HALOGENIDES OF THE ESTER OF 2,N,N,N-DIMETHYL-ALKYL-AMINO-ETHANOL WITH SUBSTITUTED ACETIC ACID AND PHARMACEUTICAL COMPOSITIONS HAVING ANTIINFLAMMATORY AND ANTISEPTIC ACTIVITIES CONTAINING SAME

Abstract: Halogenides of the ester of 2-N,N,N-dimethyl-alkyl-­ amino-ethanol with substituted acetic acid, of the general formula wherein R is hydrogen or lower C₁-C₃ alkyl, R′ is straight or branched C₈-C₁₆ alkyl, X is Cl , Br or I , Y is a radical selection from p-isobutylphenyl, 3-benzoyl-phenyl, 1-(p-chlorobenzoyl)-2-methyl-­ 5-methoxy-3-indolyl, 6-methoxy-2-naphtyl and 2-fluoro-4-biphenil, possess high antiinflamma­ tory and antiseptic properties

Description: This invention relates to halogenides of the ester of 2-N,N,N,- dimethyl-alkyl-aminoethanol with substituted acetic acid of the general formula EMI1.1 wherein: R is hydrogen or C1 -C3 lower alkyl, R' is straight or branched C8-C16 alkyl, X is C1 , Br or I Y is a radical selected from p-isobutylphenyl, 3 benzoylphenyl, 1-(p-chlorobenzoyl)-2-methyl-5 methoxy-3-indolyl ,6-methoxy--2-naphty and 2-fluoro-1- biphenyl, having high antiinflamatory and antiseptic properties. This invention also relates to pharmaceutical compositions having anti inflammatory and antiseptic properties, which contain, as the active principle, a therapeutically effective amount of the compounds of formula (I) and a pharmacologically acceptable excipient. These anti inflammatory and antiseptic compositions are only intended for topical administration. There- fore, in addition to containing a compound of formula (1), as their active principle, they also comprise appropriate excipients that are suitable for the formulation of topical compositiolls as will be explained hereinafter. Because of their powerful antiinflammatory and arlLis(?y)tic properties the compounds of formula (I) can be used particularly for the treatment of iiifcctious diseases in which in addition to reducing the inflammatory process, it is desirable to prevent or abolish the growth of pathogenic microorganisms. This kind of pathologies is typically represented by certain affections of the skin and the mucosae, such as: -distrophycally developing cutaneous ulcers (trophic, post-thrombotic, varicose, decubitus ulcers, etc..) -burns -vaginitis in the presence or absence of microbial component -inflammation of soft tissues in the oral cavity and the upper respiratory tract, in the presence or absence of microbic component (stomatitis, tonsillitis, pharyngitis, etc...) Furthermore, the compounds of formula (I) can be used in the topical treatment of painful affections of the musculoskeletal system (traumas, osteoarthrosis, etc...) due to their yood antioedema and analgesic effects. Preferred halogenides of the formula (I) are those wherein R is hydrogen or methyl, R' is octyl, decyl, dodecyl or cetyl, and X is Br or I Particularly preferred among these are: the bromide of the ester of 2-N,N,N-dimethyl-octyl-amino-ethanol with p-isobutylphenyl-alpha-methylacetic acid of the formula EMI3.1 (hereinafter referred to as AF 150); the iodide of the ester of 2-N,N,N-dimethyl-decyl -amino-ethanol with 1-(p-chlorobenzoyl)-2-methyl-5-methoxy-3-indolyl-acetic acid, of formula EMI3.2 (hereinafter referred to as RE/8): and the iodide of the ester of 2-N,N,N-dimethyl-dodecyl, aminoethanol with 1-( p-chlorobenzoyl )- 2-methyl-5-methoxy-3-irldolyl- acetic acid, of formula EMI4.1 (referred to as RE/9 hereinafter). The compounds of formula (I) are prepared by (a) esterifying dimethyl-amino-ethanol with the acid chloride of an acid having the general formula EMI4.2 in which R and Y are as defined above, and (b) reacting the thus obtained ester with an alkyl halogenide of the general formula R'X (III) in which R' and X are as defined above. Non limiting examples of acids of formula (II) are aApha- methyl-4-(2-methyl propyl) benzenacetic, 2-(3-benzoylphenyl) propionic, 1-(p-chlorobenzoyl)-5-methoxy-2-methyl-3-ind acetic,(+)-6 methoxy-alpha-methyl-2-naphthalenacetic