NO140103B - ANALOGICAL PROCEDURE FOR THE PREPARATION OF THERAPEUTICALLY EFFECTIVE PENICILLINES - Google Patents

Description

The present invention relates to the production of

hitherto unknown benzylpenicillins which have a sulfonyl function in the a-position, i.e. benzylpenicillins with the general formula:

where R denotes hydrogen or a methyl group and R^" denotes a methyl or hydroxy group, as well as salts thereof.

The penicillins of formula I exist in two isomeric forms,

and the general formula I includes these two forms and mix-

ings thereof. As examples of salts of compounds of formula I, mention can be made of metal salts, such as sodium, potassium,

calcium and aluminum salts, ammonium salts and substituted ammonium salts, e.g. salts of such amines as trialkylamines, including triethylamine, procaine, dibenzylamine, triethanolamine, 1-N-methyl-1,2-diphenyl-2-hydroxyethylamine, ethylpiperidine and other amines suitable for forming salts of benzylpenicillin.

Penicillins of the above type show interesting chemotherapeutic properties, including broad-spectrum antibacterial

erial effect, acid stability and penicillinase resistance.

According to the present invention, the compounds are produced

with formula I by N-acylating 6-aminopenicillanic acid with a phenylacetic acid of the general formula:

where R and R"*" have the meaning indicated above, or a functionally reactive derivative thereof, and, if desired, converts the obtained penicillin compound into a salt thereof. A compound which, due to its particularly beneficial effect (see the experimental results below), is preferred to be produced according to the invention, is a-sulfobenzylpenicillin, and this compound or a salt thereof is produced by reacting 6-aminopenicillanic acid with a-sulfophenylacetic acid or a functionally reactive derivative thereof, and the reaction product is converted, if desired, into a salt thereof. The N-acylation of 6-aminopenicillanic acid with the acid of general formula II can be carried out using any suitable reactive derivative of the acid of general formula II, e.g. an acid halide, an anhydride or a mixed anhydride thereof, or the acid itself can be reacted with 6-aminopenicillanic acid in the presence of a condensing agent, e.g. dicyclohexylcarbodiimide or carbonyldiimidazole. US Patent No. 3,382,238 describes penicillins which are related to the compounds of formula I, namely sulfinyl penicillins, while the present process produces sulfonyl penicillins. There is thus a significant structural difference between the sulfinyl penicillins described in the US patent and the penicillins containing a sulfonyl function produced by the present method. In the table below, results from comparison experiments with, on the one hand, 6-[3-(methylsulfinyl)-propionamido]-penicillanic acid potassium salt, prepared after a modification of example 2 in US patent no. 3,382,238, are listed in the table designated A, and on the other hand two compounds which have been prepared by the present method, namely α-methylsulfonylbenzylpenicillin (designated B) and α-sulfobenzylpenicillin (designated C). The table shows minimum inhibitory concentrations in ug/ml for the specified organisms. It appears that the compounds produced according to the invention exhibit significantly better activity than the comparison compound over the entire spectrum, and compound C in particular exhibits excellent broad-spectrum activity, especially against Pseudomonas. In order to better assess the difference in reaction for the most resistant strains of Pseudomonas bacteria to penicillins, bacterial growth experiments were carried out. The table below shows the results obtained for two known penicillins, namely ampicillin and carbenicillin and a compound produced according to the present invention, namely α-sulfobenzylpenicillin. The amount of bacterial growth is denoted as +, ++ and +++; - means no growth. Attempts were also made to illustrate the antibacterial spectrum of action for ampicillin and carbenicillin in comparison with three compounds produced according to the present invention, namely the compounds α-methylsulfonylbenzylpenicillin, in the table below denoted D, α-methyl-sulfonyl-2-methylbenzylpenicillin, in the table below designated E, and α-sulfobenzylpenicillin, designated F in the table. The results obtained are indicated in table 3 below. penicillins. The acid stability was determined in ethanolic hydrochloric acid, while the penicillinase resistance was determined in phosphate buffer solution at 37°C. The results obtained are shown in table 4 below. The new