JP2002517521A - Acylated alkylated cyclodextrin derivatives and their use as drug carriers - Google Patents

Abstract

(57) Abstract: at least one or more lower alkyl groups in the molecule, a cyclodextrin derivative is disclosed having at least one C _2-20 alkanoyl group. Also disclosed are pharmaceutical preparations in which the derivative and the drug are in intimately combined form. A cyclodextrin derivative having reduced hemolytic activity and its use as a drug agent are provided.

Description

DETAILED DESCRIPTION OF THE INVENTION

TECHNICAL FIELD The present invention relates to an acylated alkylated cyclodextrin derivative, a production method thereof, and use as a drug carrier.

(Conventional technology) Cyclodextrin (hereinafter also referred to as CyD) has a glucose residue α-1,4.
An oligosaccharide linked cyclically by a bond consisting of 6, 7, 8 glucose residues,
Those called α, β, and γ-CyD, respectively, are known. In addition, these Cy
A glucosyl group is α-1,6 bonded to one or two of the glucose units of D,
Alternatively, a so-called branched cyclodextrin in which a maltosyl group is α-1,6 linked (hereinafter, referred to as “branched cyclodextrin”)
Branched CyD) is also known.

[0003] These CyDs and branched CyDs have a high ability to include certain chemical substances, and in the fields of medicine, food and cosmetics, stabilization of unstable substances, retention of volatile substances, poor solubility in water or poor solubility in water. It is used for various purposes such as solubilization of insoluble substances.

[0004] In addition, a variety of CyD derivatives have been provided in order to utilize the physicochemical properties and inclusion ability of CyD as described above as a multifunctional drug carrier. However, in the course of these researches, it was pointed out that there were some problems in physical properties, inclusion ability, pharmacokinetics, economics, and the like, and it became clear that there were limitations on the uses thereof. In particular, drug carriers for injections and various mucosal preparations (eg, eye drops, suppositories, etc.) that require low hemolytic activity, low local irritation, and low tissue damage, ie, solubilizing agents (solubilizing agents) And its use as an agent for preventing tissue damage was extremely inconvenient.

On the other hand, α-CyD has a relatively high solubility in water of 14.5 g / 100 mL (25 ° C.), and has a lower hemolytic activity and muscle stimulating activity than β-CyD, There is a restriction that the target is limited to small molecules. Further, the price is about 30 times higher than that of β-CyD, and there is an economic disadvantage. Further, γ-CyD is the most excellent of α-, β- and γ-CyD in terms of safety such as hemolytic activity and tissue damage, and has the same inclusion ability as β-CyD, but the price is low. Since it is about 100 times as large as β-CyD, it is not used much for economic reasons. Further, glucosylated or maltosylated branched CyD is also of interest in that the water solubility is improved as compared to the corresponding unbranched CyD, but the behavior of the drug as a carrier as described above is as follows. Not always satisfactory.

[0006] Therefore, attempts have been made to improve physical properties and functionality by chemically modifying β-CyD, which is easily available. For example, heptakis (2,6-di-O-methyl) -β-CyD (hereinafter, DM-β-C) obtained by methylating the hydroxyl groups at positions 2 and 6 of glucose
Heptakis (2,3,2), in which all hydroxyl groups at positions 2, 3, and 6 are methylated.
6-tri-O-methyl) -β-CyD (hereinafter referred to as TM-β-CyD) and 2-hydroxypropyl-β-CyD (hereinafter HP) in which a hydroxypropyl group is mainly introduced into the hydroxyl group at the 6-position -Β-CyD) and the like. Szejtli et al. Suggest that DM-β-CyD can be used as a solubilizing agent for injection (J. Incl. Phenom., 1 (2) 135 (1983)).

[0007] However, DM-β-CyD is extremely soluble in water and has a strong inclusion force, but its solubility and stability constant are remarkably reduced at a high temperature side, and the dissociation of the drug from the drug inclusion complex is also easy. Therefore, there is a problem that the setting of sterilization conditions for the injection solution is hindered. Furthermore, DM-β-CyD has stronger hemolytic activity than β-CyD, and also has greater tissue damage upon intramuscular injection. This tendency is due to TM-β-C
The same applies to yD, and TM-β-CyD indicates an intermediate value between DM-β-CyD and β-CyD. On the other hand, with respect to HP-β-CyD, the solubility decrease and stability constant decrease at high temperature are greatly improved, and tissue damage such as hemolytic activity and muscle irritation is also considerably improved as compared with β-CyD. equivalent to α-CyD, natural Cy
At present, it is far from γ-CyD, which has the lowest hemolytic activity and muscle stimulating property among D.

[0008] Kamigama et al. Describe 2-hydroxyethyl-CyD having a 2-hydroxyethyl group introduced thereinto.
(Hereinafter referred to as HE-β-CyD) and 2,3-dihydroxypropyl-CyD
(Hereinafter referred to as DHP-β-CyD) is used as a drug carrier by utilizing its low hemolytic or hemolytic activity suppressing action, low tissue injury or tissue injury preventing action, low local irritation or local irritation reducing action. (JP-A-64-61430).
issue). However, the inhibitory action of HE-β-CyD and DHP-β-CyD on hemolysis is almost the same as that of γ-CyD, and further improvement will be desired for clinical use.

