GB2151203A

GB2151203A - Compositions for making vesicles

Info

Publication number: GB2151203A
Application number: GB08422629A
Authority: GB
Inventors: Hiromichi Takahashi; Kaoru Tsuji
Original assignee: Kao Corp
Current assignee: Kao Corp
Priority date: 1983-09-29
Filing date: 1984-09-07
Publication date: 1985-07-17
Also published as: FR2552666B1; GB8422629D0; DE3435517A1; JPS6072830A; JPH0326165B2; FR2552666A1; GB2151203B

Abstract

Vesicular compositions comprise the ingredients (A): a phosphate salt of formula (I> <IMAGE> in which R1 and R2 are respectively C6-C24 hydrocarbon groups and M is for example an alkali metal; and (B) one or more surface active agents selected from specific groups.

Description

SPECIFICATION Vesicular compositions iJ Field of the invention: This invention relates to vesicular compositions and more particularly, to vesicular compositions which are able to form stable vesicles over a long term.

ii) Description of the prior art: It is well known that phospholipids, particularly lecithin, which are important constituents of biomembrane form in water double-layered hollow vesicles called liposome. This liposome is a hollow lipid double-layered sphere and can contain various chemical substances in the lumen thereof and thus has a close resemblance with red cells regard to the structure. In this sense, the liposome has been studied as a model of red cell or a cellular model and plays an important role in the studies of biomembrane.

In recent years, attention has been paid to the liposome as a carrier in vivo of drugs. More particularly, the liposome which is able to contain various chemical substances in the lumen thereof may be regarded as a kind of capsule. It has been reported that when a drug is dosed after included in the liposome, the metabolism in vivo of the drug is suppressed and the drug is kept in the living body over a long term, sustaining its medical efficacy (see, for example FEBS Letters, Vol, 36. No. 3, page 292, 1973), side effects of the drug, e.g. allergy, are suppressed (see, for example, FEBS Letters, Vol.45, No. 1, page 71, 1974), and distribution of the drug into various organs changes (see, for example, Eur. J. Biochem. Vol.47, page 179, 1974).

As noted above, liposome exhibits good properties as a carrier in vivo of drug and particularly, its property of changing the distribution of drug into organs has the possibiility of enabling a drug to be selectively acted on affected organs, attracting the attention as the so-called target effect with liposome. For instance, most anticancer drugs involve side effects because they act not only on cancer cells, but also on healthy normal cells. If it is possible that an anticancer drug is selectively acted on cancerous cells administering it after incorporation into liposome, this mode of administration is considered to be very useful. In practice, some attempts give good results (for example, Collection of Summaries of Lectures to the Meeting of the Japanese Cancer Society, page 8, (1976).

On the other hand, a suitable conductive material such as alkylamine is embedded in the double-layered portion while placing a photoreductive substance such as cupric ions into the internal liquid of the liposome and photooxidative substance such as ascorbic acid into the external liquid of the liposome. Subsequently, when such liposome is irradiated with light, it becomes possible to condense a specific type of ion from the external to internal liquids depending on the property of the conductive material embedded in the double-layered membrane. For instance, it is possible to collect, as resources, specific types of elemental ions from sea water.

As noted above, liposome will bring about an epoch-making, new technique such as in administration of drugs. However, because phospholipids capable of forming the liposome are living body-derived substances, there are great limitations on their chemical structures, so that there are disadvantages such as inconvenience in imparting various functions thereto and relatively poor chemical stability.

In order to eliminate such limitations on the phospholipid liposome, studies have been recently made to form liposome-type vesicles from synthetic surface active agents. It has been confirmed up to now that several kinds of surfactant have the ability of forming vesicle. Such surface active agent vesicles have the double-layered hollow vesicular structure similar to liposome and can thus be applicable as a carrier for drug as noted before.

However, known vesicles of surface active agents are all prepared by dispersing surface active agents in water and irradiating the dispersion, for example, with ultrasonic wave. Because the resulting vesicular structure is unstable, the vesicles cannot be preserved over a long term. More particularly, where dialkyl phosphate used in the present invention are used to obtain vesicles according to the above-described known method, the vesicular structure is broken within several days with the vesicle solution becoming turbid or gelled.

