Aminoalkylcarbamic esters of eseroline suitable for use as chlorinesterase activity inhibitors

PRIOR ART
Several memory dysfunctions, and in particular the senile dementia of Alzheimer's type, are characterized by a reduction in the acetylchyoline neurotransmitter levels in some areas of brain. In these cases, the inhibition of acetylcholinesterase, an enzyme that hydrolyzes acetylcholine, is therapeutically effective.
As known, physostigmine is a potent natural inhibitor of acetylcholinesterase which, as demonstrated by several clinical tests, has beneficial effects on patients affected by mental diseases. However, its unfavourable pharmacokinetic properties and side effects make its clinical use scarcely advisable.
As also known, heptastigmine, or heptylcarbamic ester of eseroline, is a less potent acetylcholinesterase inhibitor than physostigmine, but shows better pharmacokinetic properties and reduced side effects. Heptastigmine gave good results in a clinical pilot test on patients affected by Alzheimer's disease. However, it caused neutropenia phenomena in some, though very few, patients.
Further derivatives of eseroline were synthesized in recent times. For example, a compound consisting of eseroline carbamic ester with morpholine was disclosed in Example 30 of EPA 0253372 by Hoechst-Roussel. However, said compound produced a low acetylcholinesterase activity depression and was not further developed.
SUMMARY
It has now been surprisingly found that the compounds as per formula (I) have none of the disadvantages of the compounds already known and exhibit a high inhibition of cholinesterase activity. The present invention relates to aminoalkylcarbamic esters of eseroline as per general formula (I) ##STR3## where n is an integer from 2 to 20; R1 may be H, methyl, ethyl, propyl, and isopropyl; and R is an amine selected out of the group consisting of ##STR4## where R2 is linear or branched C.sub.1 -C.sub.4 alkyl, or acyl or benzyl; R3 and R4 are linear or branched C.sub.1 -C.sub.4 alkyls, or benzyl or methoxyethyl, methoxypropyl, methoxybutyl.
The raw material used for the preparation of the esters according to the present invention is eseroline (II) ##STR5## hereinafter also referred to a ESOH. The relative preparation process is characterized by the following steps:
a) heterocyclic compound RH, where R is as defined above, is caused to react with .OMEGA.-haloalkylphthalimide as per formula (III) ##STR6## where X is selected out of the group consisting of F, Cl, Br, and I;
b) the phthalic group of the obtained compound is removed by hydrazinolysis to form a primary amino derivative as per formula (IV)
R(CH.sub.2).sub.n NH.sub.2 (IV)
c) carbonyl diimidazole is caused to react first with eseroline and then with the aforesaid amino derivative according to the following reactions: ##STR7## Compound (I), where R1=H, is thus obtained. For the preparation of compound (I), where R1=alkyl group, before proceeding to step c), the amino derivatives (IV) is alkylated to form the corresponding secondary amino derivative.
Compounds as per general formula (I) and their pharmacologically acceptable salts exhibit a higher cholinesterase activity depression and reduced side effects if compared with the compounds mentioned in the prior art. Therefore, the present invention also contemplates the use of compounds as per general formula (I) and of their pharmacologically acceptable salts in the preparation of pharmaceutical compositions suitable for therapeutic use because of their inhibiting acetylcholinesterase action.

DETAILED DESCRIPTION OF THE INVENTION
The characteristics of and the advantages offered by the aminoalkylcarbamic esters of eseroline suitable for use as cholinesterase activity inhibitors and the relative preparation process according to the present invention are going to be illustrated in more detail.
In the present patent the term eseroline indicates (3aS,8aR)- 1,2,3,3a,8,8a-hexahydro-1,3a,8-trimethylpyrrol[2,3-b]indol-5-ol. Said compounds have the following general formula (I) ##STR8## where n is an integer from 2 to 20; R1 may be H, methyl, ethyl, propyl, and isopropyl; and R is an amine selected out of the group consisting of ##STR9## where R2 is linear or branched C.sub.1 -C.sub.4 alkyl, or acyl or benzyl; R3 and R4 are linear or branched C.sub.1 -C.sub.4 alkyls, or benzyl or methoxyethyl, methoxypropyl, methoxybutyl.
Compounds as per general formula (I) are prepared according to the following process:
a) heterocyclic compound RH, where R is as defined above, is caused to react with .OMEGA.-haloalkylphthalimide as per formula (III) ##STR10## where X is selected out of the group consisting of F, Cl, Br, and I. The reaction is carried out in the presence of a solvent selected among ethyl ether, acetonitrile and tetrahydrofuran, at room temperature, and with a molar ratio of RH to (III) ranging from 3:1 to 1:1. The following compound is obtained: ##STR11##
b) the compound obtained under a) is treated with hydrazine aqueous solution in order to remove the phthalic group.
The reaction is carried out in methanol and the mixture is heated to reflux. Solvents are removed by evaporation under reduced pressure and the product is purified by chromatography.
A primary amino derivatives as per formula (IV)
R(CH.sub.2).sub.n NH.sub.2 (IV)
is obtained;
c) carbonyl diimidazole is caused to react with eseroline (ESOH) according to the following raction: ##STR12## The reaction is carried out in the presence of a solvent selected among methylene chloride, toluene, benzene, xylene, DMF at room temperature, and with a molar ratio of carbonyl diimidazole to eseroline ranging from 3:1 to 1:1.
The reaction mixture is added with compound (IV) to promote the following reaction ##STR13## The reaction is carried out at room temperature with an initial molar ratio of (IV) to carbonyl diimidazole ranging from 2:1 to 1:1. The solvent is removed by evaporation under reduced pressure and the product is purified by chromatography.
Compounds (I) with R1=H are obtained with the process described hereinabove. For the preparation of compounds (I), where R1=methyl, ethyl, propyl or isopropyl, compound (IV), before being used in step c), is alkylated to form the corresponding secondary amino derivative. This can be obtained by treatment of compound (IV) with dimethylphosphate or with ethyl or propyl or isopropyl halides or by monobenzylation with benzyl bromide followed by treatment with alkyl halide and subsequent debenzylation by hydrogenolysis.
Compounds as per formula (I) can be salified with pharmacologically acceptable acids selected out of the group consisting of HCl, HBr, H.sub.2 SO.sub.4, H.sub.3 PO.sub.4, HClO.sub.4, citric acid, tartaric acid, maleic acid, salicylic acid, fumaric acid, succinic acid, and oxalic acid. Salification is obtained by dissolving the compound as per formula (I) in an organic solvent, such as ethyl ether, methanol, ethanol, isopropanol, n-butanol, diisopropyl ether, and causing the obtained solution to react with the selected acid dissolved in one of the aforesaid solvents.
For salification, the preferred solvent-reagents ratios range from 5 to 25 ml solvent/g compound (I) and from 5 to 35 ml solvent/g acid. The salt obtained is purified by the usual techniques: solvent extraction, chromatography and organic compounds crystallization. The following examples, which are concerned with the preparation of the claimed compounds, are conveyed by way of indication, not of limitation.