Also, Szejtli et al. Have proposed (carboxyl) alkyloxyalkyl derivatives of CyD and pharmaceutical compositions of them and drugs (WO 9).
2/14762). However, there is no specific description as to whether or not they exhibit a sufficient inhibitory action on hemolytic activity.

DISCLOSURE OF THE INVENTION Under such circumstances, it is particularly important to provide a drug carrier having low hemolysis. In other words, if a low hemolytic drug carrier capable of administering a poorly soluble drug parenterally can be provided, it is expected to maintain a high blood concentration even for a drug which has been considered inapplicable until now. This is because it is considered to make a great contribution to the field of drug therapy. Accordingly, it is an object of the present invention to provide a CyD derivative that satisfies the above-mentioned needs, and further provides a use of such a CyD derivative as a carrier or a delivery tool for a poorly soluble drug.

[0011] The present inventors have synthesized various CyD derivatives and studied their hemolysis inhibiting action in order to solve the above-mentioned problems. As a result, a CyD derivative having a significantly lower hemolytic activity than that of a CyD having an acyl group and an alkyl group in the molecule at the same time as compared with HE-β-CyD or γ-CyD, which was conventionally considered to have a low hemolytic activity. Was found. Furthermore, it was also confirmed that these derivatives sufficiently retained the drug inclusion ability of the corresponding CyD.

Thus, according to the present invention, there is provided an acylated alkylated CyD useful as a solubilizer, adsorbent or clathrate.

The acylated alkylated CyD according to the present invention is specifically represented by the formula (I)

[0014]

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(In the formula, n is any one of integers 6 to 8, wherein positions 1 and 4 of the sugar residues at both terminals are covalently bonded to each other, and R ¹ , R ² and R ³ are Glucosyl independently represents a hydrogen atom, a lower alkyl group or a C _2-20 alkanoyl group, or optionally a hydroxy group optionally substituted with a lower alkyloxy group or a C _2-20 alkanoyloxy group. Represents a group or a maltosyl group, provided that a total of 3 × n R ¹ , R ^2, and R ³ , each consisting of n, is at least one lower alkyl group and at least one acyl group at the same time. And the remaining group, if present, is a hydrogen atom, or up to two of said glucosyl or maltosyl groups) is an acylated alkylated cyclodextrin derivative. Such derivatives may be provided as compounds having different degrees of substitution of acyl groups and / or alkyl groups or partially epimerized compounds depending on starting materials, reaction conditions, and the like for producing them. It may be convenient to provide it in the form of a mixture.
In addition, the acylated alkylated CyD derivative sufficiently satisfies the object of the present invention even in the form of a mixture, and such a mixture is also provided by the present invention.

The acylated alkylated CyD derivative or a mixture of two or more of the derivatives can be efficiently produced by an acylation reaction using the corresponding partially alkylated CyD derivative as a starting material. Therefore, according to the present invention, there is also provided a method for producing such an acylated alkylated CyD.

[0017] As described above, the acylated alkylated CyD derivative or a mixture of two or more of the derivatives is derived from HE-, which has been conventionally regarded as having low hemolytic activity, even if it is derived from β-CyD. In addition to exhibiting significantly lower hemolytic activity than β-CyD and γ-CyD, β-CyD sufficiently retains the ability of β-CyD to inherently contain drugs.
In addition, mucosal irritation in rabbits with these acylated alkylated CyD derivatives is significantly lower than that of DM-β-CyD.

Thus, according to the present invention, there is provided the use of the above-mentioned acylated alkylated CyD derivative or a derivative thereof as a carrier or a delivery tool for a drug that is hardly soluble or insoluble in water. A specific embodiment of this use is as described above in the acylated alkylated Cy.
There is provided a pharmaceutical preparation in which the D derivative or a mixture of two or more of the derivatives and the above-mentioned drug are tightly mixed. Also provided are methods for preparing such pharmaceutical formulations.

(Detailed Description of the Invention) The term “acylated alkylation” as used in the present invention means a state in which an acyl group and an alkyl group are simultaneously present on the same molecule. Therefore, the acylated alkyl group CyD of the present invention
The derivative has at least one of the hydroxyl groups in the CyD molecule converted to an acyl ester and at least one of the other hydroxyl groups converted to an alkyl ether.

Such a CyD derivative having an acyl group and an alkyl group on the CyD molecule at the same time has a surprisingly significant decrease in hemolytic activity as compared with the corresponding CyD, as described above. However, from the viewpoint that the hemolytic activity is significantly reduced, in the above formula (I), a total of 2 × n (eg, 50% of 14 R ¹ and R ³ according to β-CyD (eg, 7 or more according to β-CyD) are lower alkyl groups, the remaining R ¹ and R ³ and R ² are at least one acyl group, and if present, the remaining R ¹ , R ^A CyD derivative or a mixture of two or more of these derivatives is preferably a hydrogen atom for ² and R ^3. More preferably, 50% or more, particularly about 100%, of a total of 2 × n R ¹ and R ³ is a lower alkyl group. And n
Examples thereof include a CyD derivative in which 50% or more, in particular, about 100% of R ² is a C _2-20 alkanoyl group, or a mixture of two or more of such derivatives.

