JPS59101489A - Iron-porphine complex having one phosphorylcholine group - Google Patents

Abstract

NEW MATERIAL:The compound of formula I (n is 1-20). EXAMPLE:Iron(III)-5, 10, 15, 20-{tri(alpha, alpha, alpha-o-pivalamidophenyl)-alpha-o-[12-(2'-trimethylaminoethyl )phosphoryloxy-2, 2-dimethyldodecanamido ]phenyl }-porphyrin complex. USE:Agent for adsorption and desorption of oxygen. PROCESS:The objective compound can be prepared, e.g. by (1) reacting the carboxylic acid chloride of formula II with 5,10,15,20-tetra alpha,alpha,alpha,alpha-o-aminophenyl) porphine and pivaloyl chloride to obtain the compound of formula III, (2) reacting the compound with anhydrous aluminum chloride in the presence of anisole, (3) reacting the obtained compound of formula IV with ferrous bromide in the presence of pyridine, and (4) reacting the resultant compound of formula Vwith 2-chloro-2-oxo-1,3,2-dioxaphosphorane and trimethylamine. The compound of formula II is also novel, and is prepared by reacting omega-benzyloxyalkyl halide with dilithium anion of 2-methylpropionic acid, and reacting the product with thionyl chloride.

Description

Detailed Description of the Invention The present invention relates to iron-5, 10, 15 iron-5, 10, 15 having one phosphorylcholine group represented by the general formula .

20-[α, α, α, α-0-(Substituted amido)phenyl]phorphine complex.

Many iron-porphyrin complexes that can form oxygen complexes in organic solvents at room temperature have been reported.

Especially J, P, Col1man et al.\Journal o
f the American Chemical 5o
ciety r 97 + 1427 (1975)
Iron(II) -5,10,15,20 reported in
The -tetra[α,α,α,α-0-(pivalamido)phenyl]porphine complex is useful in that it forms a very stable oxygen complex in anhydrous aprotic solvents, but in the presence of even small amounts of water. The disadvantage is that it oxidizes quickly. As a useful oxygen adsorbing and desorbing agent, especially when considering application to living organisms, an iron-porphyrin complex that effectively adsorbs and desorbs oxygen in water at room temperature or 37° C. is desired. E, Hase
-gawa et al.% Biochemical and B
lophysicalResearchCommun
icationa * 105 y 1416 (19
Iron (
II) -5,10,15,20-tetra [α, α, α
, α-o-(-paramido)phenyl]porphine complex, it has become possible to generate an oxygen complex from a room source in an aqueous medium. However, although this iron-porphyrin complex has a great stabilizing effect on the oxygen complex due to its four bipalamide groups, it has a drawback in terms of the amount that can be embedded in liposomes due to its structure.

The purpose of the present invention is to obtain iron-5,10,15,20-tetra [α
.

To provide an iron-porphyrin complex that can be easily embedded in liposomes made of phospholipids while retaining its oxygen adsorption/desorption function. be. A general type that has a partial structure of a vivaramide group necessary to obtain a stable oxygen complex, and also has a phosphorylcholine group similar to the hydrophilic part of natural or synthetic phospholipids, especially phosphatinorcholine, as a hydrophilic part at its terminal end. This object can be achieved by the novel compound represented by formula (1). The compound of the present invention has a tetraphenylporphyrin core containing three bipalamide groups as a hydrophobic part and a terminal phosphorylcholine part as a hydrophilic part. Similar embedding with hydrophobic parts and hydrophilic parts such as phosphorylcholine parts is facilitated. Moreover, it is a very useful compound in that it has no effect on the stability of oxygen complexes.

According to the present invention, the novel iron-tetraphenylporphine complex represented by the general formula I is prepared by converting 1 equivalent of a carboxylic acid chloride represented by the formula ■H3 (where n is an integer from 1 to 20) into a known 5.10 , 15.20-tetra [
α, α, α, α-〇-aminophenyl) porphine (hereinafter abbreviated as H2TamPP) 1 equivalent and further reacted with 3 equivalents of piparoyl chloride to obtain 2■ (where n is defined earlier) ) is reacted with anhydrous aluminum chloride in the presence of anisole, and the resulting compound represented by the formula ■CR5 1 (where n is the same as defined above) is reacted with pyridine. 2-chloro-2-oxo-2-chloro-2-oxo- P can be produced by reacting with 1°3,2-dioxaphosphorane and further reacting with excess trimethylamine.