For utilization of the surface active agent vesicles as the drug carrier or for other purposes, it would be essential to improve the stability and thus there is a strong demand of developing techniques for the improvement.

Summary of the invention In view of the above circumstance, the present inventors have made intensive studies to obtain vesicles stable over a long term and, as a result, found that vesicles obtained from a system of a specific type of phosphoric acid ester having the formabilty of vesicle, a kind of surface active agent and water is stable over a long term. The present invention is accomplished on the basis of the above finding.

Accordingly, an object of this invention is to provide a vesicular composition which comprises the following two ingredients (A) and (B), the ratio by weight of (A) and (B) ingredients being in the range of 100:1 to 100:100.

(A) a phosphate salt of the general formula (I)

in which R1 and R2: hydrocarbon group having from 6 to 24 carbon atoms, and M: alkali metal, alkanolammonium, tetraalkylammonium hydroxide, lysine, alginine, histidine or morpholine, (B) one or more surface active agents selected from the group consisting of compounds of the formulas (a) to (w), R3COOM (a)

R3OSO3M (d) R3O(AO)nSO3M (e)

R3O(AO)nH (h)

HO(AO)p(C3HsO)q(AO)rH (j)

R3COO(AO)nR6 (I)

AOH () R3CON (r) AOH r

in which, R3: hydrocarbon group having from 6 to 36 carbon atoms, R4 and R5: hydrocarbon group having from 5 to 23 carbon atoms, Re, R7 and R8: hydrogen atom or acyl group having from 6 to 24 carbon atoms provided that if R7 and R8 are both contained in one molecule, at least one of them is hydrogen atom, Rg and R10: hydrogen atom or hydrocarbon group having from 6 to 24 carbon atoms, at least either one of them being hydrogen, R11, R12 and R13: hydrocarbon group having from 1 to 24 carbon atoms, R14: hydrocarbon group having from 9 to 23 carbon atoms, R15: hydrocarbon group or hydroxyhydrocarbon group having from 1 to 24 carbon atoms, R16 and R17: hydrocarbon group or hydroxyhydrocarbon group having from 1 to 4 carbon atoms, or benzyl group, A: alkylene group having from 2 to 4 carbon atoms, M: same meaning as defined before, I, m and n: 0 oran integeroffrom 1 to 150, p, q and r: integer of from 1 to 150, t, integer of from 1 to 4.

It is known that the phosphate salt (I), which is (A) ingredient, forms vesicles as reported, for example, in J. Am. Chem. Soc., Vol. 101, page 2231 (1979). However, it is not known that the vesicles are kept stable over a long term by incorporating (B) ingredient into (A) ingredient. This has been uncovered for the first time by the present inventors.

Detailed description of the invention and preferred embodiments The phosphate salt being (A) ingredient of the present invention can be obtained by neutralizing a phosphate represented by the formula (I')

in which R1 and R2 have, respectively, the same meanings as defined before, with a base compound. The base compounds used for the neutralization preferably include alkali metal-containing base compounds such as sodium hydroxide, sodium carbonate and the like, alkanolamines, tetraalkylammonium hydroxides, lysine, alginine, histidine, and morpholine. Among alkanolamines, there are more preferably used those having alkanol moieties such as monoethanol, diethanol, triethanol, monopropanol, dipropanol and tripropanol. More preferable tetraalkylammonium hydroxides are those having alkyl moieties such as methyl, ethyl, propyi, butyl and pentyl.Examples of (A) ingredient include didecyi phosphate triethanolamine salt, didodecyl phosphate triethanolamine salt, dihexadecyl phosphate triethanolamine salt, dioleyl phosphate triethanolamine salt, dioctadecyl phosphate sodium salt, dioctadecyl phosphate monoethanolamine salt, dioctadecyl phosphate diethanolamine salt, dioctadecyl phosphate diisopropanolamine salt, dioctadecyl phosphate tetramethylammonium hydroxide salt, dioctadecyl phosphate lysine salt, doictadecyl phosphate alginine salt, dioctadecyl phosphate morpholine salt, and the like. The two hydrophobic chains (R1, R2) of these phosphate salts may have unsaturated bonds, but are preferred to be saturated hydrocarbon groups.