In consideration of the harmony between the inclusion ability of the drug and the economy, n = 7 in the formula (I), that is, β
Preferred are those corresponding to -CyD and having no glucosyl group or maltosyl group as a branched sugar residue. Accordingly, even more preferred acylated alkylated CyD derivatives include 14 or more of R ¹ and R ³ are lower alkyl groups, and 4 or more of 7 R ² are C _2-20 alkanoyl groups. A certain β-CyD derivative or a mixture of two or more of the derivatives is mentioned. The mixture referred to here is, for example, R ¹
And a mixture of two or more compounds selected from the group consisting of compounds in which 7 to 14 of R ³ are lower alkyl groups. In this case, the number of R ² is a C _2-20 alkanoyl group can be constant or different.

However, most preferably, β-CyD derivatives wherein all of R ¹ and R ³ are lower alkyl groups and all of R ² are C _2-20 alkanoyl groups, and such derivatives are primarily (ie, , More than 50%).

The lower alkyl group includes a straight-chain or branched alkyl group having 1 to 6 carbon atoms, preferably, methyl, ethyl, n-propyl, iso.
-Propyl, n-butyl and sec-butyl groups, among which the most preferred is a methyl group.

In the above C _2-20 alkanoyl group, the alkyl portion may be linear or branched, but preferred are acetyl, n-propanoyl, n-butanoyl, n-pentanoyl (or valeryl), n-hexanoyl (or caproyl), n-heptanoyl (or enanthryl), n-octanoyl (or capryl), n-dodecanoyl (or lauryl), n-tetradecanoyl (or myristylyl) and n-octadecanoyl (or stearyl) And the like, among which acetyl, n-propyl, n-butanoyl and n-hexanoyl are preferred, of which the most preferred is the acetyl group.

Thus, an acylated alkylated CyD derivative according to the present invention or said derivative 2
A mixture of at least three kinds, particularly preferably, has a substitution degree (DS) at the 3-position of 7 or more.
Heptakis (2,6-di-O-methyl 3-acetyl) -β-CyD (hereinafter D
MA-β-CyD, the following abbreviations will be followed) and mixtures containing predominantly said DMA-β-CyD or acetylated DA-β with a low degree of substitution at position 3 (DS: 3.5-6) -CyD mixtures.

The acylated alkylated CyD of the present invention as described above is another embodiment of the present invention, which is represented by the formula (II):

[0027]

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(In the formula, n is any one of integers 6, 7, and 8, wherein the 1- and 4-positions of the sugar residues at both ends are covalently bonded to each other, and R ⁴ , R ⁵ and R ⁶ independently represents a hydrogen atom, a lower alkyl group, a glucosyl group or a maltosyl group, provided that a total of 3 × n R ⁴ , R ⁵ and R ⁶ consisting of n are simultaneously at least 1 Lower alkyl groups and at least one hydrogen atom, and a maximum of two glucosyl groups and maltosyl groups) or a mixture of such derivatives in a polar solvent. Reacting with the activated C _2-20 alkanoic acid in the presence of the agent,
Can be manufactured.

As the partially alkylated CyD derivative of the formula (II), those known per se or commercially available, or those produced according to a method known per se can be used. As the activated C _2-20 alkanoic acid, an acid anhydride or an acid halide (chloride, bromide) of an alkanoic acid can be used, and an acid anhydride is preferable. When an acid halide is used, it is preferable to use a basic organic amine such as triethylamine as a scavenger for hydrogen halide, but it is advantageous to use pyridine as a solvent and a scavenger or condensing agent for hydrogen halide. is there.

When an acid anhydride is used as an acylating reactant and pyridine is used as a solvent, an amount of pyridine in which a reactant of the compound of the formula (II) and the acid anhydride is sufficiently dissolved is used, usually around 80 ° C. By reacting at a temperature of several hours to 72 hours, a desired number of acyl groups can be introduced onto CyD. From the reaction mixture thus obtained, the objective acylated alkylated CyD derivative can be isolated and purified by using solvent extraction, various types of chromatography and recrystallization known per se. The mixture of two or more derivatives may be separated from the reaction solvent and unreacted reactants or by-products. Usually, after completion of the reaction, the reaction mixture is dropped into ice water to decompose excess acid anhydride, and then the desired CyD derivative is extracted with chloroform. The extract is added with sodium carbonate, desalted, separated and purified on a silica gel column, and if necessary, recrystallized from a suitable solvent. The obtained substance is concentrated to dryness to obtain the desired CyD derivative. Further, the structure of the obtained substance can be confirmed by a mass spectrum, elemental analysis and the like.

As described above, the acylated acetylated CyD derivative of the present invention or the mixture of the derivatives obtained as described above has a significantly reduced hemolytic activity and a lower room temperature than conventional CyDs as described above. And has an action of solubilizing a drug that is hardly soluble or substantially insoluble in water, and thus is useful as a carrier or delivery tool for such drugs.