The starting material of general formula (1) is a new compound, which was synthesized by the following method. Ge ω-benzyloxyalkyl halide
orge R, Nevrkome et al.
g, 1975.517. 2 generated according to the report of
-Solithium anion of methylpropionic acid is first heated at low temperature (-70℃ to -20℃) and then heated to 30 to 45℃.
The reaction was carried out at ℃. The reaction mixture was then decomposed with cold dilute hydrochloric acid,
The crude product extracted with a solvent is recrystallized in a non-polar solvent such as petroleum ether, n-hexane, n-hebutane to obtain ω-
Pencyloxy-2,2-dimethylalkanoic acid was obtained as colorless crystals. This is reacted with an excess amount of thionyl chloride in a nonpolar solvent, preferably benzene or carbon tetrachloride, or in the absence of a solvent, and then concentrated under reduced pressure. Acid chloride was obtained. The ω-penzyloxyalkylno-1lide represented by the formula of the raw material ■ (where n has the same meaning as defined above, and ``g'' represents chlorine or bromine) is given by n-1'x'=ct. If A,
J, Hi 11 et al., Journal of the A
merican Chemical 5ociety
+48+257 (1926), as reported by
If n = 2, X' = Br, then S, Creme
r et al. Journal of theAmerica
n Chemical 5ociety + 86
+4197 (1964). Also, for n-3 to 20, X'=Br, α, ω
- Obtained by refluxing a diglomoalkane with one equivalent of sodium benzyl oxide in benzene.

One equivalent of carboxylic acid chloride of general formula H is added to known H2Ta
mPP (J, P, Col 1man et al., Jour
nal of the Amertcan Chemical
al 5ociety + 97 r 1427 (19
75)) in an anhydrous aprotic solvent, preferably tetrahydrofuran, chloroform, nochloromethane, N,N-trimethylformamide or acetone, containing an excess of base, preferably bilysin, triethylamine, 4- After reacting in the presence of (N,N-dimethylamino)pyridine for 1 to 24 hours at 0°C to room temperature, 3 equivalents or more of piparoyl chloride was added and reacted for 1 to 24 hours at room temperature. . The reaction solution was poured into water and extracted with ether, chloroform or dichloromethane. After washing with water, the separated organic solvent was dried with Glauber's salt and evaporated to dryness. The resulting residue was purified by silica gel chromatography to obtain a compound represented by the general formula (■). A compound was obtained.

To remove the benzyl protecting group of the cono compound, an excess of anhydrous aluminum chloride was added in a mixed solvent of sochloromethane di-dinitromethane in the presence of an excess amount of anisole at a temperature of -5°C to 30°C, preferably 15°C to 25°C. The reaction was allowed to proceed for 1 to 12 hours. The reaction mixture was poured into ice water and extracted with chloroform. The extract was washed with water and then with a quaternary sodium bicarbonate aqueous solution. The separated organic solvent layer was dried with Glauber's salt, and the residue obtained by evaporating the chloroform was subjected to silica gel chromatography. or recrystallization from a suitable solvent such as benzene, petroleum ether, ether, methanol, or a mixed solvent thereof, the alcohol compound represented by the general formula (2) was obtained.

To introduce iron into the obtained compound of general formula ＼J,P
, Col1man et al.XJournal of the
Anerican Chemical 5ocjety
, 97, 1427 (1975), with excess ferrous bromide in the presence of pyridine in anhydrous tetrahydrofuran refluxed under a nitrogen gas atmosphere. The obtained compound of general formula V has one bromide ion as a counter ion in a central iron trivalent state.

Phosphorylcholine of the compound of general formula (■) N, S,
Chandrakumar et al., Tetrahedron
The reaction of phosphorylcholination of alcohol reported in Letters + Dan, 1043 (1982) was improved. The compound of general formula (2) is prepared by adding 2-chloro-2-oxo-1,3,2-nooxa in an anhydrous, non-70-rotonic solvent such as nochloromethane, chloroporm or benzene in the presence of 2 to 2 equivalents of biriline to the alcohol group. 1 to 2 equivalents of phosphorane were added dropwise at room temperature (not 0° C.) and then reacted at room temperature for 6 to 24 hours. The reaction solution was distilled off under reduced pressure, and the resulting residue was evaporated with acetonitrile or N
, dissolved in N-trimethylformamide, added trimethylamine, and heated at 50°C to 65°C in a stainless steel sealed container.
The reaction was carried out for 12 to 24 hours. The reaction solution was concentrated under reduced pressure, the resulting residue was dissolved in a solvent such as acetone, methanol, acetonitrile, chloroform, or sochloromethane, and the insoluble portion was filtered off. (manufactured by Pharmacia) or Toyopearl IFIW-40 (manufactured by Toyo Soda Co., Ltd.) by column chromatography to obtain the desired general formula (1).
A compound represented by was obtained.