The surface active agents which are used as (B) ingredient in the practice of the invention are considered to have the action of stabilizing vesicles. The surface active agents enumerated in (a) through (g) are anionic active agents, those agents of (h) through (s) are nonionic active agents, and those agents of (t) through (w) are amphoteric active agents. In these (B) ingredients, the alkylene oxide added to the nonionic active agent of (h) through (s) is preferably ethylene oxide.

In the vesicular composition of the invention, the mixing ratio by weight of (A) and (B) ingredients is important and should be in the range offrom 100:1 to 100:100. Outside the above range, no vesicles are formed, or even if vesicles are formed, they are unstable.

For the preparation of the vesicular composition according to the invention, (A) and (B) ingredients are dissolved in solvents capable of dissolving both ingredients, followed by agitating to obtain a uniform solution and removing the solvent therefrom according to any known procedures.

In order to obtain vesicles from the thus obtained vesicular composition of the invention, it is sufficient to suspend the vesicular composition in water and subject the suspension to ultrasonic irradiation. Aside from the above procedure, vesicle solutions may be obtained, for example, by a procedure in which the vesicular composition is dissolved in a solvent soluble in water such as ethanol and the aqueous solution is intensely injected into water, or by a procedure in which the vesicular composition is solubilized using aqueous surface active agents and vesicles are formed while removing the surface active agent by dialysis.

The vesicle solution prepared from the vesicular composition of the invention should preferably have a concentration of from 1 to 50 wt% (hereinafter referred to simply as %), more preferably from 5 to 30%.

When the concentration exceeds 50%, its viscosity becomes too high, which may cause disadvantages in the preparation and use of vesicles. In case where the concentration is less than 1%, no troubles in the preparation and in the use take place but the costs of transportation and container for the vesicle solution increase, thus being not economical.

The most reliable method, currently known in the art, of confirming vesicles in the thus prepared vesicle solution is an electron-microscopic observation using a negative staining technique. The negative staining technique is a method in which the electron density of the hydrophilic moieties of surface active agents capable of forming vesicles is increased by means of phosphotungstic acid or uranyl acetate and the moieties are stained black. In the practice of the invention, formation of vesicles was observed through the microscopic observation. The vesicle solution containing vesicles are transparent with good fluidity. On the other hand, the solution in which the compounds used do not form vesicles but give a multi-layered structure is gelled and opaque with very poor fluidity.Accordingly, in the stability test of vesicles (described hereinafter in detail), it is possible to judge the stability of a vesicular composition by preparing a vesicle solution by the ultrasonic method and observing transparency and fluidity of the solution as a function of time. One of known supplementary simple methods of confirming vesicles is a NMR method. The relaxation time or width of absorption line of 1H- or 3C-NMR of vesicles is largely different from that of cases other than vesicles. More particularly, with vesicles, the relaxation time is prolonged with a narrow, sharp width of absorption line. In cases other than vesicles, the relation time is short with a large width of absorption line.

However, the presence of the double-layered structure by which vesicles are characterized cannot be directly confirmed by the NMR method. In order to strictly confirm the presence of vesicles, the observation through an electron microscope should be made.

The present invention is illustrated in more detail by way of examples, which should not be construed as limiting the present invention.

Example 1 Vesicular compositions were prepared from phosphate salts and various surface active agents indicated in Table 1 to check formation and stability of vesicles.

[Preparation of vesicularcompositions] Ten grams of a respective phosphate salt dissolved in ethanol and 1 g of a surface active agent were mixed and agitated until a uniform solution was obtained. Subsequently, the ethanol was removed using an evaporator to obtain a powder of the vesicular composition.