Therefore, in another aspect of the present invention, the above-mentioned acylated alkylated CyD
There is provided a pharmaceutical preparation in which a derivative or a mixture of two or more of the derivatives and the above-mentioned drug are in a tightly mixed state. The “closely mixed state” means a state in which the CyD derivative and the drug are homogeneously mixed or a state in which the drug forms an inclusion complex with the CyD derivative.

The preparation in such a state can be prepared by suspending or dissolving a CyD derivative and a drug in an aqueous solvent (including a mixed solvent of methanol, ethanol, acetonitrile, dimethylformamide and the like and water). Can be prepared by sufficiently kneading using a kneader or the like commonly used for the preparation of

The above pharmaceutical preparations can be in the form of parenteral administration, that is, intravenous injection, intramuscular injection, subcutaneous injection, topical administration to skin or mucous membrane, etc., but are not limited thereto. However, an oral dosage form can be used.

The drug or active ingredient applicable to the preparation according to the present invention may be any drug, including water-soluble and poorly soluble, as long as the purpose of the present invention is met.
In general, it may be a drug that is hardly soluble or insoluble in water or an unstable drug.

The active ingredient has a local physiological effect, and, in the case of permeation through the mucous membrane or in the case of oral administration, has an effect on the whole body after being transported to the gastrointestinal tract together with saliva. Dosage forms prepared from the conjugates according to the invention are particularly suitable for those in which the active ingredient is active over a long period of time (ie, a drug with a half-life of at least several hours). For example, analgesics and anti-inflammatory agents (NSAIDs, flurbiprofen, fentanyl, indomethacin, ketoprofen, napmetone, paracetamol, piroxicam, tramadol), antiarrhythmic agents (procainamide, quinidine, verapamil), antibacterial agents and protozoal agents (amoxylin, ampicillin, benzab) Timpenicillin, Benzylpenicillin, Cefaclor, Cefadroxil, Cefprozil,
Cefuroxime acetill, cephalexin, chloramphenicol, chloroquine, ciprofloxacin, clarithromycin, clavulanic acid, clindamycin, doxycycline, erythromycin, flucloxacillin, halofantaline, isoniazid, kanamycin, lincomycin , Mefloquine, minocycline, nafcillin, neomycin, norfloxacin, ofloxacin, oxacillin, phenoxymethylpenicillin, pyrimethamine sulfadoxime, streptomycin), blood coagulant (warfarin), antidepressants (amitributyline, amoxapine, butriptyline, clomipramine, clomipramine, clamipramine Doxepin, fluoxetine, gepirone, imipramine, lithium carbonate, mianserin, milnacipura , Nortriptyline, paroxetine, sertraline, 3-
[2- [3,4-dihydrobenzofuro [3,2-c] pyridin-2 (1H) -yl] ethyl] -2-methyl-4H-pyrido [1,2-a] pyrimidin-4-one)
), Diabetes drugs (glibenclamide, metformin), antiepileptic drugs (carbamazepine, clonazepam, ethosuximide, phenobarbital, phenytoin,
Primidone, topiramate, valpromide), antifungal agents (amphotericin B, chlorimazole, econazole, fluconazole, flucytosine, griseofulvin, itraconazole, ketoconazole, miconazole nitrate, nystatin, terbinafine, policonizoledin, astrominazole, econazole, econazole, econazole, econazole, econazole, econazole, econazole, econazole, estrazole, econazole, estrazole, estrazole, econazole, estrazole) , Fexofenadine,
Lunarizine, levocabastine, loratadine, norastemizole, oxatomide, promethazine, terfenadine), antihypertensives (captolyl, enalapril, ketanserin, lisinopril, minoxididine, prazosin, ramipril, reserpine, terazosin), antimuscarinic agents (atropine sulfate, antihistin) Viral agents (acyclovir, AZT, ddc, ddI, ganciclovir, roviride, tivirabine, 3TC, delavirdine, indinavir, nelfinavir, ritonavir, saquinavir), antitumor agents (adriamycin, cladribine, cisplatin,
Dactinomycin, daunorubicin, doxorubicin, etoposide, irinotecan, mitomycin, mitoxantrone, tamoxifen, taxol, taxotere, topotecan, trimetrexate, vincristine, vinblastine), migraine drugs (almotriptan, arniditan, eletriptan, frovatan) , Naratriptan, rizatriptan, sumatributane, zolmitributane), Parkinson's disease drugs (promocribtin mesylate, levodopa, selegiline)
, Antipsychotics (alprozolam, buspirone, chlordiazepoxide, chlorpramazine, diazepam, flupentixol, fluphenazine, flurazepam,
9-hydroxyrisperidone, lorazepam, mazapertin, olanzapine, oxazepam, pimozide, pipamperone, piracetam, promazine, risperidone,
Cellhotel, selocale, sertindole, sulpiride, temazepam, thiothixene, triazolam, trifluperidol, ziprasidone, zolpidem), anti-stroke (ruberzol, ruberzol oxide, riluzole, aptiganel,
Eliprodil, remasemide), antitussives (dextromethorphan, lepodropropidine), β-blockers (atenolol, carbediol, metoprolol, nebivolol, prapanol), inotropic agents (amrinone, digitoxin, digoxin, milrinone), adrenal hormones (propion) Beclomethasone acid, betamethasone, budesonide, dexamethasone, hydrocortisone, methylblednisolone, prednisolone, prednin, triamcinolone), disinfectant (chlorhexidine), diuretic (acetazolamide, fursemide, hydrochlorothiazide, isosorbide), enzyme (lysozyme, lysozyme, lysozyme, lysozyme, lysozyme) (Anethole, anise oil, caraway, cardamom, cassia oil, cineole, clove oil, cilantro oil, dementholmi Oil, inondo oil, eucalyptus oil, eugenol, ginger oil, lemon oil, mustard oil, neroli oil, nutmeg oil, orange oil, peppermint, sage, spearmint, teripineol, thyme, digestive system drugs (cimetidine, cisapride, cleboprid) , Diphenoxylate, dombergidone, famotidine, lansoprazole, loveramide, loperamide oxide, mesalazine,
Metoclopramide, mosapride, nizatidine, norcisapride, olsalazine,
Omeprazole, pantoprazole, perprazole, prucalpride, ranitidine, rabeprazole, lidogrel, sulfasalazine), hemostatic agent (aminocaproic acid), hyperlipidemic agent (atorvastatin, lovastatin, pravastatin, probucol, simvastatin), local anesthetic (benzocaine, Lignocaine),
Opioid analgesics (buprenorphine, codeine, dextromoramide, dihydrocodeine, hydrocodone, oxycodone, morphine), parasympathetic stimulants (epatastigmine, galantamine, metrifonate, neostigmine, physostigmine, tacrine, donepezil, rivastigmine, mirinstigrin, mivastigrin, mivastigmine, mirabigline, mevaline) , Talsacridine, xanomeline, memantine, lazabemide), sex hormones (estrogen inclusions for estrogens, ethinylestradiol, mestranol, estradiol, estriol, estriol, estrone, progestogens chlormadinone acetate, mestranol acetate, 17-deacetylnorgestiate) Mate, desogestrail, dienogest, dydrogesterone, ethinodiol acetate, gestodene 3-ketodesogestrail, levonorgestrail, linenestrenol, metroxyprogesterone acetate, megestrol, norethindrone, norethindrone acetate, norethisterone, norethisterone acetate, norethinidrone, norethynidrone acetate, norethisterone, norethisterone acetate, norethynodrel, norgestinolel Norgestrienone, progesterone, kinggestanol acetate, stimulant (sildenafil), vasodilator (amlodibin, bufromezil, amyl nitrate, zirdiazem, dipyridamole, nitroglycerin, isosorbide dinitrate, lidofrazine, molsidomine, nicardipine, nifedipine, oxypentofylline , Erythritol nitrate)
Is mentioned.

