Penicillins side chain precursors

Chemical Modification. The chemistry and synthetic strategies used in the commercial synthesis of cephalosporins have been reviewed (87) and can be broadly divided into ( /) Selection of starting material penicillin precursors must be rearranged to the cephalosporin nucleus (2) cleavage of the acyl side chain of the precursor (2) synthesis of the C-7 and C-3 side-chain precursors (4) acylation of the C-7 amino function to introduce the desked acylamino side chain (5) kitroduction of the C-3 substituent and 6) protection and/or activation of functional groups that may be requked. 

D-p-Hydroxyphenylglycine and its derivatives are important as side-chain precursors for semisynthetic penicillins and cepharosporines. Yamada and coworkers of our laboratory found that these amino acids can be efficiently prepared from the corresponding 5-monosubstituted hydantoins using the microbial enzyme D-hydantoinase [4]. 

Semi-Synthetic Antibiotics. In 1959, Batchelor and coworkers in the Beecham Research Laboratories in England discovered that the penicillin nucleus, 6-aminopenicil-lanic acid (6-APA), accumulated during fermentation when side chain precursors were omitted. This 6-APA could be used for the chemical synthesis of entirely new types of penicillin by coupling with new side chains. Shortly thereafter, several sources of penicillin amidase were found that would cleave the phenylacetyl side chain from penicillin G, thus producing a more economical source of 6-APA. A vast number of synthetic penicillins have been generated, and a few have achieved clinical importance. Several objectives were sought ... 

The natural penicillins, primarily G and V, have a relatively narrow spectrum. They act mostly on gram-positive organisms. The fact that proper selection of precursors could lead to new variations in the penicillin side chain offered the first source of synthetic penicillins. Penicillin V, derived from a phenoxy-acetic acid precursor, attracted clinical use because of its greater acid tolerance, which made it more useful in oral administration. Also, the widespread use of penicillin eventually led to a clinical problem of penicillin-resistant staphylococci and streptococci. Resistance for the most part involved the penicillin-destroying enzyme, penicillinase, which attacked the beta-lactam structure of the 6-aminopenicillanic acid nucleus (6-APA). Semisynthetic penicillins such as ampicillin and carbenicillin have a broader spectrum. Some, such as methicillin, orafi-cillin, and oxacillin, are resistant to penicillinase. In 1984, Beecham introduced Augmentin, which was the first combination formulation of a penicillin (amoxicillin) and a penicillinase inhibitor (clavulanic acid). Worldwide production of semisynthetic penicillins is currently around 10,000 tons/year, the major producers are Smith Kline Beecham, DSM, Pfizer, and Toyo Jozo. 

Implicit in the isolation of penicillins with dilferent side-chains is the formation of a common core. Indeed, dilferent penicillins, such as penicillin V could be produced biosynthetically by adding dilferent side-chain precursors, e.g. phenoxyacetic acid, to the fermentation medium. The common core,... 

The discovery that addition of side-chain precursors to the culture medium favoured production of the penicillin that had incorporated this side-chain allowed the production of a whole range of novel penicillins. Although around 100 such structures were prepared, only one had more desirable properties than penicillin G, and this was christened penicillin V or phe-noxymethylpenicillin. Although this compound was not as potent as penicillin G, it was much more stable to acid (in the stomach), and so could be administered by mouth rather than requiring injection like penicillin G. This was a major advantage for patients, and penicillin V rapidly became the antibiotic of choice for GPs. 

There was a general lack of interest in the penicillins in the 1950s after the exciting progress made during World War II. By that time, it was realized that P. chrysogenum could use additional acyl compounds as side-chain precursors (other than phenylacetic acid for penicillin G) and produce new penicillins, but only one of these, penicillin V (phenoxymethylpenicillin), achieved any... 

During the same period, Sheehan was working toward a total synthesis of penicillins. In 1958, he announced the synthesis of 6-amino-penicillanic acid (6-APA) and its utility for the preparation of new penicillins by acylation (67, 68). (Almost 10 years earlier, this substance had been postulated to be an intermediate in the biosynthesis of penicillins (69, 70). Prior Japanese literature also contained clear suggestions that it had been formed by enzymatic hydrolysis of benzylpenicillin (71) and in fermentations carried out in the absence of side chain precursors... 