[Formation and confirmation of vesicles] Ten grams of the resulting vesicular composition in the form of powder was admixed with 90 g of water, followed by agitating to obtain a viscous, opaque gel-like composition. The gel-like composition was kept at 60 C and subjected to ultrasonic irradiation at 100W and 25 KHz for about 1 hour. The resulting solution was observed through an electron microscope by which formation of vesicles was confirmed. It will be noted that solutions in which vesicles were formed were substantially transparent and fluid.

[Stability test of vesicles] The vesicle solutions thus obtained were preserved for 3 months in a thermostatic chamber of 20"C. The stability was evaluated by comparing the state of each solution after 3 months with the state thereof immediately after the preparation with regard to transparency and fluidity. The evaluation standards for the formation and stability of vesicles are as follows.

A. The state after the preservation is completely same as the state immediately after the preparation with the vesicular structure being perfectly kept.

B: As compared with the solution immediately after the preparation, only a slight increase of viscosity is recognized with the vesicular structure being substantially perfectly kept.

C: The solution increases in viscosity and is considerable in opacity with little vesicular structure existing.

D: The solution immediately after the preparation is completely turned into a gelled and opaque with no vesicular structure being recognized.

*: Even though the ultrasonic irradiation is effected, no vesicles are formed.

TABLE 1 Results of Observation Surface Surface Surface Surface Active Active Active Active Agent* Agent 1 Agent2 Agent3 Phosphate salt Didecyl phosphate triethanolamine salt A A B Didoceyl phosphate triethanolamine salt A A B Ditetradecyl phosphate triethanolamine salt A A A Dihexadecyl phosphate triethanolamine salt B A A Dioctadecyl phosphate triethanolamine salt B A A Dieicocyl phosphate triethanolamine salt B B B Dioleyl phosphate triethanolamine salt B A A *:Surface active agents Surface active agent 1

(dodecyl sulfate triethanolamine salt) Surface active agent 2 C18H3-O-(CH2CH2O)1 00H (polyoxyethylene oleyl ether, N'=100) Surface active agent 3

(distearyl aminimide) Comparative Example 1 10 g of each of phosphate salts indicated in Table 1 was dissolved in 90 g of water, followed by forming vesicles according to the procedure of Example 1. The vesicle solutions were preserved in a thermostatic chamber of 20"C for 3 months to check the state thereof after the preservation. In all cases of the seven phosphate salts, vesicles were found to be formed, but after the 3 month-preservation, the solutions were turned gelled and opaque with a loss of the vesicular structure.

Example 2 Dioctadecyl phosphate triethanolamine salt and surface active agents indicated in Table 2 were used to prepare vesicular compositions according to the procedure of Example 1. The stability of vesicles obtained from the compositions were determined. The results are shown in Table 2.

TABLE 2 Results of Surface Active Agent Observation Sodium stearate B Sodium linear nonylbenzenesulfonate B Dodecylphosphate triethanolamine salt A Dodecylsulfate treithanolamine salt B Polyoxethylene dodecyl ether sulfate triethanolamine salt (n=8) A Sodium polyoxyethylene nonylphenyl ether sulfate B Sodium distearylsulfosuccinate B Polyoxyethylene stearyl ether (n=10) A Polyoxyethylene nonylphenyl ether (n=10) A Stearyl alcohol B Polyoxyethylene polyoxypropylene block polymer (p=10, q=10, r=10) B Polyoxyethylene sorbitan stearate (I +m+n=5) B Sorbitan stearate B Polyoxyethylene stearate (n=7) A Polyoxyethylene glycerine stearate (I +m +n=8) A Stearic acid monoglyceride A Stearyl glyceryl ether A Pentaerythritol stearate A Polyoxyethylene stearylamine (m+n=20) A Stearic acid diethanolamide A Stearyl dimethylamine oxide A Distearyl aminimide A Stearyl dimethylcarbobetaine (t=1) A Sodium dodecylaminocarboxylate (t= 1) B Stearyl dimethylsulfobetaine (t=3) A Example 3 Fifteen phosphate salts in total indicated in Table 3 and the three surface active agents used in Example 1 were used in combination to prepare 45 vesicular compositions according to the procedure of Example 1 in order to check the stability of vesicles obtained from the respective compositions.