In the preparation according to the present invention, the compounding ratio of the acylated alkylated CyD and the drug can be any ratio as long as it meets the purpose, but from the viewpoint of controlling the release of the drug from the preparation. The acylated alkylated CyD to drug can be selected from 1: 4 to 4: 1, preferably 1: 2 to 2: 1 on a molar ratio basis.

[0038] The preparation of the present invention may further contain, if necessary, other pharmaceutically acceptable auxiliaries or additives in a range that does not adversely affect the purpose of the present invention. Such auxiliaries or additives include stabilizers, solubilizers, suspending agents, emulsifiers, buffers, preservatives, isotonic agents, or other suitable additives commonly used in the art. Agents can be mentioned.

Hereinafter, the present invention will be described specifically with reference to examples. Note that the present invention is not limited to these examples.

Example 1 Synthesis of DMA-β-CyD DM-β-CyD (12 g) was dissolved in 60 mL of anhydrous pyridine, and 25 mg of 4-dimethylaminopyridine was added. Further, 12 mL of acetic anhydride was slowly dropped, and the reaction was performed at 80 ° C. for 24 hours. After completion of the reaction, the mixture was dropped into ice water to decompose excess acid anhydride and extracted with chloroform. The extract was added with sodium carbonate, desalted, separated and purified by a silica gel column. The obtained substance was concentrated to dryness to obtain the desired DMA-β-CyD (DS.7). This CyD derivative has a melting point of 113 to 117 ° C. and a solubility in water at 25 ° C. of more than 50 mg / dl. The resulting DMA-β-CyD (DS.7) was recrystallized from water to give white crystals having a melting point of 126 ° C. The results of the mass spectrum and the ¹ H-NMR spectrum are shown in FIGS. 1 and 2, respectively.

FAB MS (negative mode) m / z 1777 [M + m-nitrobenzyl alcohol (matrix) -H); ¹ H-NMR (CDCl ₃ ) d 5.16 (
t, 1H, CyDH-3), 5.00 (d, 1H, CyDH-1), 3.9.
1-3.87 (m, 2H, CyDH-5 and H-6b), 3.79 (t
, 1H, CyDH-4), 3.54 (d, 1H, CyDH-6a), 3.3.
7 (s, 3H, 6- CH 3), 3.33 (s, 3H, 2-CH 3), 3.21 (d
d, 1H, CyDH-2), 2.04 (s, 3H, 3-CH).