EXAMPLE 1
Preparation of Morpholinylethylcarbamoyleseroline
a) Preparation of 2-morpholinylethylphthalimide
2-Bromoethylphthalimide (508 mg; 2.0 mmoles) and morpholine (0.35 ml; 4.0 mmoles) were dissolved in anhydrous ethyl ether (5 ml). The mixture was allowed to stir at room temperature for 30 hours approx. The reaction was discontinued by solvent evaporation under reduced pressure. The obtained residue was purified by chromatography on silica gel (CH.sub.2 Cl.sub.2 /MeOH 95:5) yielding 271 mg of pure product.
Yield 52%.
Analysis: calcd.: C, 64.60; H, 6.20; N, 10.76. found: C, 64.51; H, 6.25; N, 10.98.
b) Preparation of 2-morpholinylethylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 2-morpholinylethylphthalimide (208 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. After one hour, the solvents were evaporated under reduced pressure; the obtained product was taken up with CHCl.sub.3 and the collected precipitate was filtered CHCl.sub.3 was evaporated under reduced pressure yielding a solid (115 mg). The product was purified by silica gel chromatography with gradient, eluting with CHCl.sub.3 /MeOH/H.sub.2 O, 65:35:5, to remove the products migrating almost at the eluent front, followed by eluting with CHCl.sub.3 /MeOH/NH.sub.4 OH, 65:35:2, to isolate the desired product of very intense colour by ninhydrin and iodine reactions.
65 mg of pure product were obtained. Yield 62%.
Analysis: calcd.: C, 55.36; H, 10.84; N, 21.51. found: C, 55.51; H, 10.55; N, 21.98.
c) Preparation of N-morpholinylethylcarbamoyleseroline
Carbonyl diimidazole (36 mg; 0.22 mmoles) was added to a degassed solution of eseroline (25 mg; 0.11 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then added with 2-morpholinylethylamine (43 mg; 0.33 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (10 ml). Stirring at room temperature was continued for additional 2 hours approx. after the addition of 2-morpholinylethylamine. The reaction course was controlled by TLC (CHCl.sub.3 /MeOH 87.13). At the end of the reaction, the solvent was evaporated under reduced pressure yielding an oil (50 mg), which was purified by column chromatography on silica gel eluting with acetone/MeOH 8:2.
8 mg of pure product were obtained. Yield 20%.
.sup.1 H-NMR (200 MHz, CDCl.sub.3): .delta. 6.60 (m, 2H); 6.35 (d, 1H); 5.45 (s broad, 1H); 4.16 (s, 1H); 3.70 (m, 4H); 3.37 (m, 2H); 2.93 (s, 3H); 2.55 (s, 3H); 2.60-2.45 (m, 8H); 1.98 (m, 2H); 1.43 (s, 3H).
Analysis: calcd.: C, 64.15; H, 8.08; N, 14.95. found: C, 64.52; H, 8.25; N, 14.98.
EXAMPLE 2
Preparation of Morpholinylpropylcarbamoyleseroline
a) Preparation of 3-morpholinylpropylphthalimide
3-Bromopropylphthalimide (536 mg; 2.0 mmoles) and morpholine (0.35 ml; 4.0 mmoles) were dissolved in anhydrous ethyl ether (5 ml). Reaction times and process as per Example 1.
291 mg of pure product were thus obtained. Yield 53%
Analysis: calcd.: C, 65.68; H, 6.61; N, 10.21. found: C, 65.61; H, 6.75; N, 10.18.
b) Preparation of 3-morpholinylpropylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 3-morpholinylpropylphthalimide (219 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
75 mg of pure product were obtained. Yield 65%.
Analysis: calcd.: C, 58.30; H, 11.18; N, 19.42. found: C, 58.00; H, 11.25; N, 19.88.
c) Preparation of N-morpholinylpropylcarbamoyleseroline
Carbonyl diimidazole (36 mg; 0.22 mmoles) was added to a degassed solution of eseroline (25 mg; 0.11 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 3-morpholinylpropylamine (48 mg; 0.33 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 3-morpholinylpropylamine. Reaction process as per Example 1.
16 mg of product were obtained. Yield 38%.
.sup.1 H-NMR (200 MHz, CDCl.sub.3): .delta. 6.85 (m, 2H); 6.50 (d, 1H); 6.27 (s broad, 1H); 4.70 (s, 1H); 3.78 (t, 4H); 3.38 (m, 2H); 3.02 (s, 3H); 2.65 (s, 3H); 2.70-2.30 (m, 8H); 2.15 (m, 2H); 1.77 (m, 2H); 1.55 (s, 3H).
Analysis: calcd.: C, 64.93; H, 8.30; N, 14.41. found: C, 64.81; H, 8.15; N, 14.70.
EXAMPLE 3
Preparation of Morpholinylbutylcarbamoyleseroline
a) Preparation of 4-morpholinylbutylphthalimide
4-Bromobutylphthalimide (564 mg; 2.0 mmoles) and morpholine (0.35 ml; 4.0 mmoles) were dissolved in anhydrous ethyl ether (5 ml). Reaction times and process as per Example 1.
317 mg of pure product were obtained. Yield 55%
Analysis: calcd.: C, 66.65; H, 6.99; N, 9.71. found: C, 66.41; H, 6.77; N, 9.99.
b) Preparation of 4-morpholinylbutylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 4-morpholinylbutylphthalimide (231 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
86 mg of pure product were obtained. Yield 68%
Analysis: calcd.: C, 60.73; H, 11.47; N, 17.70. found: C, 60.53; H, 11.27; N, 17.45.
c) Preparation of N-morpholinylbutylcarbamoyleseroline
Carbamoyl diimidazole (36 mg; 0.22 mmoles) was added to a degassed solution of eseroline (25 mg; 0.11 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 4-morpholinylbutylamine (52 mg; 0.33 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 4-morpholinylbutylamine. Reaction times and process as per Example 1.
14 mg of pure product were obtained. Yield 32%.
.sup.1 H-NMR (200 MHz, CDCl.sub.3): .delta. (m, 2H); 6.34 (d, 1H); 5.92 (s broad, 1H); 4.15 (s, 1H); 3.73 (t, 4H); 3.25 (m, 2H); 2.88 (s, 3H); 2.55 (s, 3H); 2.80-2.33 (m, 8H); 1.95 (m, 2H); 1.75 (m, 2H); 1.58 (m, 2H); 1.45 (s, 3H).
Analysis: calcd.: C, 65.65, H, 8.51; N, 13.91. found: C, 65.55; H, 8.33; N, 14.00.
