8- Aminopenicillanic acid

Many semi-synthetic penicillins are made from 6-aminopenicillanic acid (6-APA, R = NH2>. 

To this there is added dropwise with continued cooling and stirring a solution of ethyl chlorocarbonate (0.1 mol). After approximately 10 minutes, the acylating mixture is cooled to about -5°C and then is slowly added to a stirred ice-cold mixture of 6-aminopenicillanic acid 0.1 mol), 3% sodium bicarbonate solution (0.1 mol) and acetone. This reaction mixture is allowed to attain room temperature, stirred for an additional thirty minutes at this temperature and then is extracted with ether. 

The known methods for the preparation of D- -)-a-aminobenzylpenicillin by the acylation of 6-aminopenicillanic acid result in the preparation of aqueous mixtures which contain, in addition to the desired penicillin, unreacted 6-aminopenicillanic acid, hydrolyzed acylat-ing agent, and products of side reactions such as the products of the acylating agent reacted with itself and/or with the desired penicillin, as well as other impurities. 

Example 1 ot-AzidobenzylpenicWin via the Mixed Anhydride — A solution of o-azido-phenylacetic acid (8.9 grams, 0.05 mol) of triethylamine (5.1 grams, 0.05 mol) in 50 ml of dry dimethylformamide was stirred and chilled below -5°C. At this temperature ethyl chloroformate (4.7 ml) was added in portions so that the temperature was never above -5°C. After the mixture had been stirred for 20 minutes, dry acetone (100 ml), chilled to -5°C, was added in one portion, immediately followed by an ice-cold solution of 6-aminopenicillanic acid (10.8 grams, 0.05 mol) and triethylamine (5.1 grams, 0.05 mol) in 100 ml of water, and the stirring was continued for VA hours at 0°C. 

Example 2 a-Azidobenzylpenicillin via the Acid Chloride — 6-aminopenicillanic acid (18.5 grams, 0.085 mol) and sodium bicarbonate (21 grams, 0.025 mol) were dissolved in 200 ml of water and 100 ml of acetone. To this solution, chilled in ice, was added a-azidophenyl-acetyl chloride (16.6 grams, 0.085 mol), diluted with 10 ml of dry acetone. The temperature is held at 0° to 5°C and the reaction mixture was stirred for 2 A hours. 

B) Acylation of 6-Aminopenicillanic Acid To a solution of the aryl halocarbonyl ketene (0.1 mol) in methylene chloride (sufficient to provide a clear solution and generally from about 5 to 10 ml per gram of ketene) there is added the proper alcohol RjOH (0.1 mol), in this case 5-indanyl alcohol. The reaction mixture is maintained under an atmosphere of nitrogen and stirred for a period of from 20 minutes to 3 hours, care being taken to exclude moisture. The temperature may range from about -70° to about -20°C. The infrared spectrum of the mixture is then taken to determine and confirm the presence of the ketene ester. A solution of 6-aminopenicillanic acid-triethylamine salt (0.1 mol) in methylene chloride (50 ml) is added and the mixture stirred at -70° to -20°C for 10 minutes. The cooling bath is then removed and the reaction mixture stirred continuously and allowed to warm to room temperature. 

The reaction between 6-aminopenicillanic acid (6.5 g) and 3-o-chlorophenyl-5-methyllsoxa2ole-4-carbonyl chloride (7.66 g) gave the sodium salt of 3-o-chlorophenyl-5-methyl4-isoxa2olyl-penicillin (9.9Bg) asa pale yellow solid. Colorimetric assay with hydroxy lamina against a ben2-ylpenicillin standard indicated a purity of 6B%. 