Example 2 Synthesis of acetylated DM-β-CyD having a low degree of substitution at the 3-position A small amount of acetic anhydride (4 g) was added to DM-β-CyD (10 g, 7.5 mmol).
. (6 g, 45 mmol) to produce acetylated DM-β-CyD. The acetylated DM-β-CyD obtained is various D. S. Thus, recrystallization was not carried out, but other production conditions were the same as those described in Example 1. D. S. The value was determined by the peak ratio between the anomeric proton (H-1) of CyD and the methyl proton of the acetyl group in the ¹ H-NMR spectrogram (see FIG. 3), and was 3.8. Hereafter, this mixture is referred to as DMA4-β-Cy
It is called D.

Example 3 Synthesis of butyrylated DM-β-CyD DM-β-CyD (5 g) was dissolved in 25 mL of anhydrous pyridine, 9 mL of n-butyric anhydride was added, and the mixture was reacted at 80 ° C. for 24 hours. After completion of the reaction, the reaction mixture was dropped into ice water to decompose excess acid anhydride and extracted with chloroform. The extract was added with sodium carbonate, desalted, separated and purified by a silica gel column.
The obtained substance was concentrated to dryness to obtain the target butyrylated DM-β-CyD. This C
The melting point of the yD derivative was 108 to 111 ° C. FIG. 4 shows the results of ¹ H-NMR.

Example 4 Synthesis of Octanoylated DM-β-CyD DM-β-CyD (5 g) was dissolved in 25 mL of anhydrous pyridine, 16 mL of octanoic anhydride was added, and the mixture was reacted at 80 ° C. for 24 hours. After completion of the reaction, the reaction mixture was dropped into ice water to decompose excess acid anhydride and extracted with chloroform.
The extract was added with sodium carbonate, desalted, separated and purified by a silica gel column. The obtained substance was concentrated to dryness to obtain the desired octanoylated DM-β-CyD. This CyD derivative is an oil at 25 ° C. FIG. 5 shows the result of ¹ H-NMR. Characteristic test: (1) Measurement of stability constant of DMA-β-CyD In this test, the inclusion characteristic of DMA-β-CyD was determined by comparing the inclusion properties of its raw materials, β-CyD,
It was compared with those of DM-β-CyD and TM-β-CyD.

The stability constants of DMA-β-CyD and flurbiprofen were determined by the solubility method (ie, Higuchi et al., Adv. Anal. Chem.
Inatr. 1965, 4, 117-212. ). For comparison,
In addition, the stability constants of β-CyD, DM-β-CyD, TM-β-CyD and flurbiprofen were determined, and are summarized in Table 1.

[0046]

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Table 1: Compound Stability constant (M ^-1 ) β-CyD 3613 DM-β-CyD 8055 TM-β-CyD 1655 DMA-β-CyD 1212 From these results, the inclusion ability of DMA-β-CyD is DM-β-CyD. Inferior to that of CyD, but almost equal to the inclusion capacity of TM-β-CyD. (2) Hemolytic test of DMA-β-CyD and DMA4-β-CyD In the hemolytic test, 6 to 7 mL of blood was collected from the auricular vein of a white rabbit, and the erythrocyte preservation solution 1 was collected.
mL was added and mixed gently. To this is added a 10 mM isotonic phosphate buffer (pH 7.4).
4) 4-5 mL was added, mixed gently, centrifuged at 1000 g for 5 minutes, and the supernatant was removed. This washing operation was repeated three times, and 10 mL of 10 mM isotonic phosphate buffer (pH 7.4) was added to 1 mL of this solution to make 20 mL, thereby preparing a 5% erythrocyte suspension.

Various concentrations of CyD derivatives were diluted with 10 mM isotonic phosphate buffer (pH 7.4) and incubated at 37 ° C. 4 mL of this solution was taken, 0.2 mL of a 5% erythrocyte suspension was added, and the mixture was incubated at 37 ° C. for 30 minutes. After centrifugation at 1,000 g for 5 minutes, the absorbance of 3 mL of the supernatant at 543 nm was measured to determine the hemolytic activity. The specimen was visually observed using a microscope. The results obtained are shown in FIG. 100 mM DMA-β-CyD (DS7) in 5% erythrocyte suspension
, DMA4-β-CyD (DS 3.8), β-CyD, DM-β-CyD,
TM-β-CyD, 2-HP-β-CyD (DS 4.8) (Shiotan
i, K. et al., Pharm. Res. 1995, 12, 78-84,
) Or sulfobutyl ether β-CyD (DS.3.5) (see the above-mentioned literature), and the above treatment was carried out.