EXAMPLE 4
Preparation of Morpholinylpentylcarbamoyleseroline
a) Preparation of 5-bromopentylphthalimide
Potassium phthalimide (5.4 mmoles) was added to a solution of 1.5-dibromopentane (3.72 g; 16.2 mmoles) in dimethylformamide (8 ml) and heated to 80.degree. C. approx. under stirring. After 2 hours, water was added and the solution was repeatedly extracted with ether. The collected organic phases were washed with water, dried on Na.sub.2 SO.sub.4 and evaporated under reduced pressure to give a solid (4.50 g), which was purified by chromatography on silica gel (Etp/Et.sub.2 O 8:2) yielding 2.38 g of a white solid. Yield 68%.
Analysis: calcd.: C, 72.20; H, 6.53; N, 6.47. found: C, 72.01; H, 6.55; N, 6.32.
b) Preparation of 5-morpholinylpentylphthalidmide
5-Bromopentylphthalimide (432 mg; 2.0 mmoles) and morpholine (0.35 ml; 4.0 mmoles) were dissolved in anhydrous ethyl ether (5 ml). Reaction times and process as per Example 1.
326 mg of pure product were obtained. Yield 54%.
Analysis: calcd.: C, 67.53; H, 7.33; N, 9.26. found: C, 67.51; H, 7.25; N, 9.28.
c) Preparation of 5-morpholinylpentylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 5-morpholinylpentylphthalimide (242 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
81 mg of pure product were obtained. Yield 59%.
Analysis: calcd.: C, 62.75; H, 11.70; N, 16.25. found: C, 62.87; H, 11.83; N, 16.38.
d) Preparation of N-morpholinylpentylcarbamoyleseroline
Carbamoyl diimidazole (36 mg; 0.22 mmoles) was added to a degassed solution of eseroline (25 mg; 0.11 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 8-morpholinyloctylamine (57 mg; 0.33 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 8-morpholinyloctylamine. Reaction times and process as per Example 1.
27 mg of pure product were obtained. Yield 60%.
Analysis: calcd.: C, 66.32; H, 8.71; N, 13.44. found: C, 66.31; H, 8.88; N, 13.44.
EXAMPLE 5
Preparation of Morpholinylhexylcarbamoyleseroline
a) Preparation of 6-bromohexylphthalimide
Potassium phthalimide (5.4 mmoles) was added to a solution of 1,6-dibromohexane (3.95 g; 16.2 mmoles) in dimethylformamide (8 ml) and heated to 80.degree. C. approx. under stirring. Reaction times and process as per Example 4.
2.46 g of pure product were obtained. Yield 66%.
Analysis: calcd.: C, 73.02; H, 7.00; N, 6.08. found: C, 72.83; H, 7.25; N, 5.98.
b) Preparation of 6-morpholinylhexylphthalimide
6-Bromohexylphthalimide (633 mg; 2.0 mmoles) and morpholine (0.35 mg; 4.0 mmoles) were dissolved in anhydrous ethyl ether (5 ml). Reaction times and process as per Example 1.
342 mg of pure product were obtained. Yield 54%.
Analysis: calcd.: C, 68.33; H, 7.65; N, 8.85. found: C, 68.22; H, 7.75; N, 8.99.
c) Preparation of 6-morpholinylhexylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 6-morpholinylhexylphthalimide (253 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
94 mg of pure product were obtained. Yield 63%.
Analysis: calcd.: C, 64.48; H, 11.90; N, 15.03. found: C, 64.52; H, 12.01; N, 14.97.
d) Preparation of N-morpholinylhexylcarbamoyleseroline
Carbamoyl diimidazole (36 mg; 0.22 mmoles) was added to a degassed solution of eseroline (25 mg; 0.11 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 6-morpholinylhexylamine (61 mg; 0.33 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 6-morpholinylhexylamine. Reaction times and process as per Example 1.
24 mg of pure product were obtained. Yield 51%.
Analysis: calcd.: C, 66.95; H, 8.90; N, 13.01. found: C, 67.02; H, 8.99; N, 13.21.
EXAMPLE 6
Preparation of Morpholinylheptylcarbamoyleseroline
a) Preparation of 7-bromoheptylphthalimide
Potassium phthalimide (5.4 mmoles) was added to a solution of 1,7-dibromoheptane (4.18 g; 16.2 mmoles) in dimethylformamide (8 ml) and heated to 80.degree. C. approx. under stirring. Reaction times and process as per Example 4.
2.73 g of pure product were obtained. Yield 69%.
Analysis: calcd.: C, 73.74; H, 7.43; N, 5.73. found: C, 73.70; H, 7.24; N, 5.77.
b) Preparation of 7-morpholinylheptylphthalimide
7-Bromoheptylphthalimide (489 mg; 2.0 mmoles) and morpholine (0.35 ml; 4.0 mmoles) were dissolved in anhydrous acetonitrile (5 ml). Reaction times and process as per Example 1.
324 mg of pure product were obtained. Yield 49%.
Analysis: calcd.: C, 69.01; H, 7.93; N, 8.47. found: C, 68.91; H, 8.06; N, 8.57.
c) Preparation of 7-morpholinylheptylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 7-morpholinylheptylphthalimide (264 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
99 mg of pure product were obtained. Yield 62%.
Analysis: calcd.: C, 65.96; H, 12.08; N, 13.98. found: C, 66.13; H, 11.89; N, 13.98.
d) Preparation of N-morpholinylheptylcarbamoyleseroline
Carbonyl diimidazole (36 mg; 0.22 mmoles) was added to a solution of eseroline (25 mg; 0.11 mmoles) in anhydrous toluene (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 8-morpholinyloctylamine (66 mg; 0.33 mmoles) in anhydrous toluene (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 8-morpholinyloctylamine. Reaction times and process as per Example 1.
24 mg of pure product were obtained. Yield 51%.
Analysis: calcd.: C, 67.54; H, 9.07; N, 12.60. found: C, 67.51; H, 9.15; N, 12.97.
EXAMPLE 7
Preparation of Morpholinyloctylcarbamoyleseroline
a) Preparation of 8-bromooctylphthalimide
Potassium phthalimide (5.4 mmoles) was added to a solution of 1,8-dibromooctane (4.41 g; 16.2 mmoles) in dimethylformamide (8 ml) and heated to 80.degree. C. approx. under stirring. Reaction times and process as per Example 4.
2.8 g of pure product were obtained. Yield 67%.
Analysis: calcd.: C, 74.39; H, 7.80; N, 5.42. found: C, 74.54; H, 7.78; N, 5.08.
b) Preparation of 8-morpholinyloctylphthalidmide
8-Bromooctylphthalimide (676 mg; 2.0 mmoles) and morpholine (0.35 ml; 4.0 mmoles) were dissolved in anhydrous acetonitrile (5 ml). Reaction times and process as per Example 1.
331 mg of pure product were obtained. Yield 48%.
