DE4402748A1 - Di:methyl amino ethyl phenyl propionate(s) - Google Patents

Abstract

N,N-Dimethyl amino ethyl beta (4-hydroxy-3,5-di tert butyl phenyl) propionates of formula (I) are new. R = H, or 1-16C alkyl. X = Cl, Br, or I.

Description

The present invention relates to new compounds more precisely N, N-dimethylaminoethyl-β- (4-hydroxy-3,5-di-tert.- butylphenyl) propionate, its halogen hydrate and tertiary and quaternary salts of the general formula

wherein

C (CH₃) ₃,
RH or alkyl,
X Cl, Br, J

mean. These connections can serve as the basis for Preparations are used which are an anticholinesterase and have antivirus activity.

These connections were previously unknown their properties are neither in the patent literature, nor described in scientific and technical publications.

The use of inhibitors of acetylcholinesterase as a medicinal preparation is one of the essential procedures in the treatment of Alzheimer's disease. Anticholinesterase preparations increase the concentrations a neuromediator, d. H. of acetylcholine, by Inhibition of its enzymatic hydrolysis. So reinforce  the preparations used have the function of cholinergic Systems at the central level (central nervous system) and on the peripheral levels (synapses). Therapeutic Preparations of the type of action mentioned are in one Review article by Giacobini (M. Pomponi, E. Giacobini, M. Brufani "Present state and future development of the therapy of Alzheimer's disease aging ", vol. 2, No. 2, pp. 125-153, 1990). The most effective of choloergic preparations are Metriphonate, Huperzine A, Tacrine bethanechol, Arecholine, Oxotremorine u. a. The disadvantages of these preparations are Toxicity, side effects, no antioxidant activity (AOA), the latter for treatment Alzheimer's disease is very important Characterized enlargement of the formation of free radicals becomes. Therefore, the development of the invention proposed new synthetic preparations from anticholinesterase effect, which is also an additional AOA necessarily have a new class of inhibitors with an additional current pharmacological Activity.

It is a substance (physostigmine) known to be a Dihydroizocholine derivative, which acts as a cholinesterase inhibitor in the treatment of Alzheimer's disease is used. (Becher R. E., Giacobini E. Mechanism of cholinesterase inhibition in senile dementia of Alzheimer type: clinical, pharmacological and therapeutic, phar. "Drug  Development Res., "12, 1988, pp. 163-195).

However, this substance is toxic, has side effects and has a short duration of action.

The connection to be registered comes closest their chemical structure is a potassium salt of phenosanic acid, that's a structure

has, where R is C (CH₃) ₃ and this salt in medicine and agriculture as a medicine against Burn injuries and stress as well as growth material ("aging", vol. 2, No 2, pp. 125-153, 1990) can be used.

The disadvantages of the salt mentioned are its minor Effectiveness, poor water solubility of salts with the formation of unstable solutions are easily exposed to hydrolysis, creating an alkali and a starting acid is formed, which is the area of application of the above salt in medical practice significantly limited. This substance has a low level anticholinesteras activity what they use as a potential neurological substance makes it less effective.

In some areas of medicine is considered a current problem also the cytomegalovirus infection (ZMV infection) because this developed in the background of an acquired immunodeficiency that is through chemotherapy and radiation treatment of tumors, through immunosuppressive effects  in organ transplantation or some other viral infections (including AIDS) arises. The cytomegalovirus infection is very common the cause of exitus letalis in the above Examples as well as for early childhood lethality.

As the only known means of fighting this Infection is ganciclovir (guanine), which the Synthesis of infectious deoxyribonucleic acid (DNA), d. H. inhibits a virus (Biron K., Stanat S.C., Sorell J. et. al. Metabolic activation of the nucleoside analog 9 - {[2-hydroxy- 1- (hydroxymethyl) ethoxy] methyl} guanine in human diploid fibroblasts infected with CMU Pr. Nat. Ac. Sci. 1985, 82, 2473-2477).

A negative property of the preparation ganciclovir However, there are some variants of viruses are present that are insensitive to this preparation are, and that in the organism ganciclovir-resistant Strains emerge quickly, which is at the same time the treatment effectiveness diminishes or nullifies (Erice A., Chou S., Biron K., Stanat S.C. et. al. Progressive disease due to ganciclovirresistant CMU in immunocompromised patents. The New Engl. J. of Med. 1989, 320, 5, 289-293).

The registered invention is based on the object to find new antivirus products, which the reproduction of a number of viruses containing DNA and RNA influence.  

Disclosure of the invention

The invention aims at the production of dimethylaminoethyl propionate, its halogen hydrates and quaternary Ammonium salts for the synthesis of compounds which as Basis (basis) for preparations which are an anticholinesterasic, antiviral,. . . Possess activity.

