannan derivative additives An aqueous dispersion of micelles can be prepared according to a known method for preparing micelles.

The lipid membrane structure of the present invention is, after oral administration,
The retained drug can be efficiently delivered to the Peyer's patches in the digestive tract. Peyer's patches are aggregates of lymph node 1 in the small intestinal mucosa, which is one of the tissues involved in the absorption of drugs. It is a very important organization. Therefore, by allowing the lipid membrane structure of the present invention to retain a drug that has conventionally been poorly absorbed by oral administration, its absorbability can be improved.

EXAMPLES The present invention will be described with reference to examples and test examples, but the present invention is not limited thereto. Since the basic method for preparing liposomes described here and the experiment of the transferability of liposomes to Peyer's patches are common, they will be collectively described at the beginning. Method for Preparing Liposomes (Examples 1 to 4, Control Examples 3 to 4) Lipids having the molar ratios shown below were taken in eggplant-type Kolben as a total lipid amount of 50 μmol, and once completely dissolved in a chloroform-methanol mixed solution. The organic solvent was distilled off using an evaporator under reduced pressure. Further, the organic solvent was completely removed in a desiccator under reduced pressure to dry the lipid mixture.
Next, 2 ml of Tris-hydrochloric acid buffered physiological saline solution (pH 7.4) containing 10 mg / ml of 6-carboxyfluorescein (6-CF) or 10 mg / ml of phenol red, which is an internal aqueous phase marker, was added, and the mixture was heated to 50 ° C. And agitated with a Portex mixer. Next, a polycarbonate membrane filter having a pore size of 0.4 μm was passed at 50 ° C. to obtain a liposome dispersion liquid having a particle size of 0.4 μm or less. For the internal aqueous phase marker not encapsulated in liposomes, the liposome dispersion obtained above is passed through a Sephadex G-100 column (3 cmφ × 45 cm, eluent: Tris-hydrochloric acid buffered saline, pH 7.4). It was removed by. The liposome fraction in which the inner aqueous phase marker is encapsulated is finally diluted with Tris-hydrochloric acid buffered physiological saline (pH 7.4) to give a 6-CF concentration of 4.
400 μg as phenol red when 0 μg / ml
/ Ml (final total lipid concentration is about 1 mM = 1 μmol / ml)
Was adjusted so that The mannan-modified liposome of Example 4 had the same PC / PS / CHOL = as in Example 2.
After preparing a 7/3/2 (molar ratio) liposome, the liposome and the mannan-cholesterol derivative were added according to the method of Sunamoto et al. (The addition amount of the mannan-cholesterol derivative was 1 μmol of the phospholipid in the liposome). It was prepared by reacting at 43.1C for 2 hours as mannan (53.1 µg). Control Example 3: PC / PS / CHOL = 7/0/2 (molar ratio) Example 1: PC / PS / CHOL = 7/1/2 (molar ratio) Example 2: PC / PS / CHOL = 7 / 3/2 (molar ratio) Example 3: PC / PS / CHOL = 7/5/2 (molar ratio) Example 4: PC / PS / CHOL / Man-CHOL Control Example 4: PC / SA / CHOL: 7 / 3/2 (molar ratio) [abbreviation] PC: egg yolk phosphatidylcholine PS: hydrogenated soybean phosphatidylserine CHOL: cholesterol Man-CHOL: mannan-cholesterol derivative [N- (2-N-cholesterylcarboxyaminoethyl) carbamylmannan, Product Name: Chol-AECM
-Mannan (manufactured by Wako Pure Chemical Industries, Ltd.)] SA: Transferability experiment of stearylamine liposomes to Peyer's patches (Test Examples 1 to 1)
6) The transfer of liposomes to Peyer's patches was examined by the in situ loop method. Wistar male rat (25
(About 0 g) was fasted for 18 hours, the abdomen was opened under urethane anesthesia, and the small intestine was ligated into four sections at intervals of about 15 cm to prepare a loop. Then 1.0 ml of various test solutions in this loop
(4.0 μg as 6-CF, 400 μg as phenol red, about 1 μmol as total lipid) was injected, and after a certain period of time, the intestinal tract was removed and washed with physiological saline. Next, the structure of the Peyer's plate and the vicinity of the Peyer's plate (non-Peyer's plate) 20
mm ² was excised, and the internal aqueous phase marker that had migrated to both tissues was extracted with isoamyl alcohol and quantified. Test Example 1 As a test solution, Control Example 1 (4.0 μg of free 6-CF /
ml), Control Example 3 (PC-liposomes) and Example 2
Using PS-liposomes, a transfer experiment to Peyer's patches was performed. The results of 2 hours after the administration of the test solution are shown in Fig. 1 left and Table 1, and compared to free 6-CF, transfer of 6-CF encapsulated in PC-liposomes and PS-liposomes to Peyer's patches and non-Peyer's patches. Was found to increase. Furthermore, it was also found that PS-liposomes had significantly higher migration to Peyer's patches than to non-Peyer's plates.