Analysis: calcd.: C, 69.74; H, 8.19; N, 8.13. found: C, 70.05; H, 8.25; N, 8.00.
c) Preparation of 8-morpholinyloctylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 8-morpholinyloctylphthalimide (275 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
102 mg of pure product were obtained. Yield 60%.
Analysis: calcd.: C, 67.27; H, 12.23; N, 13.06. found: C, 67.02; H, 12.25; N, 13.08.
d) Preparation of N-morpholinyloctylcarbamoyleseroline
Carbonyl diimidazole (36 mg; 0.22 mmoles) was added to a degassed solution of eseroline (25 mg; 0.11 mmoles) in anhydrous toluene (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 8-morpholinyloctylamine (70 mg; 0.33 mmoles) in anhydrous toluene (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 8-morpholinyloctylamine. Reaction and process as per Example 1.
36 mg of pure product were obtained. Yield 72%.
.sup.1 H-NMR (200 MHz, CDCl.sub.3): .delta. 6.78 (m, 2H); 6.32 (d, 1H); 4.92 (s broad, 1H); 4.18 (s, 1H); 3.72 (m, 4H); 3.20 (m, 2H); 2.91 (s, 3H); 2.55 (s, 3H); 2.48-2.23 (m, 8H); 1.92 (m, 4H); 1.45 (s, 3H); 1.31 (m, 12H).
Analysis: calcd.: C, 68.09; H, 9.23; N, 12.21. found: C, 68.00; H, 9.00; N, 12.22.
EXAMPLE 8
Preparation of Morpholinyldodecylcarbamoyleseroline
a) Preparation of 12-bromododecylphthalimide
Potassium phthalimide (1 g; 5.4 mmoles) was added to a solution of 1,8-dibromododecane (5.31 g; 16.2 mmoles) in dimethylformamide (8 ml) and heated to 80.degree. C. approx. under stirring. Reaction times and process as per Example 4.
3.46 g of pure product were obtained. Yield 68%.
Analysis: calcd.: C, 76.40; H, 8.98; N, 4.45. found: C, 76.51; H, 9.12; N, 4.06.
b) Preparation of 12-morpholinyldodecylphthalimide
12-Bromododecylphthalimide (623 mg; 2.0 mmoles) and morpholine (0.35 ml; 4.0 mmoles) were dissolved in anhydrous ethyl ether (5 ml). Reaction times and process as per Example 1.
392 mg of pure product were obtained. Yield 49%.
Analysis: calcd.: C, 71.97; H, 9.06; N, 6.99. found: C, 72.07; H, 8.94; N, 7.17.
c) Preparation of 12-morpholinyldodecylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 12-morpholinyldodecylphthalimide (320 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
147 mg of pure product were obtained. Yield 68%.
Analysis: calcd.: C, 71.06; H, 12.67; N, 10.35. found: C, 70.89; H, 12.33; N, 10.43.
d) Preparation of N-morpholinyldodecylcarbamoyleseroline
was added to a degassed solution of eseroline (25 mg; 0.11 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 12-morpholinyldodecylamine (89 mg; 0.33 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 12-morpholinyldodecylamine. Reaction times and process as per Example 1.
27 mg of pure product were obtained. Yield 48%.
Analysis: calcd.: C, 70.00; H, 9.79; N, 10.88. found: C, 70.01; H, 9.99; N, 10.98.
EXAMPLE 9
Preparation of N-Methylpiperazinylethylcarbamoyleseroline
a) Preparation of 2-(N-methylpiperazinyl)ethylphthalimide
2-Bromoethylphthalimide (508 mg; 2.0 mmoles) and N-methylpiperazine (0.44 mg; 4.0 mmoles) were dissolved in anhydrous ethyl ether (5 ml). Reaction times and process as per Example 1.
272 mg of pure product were obtained. Yield 50%.
Analysis: calcd.: C, 66.16; H, 6.66; N, 15.42. found: C, 65.97; H, 6.65; N, 15.55.
b) Preparation of 2-(N-methylpiperazinyl)ethylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 2-(N-methylpiperazinyl)ethylphthalimide (218 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
73 mg of pure product were obtained. Yield 64%.
Analysis: calcd.: C, 58.71; H, 11.97; N, 29.33. found: C, 58.91; H, 12.00; N, 29.35.
c) Preparation of N-(N-methylpiperazinylethyl)carbamoyleseroline
Carbonyl diimidazole (36 mg; 0.22 mmoles) was added to a solution of eseroline (25 mg; 0.11 mmoles) in anhydrous benzene (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 2-(N-methylpiperazinyl)ethylamine (47 mg; 0.33 mmoles) in anhydrous benzene (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 2-(N-methylpiperazinyl)ethylamine. Reaction times and process as per Example 1.
19 mg of pure product were obtained. Yield 45%.
Analysis: calcd.: C, 65.09; H, 8.58; N, 18.06. found: C, 65.12; H, 8.73; N, 18.22.
EXAMPLE 10
Preparation of N-Methylpiperazinylpropylcarbamoyleseroline
a) Preparation of 3-(N-methylpiperazinyl)propylphthalimide
3-Bromopropylphthalimide (536 mg; 2.0 mmoles) and N-methylpiperazine (0.44 ml; 4.0 mmoles) were dissolved in anhydrous ethyl ether (5 ml). Reaction times and process as per Example 1.
292 mg of pure product were obtained. Yield 51%.
Analysis: calcd.: C, 67.12; H, 7.04; N, 14.67. found: C, 66.99; H, 7.00; N, 14.87.
b) Preparation of 3-(N-methylpiperazinyl)propylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 3-(N-methylpiperazinyl)propylphthalimide (229 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
80 mg of pure product were obtained. Yield 64%.
Analysis: calcd.: C, 61.11; H, 12.18; N, 26.71. found: C, 61.01; H, 12.35; N, 27.02.
c) Preparation of N-(N-methylpiperazinylpropyl)carbamoyleseroline
Carbonyl diimidazole (36 mg; 0.22 mmoles) was added to a solution of eseroline (25 mg; 0.11 mmoles) in anhydrous benzene (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 3-(N-methylpiperazinyl)propylamine (52 mg; 0.33 mmoles) in anhydrous benzene (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 3-(N-methylpiperazinyl)propylamine. Reaction times and process as per Example 1.
8 mg of pure product were obtained. Yield 19%.
Analysis: calcd.: C, 65.81; H, 8.79; N, 17.43. found: C, 65.51; H, 9.05; N, 17.77.
EXAMPLE 11
Preparation of N-Methylpiperazinylbutylcarbamoyleseroline
a) Preparation of 4-(N-methylpiperazinyl)butylphthalimide
4-Bromoethylphthalimide (564 mg; 2.0 mmoles) and N-methylpiperazine (0.44 ml; 4.0 mmoles) were dissolved in anhydrous ethyl ether (5 ml). Reaction times and process as per Example 1.
288 mg of pure product were obtained. Yield 48%.
