JPS6081192A - Phospholipid compound - Google Patents

Abstract

NEW MATERIAL:A compound of formula I (R is H or methyl; n is an integer 8-16; m is an integer 13-21). EXAMPLE:rac-1-[( 2-Methacryloyloxy )dodecanoyl]-2-O-stearyl-glycero-3-phosphocholine of formula II. USE:A material for obtaining chemically and physically stable microcapsules not subjected to hydrolysis with lipase. PREPARATION:An omega-hydroxyalkanoic acid of formula III, e.g. 2-phenyl-5-m- dioxanol, is reacted with stearyl bromide, etc. to give an omega-(meth)acryloyloxyalkanoic acid of formula IV, e.g. 12-(methacryloyloxy)dodecanoic acid, which is then reacted with succinyl chloride and converted into an acid chloride of formula V. The resultant acid cloride is then condensed with a glycerol-2-alkyl ether of formula VI in the presence of a dehydrochlorinating agent to afford a condensation product of formula VII, which is further converted into phosphochlorine by the conventional method.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to phospholipid compounds, and particularly to phosphatidylcholine type phospholipid compounds having a polymerizable group.

At present, various attempts are being made to provide pharmaceutical substances and 111% additives in microscopic capsules in the form of 8 products for each person.

Like this 7. The 49th method of c microcapsule ratio is:
Encapsulation of 1-3 molecular compounds by emulsification method (Japanese Patent Publication No. 45-2757 and Japanese Patent Publication No. 17950-1973) and interface 49. The main reaction is the lG1'i combination reaction, +13 combination (
This is encapsulation (Japanese Patent Publication No. 7395/1983 and Japanese Patent Publication No. 7396/1983) which has increased the production of polyamide (polyamide, etc.) by 1,000 times. However, in these methods, for example, when a safe organic solvent (e.g., nooxane, acetone, xylene, benzene) is added to the capsule during the synthesis process, Toxicity due to remaining capsules and large particle size (several μm)
to 1000 μm), which poses a problem in that it may cause problems such as blood clots, making it difficult to use the obtained capsules as medical products.

By the way, the main purpose of encapsulating pharmaceutical substances and microcapsules is to maintain the activity of pharmaceutical substances and enzymes that are unstable in vivo for a long period of time. therefore,! Medicine for samurai 5. When aiming to use microcapsules as a four-product product, the following conditions are particularly important, including the problems of the conventional techniques described above. In other words, (1) to reduce the size of microcapsule particles (several μm or less), (2) to improve the stability of capsules in vivo, and (3) to improve the characteristics of capsule usage. It is to give a relative's name.

As a technique to add ii4 to the degree of sufficiency Vc of the above conditions, various phospholipids, which are components of biological membranes, form minute spherical aggregates (liposomes, particle size U, 05 to 1 ()μ) in water.
7n) Taking advantage of the total formation, first synthesize a phospholipid compound having a polymerizable group, and polymerize the liposome thus formed while maintaining its original shape.
Techniques for obtaining microcapsules have been recently reported. For example, S, L.

Journal of the A by Regen et al.
inerical Chemical 5ociety
, 104.791-795 (1982) (hereinafter referred to as document (1)), 14-Il, An by Hub et al.
gewantqCltemie International
onal Edition English version, Dan, 938-94
0 (1980) (hereinafter referred to as literature (11)), and Angewante e by A. Akimoto et al.
hemie International Editi
on English 1fη version, main 0.9O-91 (1981) (hereinafter referred to as document (110)).

The synthetic phospholipid compound forms liposomes (or vehicles) in water. In order to microencapsulate this liposome as it is, the polymerization group in each compound (i.e., vinyl group in the compound of Reference (1), diacetylene group in the compound of Reference (II), diacetylene group in the compound of Reference (II), In the compound, butadiene group) is polymerized.

However, when considering that these microcapsules are to be administered into a living body as part of a pharmaceutical product, it is necessary that the microcapsules G are not only physically stable but also chemically stable. In particular, when manufacturing microcapsules using phospholipid derivatives, chemical decomposition of microcapsules due to the hydrolysis reaction of the phosphorus J1m acyl moiety, which is induced by various glues iR, is suppressed. It is necessary to devise ways to do so.

However, conventional phospholipid compounds have not been developed in consideration of this point, and greater chemical stability is desired.