With respect to 1 equivalent of this compound, 1 to 300 equivalents, preferably 5
~150 equivalents and phospholipids such as egg yolk lecithin, soybean lecithin, egg yolk phosphatidylethanolamine, sphingomyelin, sonoglumitoyl phosphatisolcholine,
Single or mixed substances such as distearoylphosphatinorcholine and simyristoylphosphatisolcholine 30~
1000 equivalents of the organic solvent solution was dried under reduced pressure to form a thin film on the wall of the container. Add to this an aqueous medium such as pure water, buffered aqueous solution (pH 5-9), physiological saline, etc.
In addition, the membrane was peeled off by shaking and treated with ultrasound to obtain a liposome dispersion aqueous solution containing the iron (IID porphyrin complex) represented by the general formula I.The central iron ( i: To reduce trivalent to divalent, an equivalent to a slight excess of sodium dithionite, sodium borohydride or ascorbic acid,
It may be added to this liposome dispersion solution which has been made into a nitrogen atmosphere by blowing nitrogen gas into it for a while. Or E for reduction
, Hasegawa et al., Biochemical
al and Blo-physical Re5ea
rch Communications +- Lily 2
793 (1982) can also be used. When oxygen is blown into the IJ,j''some dispersion solution of the iron (n)-porphyrin complex thus obtained at room temperature, it becomes an oxygen complex, and when nitrogen is blown into this, it is reversible to the original iron (II) complex -\. It was confirmed that it acts as an adsorption/desorption agent for oxygen, as the visible absorption spectrum changes.Especially, even when the molar ratio of the compound represented by general formula I to phospholipid is 1:75 to 1:40, The advantage of conventional iron-5,10 is that liposome solutions can be easily produced.
,15゜20-tetra[α,α,α,α-o-(pivalamido)phenyl]porphine complex and phospholipid in a ratio of at least 1:100, preferably 1:150 or more is necessary to form a liposome dispersion solution. This is a big improvement in some respects.

The present invention will be explained in more detail with reference to the following examples, which are not intended to limit the invention.

Reference example 1 10-benzyloxydecanyl bromide is l.

10-nobromodecane 100I and an equivalent amount of sodium benzyl oxide are reacted under reflux in tetrahydrofuran,
The precipitate was filtered, concentrated, and then distilled under reduced pressure. Yield 4611 points 1
85-189℃/3 screams of Hg.

George R, Newkome and others, 5ynthes
i s r 1975 +517, the lithium soanion of 2-methylzolopionic acid was generated with lithium soisoprobylamide in tetrafuran under a nitrogen atmosphere, and 10-benzyloxydecanyl bromide was prepared at −20°C. After dropping 18 g, the mixture was reacted at 45° C. for 2 hours. The reaction mixture was added to cold diluted hydrochloric acid and extracted with ether, and the separated ether layer was washed with diluted hydrochloric acid and then with water, separated and dried over Glauber's salt. The crude oil obtained by evaporation to dryness was recrystallized from petroleum ether, yielding colorless crystals of 12-pentyloxy-2,2-tsumethyldodecanoic acid, yield 8.4,9.
．． Obtained with a yield of 46%. Melting point 53-55℃, elemental analysis: C
Calculated value (chi) as 21H5405; C75,4,I(
10,25, analysis value (related); C75,64°Hlo
, 09° proton nuclear magnetic resonance spectrum (CDC65
) δppm: 1.18 (6H,s, -CH3
), 1.26 (16H, s, -CH2-), 3.
46 (2H, t.

PhCI (20CJ bit CFI2-), 4.51 (2H
, s, PhC 0-), 7.33 (5H, s, phenyl proton).

3.34 fl of this obtained carboxylic acid was mixed with 5 liters of anhydrous benzene.
ml, added 1,2 M of thionyl chloride, reacted at room temperature for 12 hours, and dried under reduced pressure to yield 12-benzyloxy-2,2-methyl-dorikanoyl chloride as a colorless oil in a yield of 3.53 g. Obtained. Red-? External absorption-vector (CC64) I/1790-(-o
-ot).

Proton nuclear magnetic resonance spectrum (CDCl2) δppm
:1, 28 (22H=s, -CH3 and -CH2
-), 3, 46 (2H, t, , PhCH20CH
2CH2−), 4.50(2H*s, phca2o
-), 7.32 (5Hr s r phenyl proton).