Analysis: calcd.: C, 67.98; H, 7.38; N, 13.98. found: C, 68.14; H, 7.57; N, 14.07.
b) Preparation of 4-(N-methylpiperazinyl)butylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 4-(N-methylpiperazinyl)butylphthalimide (240 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
90 mg of pure product were obtained. Yield 66%.
Analysis: calcd.: C, 63.12; H, 12.36; N, 24.52. found: C, 63.31; H, 12.56; N, 24.44.
c) Preparation of N-(N-methylpiperazinylbutyl)carbamoyleseroline
Carbonyl diimidazole (36 mg; 0.22 mmoles) was added to a solution of eseroline (25 mg; 0.11 mmoles) in anhydrous benzene (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 4-(N-methylpiperazinyl)butylamine (56 mg; 0.33 mmoles) in anhydrous benzene (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 4-(N-methylpiperazinyl)butylamine. Reaction times and process as per Example 1.
20 mg of pure product were obtained. Yield 44%.
Analysis: calcd.: C, 66.48; H, 8.98; N, 16.85. found: C, 66.33; H, 9.02 N, 17.00.
EXAMPLE 12
Preparation of N-Methylpiperazinylhexylcarbamoyleseroline
a) Preparation of 6-(N-methylpiperazinyl(hexylphthalimide
6-Bromohexylphthalimide (460 mg; 2.0 mmoles) and N-methylpiperazine (0.44 ml; 4.0 mmoles) were dissolved in anhydrous ethyl ether (5 ml). Reaction times and process as per Example 1.
302 mg of pure product were obtained. Yield 46%.
Analysis: calcd.: C, 69.49; H, 7.98; N, 12.98. found: C, 69.25; H, 8.13; N, 13.08.
b) Preparation of 6-(N-methylpiperazinyl)hexylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 6-(N-methylpiperazinyl)hexylphthalimide (263 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
112 mg of pure product were obtained. Yield 70%.
Analysis: calcd.: C, 66.29; H, 12.64; N, 21.07. found: C, 66.07; H, 12.77; N, 20.99.
c) Preparation of N-(N-methylpiperazinylhexy)carbamoyleseroline
Carbonyl diimidazole (36 mg; 0.22 mmoles) was added to a solution of eseroline (25 mg; 0.11 mmoles) in anhydrous benzene (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 6-(N-methylpiperazinyl)hexylamine (66 mg; 0.33 mmoles) in anhydrous benzene (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 6-(N-methylpiperazinyl)hexylamine. Reaction times and process as per Example 1.
30 mg of pure product were obtained. Yield 60%.
Analysis: calcd.: C, 67.69; H, 9.32; N, 15.78. found: C, 68.00; H, 9.33; N, 15.97.
EXAMPLE 13
Preparation of N-Methylpiperazinyloctylcarbamoyleseroline
a) Preparation of 8-(N-methylpiperazinyl)octylphthalimide
8-Bromooctylphthalimide (516 mg; 2.0 mmoles) and N-methylpiperazine (0.44 ml; 4.0 mmoles) were dissolved to anhydroius ethyl ether (5 ml). Reaction times and process as per Example 1.
356 mg of pure product were obtained. Yield 50%.
Analysis: calcd.: C, 70.76; H, 8.48; N, 11.78. found: C, 70.80; H, 8.53; N, 11.83.
b) Preparation of 8-(N-methylpiperazinyl)octylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 8-(N-methylpiperazinyl)octylphthalimide (285 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
118 mg of pure product were obtained. Yield 65%.
Analysis: calcd.: C, 68.67; H, 12.86; N, 18.47. found: C, 68.50; H, 12.66; N, 18.54.
c) Preparation of N-(N-methylpiperazinyloctyl)carbamoyleseroline
Carbonyl diimidazole (36 mg; 0.22 mmoles) was added to a solution of eseroline (25 mg; 0.11 mmoles) in anhydrous benzene (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 8-(N-methylpiperazinyl)octylamine (75 mg; 0.33 mmoles) in anhydrous benzene (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 8-(N-methylpiperazinyl)octylamine. Reaction times and process as per Example 1.
30 mg of pure product were obtained. Yield 57%.
Analysis: calcd.: C, 68.76; H, 9.62; N, 14.84. found: C, 68.43; H, 9.48; N, 14.88.
EXAMPLE 14
Preparation of Pyrrolidinyloctylcarbamoyleseroline
a) Preparation of 8-pyrrolidinyloctylphthalimide
8-Bromooctylphthalimide (517 mg; 2.0 mmoles) and pyrrolidine (0.33 ml; 4.0 mmoles) were dissolved in anhydrous ethyl ether (5 ml). Reaction times and process as per Example 1.
348 mg of pure product were obtained. Yield 53%.
Analysis: calcd.: C, 73.14; H, 8.59; N, 8.53. found: C, 73.11; H, 8.75; N, 8.54.
b) Preparation of 8-pyrrolidinyloctylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 8-pyrrolidinyloctylphthalimide (263 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
105 mg of pure product were obtained. Yield 66%.
Analysis: calcd.: C, 72.67; H, 13.21; N, 14.12. found: C, 72.72; H, 13.25; N, 14.07.
c) Preparation of N-pyrrolidinyloctylcarbamoyleseroline
Carbonyl diimidazole (36 mg; 0.22 mmoles) was added to a solution of eseroline (25 mg; 0.11 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 8-pyrrolidinyloctylamine (65 mg; 0.33 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 8-pyrrolidinyloctylamine. Reaction times and process as per Example 1.
24 mg of pure product were obtained. Yield 48%.
Analysis: calcd.: C, 70.55; H, 9.56; N, 12.65. found: C, 70.51; H, 9.55; N, 12.74.
EXAMPLE 15
Preparation of Piperidinyloctylcarbamoyleseroline
a) Preparation of 8-piperidinyloctylphthalimide
8Bromooctylphthalimide (517 mg; 2.0 mmoles) and piperazine (0.40 ml; 4.0 mmoles) were dissolved in anhydrous ethyl ether (5 ml). Reaction times and process as per Example 1.
431 mg of pure product were obtained. Yield 63%.
Analysis: calcd.: C, 73.65; H, 8.83; N, 8.18. found: C, 73.55; H, 8.95; N, 8.08.
b) Preparation of 8-piperidinyloctylamine
Hydrazine (aqueous solution at 35% by wt.) (0.15 ml; 1.6 mmoles) was added to 8-piperidinyloctylphthalimide (274 mg; 0.8 mmoles) in methanol (5 ml) and the resulting solution was refluxed. Reaction times and process as per Example 1.
119 mg of pure product were obtained. Yield 70%.