By the way, one of the decomposition processes of phosphorus protein by linose is the hydrolysis of the acyl group at the 2-position of glycerol.
:1'-. Therefore, it is essential that the acyl ester moiety at position 2 be made into a +(9) structure that is not subject to hydrolytic attack by lipase.

Therefore, the purpose of this coating is to provide microcapsules that are physically stable and resistant to hydrolysis by lipase.]1? An object of the present invention is to provide a phospholipid compound having a structure.

The present inventors investigated phosphatitulcholine-type phosphorus J1 base compounds and found that glycerin-2-alkyl ether-type compounds are more stable against decomposition by lipase! Therefore, the present invention was developed.

That is, according to this 9 ming, 6; il, general formula % formula %) (where R is a hydrogen atom or a methyl group, n is an integer of 8 to 16, m is an integer of 13 to 21). Phosphatidylcholine type phospholipid compounds are provided.

The phosphatidylcholine type phospholipid compound of the present invention represented by the above W+ general formula (1) can be synthesized, for example, by the following process. That is, the formula 0 R obtained by reacting an ω-hydroxyalkanoic acid represented by the formula % formula % () (where ゎ is the same as above) with methacryloyl chloride or acryloyl chloride (however, R and R are the same as above) The ω-methacryloyloxyalkanoic acid compound represented by
is converted into the acid chloride represented by the formula, or 2 moles thereof are converted to the acid anhydride represented by the formula by reacting with dicyclohexylcarboimide (hereinafter abbreviated as DCC) etc. It is activated by KotoK. Next, the compound side or (b) is the glycerol-
The 2-alkyl ether is reacted with Kon1 to obtain a condensation product represented by the formula %2. This condensation involves combining the compound of formula (Q or I) and the compound of formula (6) in a molar ratio of 0.5:1.
or 3:1 ratio, and an appropriate amount of dehydrochlorination agent (pyridine, p-(LN-dimethylamino)giricin, etc.)
in a suitable solvent (e.g., anhydrous tetrahydrofuran, chloroform, carbon tetrachloride, etc.) containing
Perform at a temperature of 0°C. The reaction time is usually 1 hour to 20 hours.

Finally, the condensation product of formula (111) is converted to Yon R@Hi
Pharm, Acta He1v-+ by rt et al.
33 + 349 (1958) or the method described in Te
trahedr on Letters+23.104
The desired phosphatidylcholine type phospholipid compound of the general formula (I) can be obtained by the reaction according to the method described in 3 (1982).

The phospholipid compounds of the present invention can be prepared using common methods for obtaining liposomes from natural phospholipids (e.g., XD, Pa
Bioehimi by pahadjopoulos et al.
a et BiophyaLa Acta+ 135
+639-652 (1967), etc.) to form liposomes with a particle size of 0.02-1μ in water. The ease of forming this liposome is determined by m,
The larger the value of n, the higher the value. The thus obtained liposome dispersion aqueous solution is exposed to all ultraviolet light and ultraviolet monochromatic light (for example, 240
When irradiated with light (nm), the polymerization reaction proceeds rapidly due to K, and the microcar f-+f is irradiated. Small Mai
Ro91de7ru1. - Since it is composed of tt7 by a polymer, it is physically stable, and the phosphatinorcholine type phosphorus compound of the general formula (1), which is a monomer of the polymer, is susceptible to hydrolysis by lipase. Hateful(
・Because it has a %7 structure, it is chemically stable. Suitable as a carrier.

Examples of this invention will be described below.

Example 1 (A) J, Amer by H, Hibbert et al.
Cbem.

Soc, 62.1601-1613 (1929) 2-phenyl-5-m-nooxanol fl) obtained according to the method described in Arnold et al., Jug, L.
ieb.

Ann, Chem, +709, 234-239 (19
67), 2-stearylJ-ruoxy-1,3-deronogundiol was synthesized by reacting with stearyl bromide.

