Penicillin 6-aminopenicillanic acid

Aminopenicillanic acid Penicillin G Penicillin V Ampicillin Penicillin N... 

Cole, M. Formation of 6-Aminopenicillanic Acid, Penicillins and Penicillin Acylase by Various Fungi. Applied Microbiol. 14, 98 (1966). 

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

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. 

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. 

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. 

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.

Before we leave our discussion of preparing 6-aminopenicillanic acid for use as a starting material in the manufacture of semi-synthetic penicillins, we should point out that similar processes are used in the manufacture of semi-synthetic cephalosporins. Here tire key intermediate is 7-aminodeacetoxycephalosporanic add (7-ADCA). We have drawn outline schemes comparing the production of semi-synthetic penicillins and cephalosporins in Figure 6.15. You will see that the two schemes are very similar. 

Immobilized system the air circulates over a film of microorganisms that grows on a solid surface. In an immobilized bioreactor, particulate biocatalysts for enzyme production and conversion of penicillin to 6-aminopenicillanic acid are used. 

Penicillin G acylase (PGA, EC 3.5.1.11, penicillin G amidase) catalyzes the hydrolysis of the phenylacetyl side chain of penicillin to give 6-aminopenicillanic acid. PGA accepts only phenylacetyl and structurally similar groups (phenoxyacetyl, 4-pyridylacetyl) in the acyl moiety of the substrates, whereas a wide range of structures are tolerated in the amine part [100]. A representative selection of amide substrates, which have been hydrolyzed in a highly selective fashion, is depicted in Figure 6.36. 

Fig. 27. Activity loss a/a of Acylase enzym resign with the reaction time t of the enzymatic deaccylation of Penicillin G to 6-Aminopenicillanic acid (reactor design see Fig. 28)...

Fig. 13. Outlet concentration profiles from a batch chromatographic bioreactor for enzyme catalyzed hydrolysis. Both products, i. e., 6-aminopenicillanic acid (open triangle) and phenylacetic acid (open circle), are separated resulting in a very high conversion. The penicillin G profile is presented as a dashed line (Reprinted with permission from [183])...

Semi-synthetic penicillins are accessed from 6-aminopenicillanic acid, (6-APA), derived from fermented penicillin G. Starting materials for semi-synthetic cephalosporins are either 7-aminodesacetoxycephalosporanic acid (7-ADCA), which is also derived from penicillin G or 7-aminocephalosporanic acid (7-ACA), derived from fermented cephalosporin C (Scheme 1.10). These three key building blocks are produced in thousands of tonnes annually worldwide. The relatively labile nature of these molecules has encouraged the development of mild biocatalytic methods for selective hydrolysis and attachment of side chains. 

Let us look at the common substracture of the penicillin antibiotics, namely 6-aminopenicillanic acid, to illustrate some aspects of working out whether a chiral centre is allocated the I or 6 configuration. 

These defects have spurred attempts to prepare analogs. The techniques used have been (1) natural fermentation (in which the penicillin-producing fungus is allowed to grow on a variety of complex natural nutrients from which it selects acids for incorporation into the side chain), (2) biosynthetic production (in which the fermentation medium is deliberately supplemented with unnatural precursors from which the fungus selects components for the synthesis of "unnatural" penicillins), (3) semisynthetic production (in which 6-aminopenicillanic acid (2) is obtained by a process involving fermentation, and suitably activated acids are subsequently reacted chemically with 6-APA to form penicillins with new side chains) and (4) total synthesis (potentially the most powerful method for making deep-seated structural modifications but which is at present unable to compete economically with the other methods). 

For years the most popular penicillin was a natural one, penicillin G, but it is not acid stable and is absorbed poorly through the intestine. Penicillin G can be hydrolyzed in the laboratory to 6-aminopenicillanic acid, which can... 

An extremely important progress in the development of penicillins took place in 1959, when the penicillin nucleus, 6-aminopenicillanic acid (6-APA), was removed from the side chain and isolated from a culture of Penicillium chrysogenum. 

Penicillins refer to a family of both natural and semi-synthetic antibiotics. Although all exhibit a 6-aminopenicillanic acid core ring structure (Figure 1.15), they differ in the structure of their side-chains. Naturally produced penicillins include penicillins G and V. Semi-synthetic penicillins can be manufactured by enzymatic removal of a natural penicillin side-chain (using... 

Figure 1.15. Structure of 6-aminopenicillanic acid (a) generalized penicillin structure (b) and side-groups present in two natural penicillins and two semisynthetic penicillins (c)...