compounds, produced according to the present invention, have an acid stability which is far superior to that of penicillin G and carbenicillin. Furthermore, the penicillins produced according to the invention show good stability against Bacillus licheniformis penicillinase which was used in the penicillinase resistance test reproduced above. Furthermore, from the above data, it appears that the penicillin compounds produced according to the present invention possess a broad spectrum of action, both against gram-positive and gram-negative bacteria. The following examples illustrate the invention and the production of starting materials. ;Example 1 ;1) α-methylsulfonylphenylacetic acid ;To a solution of 21.0 g (0.115 mol) α-methylthiophenyl acetic acid in 70 ml of glacial acetic acid, 29.68 ml (0.212 mol) of 30% hydrogen peroxide are added over 45 minutes . The temperature is kept below 25°C. The solution is allowed to stand at room temperature for 11 days, after which 70 ml of water and 40 ml of 6N sulfuric acid are added, and excess hydrogen peroxide is removed by titration with a 0.1N solution of potassium permanganate (78 ml). The reaction mixture is evaporated to dryness and the residue is extracted with 175 ml of ethanol. The insoluble salt is removed by filtration and the filtrate is evaporated in vacuo. The residue is dissolved in 175 ml of chloroform, and the desired compound is precipitated by adding petroleum ether (boiling range 40-80°C). After two further crystallizations from chloroform-petroleum ether, the yield of the analyte α-methylsulfonylphenylacetic acid is 13.4 g (54% of the theoretical). Melting point 131-132°C. ;b) The potassium salt of α-methylsulfonylbenzylpenicillin ;To a mixture of 4.32 g (0.020 mol) 6-aminopenicillanic acid and 6.4 ml (0.046 mol) triethylamine in 40 ml dry chloroform, a solution of 0.020 mol of α-methyl-sulfonylphenyl acetyl chloride (prepared from α-methylsulfonylphenylacetic acid and thionyl chloride) in 20 ml of chloroform. The temperature is kept at 5°C during the addition. The mixture is stirred at this temperature for 30 minutes and then for 2 hours at room temperature. The mixture is cooled to 10°C, 120 ml of water is added and acidified to a pH value of 2 with 3N sulfuric acid. The phases are separated, and the aqueous phase is extracted with two portions of chloroform of 40 ml each. The combined chloroform extracts are cooled to 10°C and covered with 120 ml of water. The pH value is brought up to 9 with 1.5N sodium carbonate solution, and the phases are separated. The aqueous extract is washed with 100 ml of ether, covered with 200 ml of ether, cooled to 10°C and acidified to pH 2 with 3N sulfuric acid. The phases are separated and the aqueous phase is extracted with two portions of ether of 100 ml each. The combined ether extracts are washed with 100 ml of water, dried over MgSO 4 and diluted with 150 ml of dry ether. When treated with 1 equivalent of potassium 2-ethylhexanoate in n-butanol, 5.40 g (60%) of the penicillin is obtained in the form of a white, solid substance which, according to colorimetric determination with hydroxylamine, has a degree of purity of 91%. IR spectrum showed bands at: 690, 840, 960, 1120, 1300, 1340, 1380, 1450, 1500, 1610, 1670, 1760, 2900 and 3300 cm"<1>. ; Example 2 ; a) g- methylthio- 4-methylphenylacetic acid; 15.9 g of α-bromo-4-methylphenylacetic acid is reacted with methyl mercaptan. 12.8 g (99% of the theoretical) α-methylthio-4-methylphenylacetic acid is obtained which is recrystallized from petroleum (boiling range 80-110° C). 42%; H 6.29%; S 16.18%. ;The starting compound, α-bromo-4-methylphenylacetic acid, is prepared as follows: To a mixture of 10.0 g (0.066 mol) 4-methylphenylacetic acid and 0.2 g of red phosphorus at 100°C are added over the course of 2 hours with 12.5 g (0.078 mol) of bromine. The reaction mixture is allowed to cool and 50 ml of water is added. The mixture is extracted with two portions of diethyl ether of 50 ml each, and the combined ether extracts washed with 20 ml of a 5% solution of sodium bisulphite and with 20 ml of water. After drying over MgSO4, the ether is evaporated. The remainder is distilled in vacuum. The fraction (6.13 g, 41% of theoretical) boiling at 141-157°C/0.55 mm Hg is collected and recrystallized from benzene-petroleum (boiling range 80-110°C), thereby obtaining the the analyzers compound with a melting point of 86-89°C. ;Analysis: ;Calculated for CgHgBrC^: C 47.18%;- H 3.96%; Br 34.89% ;Found: C 47.15%; H 4.02%; Br 34.71%. ;b) g-methylsulfonyl-4-methylphenylacetic acid; To a solution of 8.60 g. (0.0447 mol) a-methylthio-4-methylphenylacetic acid in 40 ml of glacial acetic acid, add 12.7 ml (0.112 mol) of 30% hydrogen peroxide within 45 minutes. The temperature is kept below 25°C. The solution is left for 12 days at room temperature. 