Incorporation of acetate into cephalosporin C.

Cephalosporin C, an antibiotic with structure (I) (Abraham & Newton, 1961; Hodgkin & Maslen, 1961), was first isolated from an impure preparation of the chemically related penicillin NJ [(D-4amino-4-carboxybutyl)penicillin] that had been obtained from the culture fluid of a Cephalosporium sp. (Newton & Abraham, 1956; Abraham & Newton, 1956). The amount of cephalosporin C produced by this organism, which had been isolated by Brotzu (1948), was too low for the antibiotic to be detected by antibacterial assay in the culture fluid. Much higher yields of the antibiotic were subsequently obtained from a mutant of the original Cephalosporium sp. (Miller, Kelly & Codner, 1961), and the use of the mutant strain enabled workable methods to be devised for the isolation of cephalosporin C from fermentation liquors. With the original strain of Cephalosporium sp. the production of penicillin N in a medium containing sucrose and corn-steep liquor was increased by the addition to the medium of ammonium acetate (Florey et al. 1956). A similar culture medium containing ammonium acetate was shown to be useful for the production of cephalosporin C by the mutant strain by Miller et al. (1961). These findings raised the questions whether acetate was incorporated from the medium into penicillin N and whether it played a part in the biosynthesis of cephalosporin C other than that of providing the acetoxy group in the molecule. When benzylpeni* Present address: The Radiochemical Centre, Amersham, Bucks. t Present address: Microbiological Research Establishment, Porton, near Salisbury, Wilts. t We have agreed to adopt the name 'penicillin N' for the antibiotic previously known as 'cephalosporin N' or ' synnematin B'. cillin was produced in a synthetic medium containing [1_-4C]acetate, 14C was incorporated into the carboxyl group of the valine fragment of the molecule and also into the C O group of the f-lactam ring (Tome, Zook, Wagner & Stone, 1953; Stevens & DeLong, 1958; Demain, 1959). This paper describes the isolation of cephalosporin C produced by laboratory-scale fermentations in complex media containing [1-14C]acetate and records the results of degradations designed to provide information about the distribution of 14C in the labelled compound.