CN1082511C - Phophorus contg. cythesine derivs. - Google Patents

Abstract

The present invention discloses cethysine derivatives synthesized recently, which comprises phosphor and has a general formula (I), wherein n is equal to 0 or 1. When the n is equal to 0, X is sodium, and R is CH3, C2H5, C3 H7, 1-C3H7 and C4H9. When the n is equal to 1, the R is CH3, R1 is 1-C3H7, C6H5 and right-CH3O-. The present invention displays that the compound has high liver protective activity and high antienzyme activity.

Description

The application of phosphorated Tocosamine derivative

Technical field

The present invention relates to protect the liver application with the active phosphorous Tocosamine derivative of antienzyme.

Background technology

Known organism alkali Tocosamine (1) extracts in the seed of Flower of Chinese Peashrub and Thermopsis lupinoides (L.) Link. at first, it is now by widely as medical, with the state of 0.15% aqueous solution (Tocosamine) as stimulant (anaelepticagent) (Mashkovsky, M.D., " pharmacological agent (Medicine remedies) ", M., 1977, volume I, 123 pages)

R=C wherein _2-5

The typical character of Tocosamine is that it can cause breathing, and this is with relevant to the reflex irritation of respiratory centre by beating of strengthening from intercarotid body.Stimulate sympathetic nerve knot and suprarenal gland body to cause arteriotony to rise simultaneously.The case that breathing and cardiomotility stopped when in the case, Tocosamine was used to poison.

Tocosamine extensively is present in nature.Have found that many plants and be the major industry approach that obtains Tocosamine with the extract that ion-exchange techniques obtains from material of vegetable origin.

At works (report of Uzbek SSR academy of sciences, 1978, N9, p.p.39-42; Uzbek SSR academy of sciences report, 1977, N7 has reported phosphorated N-[β-(dialkoxy phosphinyl (dialkocsiphosphenyl)) mercaptoethyl in p.p.40-43)] Tocosamine (2) and their iodomethylation thing (3).

The author shows that all synthetic compounds have irreversible inhibition activity to acetylcholinesterase, and manifests the selective inhibitory to the butyrylcholine esterase catalytic activity.

We do not find other phosphorated Tocosamine derivative in the literature.Summary of the invention

Because Tocosamine alkaloid has originally shown tangible biological activity, people can infer that other phosphorated Tocosamine derivative also has confers similar advantages.

The objective of the invention is to obtain having the application of the phosphorated Tocosamine derivative of the bioactive novelty that increases.

Reached purpose of the present invention like this: the phosphorated Tocosamine derivative that obtains following general formula preparation altogether patient resist application in the hepatic of hepatotropic poison: N=0 wherein, 1;

X＝S：O

R=CH ₃(a); C ₂H ₆(b); C ₃H ₇(c); I-C ₃H ₇(d); C ₄H ₉(e) R ¹=i-C ₃H ₇(a); C ₆H ₅(6);

When condition is n=1, X=0.

By making dialkyl phosphites intermediate and Tocosamine under following formula Todde-Aterton reaction conditions, react to synthesize O, O-dialkyl group-N-Flower of Chinese Peashrub base phosphoric acid ester (n=0, X=O, compound 4a-e):

O, the synthetic of O-dialkyl group-N-Flower of Chinese Peashrub base (cytisinyl) phosphoric acid ester (n=0, X=S, compound 5a-e) carries out in two steps.First step, at Tocosamine and dialkyl phosphites step of reaction (this synthetic article as E.V.Nifantyev, " chemistry of P contained compound " science magazine (M.Science) 1983,85 pages described), in the benzene medium, intermediate O, inferior Flower of Chinese Peashrub base (cytisenyl) phosphorous acid ester of O-alkyl-N-, (it generates O with the reaction of molar equivalent elementary sulfur then, the inferior Flower of Chinese Peashrub base of O-alkyl-N-phosphoric acid ester) reaction in the presence of dried argon gas stream and triethylamine forms.