Elemental analysis value (weight): H12,84 (12,8)
C73,75 (73,3) (However, the value in parentheses is C21) i440? Calculated value of i) 13C-NMR spectrum δ (ppm) i11j: 1.
3 61.9 2 79.7 4 70.2 5 31.9 6 26.1 20 22.7 21 14.1 1R spectrum (KBr pellet) (,,-”):
3350.2940.2870.1470.1120.
1080.980 (B) 12-(methacryloyloxy)dodecane branch 5.
θg of oxalyl chloride in anhydrous CC42Ome.
5 ml and 1 tl dimethylformamide (DMF)
After reacting for 1.5 hours in the presence of ijl, the solvent was distilled off under reduced pressure. The residue was dried under pressure and the corresponding acid chloride "kT8" (IR Niji co(cocj, ) 1800
cm). After dissolving this in 3 oml of dry chloroform, it was added to a solution of 5.1 g of 2-stearyloxy-1,3-zolobanediol dissolved in a mixed solvent of 30 me of dry chloroform and 2.5 ml of dry pyridine at room temperature. After dropping, the mixture was stirred overnight. This reaction mixture'(c
After sequentially washing with 5N-HCl, water, 3T Na2CO3 and water, it was dehydrated with Na25Q4. This was purified using a silica dull column (diameter 3-1 length 35 cm) (effluent solvent: benzene/ether = 9/1). After collecting the target part and distilling off the bath medium, the residue was removed from benzene and freeze-dried. Yield: 3.8 grams.

'Thin) Gj chromatography (TLC): I also f =
0.22 (Bromthymol blue test J, !: ) Elemental analysis value (nail), (φ): ) 1 11.45 (11,55) C72,91 (72,74) (Values in parentheses are CsyllyaOtr CIHi-j
:61υ, 956)ill >-1irz)13C-
NMR spectrum (CDCL3) '1, 3 62.
0,62.7 4' 14.12 77.7 5' 2
2.7 4 125.0 6 ' 31.9 5 13 (3,5 618,320' 26.0 7 167.4 21' 70.5 8 64.8 9-16.7'-16' 28.6 -3
0.017 24.9 18 34.4 19 173.7 IR spectrum (mouth).

(CHCl3 solution) 3550.2940.2870.1720, 1640° 1470.1330, 1305, 1175.1115 (
C) Cl3F(0)OCH2Cf(211r'4.
5 Durum and dry chloroform 15me were cooled with water, and 1-(12-(methacryloyloxy)
rdecanyl) glycerol-2-stearyl ether (
A) 1.5 grams (2,501 moles) of bow j! 7. Dry chloroform solution/Ij, (i 5 rng ) and dry triethylamine 6 ml (chloroform 9 ml)
And 8 separate! ] Dou and Shi are down 1.1 at the same time. I11
After completing 5 hours, the mixture was left to cool with water or at room temperature overnight. After cooling on ice, 15 types of 0.5 N-KC1 aqueous solution were slowly added to irA+, and then air pumped for 2 hours. After adding 15 ili of methanol to this, the liquid was separated, and the charcoal layer was collected. After washing this organic layer with water, it was evaporated, and the remaining solid content was dried under reduced pressure on P2O5. The reaction mixture was reacted with 30 ral of dry butanone and 20 ml of trimethylamine at 50° C. for 4 hours. The reaction mixture was evaporated and the residue was dried under reduced pressure over tPO. After dissolving in 100 ml of methanol, 3 ml of silver acetate was added. After stirring in the dark for 1.5 hours, it was filtered.The filtrate was subjected to evaporation, and the remaining solid Kokin Silica Dull column was purified (effluent solvent = 1st cfIc
t5/methanol/water = 65/25/4.2nd cfx
ct3/methanol=3/L 1/1.1/4.1/9
). Yield 0.66 grams.

TLC (7 Likal"k: CHCl3/Mep)-
/l//Water=65/25/4) was in line with the value of egg yolk lecithin (Sikuma Co., Ltd.).

Elemental analysis value: N 1.64 (1,78) ((il-1 in parentheses is 8209NI Pl ('tF
I″-it; 788.097) value) 11′C-
NMR spectrum (CDCL5/CD50D) 1.3
G4.1 9 25.0 2 77.0 10 64.9 4 59.0 11 167.3 5 66.3 12 136.5 G 54.3 13 125.3 7 174.0 14 18.3 8 34. 3 15 70.2 Charcoal A'ruY No. δ (r'pH1) 组 1'a No. δ
(ppm) 16 26.1 19 14.1 17 32.0 (~, monkey) 29.3~30.018
22.7 IR spectrum, IL/ (an-1) (CHCl5
i? ? ? B',) '2940.2870.1725
,”1640.1470.1305.1230 (wide)
, 118 f), 1095.1060.975.720 14th Patent Attorney: Suzu 4 [Takehiko, Commissioner of the Patent Office, Kazuo Wakasugi, 1. ! Display of 1 yen Patent Application No. 188325/1988 2 Name of the invention Phospholipid Compound 3 Relationship with the case of the person making the amendment Patent applicant's place of origin: Shun Hide 4, Agent address: 1-chome Toranomon, Minato-ku, Tokyo 1, No. 26 No. 5, No. 17 Mori Building 6, Specification to be amended 7゛, Contents of amendment (1) Specification Section, page 10, line 10, ``Example (2) Table on page 15 of the specification. "r26.o" below the title fδ (ppm) in the right column of r26. , lj.