and 2 fluoro-alphamethyl-4-biphenylacetic acid. More particularly, a stoichiometric amount of dimetiiyl- amino-ethanol dissolved in CHCl3 is added dropwise to a solu- tion of the acid chloride of the acid (II) in CHCl3. The resul- ting reaction mixture should be maintained at a temperature iri the range of from 200 to 60 C Then, the mixture is kept under stirring at 40-500C for 2-3 hours. The mixture is cooled and the base is freed with a stoichiometric amount of soda dissolved in as great a water volume as half volume of the reaction mixture. After vigorous stirring for 30-60 min, the organic pease is separated and dried on Na 2S04 (the pH of the aqueous phase should adjusted to 8 in order to purify the ester from any unreacted acid). The organic phase is concentrated to dryness and the residue taken up with ethyl acetate, whereupon twice the stoichiometric amount of the alkyl halogenide (III) is added thereto. The mixture is refluxed for 18-24 hours, then allowec to cool. The liquid phase is filtered off, and the compound (I) washed with ethyl acetate. The following non-limiting examples illustrate the preparation of some compounds of formula (II). example 1 - Preparation of compound AF 150. A suspension of the acid chloride of alpha-methyl-4-(2- methyl propyl) benzenacetic acid (50 g; 0.218 moles) in 50 mi of CHCl3 was prepared. A solution of dimethylaminoethanol (19,6 g; 0, 218 moles) in 20 ml of CHCl3 was added to thge suspension over a hour and a half period. The end temperature of the reaction mixture was 420C. The reaction mixture was kept under stirring for 2 hours at 450C, whereupon it was allowed to cool and, then, the stoichiometric quantity of NaOH dissolved in 60 ml water was added thereto; the resulting heterogeneous mixture was cold stirred for 30 min, the organic phase separated, dried on Na2S04 and concentrated to dryness, thus giving the ester as a chromatographically pure oil (45g, 73% yield). Eluent: CHCl /MeOH/NH 40:9:1 3 3 Plates: Merck 5554 Octyl bromide, 43 grams, was added to a solution of 31 grams of the ester dissolved in ethyl acetate and the resulting reaction mixture refluxed for 10 hours. The resulting clear mixture was taken up with 100 ml ethyl acetate, heated to its boiling temperature and then allowed to cool. Upon filtration, 26 grams (49% yield) of the title compound were obtained as a white crystalline compound. Melting point : 104-1090C. Example 2 - Preparation of the compound RE/8 The procedure of Example 1 was followed except that 28.7 g (0.076 moles) of the acid chloride of 1-(p-chlorobenzoyl)-5 methoxy-2-methyl-indole-3-acetic acid was used for preparing the ester. The ester was reacted with decyl iodide to obtain the title compound as a creamwhite crystalline product (yicid: 57%). Melting point: 187-1940C. Example 3 - Preparation of the compound RE/9 The same procedure as that in Example 2 was followed excel)t , that the ester was reacted with dodecyl iodide. The title compound was obtained (60% yield) as a light yellow crystalline product. Melting point: about 1600C. PHARMACOLOGICAL TESTS There are reported hereinbelow the results of pharmacological tests on some of the compounds of formula (I) which show microbiological activity comparable to that of the most widely used antiseptics, associated to significant antiinflammaLory effect. In the pharmacological tests, both female and male Swiss mice (25-30 g) and Wistar rats (140-170 g) were used. Since no significant differences in response between the male and female animal groups was noted, the test results were pooled. Chlorexidine gluconate and Ibuprofen (acid) were used as reference products. All of the doses and concentrations are referred to the salts When administered via the intraperitoneal and oral routes, 10 ml/kg of 0,5% suspensions in methylcellulose were administered. When administered via the intravenous route, the products were dissolved in saline solution(0,,9% NaCl). The variance analysis and the Student's t-test were used for statistical processing of the results. The following tests were carried out: (1) Topical tolerability The test was carried out on rats using two different test procedures. The products were tested at 1% concentration. According to the first procedure, 1 ml of the test suspensions were injected subcutaneously (s.c.) in the side of animals. 