Analysis: calcd.: C, 73.53; H, 13.29; N, 13.18. found: C, 73.51; H, 13.25; N, 13.08.
c) Preparation of N-piperidinyloctylcarbamoyleseroline
Carbonyl diimidazole (36 mg; 0.22 mmoles) was added to a solution of eseroline (25 mg; 0.11 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then 8-piperidinyloctylamine (70 mg; 0.33 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of 8-piperidinyloctylamine. Reaction times and process as per Example 1.
27 mg of pure product were obtained. Yield 54%.
Analysis: calcd.: C, 71.02; H, 9.71; N, 12.26. found: C, 71.01; H, 9.99; N, 12.06.
EXAMPLE 16
Preparation of N-(Morpholinyloctyl),N-Methylcarbamoyleseroline
a) Preparation of N-(morpholinyloctyl),N-methylamine
Dimethylsulphate (832 mg; 0.66 mmoles) was slowly added to 8-morpholinooctylamine (141 mg; 0.66 mmoles) under stirring, the temperature being kept below 70.degree. C. throughout the addition. Stirring was continued overnight. Then the mixture was treated with sodium hydroxide solution (880 mg) in water (15 ml) and heated to boiling. After two hours, the mixture was cooled to room temperature. An oil was separated which was combined with the water layer benzene extracts: the mixture obtained was dried on Na.sub.2 SO.sub.4 and evaporated under reduced pressure. The collected residue was purified by silica gel chromatography, eluting with CHCl.sub.3 /MeOH/NH.sub.4 OH, 65:35:2, to separate the desired product of very intense colour by ninhydrin and iodine reactions.
75 mg of pure product were obtained. Yield 50%.
Analysis: calcd.: C, 68.37; H, 12.36; N, 12.26. found: C, 68.33; H, 12.35; N, 12.28.
b) Preparation of N-(morpholinyloctyl),N-methyl carbamoyl eseroline
Carbonyl diimidazole (36 mg; 0.22 mmoles) was added to a degassed solution of eseroline (25 mg; 0.11 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (5 ml). The solution was was allowed to stir at room temperature for 1 hour approx., then N-(morpholinyloctyl),N-methylamine (75 mg; 0.33 mmoles) in anhydrous CH.sub.2 Cl.sub.2 (10 ml) was added. Stirring at room temperature was continued for additional 2 hours approx. after the addition of N-(morpholinyloctyl),N-methylamine. Reaction process as per Example 1.
36 mg of pure product were obtained. Yield 68%.
Analysis: calcd.: C, 68.61; H, 9.38; N, 11.85. found: C, 68.52; H, 9.28; N, 11.87.
EXAMPLE 17
Preparation of 8-(cis-2,6-Dimethyl)Morpholinyloctylcarbamoyl Eseroline
a) Preparation of 8-(cis/trans-2,6-dimethyl)morpholinyloctyl phthalimide
8-Bromooctylphthalimide (9 g; 26.60 mmoles) and 2,6-dimethyl morpholine (cis-trans isomers mix) (9.8 ml; 79.5 mmoles) were dissolved in anhydrous acetonitrile (50 ml). The mixture was allowed to stir at room temperature for 18 hours approx. The reaction was discontinued by solvent evaporation under reduced pressure. The collected residue was purified by chromatography on silica gel (CH.sub.2 Cl.sub.2 /MeOH 95:5). The first product isolated (Rf =0.7) is a light yellow oil (A trans) (2.04 g), the second (Rf =0.4) is a yellow oil (B cis) (3.4 g). Total yield 54.8%.
Analysis: calcd.: C,70.93; H,8.66; N,7.52. Compound A (trans isomer) found: C,71.14; H,8.58; N,7.59. Compound B (cis isomer) found: C,71.07; H,8.53; N,7.65.
b) Preparation of 8-(cis-2,6-dimethyl)morpholinyloctylamine
Hydrazine (aqueous solution at 35% by wt.) (7 ml) was added to 8-(cis-2,6-dimethyl)morpholinyloctylphthalimide B (3.4 g; 9.12 mmoles) in methanol (25 ml) and the resulting solution was refluxed. After one hour, thte solvents were evaporated under reduced pressure, the obtained product was taken up with CHCl.sub.3 and the collected precipitate was filtered. CHCl.sub.3 was evaporated under reduced pressure yielding a light yellow oil (1.614 g). Yield 73%.
Analysis: calcd.; C,69.37; H,12.47; N,11.46. found: C,69.50; H,12.45; N,11.63.
c) Preparation of 8-(cis-2,6-dimethyl)morpholinyloctylcarbamoyl eseroline
Carbonyl diimidazole (719 mg; 4.44 mmoles) was added to a solution of eseroline (484 mg; 2.22 mmoles) in benzene (5 ml). The solution was allowed to stir at room temperature for 1 hour approx., then added with 8-(cis-2,6-dimethyl) morholinyloctylamine (1.614 mg; 6.66 mmoles) in anhydrous benzene (2 ml). Stirring at room temperature was continued for additional 2 hours approx. after the addition of 8-(cis-2,6-dimethyl)morpholinyloctytlamine. The reaction course was controlled by TLC (CHCl.sub.3 /MeOH 9:1). At the reaction end, the solvents were evaporated under reduced pressure yielding an oil (2.7 g), which was purified by column chromatography on silica gel eluting first with CHCl.sub.3 /MeOH, 9:1, and then with acetone/MeOH, 7:3.
An orange coloured oil were obtained (1.6 g). Yield 55.5%.
Analysis: calcd.: C,69.10; H,9.53; N,11.51. found: C,69.23; H,9.47; N,11.59.
EXAMPLE 18
Preparation of 8-(Trans-2,6-Dimethyl)Morpholinyloctylcarbamoyl Eseroline
a) Preparation of 8-(trans-2,6-dimethyl)morpholinyloctylamine
Hydrazine (aqueous solution at 35% by wt.) (5 ml) was added to 8-(trans-2,6-dimethyl)morpholinyloctylphthalimide A (2.04 g; 5.47 mmoles), obtained as per Example 17a, in methanol (20 ml) and the resulting solution was refluxed. After 1 hour, the solvents were evaporated under reduced pressure and the obtained product was taken up with CHCl.sub.3 under reduced pressure.
976 mg of a light yellow oil were obtained. Yield 73%.
Analysis: calcd.: C,69.37; H,12.47; N,11.56. found: C,69.50; H,12.45; N,11.63.
b) Preparation of 8-(trans-2,6-dimethyl)morpholinyloctylcarbamoyl eseroline
A solution of eseroline (293 mg; 1.34 mmoles) in anhydrous benzene (5 ml) was added with carbonyl diimidazole (435 mg; 2.68 mmoles). The solution was allowed to stir at room temperature for 1 hour approx., then added with 8-(trans-2,6-dimethyl) morpholinyloctylamine (975 mg: 4.03 mmoles) in anhydrous benzene (2 ml). Stirring at room temperature was continued for additional 2 hours approx. after the addition of 8-(trans-2,6-dimethyl)morpholinyloctylamine. The reaction course was controlled by TLC (CHCl.sub.3 /MeOH 9:11). At the reaction end, the solvents were evaporated under reduced pressure yielding an oil (1.7 g), which was purified by column chromatography on silica gel eluting with CHCl.sub.3 /MeOH, 9:1, and then with actone/MeOH, 7:3. An orange coloured oil was obtained (268 mg). Yield 44%.