This is achieved through a new substance with the following chemical formula

wherein

C (CH₃) ₃,
RH or alkyl,
X Cl, Br, J

mean.

This structure is synthesized according to the following Scheme

N, N-dimethylaminoethyl-β- (4-hydroxy-3,5-di-tert-butylphenyl) - propionate is obtained by transesterification of methyl-β- (4- hydroxy-3,5-di-tert-butylphenol) propionate with N, N-dimethylaminoethanol under the conditions of an alkaline Catalysis at 130 to 140 ° C in the argon point when removed of the resulting methanol from the reaction manufactured.

The structure of the dimethylaminoethyl propionate obtained was obtained from data from elemental analysis, IR and PR spectroscopy occupied (established).

When implementing the propionate obtained in the medium of acetone, benzene or ester with hydrogen halide or with alkyl halide becomes halohydrate or a quaternary Ammonium salt of the propionate mentioned. The Structure of the salts obtained was determined by data from Elemental analysis, IR and PM spectroscopy found.

The proposed synthesis principle allows one to gain active and little toxic substance which is a hybrid Molecule represents two or more in its structure Fragments united for different physiological activities are responsible.

The hybrid structure of the material obtained, namely the presence of a fragment of the shielded phenol in their composition of the molecule next to the fragment, which is a specific or non-specific interaction with a Guaranteed neuroreceptor, determines the possibility of potential use of the present connection as neurological Substance ahead. The mechanism of their action can obviously by the anticholinesterase activity as well  be conditional, as is the case with the other known inhibitor preparations the case is (Hallak M. E., Giacobini E. "A compozition of the effects of two inhibitors on brain cholinesterase ". Neutopharmacology, 26, 1987, pp. 521-530).

It is also widely known that in a number of neurological diseases, especially in Alzheimer's Disease, which is the main cause of dementia serves elderly people, changes in cholinesterasic Activity can occur. With these pathologies became a cholinergic deficit in the central nervous system Synapses and blood erythrocytes discovered. The diminution the level of acetylcholine (i.e. the substrate of the acetylcholinesterase Reaction) in sick calls in these a correlation of the organism, which is in a erratic memory disorder, in a severe disorder of thought functions and other disorders. The disorders mentioned can be selective Inhibition of acetylcholinesterase can be reduced, which from an increase in the level of acetylcholine in the Central nervous system is accompanied.

One of the main directions in the therapy of Alzheimer's disease involves a search for pharmacological Agents that inhibit hydrolysis of Acetylcholine in the synapses due to the use of Acetylcholone esterase inhibitors cause.

As a model system for the investigation of anticholinesterasic  Activity of synthesized compounds was used an acetylcholinesterase located in the erythrocytes. Changes in plasma cholinesterase activity and in the erythrocytes one correlates with those in the Brain structures in such pathalogies as in Alzheimer's Disease (4) so that the simulation of Influences of different connections, i. H. Acetylcholinesterase inhibitors (inhibitor), on the activity of the erythrocyte membranes is adequate to determine whether their anticholinesterase effect on cholinergic functions the central nervous system (CNS) is present or missing, and thus for the screening of potential neurological substances can be exploited.

These compounds also have an antioxidant Activity (AOA). A relative AOA was determined according to the Ability of compounds to peroxidize lipids inhibit in the homogenate of the brain of the mice. Man assesses the rate of oxidation after formation of products resulting from the peroxidation of lipids of the malondialdehyde type. The compounds to be examined decrease the rate of peroxidation of Lipids, i.e. H. act as an antioxidant.

There is a reliable correlation between the antioxidative activity (AOA) of the compounds to be investigated and their ability to inhibit acetylcholinesterase in human blood red cells. An associated equation for a regression line is: y = -0.127 + 0.146 x, where y = I _C50 , x = 1 / AOA, the correlation factor is 0.9925 and p <0.001.

Table 1 shows comparative values of the anticholinesterase activity of the derivatives of shielded phenols (based on the parameter C _A50 , ie the concentration of the substance in mM, during the treatment of which a 50% inactivation of the acetylcholinesterase in the erythrocytes takes place) and the antioxidative activity (in relative units ) summarized.