[Table 1] Test Example 2 As a test solution, Control Example 2 (free phenol red 40
0 μg / ml) and Control Example 4 (SA-liposome) were used to perform a transfer experiment to Peyer's patches. The results of 2 hours after the administration of the test solution are shown in the right side of FIG. 1 and Table 2, and the migration of both Peyer's patches and non-Peyer's patches was equal to the value when free phenol red was administered, and no significant difference was observed. .

[Table 2] Test Example 3 Next, the effect of the amount of PS added on the transferability of 6-CF to Peyer's patches was examined. That is, as a test liquid,
Using the control example 3 (PC-liposomes) and the PS-liposomes of Examples 1, 2 and 3, transfer experiments to Peyer's patches were performed. The results 2 hours after the administration of the test solution are shown in FIG. 2 and Table 3, where the PS-liposomes were encapsulated as the PS content increased as compared to PC-liposomes.
An increase in the transfer of CF to Peyer's patches was observed. In addition, no significant difference was observed between the three types of PS-liposomes (Example 1, Example 2 and Example 3) having different PS contents.

[Table 3] Test Example 4 The incubation time on the transfer of PS-liposomes to Peyer's patches and non-Peyer's plates was examined. That is, using Example 2 (PS-liposomes) as a test solution, a transfer experiment to Peyer's patches was performed. The time course after administration of the test solution is shown in FIG. 3 and Table 4, and it was found that the transfer to Peyer's patches increased with the incubation time and reached saturation in about 2 hours. The transition to non-Peyer's plate was low and was almost constant.

[Table 4] Test Example 5 Using Example 2 (PS-liposomes) as a test solution, the intestinal site specificity in the transferability to Peyer's patches and non-Peyer's patches 2 hours after administration of the test solution was examined. The results are shown in Fig. 4, but no site difference was observed. Test Example 6 As a test solution, Example 2 (PS-liposomes) and Example 4 (mannan-modified PS-liposomes) were used to perform a transfer test to Peyer's patches. The results are shown in Fig. 5 and Table 5 left, and it was found that by further modifying the PS liposome with mannan, the transferability to Peyer's patches was significantly increased and the transferability to non-Peyer's patches was significantly decreased. . In addition, Control Example 5 in which an excess amount of mannan (100 times the amount added to the liposome) was co-administered with this mannan-modified PS-liposome, and an empty PS-liposome that does not encapsulate an inner aqueous phase marker with the mannan-modified PS-liposome (The film composition was the same as in Example 2, and 100 times the amount of 100
The results of Control Example 6 in which μmol of lipid was administered) are shown in the right side of FIG. 6 and the right side of Table 5. The combination of these significantly inhibited the transferability of mannan-modified PS-liposomes to Peyer's patches, and the transferability to non-Peyer's patches was not particularly affected. Therefore, the liposome according to the present invention selectively It has been proved to have the property of transferring to a plate.

[Table 5]

[Brief description of drawings]

FIG. 1 shows the results of transfer to Peyer's patches using the PS-liposomes and SA-liposomes of the present invention.

FIG. 2 shows the effect of PS addition amount on the transfer to Peyer's patches using the PS-liposomes of the present invention.

FIG. 3 shows time-course changes after administration of a test solution in transfer to Peyer's patches using the PS-liposome of the present invention.

FIG. 4 shows intestinal site specificity in transferability to Peyer's patches and non-Peyer's patches using the PS-liposome of the present invention.

FIG. 5 shows an increase in transferability to Peyer's patches by modifying the PS-liposomes of the present invention with mannan.

FIG. 6 shows increased translocation to Peyer's patches by modifying the PS-lipolymes of the invention with mannan.

Claims (4)

[Claims] 1. A lipid membrane structure for oral administration, wherein the membrane component contains phosphatidylserine. 2. A lipid membrane structure for oral administration, wherein the membrane component contains a mannose derivative or a mannan derivative. 3. A lipid membrane structure for oral administration, wherein the membrane component contains phosphatidylserine and a mannose derivative or a mannan derivative. 4. The lipid membrane structure for oral administration according to claim 1, which is a liposome, an emulsion or a micelle.