React under the Kabachnic-Fild reaction conditions by Tocosamine and aldehyde and dimethyl phosphite and to obtain dimethyl-(the inferior Flower of Chinese Peashrub base of N-) alkyl (aryl) phosphonic acid ester (n=1, X=0, R-CH ₃Compound 6 _A-C):

Implement variant the following example of the present invention more detailed ground explain the present invention, but be not its restriction.

Embodiment: 1. synthetic O, O-dimethyl-N-Tocosamine phosphoric acid ester (4a) will contain 2.86 gram (0.026 mole) dimethyl phosphites and the mixture of 15.5 milliliters of (0.16 mole) tetracol phenixin in 100 milliliters of benzene put in the flask, during stirring, add and to contain 5 gram (0.026 mole) Tocosamines and the mixture of 2.63 gram (0.026 mole) triethylamines in 230 milliliters of dry-out benzene so that solution temperature is no more than 20 ℃ speed.

After reinforced the finishing, reaction mixture was at room temperature stirred 6 hours.Leach the Triethylammonium chloride crystal of separating out, boil off solvent on the vacuum-evaporator.Crystalline substance (from benzene cross crystallization) have 5.64 grams (for theoretical yield 72.5%), fusing point is Tm:158-159 ℃.

IR spectrum, ν cm ^-1: 1270 (P-O), 1050 (P-O-C),

830(P-N-C)，1658(N-CO)。

EPR spectrum, δ, m, d 6.23d (1H, CH _α, ³J _NH5.0Hz),

7.23d，d，(！H，CH _β， ³J _NH3.2Hz)，

6.12d(1H，CH _γ)

3.34d(6H，CH ₃O，J _CH3P13.2Hz)

NMR ³¹P；δp＝9.1m.d.

Measured value, %:C 52.05; H 6.24; N 9.30; P 10.36;

C ₁₃H ₁₉N ₂O ₄O ₄P

Calculated value, %:C 52.36; H 6.37; N 9.40; P 10.40.

According to the data that X-ray diffraction is analyzed, at O, the coordination of the phosphorus atom in O-dimethyl-N-Tocosamine phosphoric acid ester molecule has been twisted, and is tetrahedron for phosphoric acid ester usually.The flattened precision to ± 0.01A of dihydropyridine ring, carbonylic oxygen atom departs from this plane (0.07A) slightly.

Tetrahydro pyridine ring is got chair conformation (△ C ⁸ _s2.7 °), the outlet of its path e atom with from other C atom difference 0.75A of mid-plane.It almost is that the ideal chair is got type conformation (△ C that the hexahydropyridine ring has ⁸ _s0.5).Methyl CH ₃Group has the Rosh orientation with respect to corresponding P-O and P-O key.

Can similarly obtain other compound 4 of this series _B-c(table 1).

2. O, O-dimethyl-N-Flower of Chinese Peashrub base thiophosphatephosphorothioate (5a) synthetic

5 gram (0.026 mole) Tocosamines and the 2.63 gram mixtures of (0.026 mole) triethylamines in 200 milliliters of dry-out benzene are put in the flask, dry argon gas flow down with room temperature under the intensively stirred while in this mixture, adds 3.33 and restrains (0.026 mole) dimethyl chloride phosphorous acid esters.Mixture was stirred 5 hours.Leach the triethylamine hydrochloride precipitation of gained, wash in batches with dry-out benzene.Sulphur 0.83 gram (0.026 mole) of filtrate and calculated amount is placed in the flask, at room temperature continues to stir 2 hours again.Then, evaporating mixture on rotatory evaporator, throw out benzene recrystallization.Obtain 8.01 gram (yield 97%) crystalline substances, Tm is a 164-165IR spectrum, ν cm ^-1: 1030 (P-O-C), 1670 (N-CO), 820 (P-S).EPR spectrum, δ, m, d 6.23d (1H, CH _α, ³J _NH6.0Hz),

7.16d，d，(1H，CH _β， ³J _NH3.2Hz)，

5.8d(1H，CH _γ)

3.40d (6H, CH ₃O, J _CH3P12.0Hz) NMR ³¹P; δ p=10.22m.d. measured value, %:C 49.91; H 6.13; N 8.87; P 9.69;

C ₁₃H ₁₉N ₂O ₃The SP calculated value, %:C 49.68; H 6.05; N 8.91; P 9.87.