example 1

0.23 g (0.01 M) sodium is dissolved in 11.0 ml (0.11 M) dimethyl ethanol. 29.1 g (0.1 M) of methyl β- (4-hydroxy-3,5-di-tert-butylphenyl) propionate are added to the solution obtained. The mixture is heated to 130. . . 140 ° C in a weak stream of argon with the resulting methanol being expelled for 3 to 4 hours. The mixture is cooled, dissolved in 100 ml of ester, the ester solution is washed twice with water (2,200 ml), dried and evaporated in vacuo. The rest is dissolved in 100 ml of hexane when heated and remains in a cooler overnight. The precipitated crystals are separated off, washed with 30 ml of cold hexane and dried. 19.5 g of crystals with a slightly pale yellowish color are obtained. An additional 7.8 g of similar crystals are obtained from the hexane mother liquor by acid extraction with subsequent alkalization of the aqueous phase with 5% HOH.
Total yield: 27.3 g (78% of theory) of N, N-dimethylaminoethyl-β- (4-hydroxy-3,5-di-tert.butylphenyl) propionate, mp: 69-70 ° C.
Found,%: C 72.35; H 9.94; N 5.10; C₂₁H₃₅O₃N
Calculated,%: C 72.16; H 10.09; N 4.01.
IR spectrum (KBr): ν _OH = 3640; ν _CO = 1649 cm ^-1 .
PR spectrum (CDCl₃, δ, ppm); C (CH₃) ₃ = 1.42; CH₂ (δ) = 2.61; CH₂COO = 2.86 (J = 8.2 Hz), COOCH₂ = 2.54; CH₂N = 4.17 (J = 5.8 Hz) (CH₃) ₂N = 2.27; OH = 5.02; H _m = 6.95.

Example 2

The solution of 3.5 g (0.01 M) propionate in 25 ml dry Ester is saturated with dry HCl until absorption of the latter ceases. The failed crystals are separated, with 20 ml dry ester and 30 ml hexane washed, dried in vacuo.

3.6 g (93% of theory) of chlorohydrate of N, N-dimethylaminoethyl-β- (4-hydroxy-3,5-di-tert-butylphenyl) propionate (ICHFAN-9) are obtained, m.p .: 176- 177 ° C.
Found,%: C 65.08; H 9.31; N 3.76; Cl 8.91; C₂₁H₃₆O₃NCl.
Calculated,%: C 65.34; H 9.40; N 3.63; Cl 9.19,

Example 3

A solution of 3.5 g (0.01 M) propionate in 25 ml dry ester 1.2 g (0.011 M) ethyl bromide added. The mixture thus formed is kept at 20 to 25 ° C for 2 hours kept, the precipitated bromide residue of the quaternary ammonium salt is separated off, washed with hexane, dried,  crystallized from water.

4.6 g (98% of theory) of plate-like crystals (ICHFAN-10) with a melting point of 169 to 170 ° C. are obtained.
Found,%: C 53.76; H 7.56; N 2.73; J 25.40; C₂₂H₃₈O₃NJ.
Calculated,%: C 53.76; H 7.79; N 2.85; J 25.82
PR spectrum DCl₃, δ, ppm): C (CH₃) ₃-1.42; CH₂ (δ) -2.67; CH₂COO = 2.85 (J = 8.2 Hz); (CH₃) ₂N = 3.49; COOCH₂ = 4.09; CH₂N = 4.58; OH = 5.07; H _m = 6.95.

Example 4

A solution of 3.5 g (0.01 M) propionate in 25 ml dry Ester is added 1.2 g (0.011 M) ethyl bromide. The so educated Mixture is kept at 20 to 25 ° C for 2 hours, the precipitated bromide residue of the quaternary ammonium salt is separated, washed with hexane, dried.

This gives 4.2 g (92% of theory) of plate-like bromide crystals (ICHFAN-10-C₁ mit) with a melting point of 167 to 168 ° C.
Found,%: C 60.02; H 8.48; N 2.79; Br 17.00; C₂₃H₄₀NBr.
Calculated,%: C 60.24; H 8.79; N 3.05; Br 17.44.
PR spectrum (CDCl₃, δ, ppm): C (CH₃) ₃ = 1.41; CH₃CH₂ = 0.92; CH₃CH₂ = 3.52 (J = 7.1 Hz); CH₂ (δ) = 2.67; CH₂COO = 2.85 (J = 8.2 Hz); (CH₃) ₂N = 3.45; COOCH₂ = 4.07; CH₂N = 4.51; OH = 5.03; H _m = 6.95.

The target product, d. H. Halogen hydrate or halide of quaternary ammonium salt receives a two-stage course Synthesis in the usual chemical equipment, the Yield is up to 95%. The process can be industrialized Scale based on available raw material, d. H. Methyl- β- (4-hydroxy-3,5-di-tert-butylphenyl) propionate, without further ado  realize.

In this way, a new substance is based of shielded phenols and their synthesized Properties have been studied, which is the possibility offer the synthesized substance as the basis for the Creation of medicinal products during treatment Alzheimer's disease and other forms of dimensions to judge.

The task is also solved in that as Agent that has antivirus activity, the registered Halide of the quaternary ammonium salt of dimethylaminoethyl β- (4-hydroxy-3,5-di-tert-butylphenyl) propionate (ICHFAN-10) is used.