(3) Detailed hot water temperature "r9-16.7'-16'" under the title 1' Carbon number in the right column of the table on page 15.
Correct it to -16,7°-19'J.

(4) Between ")" and "Collection JIX" in page 16, line 19 [[] of the specification, rac-1-[12-(
methacryloyloxy)dodecanoyl]-'2-0-stearyl-glycero-3-phosphocholine was obtained. ” is inserted.

(5) The chemical formula at the beginning of page 17, line 6 of the specification is corrected as follows.

(6) Insert the following sentence in a new line after the opening of page 18, line 8 of the specification.

"X/A According to (A-1) of Example 1, 2-phenyl-
5-m-dioxanol and myristylcica bromide) and the compounds listed in 1C were synthesized. Yield 29%.

Elemental analysis value (wt%): H12,65 (12,58) C70,48 (70,78) (In parentheses (1α is the calculated value of C, H, O,) "C-
N f'vl R spectrum δ (ppm) i + fj (C
DCl2, TMS): 1.3 61.9 2 79.6 4 70.2 5 31.9 6 26.0 7-15 29.3-30.0 16 22.7 17 14.1 Mass spectrum: M" +1=289 (molecular weight 289) (
B1-[12-methacryloyloxy dodecanoid Example 1 The corresponding acid 1 prepared from 2.5 grams of 12-(methacryloyloxy)dodecane by the method of (B)
Example 1 (B)
1-[12-(methacryloyloxy)dodecamyl]glycerol myristyl ether-2 was obtained by reaction and purification according to the following procedure. No. Yield 2.2 grams. Yield 46%.

Thin layer chromatography (TLC): Rf = O,
+8 (BTB■, benzene/ether = 9/1) IR spec) Le (cm): 3800.2940, 2875
．． 1720.1840゜ (CHCI, Error 'A) 1470.1330, 1305.11?5, 11151
3C-NMR SHE') l・Jl/ (CDCI,,
8 (ppm)), ``car door δ m ′! l large i′M
-δ ml, 3 B2.0, 62.7 4' 14.1
2 77.7 5 ° 22.7 4 125.0 6 ° 31,8 5 136.5 16 ° 26.1 G 18.3 1 7 ° 70.5 7 167.4 1O-IS,? '-15'28.8-
30.08 84.8 17'24.9 ・,゛1 8 34.2 1 9 173.7 9 2B, 0 Example 1 According to the method of (C), l-[12-(methacryloyloxy)dodecanyl]- The above compound was synthesized from 1.0 g of glycerol-myristyl ether.

Yield: 0.16 grams (12% yield).

TLC (silica gel, chloroform/methanol/wood =
The Rf value of 65/25/4) is egg yolk lecithin (Sigma)
matched the value of

'3C-NMR (CD C13, TMS) 1.3 64
．． 0 12 167.3 2 77.0 13 +38.5 4 59.0 14 125.0 5 66.3 15 +8.3 fi 54.4 II (70,5 7173,7'172e, 1 8 34.3 18 31.'9 9 25.0 19 22.7 11 1114.8 20 14.1 Carbon number 1 δ (m ′, ″'shigohi δ m10 2
B, OA, 8 28.7-30.1 Elemental analysis value: N 1
．． 88 (1,95) (values in parentheses are C, H, N,
(Calculated value of 09P) Example 3 0.1 g of the phospholipid compound synthesized in Example 1 was suspended in 5 ml of heavy water and subjected to ultrasonic treatment (under water cooling, under nitrogen atmosphere) to achieve almost uniform dispersion. An aqueous solution was obtained.