4 or 24 hours following administration, the animals were sacrificed, and the degree of subcutaneous irriration estimated. The presence of erythema, oedema and necrosis was assessed based on the following arbitrary rating scale - 0 : no response - 1 : moderate response - 2 : sharp response - 3 : marked response - 4 : extreme response According to the second test procedure, 1 ml of the suen- sions were administered intra-peritoneaLy at the same tirne, 1 ml of 0,1% Blue Evans was administered i.v. to the animals. 4 hours following administration the rats were sacrifice and the peritoneum washed with 10 ml of saline solution. The washing liquid was measured, then centrifuged at 5,000 rpm for 10 iiiiri. The volumeJoptical density (O.D. ) at 640 nm and irritation coefficient (volume x O.D. at 640 nm) of each sample were noted. (2) Antiexudative activity The test was carried out in the rat. The products were orally administered 1 hour before the intraperitoneal injection of 0,5 ml of 1% acetic acid. Along with the irritant agent, 1 ml of 0,1% Blue Evans was injected i.v. to the rats. The same procedure as that described above was followed for collecting the peritoneal exudate and rating the irritation degree. (3) Acute toxicity The test was carried out in the mouse. The products were administered by intraperitoneal injection. The death rate was assessed daily over a period of 14 days following treatlnent. RESULTS (1.1) Topical tolerability The results as to tolerability in the rat subcutaneous tissues are reported in Table 1. The results of tolerability in rat peritoneum are shown in Table 2. The data denonstrate that in rat subcutaneous tissues the compounds of the invention cause a light to moderate degree of irritation, that. is, or the same order of magnitude as that observed with methylce)- lulose. The data relating to rat peritoneum are in substantial agreement with those relating to the rat subcutaneous tissues. It is to be noted that Chlorexidine causes an intense exudative reaction to occur in the rat peritoneum, insofar as it brings about an increase in response which is 442% higher than that recorded for the control group. (2.1) Antiexudative activity The results obtained for acetic acid-induced peritoneal exudation are illustrated in Table 3. Ibuprofen was used as reference product. AF 150 is the most active derivative in this test. In fact, 400 mg/kg orally and 200 mg/kg orally inhibit the inflammatory reaction to a statistically significant extent, 51% and 37% respectively. RE 8 and RE 9 derivatives exhibit a statistically significant effect even tough their activity is less potent than that of AF 150. In comparative tests, Ibuprofen already exhibits its most potent effect at 200 mg/kg orally; in fact its percent inhibition does not increase even tough the dosage exceeds this Threshold. A study of the antiinflammatory activity clearly shnwed that AF 150, RE 8 and RE 9, wtien adnini- stered orally, inhibit acetic acid-induced peritonitis. In comparative tests, Ibuprofen proved more active than the derivatives under oxamination including AF 150 whose antiinflammatory molely of the molecule is Ibuprofen. However, it has to be kept in mind that, in the products under examination, the portion pertaining to the antiinflalllrn,- tory moiety is about 50% of the whole structure. Consequently, with regard to the antiexudative activity, the doses of the compounds according to the invention, the number of mg/kg being the same in either case,are half the doses of Ibuprofen. (3.1) Acute toxicity Acute toxicity data are illustrated in Table 4. The tests were run in mouse, the