Can similarly obtain such compound 5 _B-EThe result is as shown in table 1.

Table 1

The physicochemical constant of described compound and ultimate analysis data

Compound	Tm，℃	n 20	Measured value, %				Molecular formula	Calculated value, %				Yield
																C	H	N	P	FORMULA	C	H	N	P	％
4a	158-159	-	52.08	6.27	9.48	10.26	C 13H 19N 2O 4P	52.36	6.37	9.40	10.40	72.5
													4b	-	1.5027	55.39	7.13	8.65	9.66	C 15H 23N 2O 4P	55.21	7.05	8.59	9.51	68.4
4c	-	1.5027	57.72	7.69	7.82	8.83	C 17H 27N 2O 4P	57.62	7.62	7.91	8.75	69.7
													4d	115-116	-	57.81	7.74	7.87	8.86	C 17H 27N 2O 4P	57.62	7.62	7.91	8.75	70.5
4c	-	1.4836	59.77	8.19	7.28	8.20	C 19H 31N 2O 4P	59.68	8.11	7.33	8.11	63.8
													5a	164-165	-	49.91	6.18	8.87	9.69	C 13H 19N 2O 3PS	49.68	6.05	8.91	9.87	97.0
5b	99-100	-	52.72	6.79	8.14	9.17	C 15H 23N 2O 3PS	52.63	6.72	8.19	9.06	92.5
													5c	-	1.5486	55.24	7.41	7.50	8.43	C 17H 27N 2O 3PS	55.13	7.30	7.57	8.38	87.3
5d	-	1.5463	55.19	7.38	7.49	8.47	C 17H 27N 2O 3PS	55.13	7.30	7.57	8.38	75.1
													5c	-	1.5446	57.38	7.86	7.01	7.91	C 19H 31N 2O 3PS	57.28	7.79	7.04	7.79	82.0
6a	137-139	-	57.70	7.72	8.03	8.67	C 17H 27N 2O 4P	57.56	7.58	7.90	8.52	78.1
													6b	190-192	-	61.87	6.47	7.19	7.99	C 20H 25N 2O 4P	61.85	6.44	7.21	7.98	85.2
6c	186-188	-	60.18	6.74	6.56	7.37	C 21H 28N 2O 5P	60.14	6.68	6.68	7.39	87.0

3. dimethyl-2-(N-Flower of Chinese Peashrub base)-2-isobutyl-phosphonic acid ester (6a).

Make and contain the mixture of 3.8 gram (0.02 mole) dimethylphosphites and catalytic amount 18-hats-6 in 100 milliliters of benzene and seethe with excitement together and steamed the water that forms in the reaction process in 3 hours simultaneously.After reacting end and steaming solvent, the residue hexane wash with benzene-ether mixture recrystallization, obtains 5.52 gram (yield 78.1%) crystallisates, and fusing point is 138 ℃.

IR spectrum, ν cm ^-1: 1220 (P-O), 1070 (P-O-C).

EPR spectrum, δ, m, d 6.72d (1H, CH _α, ³J _NH866Hz),

7.10d，d，(1H，CH _β， ³J _NH664Hz)，

6.51d(1H，CH _γ)

3.31d(6H，CH ₃O，J _CH3P12.6Hz)；

0.50d(3H，CH ₃-C)，

0.87d(3H，CH ₃-C)。

Measured value, %:C 57.70; H 7.72; N 8.03; P 8.67;

C ₁₇H ₂₇N ₂O ₄P

Calculated value, %:C 57.56; H 7.58; N 7.90; P 8.52.