Penicillium chrysogenum on complex solid media, with the result that they were mixtures differing from one another in the identity of the side-chain moiety. When a sufficient supply of phenylacetic acid is present in liquid media, this is preferentially incorporated into the molecule to produce mainly benzylpenicillin (penicillin G in the old nomenclature). Use of phenoxyacetic acid instead leads to phenoxymethyl penicillin (penicillin V). More than two dozen different penicillins have been made in this way, but these two are the only ones that remain in clinical use. The bicyclic penicillin nucleus itself is prepared biosynthetically via a complex process from an acylated cysteinyl valyl peptide. The complete exclusion of side chain precursor acids from the medium produces the fundamental penicillin nucleus, 6-APA, but in poor yield. By itself, 6-APA has only very weak antibiotic activity, but when substituted on its primary amino group with a suitable... 

Bifty years have elapsed since the discovery of 6APA (Rolinson and Geddes 2007). 6APA was confirmed as an intermediate in the synthesis of penicillin and detected in the fermentation when no side chain precursor was added (Batchelor et al. 

FIGURE 9 MCA of the penicillin biosynthetic pathway. Based on a kinetic model for the enzymes, in this pathway the FCCs were calculated at different stages of fed-batch cultivations. During the first part of the cultivation the flux control was mainly exerted by the first step in the pathway, i.e., the formation of the tripeptide LLD-ACV by ACV synthetase (ACVS), whereas later in the cultivation flux control shifted to the second step in the pathway, i.e., the conversion of LLD-ACV to isopenicillin N by isopenicillin N synthetase (IPNS). This shift in flux control is due to intracellular accumulation of LLD-ACV, which is an inhibitor of ACVS. The initial high isopenicillin N concentration is due to the fact that this sample was taken from the inoculum culture where the side-chain precursor phenoxyacetic acid (POA) was not present in the medium. [The data are taken from Nielsen, J., and Jorgensen, H. S. (1995). Biotechnol. Prog. 11,299-305.]... 

A systematic study of the incorporation of side-chain precursors into penicillins produced by P. chrysogenum was made in the Lilly Research Laboratories in the 1940s. During the following decade H. R. V. Amstein and others confirmed that the penicillin ring system was formed from amino acid precursors—which were described by Sir Robert Robinson as always sun clear —and took the first step toward the elucidation of the interrelated pathways in penicillin and cephalosporin biosynthesis. 

The fact that proper selection of precursors could lead to new variations in the penicillin side chain offered the first source of synthetic penicillins. Penicillin V, derived from a phenoxyacetic acid precursor, attracted clinical use because of its greater acid tolerance, which made it more useful in oral administration. 

In 1959, Batchelor and coworkers in the Beecham Research Laboratories in England discovered that the penicillin nucleus, 6-APA, accumulated during fermentation when side chain precursors were omitted. This 6-APA... 

Side-chain precursors. The incorporation of isotope from deuterophenylacetyl-valine and from phenylacetic acid-1into benzylpenicilUn by P. chrysogenum showed that added phenylacetic acid and some of its derivatives were direct precursors of the penicillin side-chain (Behrens, 1949 Sebek, 1953 Halliday and Arnstein, 1956). The use of DL-a-amino-adipic acid-2- C and 6- C has indicated that free exogenous a-aminoadipic acid can be incorporated into the side-chain of penicillin N by the Cephalosporium sp. (Abraham et al., 1964). However, the isotope from added L-a-aminoadipic acid-6- C was incorporated into the antibiotic much more efficiently than that from the corresponding D-isomer. 

The discovery that 6-aminopenicillanic acid was produced in fermentations with P. chrysogenum to which no side-chain precursor was added suggested that acylation of this compound might represent the final step in the biosynthesis of the penicilHns. Whether this is so still appears to be an open question. 6-Amino-penicillanic acid (II) is formed from benzylpenicilUn and other penicillins (though not from penicilHn N) by the action of enzymes which exist in a number of microorganisms, including P, chrysogenum (Murao, 1955 Erickson and Bennett, 1965). It may therefore be a product, rather than a precursor, of the penicillins. 

A. is a central building block of penicillins and is produced either by fermentation Penicillium chry-sogenum or Pleurotis astreatus) in the absence of suitable side-chain precursors or by chemical or enzymatic cleavage of penicilline G (phenoxy-methylpenicilline). 

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