The haemolytic activity of DMA-β-CyD is determined by β-CyD, DM-β-CyD and TM
Considerable weaker than the hemolytic activity of -β-CyD. For example, β-CyD,
DM-β-CyD and TM-β-CyD were approximately 2 mM and 0.5 mM, respectively.
At 1 and 1 mM hemolysis began, and the concentrations resulting in 50% hemolysis were 4 mM, 1 mM and 2 mM, respectively. On the other hand, DMA4-β-CyD (D.
S. The hemolysis of 3.8) started at about 12 mM, and its 50% hemolysis concentration was about 22 mM. In the case of DMA-β-CyD (DS.7), no hemolysis was observed up to 100 mM. The hemolytic activity of this DMA-β-CyD was determined to be 2-HP-β-CyD (
D. S. 4.8) and the sulfobutyl ether of β-CyD (DS 3.5)
Lower than the hemolytic activity of (3) Complete rabbits treated with DMA-β-CyD or DMA4-β-CyD
Measurement of the amount of free cholesterol from whole erythrocytes DMA-β-CyD, DMA4-β-CyD, and DM-β-CyD were each diluted with a 10 mM isotonic phosphate buffer (pH 7.4), and then diluted at 37 ° C. Incubated. Take 4 mL of these solutions, add 0.2 mL of 5% erythrocyte suspension, and add
Incubated at 30 ° C for 30 minutes. After centrifugation at 1000 g for 5 minutes, the supernatant 3
5 mL of chloroform was added to the mL, and the mixture was extracted with shaking for 30 minutes. These chloroform layers were separated and concentrated to obtain samples. The cholesterol level of this sample was measured using cholesterol E-Test Wako (manufactured by Wako Pure Chemical Industries). FIG. 7 shows the obtained result.

One of the causes of CyD-induced hemolysis is known to be in the extraction of erythrocyte cholesterol and phospholipids through the formation of inclusion complexes. Therefore, in this test, DMA-β-CyD or DMA4- The release behavior of cholesterol from rabbit erythrocytes by the addition of β-CyD was examined, and a control (equivalent solution containing no CyDs) was performed.
And DM-β-CyD were compared with the cholesterol release behavior. FIG.
2 shows the amount of cholesterol released from intact rabbit erythrocytes treated with β-CyDs in 10 mM phosphate buffer (pH 7.4) at 10 ° C. DM-β-CyD induced about 80% release of cholesterol at a concentration of 0.5 mM (see FIG. 6) where only slight hemolysis occurred. In contrast, DMA-β-CyD elicited only 10% release of cholesterol at the same concentration, which was the same as the value of control experiments performed with isotonic buffer.

[Brief description of the drawings]

FIG. 1 shows a mass spectrum (matrix: DMA-β-CyD) of DMA-β-CyD obtained in Example 1.
Methanol, glycerol and m-nitrobenzyl alcohol, and so on).

FIG. 2 is a ¹ H-NMR spectrum of DMA-β-CyD obtained in Example 1.

FIG. 3 is a ¹ H-NMR spectrum of DMA4-β-CyD obtained in Example 2.

FIG. 4 is a ¹ H-NMR spectrum of butyrylated DM-β-CyD obtained in Example 3.

FIG. 5 is a ¹ H-NMR spectrum of octanoylated DM-β-CyD obtained in Example 4.

FIG. 6 shows the results of a hemolytic activity test of various CyD derivatives. In FIG. 6, a white square (□), a black triangle (▲), a white triangle (△), a white circle (〇), a black circle (●), a white inverted triangle (
▽) and open diamonds indicate DMA-β-CyD, DMA4-β-CyD,
β-CyD, DM-β-CyD, TM-β-CyD, 2HP-β-CyD (D.
S. 4.8) and β-CyD sulfobutyl ether (DS 3.5).

FIG. 7 is a graph showing the amount of free cholesterol from complete erythrocytes when erythrocytes are brought into contact with various CyD derivatives. The vertical axis indicates the cholesterol level of the whole red blood cell as 1
The free cholesterol amount (%) is shown as 00%.

──────────────────────────────────────────────────続 き Continuation of front page (81) Designated country EP (AT, BE, CH, CY, DE, DK, ES, FI, FR, GB, GR, IE, IT, LU, MC, NL, PT, SE ), OA (BF, BJ, CF, CG, CI, CM, GA, GN, GW, ML, MR, NE, SN, TD, TG), AP (GH, GM, KE, LS, MW, SD, SL, SZ, UG, ZW), EA (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM), AE, AL, AM, AT, AU, AZ, BA, BB, BG, BR , BY, CA, CH, CN, CU, CZ, DE, DK, EE, ES, FI, GB, GD, GE, GH, GM, HR, HU, ID, IL, IN, IS , JP, KE, KG, KR, KZ, LC, LK, LR, LS, LT, LU, LV, MD, MG, MK, MN, MW, MX, NO, NZ, PL, PT, RO, RU, SD, SE, SG, SI, SK, SL, TJ, TM, TR, TT, UA, UG, US, UZ, VN, YU, ZA, ZW (72) Inventor Akira Kondo Minami Isshiki, Nagaizumi-cho, Sunto-gun, Shizuoka 600-8 Jansen Kyowa Co., Ltd.Fuji Research Laboratory (72) Inventor Hiroshi Kawaji 600-8 Jansen Kyowa Co., Ltd.Fuji Research Laboratory (72) Inventor Masaaki Ota Nagatsumicho, Nagatomachi, Sunto-gun, Shizuoka 600-8 Yansen Kyowa Co., Ltd. Fuji Research Institute (72) Inventor Yasuhiro Okamoto 600-8 Jansen Kyowa Co., Ltd. Fuji Research Laboratory F-term (reference) 4C076 CC01 CC04 CC05 CC11 EE39 FF11 4C090 AA02 BA11 BD03 CA46 DA09 DA23