Table 1

The AOA (antioxidant activity) was called

where V _control and V _AO oxidation rates in the absence and in the presence of equimolar concentrations (2 × 10 ^-5 M in the present case) denote the compounds and I _{C50 stands} for the concentration which is a 50% inhibition of the membrane-bound erythrocyte acetylcholinesterase results.

ICHPHAN-10 is a crystalline substance found in water and alcohol is soluble. It is known that ICHPHAN-10 as the basis for the creation of medicinal products to treat Alzheimer's disease and others Dementia forms thanks to its increased anticholinesterasic Properties is used.

Thus, ICHPHAN-10 was used as the one agent Has antivirus activity, but as a model virus  DNA-containing cytomegalovirus and as an RNA-containing component the vesicular stomatitis virus was used.

The rating of the effect of the preparation was carried out in the culture of diploid cells M-19 of the human embryo through which in the International Collection of Cultures (VOZ) have been registered. For this, the cells in Penicillin bottles settled and at 37 ° C for three days grown, whereupon all the cells to be examined Viruses added, an absorption performed and then different concentrations in the different bottles ICHPHAN-10 were added. The cells were at 37 ° C cultured for 2 to 3 days, after which they grow at T = -20 ° C frozen, defrosted and an infection titer in the liquid of the virus is determined by also working on the diploid cells M-19 were titrated. The infected culture served as a control without the addition of the preparation.

Example 6

Investigation of the influence of different doses the reproduction of the cytomegalovirus in the diploid cells of people with infections of different Majority were infected.

A three day culture of human diploid cells one was infected with a polyinfection (0.01 TCD / cell or 1 TCD / cell) and brought different concentrations to ICHPHAN-10. After 48 hours of breeding one determined Titer values of the cytomegalovirus (see Table 2).  

Table 2

Influence of ICHPHAN-10 on the reproduction of the cytomegalovirus (strain AD 169) in different polyinfections

The data thus obtained show that ICHPHAN-10 has an activity through which the reproduction of the Cytomegalovirus is inhibited, this activity both from the dose of the preparation used for it also depends on the amount of virus introduced. At a high polyinfection (1 TCD 50 / cell) come as effective concentration of the preparation 50 µg / ml or 100 µg / ml considering the reproduction by 4 or 6 logarithms inhibit (i.e. inhibit more than 99.99% of the virus). At  a low polyinfection (0.01 TCD 50 / Zl.) turns out as effective also a concentration of 20 µg / ml, at which inhibit more than 99% of the virus.

Similar results were also obtained on another Received series of the preparation.

Example 7

Investigation of the influence of different doses drug for reproduction of the vesicular virus Stomatitis.

The effect of the preparation was tested in two tests single-cycle or multi-cycle reproduction - examined.

1. Single cycle reproduction. On the human diploid cells M-19 became a virus with a high polyinfection (10 TDC 50 / cell), then adsorption (At 37 ° C for 1 hour) and washing the product three times non-adsorbed virus carried out on the cells were cast with a support medium that the Contains preparation. After 8 hours of breeding (cultivation) the virus-containing liquid was collected and transferred to the Titrated cells M-19.

2. Multi-cycle reproduction. On cells M-19 was a virus with a low polyinfection (0.01 TDC 50 / cell) applied in a medium containing the preparation, and the cells were incubated for 3 days, after which in the liquid titrated the virus in culture M-19 has been. Cytopatic titer values were recorded on the third day (see Table 3).  

Table 3

Influence of ICHPHAN-10 on the reproduction of the virus vesicular stomatitis in single and multi-cycle infection

The data obtained show that ICHPHAN-10 is reproductive of the vesicular stomatitis virus in both Infecting the cells with a high polyinfection (a Cycle of reproduction) as well as when infecting with a low polyinfection (multi-cycle reproduction) inhibits. The titer of a newly formed virus is both in one case and in the other case by 99.9% and more diminished. When examining two others Identical results were obtained in series of the preparation.

The tests carried out have shown that ICHPHAN-10 an antivirus activity against the cytomegalovirus and the vesicular stomatitis virus, which are both high and low Showed polyinfection. At optimal doses exceeded the index of effectiveness 99.9% of the newly synthesized Viruses.

Claims (6)

1. N, N-dimethylaminoethyl-β- (4-hydroxy-3,5-di-tert-butylphenyl) propionate of the general formula whereinB H, alkyl having 1 to 6 carbon atoms,
X Cl, Br, J mean. 2. Connection according to claim 1, characterized by the presence of halohydrate. 3. A compound according to claim 1, characterized characterized in that it is a tertiary ammonium salt contains. 4. A compound according to claim 1, characterized characterized as being a quaternary Contains ammonium salt. 5. Compounds according to claim 1, characterized characterized as an anticholinesterase Possess activity. 6. Compounds according to claim 1, characterized characterized as having antivirus activity have.