compounds being administered via the intraperitoneal route. Acute toxicity of the molecules under examination is relatively high; in fact, doses of 30-60 mg/kg i.p. bring about death of 80-100% of animals. In particular, AF 150 and RE 8 do not bring about death at 15.6 mg/kg i.p. whereas at 31,2 mg/kg i.p. they are lethal to 100% of animals. RE 9 is comparatively less toxic: at 31.2 mg/kg i.p., it does not cause death whereas at 62.5 mg/kg i.p. it is lethal to 83.3% of animals. However, it should be noted that the pharmaceutical compo- sitions having antiinflammatory and antiseptic activities according to the present invention which contain one of the compounds of formula (I) as their active principle, are only intended for topical administration. Consequently, the acute toxicity data are only significant in relation to the amounts of compound that will actually be adsorbed systemically following topical treatment. It has been found that product plasma levels that are detected following topical treatment are far below those observed following systemic treatment, and are generally less than a tenth of the plasma level attained via systemic administration. Table 1 Topical tolerability in rat subcutaneous tissue (Reference compound: METHYLCELLULOSE) Compound Conc. Rais Irritation degree % n. 4h 24h I;ethylcellulose / 10 2 1 AF 150 1 10 2 1 RE 8 1 10 2 2 RE 9 1 10 2 2 Table 2 Topical tolerability in rat peritoneum (Reference compound: CHLOREXIDINE) Compound Conc. Rats Percent incream in response % n. with respect to controls Clilorendine 1 10 441 AF 150 1 10 185 RE 8 1 10 97 RE 9 1 10 95 TABLE 3 Antiexudativo activity in rat (Reference product : lbuprofen). Dosage Rats Compound mg/kg N. % Inhibition with orally respect to controls ibuprofen 400 10 58 > 0.0011) 200 10 60 > 0,012) AF 150 400 9 51 > 0,011 200 10 37 > 0,012) RE 8 400 8 46 > 0.021) 200 10 31 > 0.052) RE 9 400 7 1 40 > 0,051) 200 10 35 > 0,052) 1) Student's t-test 2) Duncan test TABLE Acute toxicity in moune Product Dosaqe Mice Mortality mg/kg i.p. N. AF 150 125 6 5/6 83.3% 62.5 6 6/6 100 % 31.2 6 6/6 100 % 15.6 6 0/6 0 % 7.8 6 0/6 0; RE B 250 6 6/6 100 % 125 6 5/6 83.3% 62.5 6 6/6 100 X 31. 2 6 6/6 100 % 1 5 6 6 0/6 0 % 7.8 6 0/6 o % RE 9 500 6 6/6 100 % 250 6 5/6 83.3% 125 6 6/6 100 X 62.5 6 5/6 83.3% 31.2 6 0/6 0% i.p. = intraperitoneal MICROBIOLOGICAL TESTE The activity in vitro has been evaluated on the following microorganisms: Gram-Positive Bacteria: Staphylococcus aureus Staphylococcus epidermidis Streptococcus faecalis Streptococcus pyogenes Bacillus subtilis Bacillus cereus mycoides Sarcina lutea Bacillus antracis Gram-Negative Bacteria: : Escherichia coli Salmonella typhimurium Salmonella cholerae suis Klebsiella pneumoniae Proteus vulgaris Pseudomonas aeruginosa Bordetella bronchiseptica Pseudomonas maltuphylia Blastomycetes: Candida albicans Candida utilis Rhodotorula rubra Cryptococcus neoformans Saccharomyces cerevisiae Ifomycetes: Aspergillus niger Penicillium funicolosum Chlorexidine gluconate was used as reference compound. Compounds were tested at 5,10, 20, 40 and 80 g/ml. Results are shown in Tables 5, 6 and 7. As it can be seen, compounds AF 150 and RE 9 are the most active compounds, because more potent than Chlorexidine against Gram-positive bacteria, Blastomycetes and Ifomycetes, and as potent as Chlorexidine against Gram-negative bacteria. Compound RE 8 is more potent than Chlorexidine against Crampositive bacteria, Balstomycetes and Ifomycetes, and less potent than Chlorexidine on Gram-positive bacteria. TABLE 5 Determination of Minimum Inhibiting Concentratio STRAIN Repository Sample Reference compound numbers AF 150 CHLOREXIDINE GLUCO Gram Positive Bacteria Staphylococcus apreus ATCC 6530P #5,0 #5,0 Staphylococcus epidermidis BB 0223 10,0 10,0 Streptococcus faccalic ATCC 8043 20,0 20,0 Sterptococcus pyogenes ATCC 12380 20,0 10,0 Bacillus subtilis ATCC 6633 10,0 #5,0 Bacillus cereaus mycoides ISM 65/42 10,0 10,0 Sarcina lutea ATCC 9341 #5,0 20,0 Bacillus antracis ISM 68/35 10,0 20,0 Gram Ncgative Esherichia coli K 12 80,0 10,0 Salmonella typhimurium ATCC 15277 80,0 20,0 Salmonella cholerae suis ATCC 10708 80,0 10,0 Klebsiella pneumonise ATCC 10031 10,0 40,0 Proteus vulgaris ATCC 6897 10,0 40,0 Pseudomonas aeruginosa ATCC 14502 10,0 80,0 Bordetella bronchiseptica ATCC 4617 10,0 40,0 Pseudomonas maltphilia ATCC 13637 40,0 80,0 Blastomycetes Candida albicans ATCC 10231 #5,0 20,0 Candida utilis BB 0173 #5,0 40,0 Rhodotoruls rubra ISM 7198 #5,0 40,0 Cryptococcus neoformans ISM 7182 #5,0 40,0 Saccharomyces cerevisiae BB 0193 #5,0 80,0 Ifomycetes Aspergillus niger ATCC 9642 10,0 80,0 Penicillium funicolosum ATCC 9644 10,0 > 80,0 TABLE 6 Determination of Minimum Inhibiting Concentration STRAIN Repository Sample Reference compound numbers RE 9 CHLOREXIDINE GLUCONA". Positive Bacteria Staphylococcus apreus ATTC 6538P #5,0 #5,0 Staphylococcus epidermidis BB 0223 #5,0 10,0 Streptococcus fascalis ATCC 8043 #5,0 20,0 Streptococcus pyogenes ATCC 12380 #5,0 10,0 Bacillus subtilis ATCC 6633 #5,0 #5,0 Bacillus cereaus mycoides ISM 65/42 #5,0 10,0 Sarcina lutea ATCC 9341 < 5,0 20,0 Bacillus antracis ISH 68/35 #5,0 20,0 l'cgative Bacteria Escherichia coli K 12 80,0 10,0 Salmonella typhimurium ATCC 15277 20,0 20,0 Salmonolls cholcroo suis ATCC 10708 20,0 10,0 Klebsiella pneumonise ATCC 10031 80,0 40,0 Proteus vulgaris ATCC 6897 20,0 40,0 Pseudomonas aeruginosa ATCC 14502 80,0 80,0 Bordetella bronchiseptica ATCC 4617 80,0 40,0 Pseudomonas maltphilia ATCC 13637 80,0 80,0 cmycetes Candida albicans ' ATCC 10231 < 5,0 20,0 Candida utilis BB 00173 #5,0 40,0 Rhodotoruls rubra ISM 7198 #5,0 40,0 Cryptococcus neoformans ISM 7182 #5,0 40,0 Saccharomyces cerevisiae BB 0193 #5,0 80,0 ycetes Aspergillus niger ATCC 9642 #5,0 80,0 Penicillium funicolosum ATCC 9644 #5,0 > 80,0 TABLE 7 Determination of Minimum Inhibiting Concentration STRAIN Repository Sample Reference compound numbers RE 8 CHLOREXIDINE GLUC( Gram Positive Bacteria Staphylococcus apreus ATCC 6538P #5,0 #5,0 Staphylococcus epidermidis BB 0223 #5,0 10,0 Streptococcus faochlis ATCC 8043 10,0 20,0 Streptococcus pyogenes ATCC 12380 10,0 10,0 Bacillus subtilis ATCC 6633 10,0 #5,0 Bacillus cereaus mycoides ISM 65/42 10,0 10,0 Sarcina lutea ATCC 9341 10,0 20,0 Bacillus antracis ISM 68/35 10,0 20,0 Gram Negative Bacteria Escherichia coli K 12 > 80,0 10,0 Salmonella typhimurium ATCC 15277 20,0 20,0 Salmonella cholerae suis ATCC 10708 20,0 10,0 Klebsiella pneumonise ATCC 10031 > 80,0 40,0 Proteus vulgaris ATCC 6897 80,0 40,0 Pseudomonas aeruginosa ATCC 14502 80,0 80,0 Bordetella bronchiseptica ATCC 4617 80,0 40,0 Pseudomonas maltphilia ATCC 13637 80,0 80,0 Blastomycetes Candida albicans ATCC. 10231 #5,0 20,0 Candida utilis BB 0173 #5,0 40,0 Rhodotoruls rubra ISM 7198 #5,0 40,0 Cryptococcus neoformans ISM 7182 #5,0 40,0 Saccharomyces cerevisiae BB 0193 #5,0 80,0 Ifomycetes Aspergillus niger ATCC 9642 #5,0 80,0 Pcnicilliun funicolosum ATCC 9644 #5, 0 > 80, 0 The most suitable pharmaceutical preparations for topical administration of the compounds according to this invention, having regard to the therapeutic fields of application, are as follows: : In gynaecology In trauma-rheumatology - ready-to-use vaginal douche - gel - small bags - cream - concetrated solution - spray lotion - spray cream - ovules In stomatology In dermatology - collutory - ointment - paste - spray cream For burn treatment - cream - spray cream - medicated gauze Concentrations of products range from 0q5 to 1% by weight, depending on the particular pharmaceutical preparation. Choose of the most appropriate excipients depends on the specific pharmaceutical preparation and will be obvious to one of ordinary skill in the pharmaceutical technology. As should be apparent excipients compatible with human use shall be utilized, preference being given to the excipients already listed in the Italian Pharmacopoeia or the Pharmacopoeias of main European and non European countries.