A suspension of 6-aminopenicillanic acid (36.4 grams) in water was adjusted to pH 7.2 by the addition of N aqueous sodium hydroxide and the resulting solution was treated with a solution of 3-(2-chloro-6-fluorophenyl)-5-methylisoxazole-4-carbonyl chloride (46.1 grams) in isobutyl methyl ketone. The mixture was stirred vigorously for hours and then filtered through Dicalite. The layers were separated and the isobutyl methyl ketone layer was shaken with saturated brine. Then, precipitation of the sodium salt only took place after dilution of the mixture with ether. In this way there was obtained 60.7 grams of the penicillin sodium salt having a purity of 88% as determined by alkalimetric assay. 

To a stirred suspension of 6-aminopenicillanic acid (540 g) in dry alcohol-free chloroform (3.75 liters) was added dry triethylamine (697 ml), and the mixture stirred for 10 minutes at room temperature. It was then cooled in a bath of crushed Ice while a solution of 2,6-dimethoxybenzoyl chloride (500 g) in dry alcohol-free chloroform (3.75 liters) was added in a steady stream over 20 minutes. When all the acid chloride had been added the cooling bath was removed and the mixture stirred for 1 hour at room temperature. The mixture was stirred vigorously and sufficient dilute hydrochloride acid (2.3 liters of 0.87 N) was added to give an aqueous layer of pH 2.5. The mixture was filtered, the layers separated, and only the chloroform layer was retained. 

Aminopenicillanic acid 2-Ethoxy-1 -naphthoyl chloride Sodium bicarbonate... 

The acid chloride obtained as described above was dissolved in dry acetone (10 ml) and added in a steady stream to a stirred solution of 6-aminopenicillanic acid (1.08 g, 5 mmol) in a mixture of N sodium bicarbonate (15 ml) and acetone (5 ml). After the initial reaction the reaction mixture was stirred at room temperature for 45 minutes, then washed with ether (3 X 25 ml). Acidification of the aqueous solution with N hydrochloric acid (11 ml) to pH 2 and extraction with ether (3 x 15 ml) gave an ethereal extract which was decolorized with a mixture of activated charcoal and magnesium sulfate for 5 minutes. 

Aminopenicillanic acid Amoxicillin Ampicillin Azidocillin Aziocillin... 

In subsequent studies,22 Sheehan et al. demonstrated that the action of diisopropylcarbodiimide on penicilloate 24, prepared by protection of the free primary amino group in 23 with trityl chloride (see Scheme 6b), results in the formation of the desired -lactam 25 in a very respectable yield of 67 %. In this most successful transformation, the competing azlactonization reaction is prevented by the use of a trityl group (Ph3C) to protect the C-6 amino function. Hydrogenolysis of the benzyl ester function in 25, followed by removal of the trityl protecting group with dilute aqueous HC1, furnishes 6-aminopenicillanic acid (26), a versatile intermediate for the synthesis of natural and unnatural penicillins. 

Scheme 6. Azlactonization of intermediate 5 (a) and synthesis of 6-aminopenicillanic acid (b).

Different strains of micro-organisms are responsible for the production of either penicillins or cephalosporins. In penicillin-producing strains, an acyltransferase enzyme system is present which can remove the side chain from isopenirillin N to give 6-aminopenicillanic acid (6-APA), and which can subsequently acylate 6-APA to generate various penicillins, the most important ones being penicillin G and V(see section 6.3, Table 6.2). 

The strategy we hope you identified is to first produce 6-aminopenidllanic acid, then attempt to add different moieties to the 6-amino group. This can be achieved either chemically or enzymatically. In the following section we will consider the conversion of penicillin C into 6-aminopenicillanic acid and follow this by examining how 6-aminopenidllanic acid may be converted into ampirillin and amoxicillin. 

Figure 6.14 Enzymatic side chain cleavage of penicillins. 6-Aminopenicillanic acid, a valuable intermediate for the production of various semi-synthetic penicillins, can be obtained through enzyme-mediated hydrolysis of the phenylacety group of penicillin G or the phenoxyacetyl group of penicillin V. The active site of the enzyme recognises the aromatic side chain and the amide linkage, rather than the penidllin nucleus. Chemical entitles other than penicillins are therefore often good substrates, as long as they contain the aromatic acetamide moiety.

---