Can similarly obtain other compound 6 of this compounds _B-C(table 1).

Industrial applicibility

The biological activity of described compound

Pharmacological research shows that institute's synthetic compound has and protects the liver and the antienzyme activity.

1. liver-protecting activity research.

1. the anxious toxicity of compound.

The small white mouse (320, each is the 18-20 gram) that arbitrarily feeds is measured mean lethal dose LD ₅₀Preparation is placed in the physiological solution once quiet notes or is placed in 1% starch suspension oral.Observed 14 days.Check the general state of animal every day, viewing duration is weighed 3 times to animal, dissects dead animal and carries out macroscopic description.

Decide LD by analytical procedure ₅₀

As test shown in the result, have now found that the LD of the oral people's of taking the photograph compound ₅₀=4300mg/kg (3200-5800), the LD of quiet notes ₅₀=1800mg/kg (1250-2550).Arithmetical av is P=95% with putting the letter half-interval.

Liver-protecting activity to acute poisoning rat hepatitis

According to pharmacologist's suggestion, the most effective liver protectant is at present " Essentisialle " (FRG) preparation, it is used to following experiment preparation as a comparison.Said preparation is complicated material, contains flavonoid, phosphatide, niacinamide and various VITAMIN.

Use 4 groups, the body weight of arbitrarily feeding of every group of 10 samples is that the male rat of 120-150 gram is studied the liver-protecting activity of described preparation to acute toxic hepatitis.

First group, with 0.1 milliliter of muriate isotonic solution animal is carried out quiet notes, (in 1 hour) carries out intraperitoneal injection with the solution of 0.4 milliliter of 40% tetracol phenixin in the oil jelly more then.

Second group, with 0.1 milliliter of sodium-chlor isotonic solution animal is carried out quiet notes, then after 1 hour again the solution of poisonous substance-tetracol phenixin in the oil jelly with 0.4 milliliter of 40% close liver carry out intraperitoneal injection.

The 3rd group, with 0.1 milliliter of " Essentsialle " solution (50mg/kg) animal is carried out quiet notes, carry out intraperitoneal injection with the solution of 0.4 milliliter of 40% tetracol phenixin in the oil jelly again after 1 hour then.

The 4th group, with the aqueous solution (50mg/kg) of 0.1 milliliter of described material animal is carried out quiet notes, carry out intraperitoneal injection with the solution of 0.4 milliliter of 40% tetracol phenixin in the oil jelly again after 1 hour then.

All injections repeats once every day, continues 4 days, then kill animals under etherization.

Poison the biological chemistry of damage rate and the histology factor in characterize cells and change the liver-protecting activity of the described preparation of assessment and comparative formulations on the basis the rat poisoned with tetracol phenixin.

Alanine aminotransferase in the blood (alaninaminotranspherase, ALAT) active and total bilirubin level, red blood corpuscle chemoluminescence infiltration, regenerated and underfed cell in the liver tissue slices have been studied.

When the research chemoluminescence, the red blood corpuscle that extracts from blood is suspended in sodium-chlor isotonic solution (1 milliliter of 10% red blood corpuscle suspension of each sample), with 0.1 milliliter of 5%H ₂O ₂Stimulation glows, and measures the mean number of chemoluminescence curve lower integral area.

Group ALAT bilirubin chemoluminescence area under curve

Mmole hour/rise micromoles per liter

1 215 8.1 8.06 0.4 4.34

2 1828 34.6 31.12 1.09 9.395

3 1291 10.5 20.84 0.25 7.35

4 445.4 11.7 12.90 0.32 5.386

Organize malnutritive % gangrene % infiltration % regeneration %

1 2.6 0.2 0.35 1.4

2 20.5 4.3 0.6 0.652

3 12.8 0.9 0.4 1.03

4 8.2 0.5 0.3 1.48

Data show that described material has liver-protecting activity to the rat of being poisoned by tetracol phenixin, and its effect is than more powerful as correlated " Essentsiallye " preparation.