Claims: CLAIMS 1. Halogenides of the ester of 2-N,N,N,-dimethyl-alkyl-aminoethanol with substituted acetic acid, of the general formula EMI23.1 wherein R is hydrogen or lower C1-C3 alkyl, R1 is straight or branched C8-C16 alkyl, X is Cl-, Br or I Y is a radical selected from p-isobutylphenyl, 3 benzoyl-phenyl, 1 -( p-chlorobenzoyl )-2-methyl-5- methoxy3-indolyl, 6-methoxy-2-naphthyl and 2-fluoro 4-biphenyl. 2. A halogenide according to claim 1, wherein R is hydrogen or methyl, R' is octyl, decyl, dodecyl or cetyl, and X is Br or I 3. As halogenide of claim 2, the bromide of the ester of 2-N,N,N-dimethyl-octyl-aminoethanol with p-isobutylphenylalpha-methylacetic acid of formula EMI23.2 4. As halogenide of claim 2, the iodide of the ester of 2-N,N,N-dimethyl-decyl-aminoethanol with 1-( p-chlorobenzoyl )-2- methyl-5-methoxy-3-indolyl acetic acid,of the formula EMI24.1 5. As halogenide according to claim 2, the iodide of the ester of 2-N, N, N-dimethyl-dodecyl-aminoethanol with 1 -( p-chloroben zoyl )-2-methyl-5-methoxy-3-indolyl-acetic acid, of formula EMI24.2 6. A pharmaceutical composition having antiinflammatory and antiseptic activities, intended for topical administration, which comprises (a) a therapeutically effective amount of a halogenide of the ester of 2-N,N,N-dimethyl-alkyl-amino-ethanol with substituted acetic acid, of the general formula EMI25.1 wherein R is hydrogen or lower C1 -C3 alkyl, R' is straight or branched C8-C16 alkyl, X- is Cl-, Br- or I-, Y is a radical selected from p-isobutylphenyl, 3-benzoyl phenyl, 1 -( p-chlorobenzoyl )-2-methyl-5-methoxy-3- indolyl, 6-methoxy-2-naphthyl and 2-fluoro-4-biphenyl, and (b) a pharmaceutically acceptable excipient suitable for preparing compositions for topical use. CLAIM for AT, ES, and CR A process for preparing halogenides of the ester of 2-N,N,Ndimethyl-alkyl-amino-ethanol with substituted acetic acid, of the general formula EMI26.1 wherein R is hydrogen or lower C1 -C3 alkyl, R' is straight or branched C8-C16 alkyl, X is Cl , Br or I Y is a radical selected from p-isobutylphenyl, 3 benzoyl-phenyl, 1-(p-chlorobenzoyl)-2-methyl-5- methoxy-3-indolyl, 6-methoxy-2-naphthyl and 2 fluoro-4-biphenyl, characterized in that it comprises the steps of: (a) esterifying dimethyl-amino-ethanol with the acid chloride of an acid of the general formula EMI26.2 wherein R and Y are as defined above and (b) reacting the thus obtained ester with an alkyl halide of general formula R'X (III) wherein R' and X are as defined herein-above.

Inventors: REINER, ALBERTO
Applicants: SIGMA-TAU INDUSTRIE FARMACEUTICHE RIUNITE S.P.A.
Application year: 1987
Main classification: C07C219
IPC codes: C07D209