Example 1 5.10,15.20-tetra(α, α, α, α-〇-
(aminophenyl) porphyrin (hereinafter abbreviated as H2TamPP) 2.1.9 was dissolved in dichloromethane (300 m
l) Make a solution, add 1 ml of pyridine, and then add 12-pentyloxy-2,2-dimethylidodecanoic acid chloride IJ.
l-1,09,! i+ was added dropwise and reacted for 3 hours at room temperature, then pivaloyl chloride 5- (large excess) and 5 ml of pyridine were each added dropwise and reacted for an additional 2 hours, then a saturated aqueous sodium bicarbonate solution (200M) was added and separated. The organic layer was dissolved in a 4% aqueous sodium hydrogen carbonate solution (200 m),
Washed twice with water (2 ooml) and dried with Glauber's salt. Benzene/ether = 7 using the residue obtained by drying under reduced pressure as a solvent
71 (v/v) by silica gel column chromatography, and further recrystallized from an ether-methanol mixed solvent.
0,15.20-[)su(α,α,α-〇-bivaramidophenyl)-α-〇-[12-pentyloxy-2,2
Yield of phenyl]porphyrin: 1.26! 9, obtained in a yield of 33 cm. Melting point 87,
5-89℃. Magnetic field desorption mass spectrum M+1242 (C
8oH2°N805; 1242). Infrared absorption spectrum (KBr) u: 3440, 3330, 2960, 29
40°2860.1690,1580,1510.14
50.1300゜1L60.970,810,760,
740cm-1111 proton nuclear magnetic resonance spectrum (
CDCl2) δppm: -2,bO (4H+g+internal porphine ring>traced), -0,21(6H18, -CH2
C (di, cos-) 0.05, 0.09 (27H, ea
ch s , (CH3)3C-CONH-) , 3.4
6 (2H, t, J=6.4H2, phcu2ocn
2cH2-), 4.50C2H=s, PhCH2
O) 7.85 (5H, s, benzyl group phenyl proton). Elemental analysis "C80" Calculated value (%) as 9ON805: C77.26, H7.30, H9.01
, Analysis value (%): C76,89, H7,31, H8
, 88 Example 2 The benzyl ether 065I obtained in Examples was dissolved in a mixed solvent of dichloromethane (15 ml) and ditromethane (15 ml), and 1 ml of anisole and 2 g of anhydrous aluminum chloride were added thereto and reacted at room temperature for 3 hours. Ice water (200
ml) and extracted with 501 nl of chloroform. The separated chloroform layer was washed with water (100 yl) and a 4% aqueous sodium bicarbonate solution (2 x 1o On/).
Dried with Glauber's salt. The residue obtained by drying under reduced pressure was subjected to silica dull column chromatography using a mixed solvent of chloroform V ether = 10/1. By recrystallizing from a benzene-n-hexane mixed solvent, red-purple crystals of 5.10, 15.20-[tri(α, α, α-
0-pinoramidophenyl)-α-o-(12-hydroxy-2,2-dimethyldodecanamide 9)phenyl]
Porufuinwo yield 0.43. F was obtained in a yield of 72%. Melting point: 220-221°C. Infrared absorption spectrum (KBr)
y = 3650~3200 (t+road), 34
40.3330,2960.2930.2860X16
90.1580.1510.1445.1300.11
60.9701805.750.740crn.

Proton nuclear magnetic resonance spectrum (CDC43) δppm
2.60 (2H, a, porphine ring N
H), -0,19(6H,8, -CH2CH2C(di)
2-CONH-) 0.10, 0.06(27H,e
achs, (CHρ3ccoNH-), 3.79 (2
H,t.

J=6.5Hz, HOCH2CH2-), 8.8
2 (8H,s, porphyrin ring β-position proton). Elemental analysis: Calculated value (%) as C73H84N805; C76,0
1゜H7,34,H9,72, analysis value; C76,17
, H7,46° H9,43° Example 3 0.575 g of Irufin obtained in Example 2 was mixed with 0.3 ml of biliran! and dissolved in 40 g of tetrahydrofuran,
In a nitrogen gas atmosphere, 0.6 g of ferrous bromide tetrahydrate was added and reacted under reflux conditions for 4 hours.

The residue obtained by drying under reduced pressure was dissolved in chloroform/methanol-5.
The product was purified by alumina column chromatography using a 0/I mixed solvent, 17 d of 48% hydrobromic acid was added to the eluate, stirred, and then dried with Glauber's salt. After drying under reduced pressure, the residue was recrystallized from a mixed solvent of methanol/dichloromethane to give black-purple crystals of bromo(5,10,15,20-[tri(α,α,α-o-hibaramidophenyl)-α- o-(
12-Human0roxy2,2-tumethyldodecanamide 9) Phenyl]porphinato)iron (I[l) with a yield of 0.
56 g, yield 87 g. Melting point 235-237°C.