65 ml of water and 17.6 ml of 6N sulfuric acid are added and excess hydrogen peroxide is removed by titration with a 0.1M solution of potassium permanganate. The reaction mixture is evaporated to dryness in vacuo and the residue is extracted with 125 ml of ethanol. The insoluble salt is removed by filtration, and the ethanol is evaporated in vacuo. The residue is dissolved in 80 ml of chloroform, and the chloroform solution is washed with 40 ml of water and dried over MgSO 4 . When the chloroform is evaporated, a colorless oil is obtained as a residue which will not crystallize. The oil is dissolved in 300 ml of 5% sodium bicarbonate solution, and the solution is washed with two portions of ether of 100 ml each and saturated with sodium chloride. The precipitate is collected and dissolved in 75 ml of water. The aqueous solution is covered with 150 ml of chloroform and acidified with 3N hydrochloric acid. The phases are separated and the chloroform phase is washed with 50 ml of water and dried over MgSO 4 . By evaporation of the chloroform, 4.5 g (44% of the theoretical) of an oil which slowly crystallizes is obtained. By recrystallization from chloroform-petroleum ether (boiling range 40-80°C), the analyte α-methylsulfonyl-4-methylphenylacetic acid with a melting point of 118-119°C is obtained. ;Analysis: ;Calculated for <C>10<H>12<o>4<S:> C 52.62%; H 5.30%; S 14.04%; ;Found: C 52.26%; H 5.34%; S 14,22. c) The potassium salt of α-methylsulfonyl-4-methylbenzylpenicillin; 1.14 g of α-methylsulfonyl-4-methylphenylacetic acid is converted into the acid chloride, which is reacted with 6-aminopenicillanic acid in the same way as described in example lb). 0.90 g (39% of the theoretical) of the above-mentioned penicillin salt is obtained in the form of a solid which, according to colorimetric determination with hydroxylamine, has a degree of purity of 63%. ;I T U I V J ;Example 3 ;a) g-methylthlo-2-methylphenylacetic acid ;To a solution of 3.41 g (0.14 9 gatom) of sodium in ;100 ml of methanol, 4 g (0.084 mol) of methyl mercaptan are added. A solution of 17.0 g (0.0743 mol) of α-bromo-2-methylphenylacetic acid in 75 ml of methanol is then added and the reaction mixture is allowed to stand at room temperature for 1 day. The methanol is then removed in vacuo by refluxing for 3 hours, and the residue is dissolved in 140 ml of H2O. The aqueous solution is washed twice with 60 ml portions of diethyl ether and acidified with 3N hydrochloric acid. Extraction is then carried out with 300 ml portions of diethyl ether, the combined ether extracts are washed with 50 ml of water, dried over MgSO4 and evaporated to dryness. The residue is recrystallized from petroleum ether (boiling range 80-110°C), which gives 11.5 g (79%) of the analyte compound with melting point 63-64°C. The starting compound, α-bromo-2-methylphenylacetic acid, is prepared by adding 59.9 g to a mixture of 45.0 g (0.30 mol) 2-methylphenylacetic acid and 1 g of red phosphorus at 100°C over 3 hours (0.374 mole) of bromine. After a further 30 minutes at 100°C, the reaction mixture is cooled and 85 ml of water is added. Then 2 extractions were carried out, first with 200 ml, and then with 100 ml of diethyl ether; the combined ether extracts are washed with 200 ml of 5% sodium bisulfite solution and 200 ml of water, and then dried over MgSO 4 , after which the diethyl ether is evaporated. The residue is recrystallized from chloroform-petroleum ether (boiling range 40-80°C), which gives 17.01 g (25%) of the analyte α-bromo-2-methylphenylacetic acid, melting point 106-109°C. ;b) a-methylsulfonyl-2-methylphenylacetic acid ;To a solution of 9.0 g (0.0458 mol) a-methylthio-2-methylphenylacetic acid in 45 ml of glacial acetic acid, add 11.85 ml (0.105 mol) over the course of 30 minutes 30% hydrogen peroxide. The temperature is kept below 25°C, and the reaction mixture is left at room temperature for 10 days. 60 ml of water are then added and the resulting precipitate (5.08 g, 49%, melting point 192-196 C, with decomposition)<* >is filtered off and washed with water. By adding 20 ml of 6N sulfuric acid and 3 2 ml of 0.1 m potassium permanganate solution to the filtrate, evaporating the mixture and extracting with ethanol, a further portion is obtained (2.80 g, 27%, m.p. 148-170°C, with decomposition ) raw product. On recrystallization from chloroform-