Analysis: calcd.: C,69.10; H,9.53; N,11.54.
found: C,69.31; H,9.61; N,11.45.
Other compounds obtained according to the present invention are Shown in Table 1.
TABLE 1__________________________________________________________________________Further examples of compounds obtained according to the inventionStructural formula Preparation Number__________________________________________________________________________ ##STR14## MF 233 V ##STR15## MF 231 VI ##STR16## MF 232 VII ##STR17## MF 239 VIII ##STR18## MF 265 IX ##STR19## MF 252 X ##STR20## MF 275 XI ##STR21## MF 245 XII ##STR22## MF 262 XIII ##STR23## MF 261 XIV ##STR24## MF 273 XV ##STR25## MF 270 XVI ##STR26## MF 237 XVII ##STR27## MF 247 XVIII ##STR28## MF 254 XIX ##STR29## MF 266 XX ##STR30## MF 229 XXI ##STR31## MF 255 XXII ##STR32## MF 246 XXIII ##STR33## MF 240 XXIV ##STR34## MF 269 XXV ##STR35## MF 277 XXVI ##STR36## MF 253 XXVII ##STR37## MF 256 XXVIII ##STR38## MF 257 XXIX ##STR39## MF 258 XXX ##STR40## MF 271 XXXI ##STR41## MF 263 XXXII ##STR42## MF 248 XXXIII ##STR43## MF 249 XXXIV ##STR44## MF 250 XXXV__________________________________________________________________________
PHARMACOLOGICAL TESTS
The compounds prepared as per the Examples reported above and several other compounds under the present invention (Table 1) were pharmacologically tested with a view to determining their inhibitory effect on acetylcholinesterase and butyrylcholinesterase activity. The used method had been developed by Ellman et al. (Ellman, G. L., Courtney, K. D., Andres, V., Featherstone, R. M.: "A New and Rapid Colorimetric Determination of Acetylcholinesterase Activity", Biochem. Pharm. 7, 88, 1961).
The tests were conducted on acetylcholinesterase from human erythrocyte and, respectively, from electric eel, and on butyrylcholinesterase from human serum, all produced by Sigma Chemical.
Table 2 shows the results obtained and, for purpose of comparison, the results obtained with the product of Example 30 of EPA 0253372, with physostigmine and heptastigmine.
TABLE 2__________________________________________________________________________Cholinesterase InhibitionIC.sub.50 MCompound t AChE E.eel t AChE hRBC t AChE rat brain t BuChE hserum__________________________________________________________________________MF 215 SF-92-057 10' N.D. 10' 1.9 .times. 10.sup.-5 10' N.D. 10' 8.7 .times. 10.sup.-4Ex 30 (EPA0253372) 30' 1.2 .times. 10.sup.-5Hoechst-Roussel 60' 1.6 .times. 10.sup.-5MF 214 10' N.D. 10' 7.0 .times. 10.sup.-7 10' N.D. 10' 4.1 .times. 10.sup.-6Ex. 1 30' 1.4 .times. 10.sup.-7 60' 1.1 .times. 10.sup.-7MF 218 10' N.D. 10' 3.8 .times. 10.sup.-6 10' N.D. 10' N.D.Ex. 2MF 225 10' 3.3 .times. 10.sup.-4 10' 7.6 .times. 10.sup.-6 10' 5.0 .times. 10.sup.-4 10' N.D.Ex. 4 30' 2.2 .times. 10.sup.- 6MF 227 10' 1.2 .times. 10.sup.-5 10' 9.3 .times. 10.sup.-7 10' 8.2 .times. 10.sup.-6 10' 1.0 .times. 10.sup.-7Ex. 5 30' 3.8 .times. 10.sup.-7MF 224 10' 1.6 .times. 10.sup.-6 10' 3.8 .times. 10.sup.-7 10' 4.1 .times. 10.sup.-6 10' 3.0 .times. 10.sup.-7Ex. 6 30' 1.7 .times. 10.sup.-7MF 217 10' 3.3 .times. 10.sup.-7 10' 1.0 .times. 10.sup.-7 10' 8.8 .times. 10.sup.-7 10' 1.7 .times. 10.sup.-7Ex. 7 30' 7.6 .times. 10.sup.-8 60' 6.3 .times. 10.sup.-8MF 228 10' 7.0 .times. 10.sup.-8 10' 7.0 .times. 10.sup.-8 10' 3.8 .times. 10.sup.-7 10' 1.0 .times. 10.sup.-7Ex. 8 30' 6.0 .times. 10.sup.-8 60' 3.0 .times. 10.sup.-8MF 230 10' 2.0 .times. 10.sup.-7 10' 3.0 .times. 10.sup.-8Ex. 13 30' 8.0 .times. 10.sup.-8 30' 5.0 .times. 10.sup.-8MF 201 10' 5.5 .times. 10.sup.-7 10' 2.9 .times. 10.sup.-7 10' N.D. 10' 4.0 .times. 10.sup.-8Heptastigmine 30' 4.1 .times. 10.sup.-7 60' 4.4 .times. 10.sup.-7ESERINE T. 10' 1.5 .times. 10.sup.-7 10' 4.7 .times. 10.sup.-8 10' N.D. 10' 7.2 .times. 10.sup.-8Fe-A-24A 30' 2.9 .times. 10.sup.-8Physostigmine 60' 0.4 .times. 10.sup.-8MF 251 10' N.D. 10' 0.7 .times. 10.sup.-7 10' N.D. 10' N.D.Ex. 14MF 241 10' N.D. 10' 1.0 .times. 10.sup.-7 10' N.D. 10' N.D.Ex. 15MF 243 10' N.D. 10' 7.4 .times. 10.sup.-6 10' N.D. 10' N.D.Ex. 16MF 268 10' N.D. 10' 0.7 .times. 10.sup.-7 10' N.D. 10' 1.8 .times. 10.sup.-7Ex. 17MF 276 10' N.D. 10' 0.7 .times. 10.sup.-7 10' N.D. 10' 2.5 .times. 10.sup.-7Ex. 18MF 233 10' 1.4 .times. 10.sup.-7 10' 0.6 .times. 10.sup.-7 10' N.D. 10' N.D.MF 231 10' 0.6 .times. 10.sup.-7 10' 0.4 .times. 10.sup.-7 10' N.D. 10' N.D.VIMF 232 10' 0.7 .times. 10.sup.-7 10' 0.4 .times. 10.sup.-7 10' N.D. 10' N.D.VIIMF 245 10' N.D. 10' 5.5 .times. 10.sup.-6 10' N.D. 10' N.D.XIIMF 229 10' 0.4 .times. 10.sup.-7 10' 0.6 .times. 10.sup.-7 10' N.D. 10' N.D.XXIMF 255 10' N.D. 10' 0.5 .times. 10.sup.-7 10' N.D. 10' N.D.XXIIMF 246 10' N.D. 10' 2.0 .times. 10.sup.-5 10' N.D. 10' N.D.XXIIIMF 240 10' N.D. 10' 2.4 .times. 10.sup.-7 10' N.D. 10' N.D.XXIVMF 252 10' N.D. 10' 2.7 .times. 10.sup.