Transfer this solution to a quartz glass cell, replace it with nitrogen gas,
It was sealed tightly. This was irradiated with light using a low pressure mercury lamp in a 50'C bath. Two hours after the light irradiation, the formation of a polymer was confirmed for the next measurement. That is, 'H-N M R1
11, it was observed that the absorption of vinyl groups based on the phospholipid compound disappeared in the light irradiated liquid, and a polymerization reaction was confirmed. In addition, the singlet peak based on the choline methyl group was split into two trees (4δ = 0.22 ppm) by adding 10 mM Eu(No,)3a [1020].
The front and back structures of the liposomes were confirmed. Furthermore, the radius of the generated posome was calculated to be approximately 17 OA from the area ratio of the two peaks.

Example 4 0.1 g of the phospholipid compound synthesized in Example 2 was polymerized according to the method of Example 3, and the structure was confirmed in the same manner.

8 g (50 mmol) of 12-hydroxydodecane liquor was dissolved in a mixture of 150 ml of dry tetrahydrofuran and 5.5 g of dry pyridine, then cooled on ice and stirred. Add to this 75ml of dry tetrahydrofuran
After slowly dropping 4.5 g of acryloyl chloride dissolved in , the mixture was stirred at room temperature overnight. The reaction mixture was filtered to remove insoluble matter, and the filtrate was dried under reduced pressure. The residue was purified with a silica gel column (elution solvent: acetic acid ethyl ester/petroleum ether = 1/1), yield 5.2 g (yield + 38%), TLC (silica gel, acetic acid ethyl ester/petroleum ether = 1/1 BTB) ): Rf=0.
20 Elemental analysis value (weight%): H10,01 (9, B9)
CB8.20 (66,84) The value in parentheses is C,, H2604 (molecular weight 270.368
) calculated value 12-(acryloyloxy)dodecanoic acid 2
．． A solution of 12-(acryloyloxy)dodecanoic acid chloride synthesized from 5 grams (9 mmol) and 2.5 grams of silicyl acid chloride in 20 ml of dry chloroform was mixed with 2-stearyloxy-1,3-propanediol 2. 6 grams (7.5 mmol) in dry chloroform (
201) and dry pyridine (1.3ml) in a mixed solvent, the mixture was slowly added dropwise and stirred overnight. This was washed with 5N hydrochloric acid, water, 30% sodium carbonate, and water in this order, and then dried over anhydrous sodium sulfate. After filtering this, the filtrate was dried under reduced pressure, and the residue was purified with a silica gel column (elution solvent: benzene/ether, = 9/l). Yield: 1.2 grams (27% yield).

TLC (silica gel, benzene/ether = 9/1.B
TB■): Rf=0.1 Infrared absorption spectrum (chloroform solution): 3450,
2940.2870.1725.1640.1Ei20
゜1470, 1415.1300.12B5.1200
．． 1115°106G, 1190. 905 cm-
Single element analysis value (weight%): H11, B2 (11,4
8,) C72,08 (72,44) The value in parentheses is C76”490G (molecular weight 596.92
9) Calculated value of “C-N MR spectrum (CDCI,
): 9, 1-5-double,, δ 11 Charcoal=9 Ichiban-1'-
δ(pml, 3 62.0, 82.7 4' 14.1
2 77.7 5° 22.7 4 128, B 6' 31.9 5 130.3 7' 28.1 6 1813.2 8° 70.5 7 64.7 ■, ■ 28.8-30.09 24.
9 10 34.2 11 173.7 8 26.0 Example 1 According to the method of (C), l-(12-(acryloyloxy)dodecanoyl]glycerol stearyl ether-2 obtained in Example 5(B) was treated with phosphocholine. Yield: 33%. TLC (silica gel, chloroform/
The Rf value of methanol/water = 65/25/4) matched that of egg yolk lecithin (Sigma).

Elemental analysis value (claw part:%): N 1.75 (1,81)
Values in parentheses are C4, H,. N+ o, p, (molecular weight 7
74.070) Calculated value of “C-N MR spectrum (CDC13, TMS):
1.3 64.0 8' 26.0 2 77.1 9' 24.13 4 58.1 10' 34.3 5 8B, 3 11' 173.7 6 54.3 4" 14.0 4' 128 .8 5" 22.7 5' 130.4 8" 31.9 6' IH, 37" 2B, 1 7° 84.8 8" 70.4 'IjLlii meeting δ m ■, ■ 28.5-30. 1 c!1 (7) Replace "16" on the second line of page 18 of the specification with r16.
Correct it as 10'J.

Claims (1)

[Claims] A phosphatidylcholine type phospholipid compound represented by the general formula % (where lt is a hydrogen atom or a methyl group, n is an integer of 8 to 16, and m is an integer of 13 to 21).