Facilitated transport of penicilHn-G in a SLM system using tetrabutyl ammonium hydrogen sulfate and various amines as carriers and dichloromethane, butyl acetate, etc., as the solvents has been reported [57,58]. Tertiary and secondary amines were found to be more efficient carriers in view of their easy accessibility for back extraction, the extraction being faciUtated by co-transport of a proton. The effects of flow rates, carrier concentrations, initial penicilHn-G concentration, and pH of feed and stripping phases on transport rate of penicillin-G was investigated. Under optimized pH conditions, i. e., extraction at pH 6.0-6.5 and re-extraction at pH 7.0, no decomposition of peniciUin-G occurred. The same SLM system has been applied for selective separation of penicilHn-G from a mixture containing phenyl acetic acid with a maximum separation factor of 1.8 under a liquid membrane diffusion controlled mechanism [59]. Tsikas et al. [60] studied the combined extraction of peniciUin-G and enzymatic hydrolysis of 6-aminopenicillanic acid (6-APA) in a hollow fiber carrier (Amberlite LA-2) mediated SLM system. 

Penicillin amidase is used industrially to produce 6-aminopenicillanic acid (6-APA) from penicillin G or V (see section 4.5). Acid is produced during the process and this will inactivate the enzyme. One way of overcoming this problem is by using a fixed bed reactor with immobilized enzyme. The substrate is pumped very rapidly... 

When H2O deacetylates the acyl-enzyme, phenylacetic acid is formed. When nucleophiles other than H2O deacylate the acyl-enzyme, a new condensation product, in this case phenylacetyl-O-R or phenylacetyl-NH-R is formed. By definition the hydrolysis of these condensation products can be catalyzed by the same enzyme that catalyzes their formation in equation 10.1. Thus, when the acyl-enzyme is formed from phenylacetyl-glycine or phenylacetyl-O-Me, this gives rise to an alternative process to produce Penicillin G, in addition to the thermodynamically controlled (= equilibrium controlled) condensation of phenylacetic acid and 6-aminopenicillanic acid (6-APA). This reaction that involves an activated side chain is a kinetically controlled (= rate controlled) process where the hydrolase acts as a transferase (Kasche, 1986 1989). 

The optimum yield of a condensation product is obtained at the pH where Ka has a maximum. For peptide synthesis with serine proteases this coincides with the pH where the enzyme kinetic properties have their maxima. For the synthesis of penicillins with penicillin amidase, or esters with serine proteases or esterases, the pH of maximum product yield is much lower than the pH optimum of the enzymes. For penicillin amidase the pH stability is also markedly reduced at pH 4-5. Thus, in these cases, thermodynamically controlled processes for the synthesis of the condensation products are not favorable. When these enzymes are used as catalysts in thermodynamically controlled hydrolysis reactions an increase in pH increases the product yield. Penicilhn hydrolysis is generally carried out at pH about 8.0, where the enzyme has its optimum. At this pH the equiUbrium yield of hydrolysis product is about 97%. It could be further increased by increasing the pH. Due to the limited stability of the enzyme and the product 6-aminopenicillanic acid at pH>8, a higher pH is not used in the biotechnological process. 

On the other hand, the discovery of the parent amine 6-aminopenicillanic acid (9.40, 6-APA) in fermentation products constituted a major breakthrough in penicillin synthesis. 

Penicillin was originally extracted from the mould Penicillium notatum but now it is extracted from its related mould Penicillium chrysogenum due to its high yield. Penicillin consists of thiazolidine ring fused with a beta lactam ring which is essential for its antibacterial activity. These two rings forms a nucleus named as 6-aminopenicillanic acid. 

It is derived from penicillin nucleus 6-aminopenicillanic acid. It has broad spectrum of activity against both gram positive and negative organisms. It is more potent than carbenicillin against Pseudomonas. 

Penicillin is but one of a series of closely related compounds isolated from fermentation broths of Penicillium notatum. This compound, also known as penicillin G (1-1) or benzyl penicillin, is quite unstable and quickly eliminated from the body. Initial approaches to solving these problems, as noted above, consisted of preparing salts of the compound with amines that would form tight ion pairs that in effect provided a controlled release of the active dmg. Research on fermentation conditions aimed at optimizing fermentation yields succeeded to the point where penicillin G or penicillin V (26-1), in which the phenylacetyl group is replaced by phenoxyacetyl, is now considered a commodity chemical. Another result of this research was the identification of fermentation conditions that favored the formation of the deacylated primary amine, 6-aminopenicillanic acid (2-4) or 6-APA, a compound that provided the key to semisynthetic compounds with superior pharmaceutical properties than the natural material. An elegant procedure for the removal of the amide side chain proved competitive with 6-APA from fermentation. This method, which is equally applicable to penicillin V, starts by conversion of the acid to the corresponding silyl ester (2-1). Treatment of that compound with phosphoms pentachloride in the... 

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