ethyl acetate-petroleum ether (boiling range 40-80°C), an analytical

pure compound whose decomposition point varies between 180 and 195°C.

c) The potassium salt of α-methylsulfonyl-2-methylbenzylpenicillin

1.14 g of α-methylsulfonyl-2-methylphenylacetic acid is converted

to the acid chloride, which is reacted with 6-aminopenicillanic acid on

same way as described in example lb). This gives 1.00 g (43% of the theoretical) of the above-mentioned penicillin salt in the form of a solid, which according to colorimetric determination with hydroxylamine has a degree of purity of 56%. The IR spectrum showed bands at 725, 755, 960, 1125, 1300, 1400, 1460, 1500, 1550, 1600, 1670, 1760, 2900 and 3400 cm<-1>.

Example 4

a- sulfobenzylpenicillin ( dipotassium salt)

A solution of 2.52 g of α-sulfophenylacetic acid and 2.02 g (0.0200 mol) of triethylamine in 10 ml of dry acetone, which was kept at a temperature of -15°C, was added dropwise to 1.08 g (0. 0100 mol) chloroformic acid ethyl ester. The resulting mixture was stirred for 1 hour at -10°C and then for 1 hour at 0°C and then 25 ml of acetone was added and cooled to -20°C. The temperature was kept below 0°C,

and an ice-cold solution of 2.16 g (0.0100 mol) of 6-aminopenicillanic acid and 2.10 g (0.0250 mol) of sodium carbonate in 35 ml of water was added as quickly as possible. The clear solution obtained was stirred for 30 minutes at -15°C, 30 minutes at 0°C and finally, 30 minutes at room temperature. The reaction mixture was washed twice with 75 ml of ether, covered with 50 ml of ethyl acetate, cooled to 5°C and stirred and acidified with 7 ml of 3N sulfuric acid to pH 1. The layers were separated and the aqueous portion was extracted three times with 30 ml of n-butanol. The combined butanol extracts were washed with 50 ml of water and dried over MgSO 4 .

The dried solution was diluted with 100 ml of ether and treated with 2 equivalents of potassium 2-ethyl hexanoate in n-butanol. The dipotassium salt of the penicillin is excreted as a solid, white, crystalline substance (2.83 g, 58%), which, according to colorimetric determination with hydroxylamine, showed a degree of purity of approx. 76%.

IR spectrum: Peaks at 700, 1045, 1208, 1330, 1410, 1460, 1540,

1610, 1670, 1760, 3000 and 3500 cm"<1>. NMR spectrum in D 2 O: Peaks at 1.6 (6H, t), 4.32 (1H, d) 5.16

(1H, s), 5.62 (2H, d) and 7.58

(5H, m).

It used α-sulfophenylacetic acid as starting material

was produced as follows:

A solution of 100 g of α-sulfophenylacetic acid disodium salt (prepared according to W.E. Truce and C.E. Olson, J . Am .Chem. Soc .

75, 1651 (1953)) in 200 ml of water, a solution of 123 g of barium chloride hydrate in 750 ml of water was added. The mixture was stirred for 15 minutes, and the barium salt was filtered off. The salt was washed twice with 50 ml of water and dried at 90°C. A suspension

tion of 42.5 g of the thus prepared barium-α-sulfophenyl acetate in 900 ml of water, 121 ml of 1M sulfuric acid was added.