-8 10' N.D. 10' N.D.XMF 275 10' N.D. 10' 0.5 .times. 10.sup.-7 10' N.D. 10' N.D.XIMF 263 10' N.D. 10' 0.5 .times. 10.sup.-7 10' N.D. 10' N.D.XXXIIMF 266 10' N.D. 10' 0.7 .times. 10.sup.-7 10' N.D. 10' N.D.XXMF 262 10' N.D. 10' 0.3 .times. 10.sup.-7 10' N.D. 10' N.D.XIIIMF 261 10' N.D. 10' 0.2 .times. 10.sup.-7 N.D. 10' N.D.XIVMF 269 10' N.D. 10' 1.2 .times. 10.sup.-7 10' N.D. 10' N.D.XXVMF 270 10' N.D. 10' 0.6 .times. 10.sup.-7 10' N.D. 10' N.D.XVIMF 271 10' N.D. 10' 0.5 .times. 10.sup.-7 10' N.D. 10' N.D.XXXIMF 273 10' N.D. 10' 0.3 .times. 10.sup.-7 10' N.D. 10' N.D.XVMF 247 10' N.D. 10' 1.7 .times. 10.sup.-8 10' N.D. 10' N.D.XVIIIMF 250 10' N.D. 10' 0.3 .times. 10.sup.-7 10' N.D. 10' N.D.XXXVMF 253 10' N.D. 10' 0.3 .times. 10.sup.-7 10' N.D. 10' N.D.XXVIIMF 248 10' N.D. 10' 1.9 .times. 10.sup.-8 10' N.D. 10' N.D.XXXIIIMF 249 10' N.D. 10' 0.5 .times. 10.sup.-7 10' N.D. 10' N.D.XXXIVMF 239 10' N.D. 10' 0.5 .times. 10.sup.-7 10' N.D. 10' N.D.VIIIMF 237 10' 3.4 .times. 10.sup.-7 10' 1.1 .times. 10.sup.-7 10' N.D. 10' N.D.XVII 30' 1.9 .times. 10.sup.-7 30' 0.7 .times. 10.sup.-7 60' 1.2 .times. 10.sup.-7 60' 0.3 .times. 10.sup.-7MF 254 10' N.D. 10' 0.5 .times. 10.sup.-7 10' N.D. 10' N.D.XIXMF 256 10' N.D. 10' 0.8 .times. 10.sup.-7 10' N.D. 10' N.D.XXVIIIMF 257 10' N.D. 10' 0.3 .times. 10.sup.-7 10' N.D. 10' N.D.XXIXMF 258 10' N.D. 10' 1.0 .times. 10.sup.-6 10' N.D. 10' N.D.XXXMF 265 10' N.D. 10' 0.8 .times. 10.sup.-7 10' N.D. 10' N.D.IX__________________________________________________________________________
Legend
AChE E.eel: acetylcholinesterase from electric eel
AChE nRBC: acetylcholinesterase from human red blood cells
AChE rat brain: acetylcholinesterase from homogenate of in toto rat brain
BuChE hserum: butyrylcholinesterase from human serum
N.D.: not determined
t: contact time (minutes)
IC.sub.50 M: median inhibitory molar concentration
The results reported in Table 2 demonstrate that the inhibition of acetylcholinesterase activity by the claimed products is high. Particularly active against human red blood cells acetylcholinesterase are the compounds where R stands for
morpholine where n ranges from 8 to 12 (Ex.7), (V), (VI)
2,6-dimethylmorpholine where n=8 (Ex.17), (Ex.18)
thiomorpholine where n=8 (VIII)
methylpiperazine where n=8 (Ex.13), n=10 (XXII), n=12 (XXI)
ethylpiperazine where n=8 (XIII)
propylpiperazine where n=8 (XIV)
butylpiperazine where n=8 (XV)
acetylpiperazine where n=10 (X)
benzylpiperazine where n=8 (XX)
piperidine where n=8 (Ex.15), n=10 (XXXIII), n=12 (XXXIV)
1,4-dioxa-8-azaspirodecane where n=8 (IX)
pyrrolidine where n=8 (Ex.14) and n=10 (XXVII)
diethylamine where n=8 (XVII), n=10 (XVIII), n=12 (XIX)
dipropylamine where n=8 (XXVIII), n=10 (XXIX)
methylbenzylamine where n=8 (XXXII)
bis(methoxy)ethylamine where n=8 (XXXI).
Compared with the compounds of this invention, the compound of Example 30 (EPA 0253372) exhibits a much lower activity.
In the tests with BuChE hserum, the compounds of the present invention gave slightly lower results than heptastigmine and physostigmine: this proves that the claimed products--especially compared with heptastigmine--are more selective toward acetylcholinesterase.
Furthermore, the claimed compounds--compared with those already known--produce much lesser side effects, as proved by the tests conducted for their determination. Table 3 shows the results obtained by acute toxicity tests on rats.
The aforementioned properties make the claimed compounds, i.e. the compounds as per formula (I) and their salts with pharmacologically acceptable acids, suitable for the treatment of all diseases characterized by acetylcholine deficiency, such as for example the disease connected with memory dysfunctions (Alzheimer's disease), or of ischaemic type brain diseases.
For that reason, pharmaceutical compositions containing said compounds in efficacious quantities mixed with pharmacologically acceptable diluents and excipients were prepared.
The invention also contemplates the method of treatment of human diseases characterized by acetylcholine deficiency, consisting of the administration of a pharmachologically efficacious quantity of said compounds by oral or parenteral route.
The therapy envisages a daily dose of compound (I) of 5 to 150 mg for oral administration and of 2.5 to 75 mg for parenteral administration.
TABLE 3______________________________________Acute toxicity to rats DL.sub.50 os (mg/kg)______________________________________ESERINE -- 4.5MF 239 VIII 23.8MF 231 VI 40MF 230 Ex. 13 27.7MF 217 Ex. 7 23MF 256 XXVIII 18.28MF 247 XVIII 14.1MF 248 XXXIII 15.3MF 268 Ex. 17 56______________________________________

Number	Date	Country
253372	Jan 1988	EPX
513703	Nov 1992	EPX

Aminoalkylcarbamic esters of eseroline suitable for use as chlorinesterase activity inhibitors

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