Magnetic field desorption mass spectrum (M+1)+1287, (
(M+1)-Br:)” 1207(C73H82N
806, FeBr=1286, Fe=57, Br-
79). Infrared absorption spectrum (KBr): 34
40.2970, 2940.2860, 1690, 15
85°1510.1445,1300,1260,10
00.810゜760 Crn-1゜Elemental analysis: C73
H82N805, calculated value as FeBr (multiple); C
68,11, H6,42, H8,70, analysis value (chi)
; C67,84, H6,46, H8,5B.

Example 4 Compound obtained in Example 3 0.345 & triethylamine 0.1! To a solution of l' in dichloromethane (20 ml) was added 0.2 g of 2-chloro-2-oxo-1,:(,2-dioxaphosphorane, and the mixture was reacted at room temperature for 12 hours, followed by drying under reduced pressure. Dissolve in acetonitrile (20 ml), add 10 ml of trimethylamine, seal the tube, and bring to 60 ml.
After reacting at °C for 15 hours, the solvent was distilled off under reduced pressure.

The resulting residue was dissolved in 4-chloroform (100 d) and washed with water. The chloroform layer was separated and dried over Glauber's salt to dryness under reduced pressure L7. Then, it was dissolved in methanol and purified by column chromatography using Toyo Pardal) fW-40s (manufactured by Toyo Soda Co., Ltd.) using methanol as a solvent. Thus iron (■) -5, i, o, 15.
20. −(tri (α, α, α −〇 −
bivaramidophenyl)-α-o-(12-(2'-trimethylaminoethyl)phosphoryloxy-2,2-dimethyltodecanamide]phenyl)porphyrin complex (
Fe (hereinafter abbreviated as pivs.dodop)) was obtained in a yield of 0.086Fl and a yield of 23q6. Melting point 178-1
80℃. Infrared absorption (KBr) ν: 3600
~3200 (broad), 3440.2960°2
940.2860, 1690, 1582, 1510, 1
445°1300.1070,1000,810,76
0on-” 'Elemental analysis 2C78H24N, 08PφF
Calculated value as e・3H2o (%) C65,67, H7,
07, N8.83. Analysis value (tf6); C65,34,
H7,4B, N8.80 Oxygen uptake M test Example 4 Iron(III)-porphyrin complex 1
．． 24 m9, N-lauryl imidazo-/l/1.0
A chloroform-methanol mixed solution of 61n9 (5 times the amount of iron (III) complex) and egg yolk phosphatinorcholine 66 (approximately 50 times the equivalent of (iron Cm) Ri form) was applied as a thin film on the wall of the container under reduced pressure. It was dried and solidified. 0.1 M-
IJ acid buffer solution 10- was added and shaken to obtain an emulsified solution, which was further subjected to ultrasonic stirring treatment (30 minutes) under a nitrogen atmosphere to obtain a uniform aqueous solution containing dispersed liposomes containing an iron (Ill) complex. .

After blowing nitrogen gas into this solution for 20 minutes to deoxygenate it, the reductases were NADP+17Q, glucose-6-phosphate 10m9, ferredoxin 0.02~, ferrexin-NADP+reductase 0.1m9,
Catalase 0.051n9, crucose-6 to phosphate dehydrodanase o, and osrn9 were added to the mixture, and the mixture was allowed to stand at room temperature for 6 hours for reduction, thereby obtaining an iron (fl)-porphyrin complex Liporeem solution. Visible absorption spectrum '61
1 constant, and the absorption thickest wavelength of this solution is 427,533,5
After confirming that it was a reduced type with a wavelength of 60 nm, oxygen gas was then blown into it for 30 seconds. The spectrum changes immediately,
It was confirmed that the resulting oxygen complexes exhibited maximum absorption wavelengths of 424 and 538 nm. When nitrogen gas is again blown in for 2 minutes to create a nitrogen atmosphere, the original reduced spectrum (427
, 533, 560 nm), it was concluded that oxygen adsorption and desorption can occur reversibly. Further, the half-life of the generated oxygen complex was determined to be about 2 hours from the results of tracking changes over time.

Claims (1)

[Scope of Claims] New leaf tetrairon-5゜10.15.20- having one phosphorylcholine group represented by the formula (where n is an integer of 1 to 20)
Cα, α, α, α-0-(substituted amido)phenyl]porphine complex.