The mixture was stirred for 1 hour and was filtered to remove the barium sulfate formed. The filtrate was evaporated to dryness in vacuo. The evaporation residue was an oil which was dried by mixing with 100 ml of methylene chloride and adding dropwise 27.4 ml (0.382 mol) of thionyl chloride. The thionyl chloride reacted

with the water, and the a-sulfophenylacetic acid is separated as a white, crystalline product. Yield: 22.3 g.

Example 5

a- sulfobenzylpenicillin (disodium salt)

A solution of 2.0 g of α-sulfophenylacetic acid (prepared

as described in example 4) in 20 ml of tetrahydrofuran was drop-

show added 3.1 ml (0.0435 mol) thionyl chloride. The solution was kept for 1 hour at 40°C, cooled and evaporated to dryness in vacuo. The α-sulfophenylacetyl chloride obtained was dissolved in 15 ml of ether. The solution was added dropwise at 0°C to a solution of

1.78 g (0.0083 mol) of 6-aminopenicillanic acid in 14 ml of water, which contained a sufficient amount of sodium bicarbonate to maintain the pH at 6.5. The pH of the reaction mixture was kept at 6.5 during the reaction by adding solid sodium bicarbonate. To-

settling of the α-sulfophenylacetyl chloride lasted 30 minutes, and the reaction mixture was further stirred for 1 hour at 0°C.

The layers were separated and the aqueous portion was washed twice

with 15 ml of ether. By adding a cation exchanger, the pH

value for the aqueous solution set to 1.2. The mixture was filtered, and the filtrate extracted twice with 20 ml of acetic acid ethyl ester, and twice with 20 ml of n-butanol. The combined organic extracts were washed twice with 20 ml of water and extracted with a solution of 0.6 g of sodium bicarbonate in 14 ml of water. The pH value of the aqueous solution was adjusted

of 6.5 with hydrochloric acid, and the solution was washed twice with 10 ml of ether. By lyophilizing the solution, 1.6 g of disodium α-sulfobenzylpenicillanate was obtained, which, according to colorimetric determination with hydroxylamine, showed a degree of purity of approx.

92%. IR and NMR spectra were identical to those of Example 4. Example 6

a- sulfobenzylpenicillin ( dipotassium salt)

A suspension of 2.62 g of the disodium salt of α-sulfophenylacetic acid (prepared according to W.E. Truce and C.E. Olson, loe.eit.)

in 10 ml of thionyl chloride was heated under reflux for 1^ hour. Excess thionyl chloride was removed in vacuo and the residue was suspended in 30 ml of acetone. The resulting suspension was added over 30 minutes to a solution of 2.16 g (0.0100 mol) 6-aminopenicillanic acid and 2.96 g (0.0352 mol) sodium bicarbonate in 20 ml of water at -5°C After 1 hour's stirring at 0°C, the reaction mixture was extracted twice with 40 ml of ether. The aqueous portion was overpoured with -48 ml of a 1:1 mixture of ethyl acetate and n-butanol, cooled to 5°C, stirred and acidified to pH = 1 with 3N sulfuric acid. The layers were separated and the aqueous part extracted with 25 ml of ethyl acetate-butanol (1:1). The combined organic extracts were washed with 40 ml of water and dried over MgSO 4 . The dried solution was diluted with 100 ml of ether and treated with 2 equivalents of potassium 2-ethyl hexanoate in n-butanol. The dipotassium salt of penicillin is excreted as an oil. When the oil was precipitated from methanol-ether, a white, solid material was obtained (4.14 g, 85%), which, by colorimetric determination with hydroxylamine, showed a degree of purity of 76%. IR and NMR spectra were identical to those listed in Example 4.

Claims (2)

1. Analogous method for the preparation of therapeutically effective penicillins with the general formula: where R denotes hydrogen or a methyl group, and R<1> denotes a methyl or hydroxy group, as well as salts thereof, characterized by N-acylating 6-aminopenicillanic acid with a phenylacetic acid of the general formula: where R and R<1> have the meaning indicated above, or a functionally reactive derivative thereof, and, if desired, converts the penicillin compound obtained into a salt thereof. 2. Analogous method according to claim 1 for the production of α-sulfobenzylpenicillin or a salt thereof, characterized in that 6-amiopenicillanic acid is reacted with α-sulfophenylacetic acid or a functionally reactive derivative thereof, and the reaction product, if desired, is converted into a salt thereof.