Described material is to the influence of the survival of rats rate of poisoning with hepatotropic poison

In order to study the influence of preparation to the mouse survival rate of poisoning with hepatotropic poison, use arbitrarily feed male, 18-20 restrains mouse, and 10 every group, totally 3 groups.

First treated animal is by 0.1 milliliter of sodium-chlor isotonic solution of intraperitoneal injection.Second treated animal is by 0.1 milliliter of " Essentsialle " solution of quiet notes, and dosage is 50 mg/kg.The 3rd treated animal is by the aqueous solution of 0.1 milliliter of material of the present invention of intraperitoneal injection (100 mg/kg).

Injected back one hour for the first time, all animals are by 0.1 milliliter of 50% carbon tetrachloride solution in the oil jelly of intraperitoneal injection.

Injection is carried out once, observes 5 days, and the survival rate in 5 days is as follows:

Group surviving animals amount survival rate

1 0 0

2 2 20

3 8 80

The result of gained shows, The compounds of this invention is poisoned to watching for animals because by hepatotropic poison and big than " Essentsialle " of dead degree.

Like this, we find that The compounds of this invention has appropriate toxicity, and its hepatoprotective effect obviously surpasses the effect of " Essentsialle " preparation in experimentation on animals.

II. antienzyme activity research.

The antienzyme activity of research synthetic compound of the present invention, the interaction of they and acetylcholinesterase (ACT) and butyrylcholine esterase (BCE).Use specificity be 1.2 unit/milligrams from the erythrocytic ACE of human blood with from the BCE of horse serum, its specificity is 9.6 unit/milligrams.Measure ACE and BCE activity in 25 ℃ of 0.2MNa-phosphate buffered saline buffers (pH=7.5) lining with the Ellman calorimetric method.The acetyl cholinesterase iodide that " Chemapol " company sells is used as substrate.The effect of reversible transglutaminase inhibitor is measured with the A.P.Brestking method, with inhibition constant (Ki) sign of conclusionizations, in the mixed type restraining effect Ki with compete component (Ki) and do not compete component (K ' i) relevant, equation is as follows:

1/Ki＝1/Ki+1/K’i

Measure P450 cytopigment activity with two kinds of methods: for 3, the hydroxylation speed of 4-benzo/a/ pyrene, by measure the 4-hydroxy benzo/a/ pyrene of formation with fluorometry, for the speed of 7-ethoxy coumarin (qumarine) diethylization, use fluorometry to measure formed umbelliferone.

Pass through pJ ₅₀The material that (making the active 50% o'clock logarithmic negative of reagent concentration that reduces) evaluates to be studied is to the inhibition influence of P450 cytopigment.

Having now found that Tocosamine phosphoric acid ester and Tocosamine thiophosphatephosphorothioate are the inhibitor that ACE has average effectiveness, is extremely effective BCE inhibitor.Observed maximum selectivity (pK1=6.7 and pK1=6.1) for phosphorothioate derivative with respect to BCE.

The present invention has also disclosed the energy inhibitor of P450 cytopigment, and it is practical that they have life to supply, pJ ₅₀=5.55 and pJ ₅₀=5.70 compound is among this.

Claims (2)

1. the application of phosphorated Tocosamine derivative in the hepatic of preparation patients opposing hepatotropic poison with following general formula: N=0 wherein, 1,

X＝S；O

R＝CH ₃；C ₂H ₅；C ₃H ₇；i-C ₃H ₇；C ₄H ₉；R ¹＝i-C ₃H ₇；C ₆H ₅；

When condition is n=1, X=0.

2. application according to claim 1, wherein said phosphorous Tocosamine derivative has following general formula: N=0 wherein, 1.