Claims (13)

[Claims] 1. A compound of the formula (I) (In the formula, n is any of integers 6, 7 and 8, wherein the 1- and 4-positions of the sugar residues at both ends are covalently bonded to each other, and R ¹ , R ² and R ³ are each independently Represents a hydrogen atom, a lower alkyl group or a C _2-20 alkanoyl group, or a glucosyl group optionally substituted with a lower alkyloxy group or a C _2-20 alkanoyloxy group for a hydroxyl group. Or a maltosyl group, provided that any one of a total of 3 × n R ¹ , R ² and R ³ consisting of n is at least one lower alkyl group and at least one C _2-20 An acylated alkylated cyclodextrin derivative represented by the formula: or two of said derivatives, which is an alkanoyl group, and the remaining group, if present, is a hydrogen atom or up to two glucosyl groups or maltosyl groups. Mixed over object. 2. 50% or more of a total of 2 × n R ¹ and R ³ are lower alkyl groups, and the remaining R ¹ and R ³ and R ² are at least one C _2-20 alkanoyl group. and if present, the remaining R ^1, R ² and R ³ according to claim 1 acylation alkylated cyclodextrin derivative or the derivative mixture of two or more, wherein a hydrogen atom. Wherein in which R ¹ and R ³ is a lower alkyl group, wherein if more than 50% of R ² is C _{2 -20} alkanoyl groups, and present, the remaining R ² is a hydrogen atom Item 4. The acylated alkylated cyclodextrin derivative according to item 1, or a mixture of two or more of such derivatives. 4. The acylated alkylated cyclodextrin derivative according to claim ^1, wherein R ¹ and R ³ are lower alkyl groups, and 50% or more of the n R ² groups are acyl groups. mixture. 5. The lower alkyl group is methyl, ethyl, n-propyl, iso-
Propyl, n-butyl and sec-butyl, wherein the alkanoyl group is acetyl,
n-propanoyl, n-butanoyl, n-pentanoyl, n-hexanoyl, n
-Heptanoyl, n-octanoyl, n-dodecanoyl, n-tetradecanoyl and n-octadecanoyl. 5. The acylated alkylated cyclodextrin derivative according to claim 1, or a mixture of two or more of the derivatives. . 6. The acylated alkylated cyclodextrin derivative according to claim 1, wherein the lower alkyl group is a methyl group and the alkanoyl group is an acetyl group, or a mixture of two or more of the derivatives. 7. The acylated alkylated cyclodextrin derivative according to claim 1, wherein n in the formula (I) is 7, or a mixture of two or more of such derivatives. 8. Heptakis (2,6-di-O-methyl-3-O-acetyl)
-Β-cyclodextrin, heptakis (2,6-di-O-methyl-3-O-butyryl) -β-cyclodextrin, or heptakis (2,6-di-O-methyl-3-O-otenoyl)- β-cyclodextrin. 9. A mixture of acetylated heptakis (2,6-di-O-methyl) -β-cyclodextrins having an average degree of acetyl substitution at the 3-position of about 3.8. 10. A process for producing an acylated alkylated cyclodextrin derivative represented by the formula (I) according to claim 1 or a mixture of two or more of the derivatives, wherein the formula (II) is (In the formula, n is any one of integers 6, 7 and 8, wherein the 1- and 4-positions of the sugar residues at both terminals are covalently bonded to each other, and R ⁴ , R ⁵ and R ⁶ are Independently represents a hydrogen atom, a lower alkyl group, a glucosyl group or a maltosyl group, provided that a total of 3 × n R ⁴ , R ⁵ and R ⁶ consisting of n are simultaneously at least one lower An alkyl group and at least one hydrogen atom, and a maximum of two glucosyl groups and maltosyl groups) or a mixture of such derivatives in a polar solvent, optionally with a condensing agent. Reacting with an activated C _2-20 alkanoic acid in the presence of to acylate the compound of formula (II). 11. A pharmaceutical preparation, wherein the drug is a compound of the acylated alkylated cyclodextrin derivative according to any one of claims 1 to 9 or a mixture of two or more of the derivatives and a drug. Formulation. 12. The drug according to claim 11, wherein the drug is selected from the group consisting of non-steroidal rheumatic drugs, steroids, cardiac glycosides, benzodiazepine derivatives, benzimidazole derivatives, piperidine derivatives, piperazine derivatives, imidazole derivatives and triazole derivatives. Pharmaceutical preparations. 13. A kneaded mixture of the acylated alkylated cyclodextrin derivative or a mixture of two or more of the derivatives according to claim 1 with a drug in an aqueous solvent, and then, if necessary, removing the solvent. The method for preparing a pharmaceutical preparation according to claim 11 or 12, wherein the preparation is removed.