The results are shown in Table 3.

TABLE 3 Results of Observation Surface Surface Surface Surface Active Active Active Active Agent Agent Agent2 Agent3 Phosphate Salt Sodium dioctadecyl phosphate B B B Potassium dioctadecyl phosphate B B B Dioctadecyl phosphate monoethanolamine salt B A A Dioctadecyl phosphate diethanolaminesalt B A A Dioctadecyl phosphate triethanolamine salt B A A Dioctadecyl phosphate monoisopropanolaminesalt B A A Dioctadecyl phosphate diisopropanolaminesalt A A A Dioctadecyl phosphate triisopropanolaminesalt A B A Dioctadecyl phosphate tetramethylammonium hydroxide salt B A A Dioctadecyl phosphate tetraethylammonium hydroxide salt B A A Dioctadecyl phosphate tetrapropylamminium hydroxide salt A B A Dioctadecyl phosphate tetrabutylammonium hydroxide salt A B A Dioctadecyl phosphate lysinesalt B A B Dioctadecyl phosphate alginine salt B A B Dioctadecyl phosphate morpholine salt B B B Example 4 Dioctadecyl phosphate triethanolamine salt and the surface active agents used in Example 1 were mixed in varied ratios indicated in Table 4, followed by preparing 39 vesicular compositions in total according to the procedure of Example 1. The compositions were used to determine their formation and stability of vesicles.

The results are shown in Table 4.

TABLE 4 Results of Observation Surface Active Mixing Ratio Agent (by weight) Surface Surface Surface Active Active Active Dioctadecyl Agent 1 Agent2 Agent3 Phosphate Surface Triethanolamine Active Salt Agent 100/0.5 D C C 100/1 B B B 100/1.25 B B B 100/1.43 B A B 100/1.67 B A A 100/2 B A A 100/2.5 B A A 100/5 B A A 100/10 B A A 100/20 A A A 100/50 A A A 100/100 B B A 100/200 * * *

Claims

1. A vesicular composition comprising the following two ingredients (A) and (B), the ratio of (A) ingredient to (B) ingredient ranging from 100:1 to 100:100, (A) a phosphate salt of the general formula (I)

R2OSO3M (d) R3O(AO)nSO3M (e)

R3(AO)nH (h)

HO(AO)p(C3H6O)q(AO)rH (j)

R3COO(AO)nR6 (I)

in which, R3: hydrocarbon group having from 6 to 36 carbon atoms, R4 and R5: hydrocarbon group having from 5 to 23 carbon atoms, R6, R7 and R8: hydrogen atom or acyl group having from 6 to 24 carbon atoms provided that if R7 and P8 are both contained in one molecule, at least one of them is hydrogen atom, R8 and Rlo: hydrogen atom or hydrocarbon group having from 6 to 24 carbon atoms, at least either one of them being hydrogen, R11, R12 and R13: hydrocarbon group having from 1 to 24 carbon atoms, R14: hydrocarbon group having from 9 to 23 carbon atoms, R15: hydrocarbon group or hydroxyhydrocarbon group having from 1 to 24 carbon atoms, R16 and R17: hydrocarbon group or hydroxyhydrocarbon group having from 1 to 4 carbon atoms, or benzyl group, A: alkylene group having from 2 to 4 carbon atoms, M: same meaning as defined before, I, m and n: 0 or an integer of from 1 to 150, p,q and r: integer of from 1 to 150, t: integer of from 1 to 4.

2. The composition according to Claim 1,whereinthealkanolammonium in the group M of (B) ingredient is monoethanolammonium, diethanolammonium, triethanolammonium, monoisopropanolammonium, diisopropanolammonium or triisopropanolammonium, and the tetraalkylammonium is tetramethylammonium, tetraethylammonium, tetrapropylammonium, tetrabutylammonium ortetrapentylammonium.

3. A composition as claimed in claim 1 and substantially as described in any one of the specific examples hereinbefore set forth.

4. Each and every novel embodiment herein set forth either separately or in any combination.