For many drugs, only racemic mixtures are available for clinical use. Because different stereoisomers of drugs often cause different physiological responses, the use of pure isomers could elicit more exact therapeutic effects. Differential complexation of a variety of drug stereoisomers by immobilized beta-cyclodextrin was investigated. Chiral recognition and racemic resolution were observed with a number of compounds from such clinically useful classes as beta-blockers, calcium-channel blockers, sedative hypnotics, antihistamines, anticonvulsants, diuretics, and synthetic opiates. Separation of the diastereomers of the cardioactive and antimalarial cinchona alkaloids and of two antiestrogens was demonstrated as well. Three dimensional projections of beta-cyclodextrin complexes of propanolol, which is resolved by this technique, and warfarin, which is not, are compared. These studies have improved the understanding and application of the chiral interactions of beta-cyclodextrin, and they have demonstrated a means to measure optical purity and to isolate or produce pure enantiomers of drugs. In addition, this highly specific technique could also be used in the pharmacological evaluation of enantiomeric drugs.

Aromatic boronic acids are reversible inhibitors of the recently classified class C beta-lactamases. The boronic acids studied include ortho-, meta- and para-methyl-, -hydroxymethyl- and -formyl-phenylboronic acid. The beta-lactamases were chromosomally-encoded enzymes, one from Pseudomonas aeruginosa, and the other specified by the ampC gene of Escherichia coli. The inhibition may be correlated with our finding that these beta-lactamases are serine enzymes, i.e. their function entails the hydroxy group of a serine residue acting as a nucleophile.

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SUMMARYOsmotically fragile protoplasts have been prepared by the action of lytic enzymes on the mycelium of Penicillium chrysogenum and Cephalosporium acremonium.The yield of protoplasts, based on DNA content, was up to 18 %. Pretreatment of the mycelium with a thiol compound was necessary with Cephalosporium but not with Penicillium. Electron micrographs indicated that the protoplasts contained all the intracellular organelles of the mycelium and were bounded only by a cytoplasmic membrane. The metabolic activity of the protoplasts, as measured by their respiration, ability to maintain intracellular amino-acid pools, and antibiotic production, was similar to that of control mycelium. L-Valine and L-a-aminoadipic acid, which are precursors of penicillin N and cephalosporin C, were taken up and concentrated by the protoplasts of C. acremonium, although the latter transported L-valine less rapidly than did the corresponding mycelium. Protoplasts of P. chrysogenum took up ~-valine but not L-a-aminoadipic acid or its 8-ester. There was little or no transport of the corresponding D-amino acids by protoplasts of either organism. I N T R O D U C T I O NOne of the difficulties encountered in studies of penicillin and cephalosporin biosynthesis has been associated with the permeability barriers of mycelium. Thus, the interpretation of some experiments has been limited by the failure of a substance added to mycelial suspensions to reach the site of synthesis. For example, no detectable amount of [14C]penicillin N was found to enter the mycelium of Cephalosporium acremonium from the extracellular fluid (Smith, Warren, Newton & Abraham, I 967). The dipeptide 6-(a-aminoadipyl)-cysteine was not taken up intact by the mycelium but hydrolysed to its constituent amino acids (Loder, Abraham & Newton, 1969).A fragmented hyphal preparation has been obtained by ultrasonic treatment of the mycelium of Cephalosporium acremonium which is able to synthesize a possible biosynthetic intermediate, the tripeptide 8-(a-aminoadipyl)cysteinylvaline, from 6-(L-a-aminoadipyl)-Lcysteine and ~~-[ l~C ] v a l i n e (Loder & Abraham, 1971). However, no indication of penicillin or cephalosporin formation was obtained in fragmented hyphal systems of this type and it seemed that a study should be made of gentler methods of disrupting the mycelium. The first step of a possible approach was the enzymatic removal of the hyphal walls to form osmotically fragile protoplasts. For example, a bacterium-free extract was prepared from protoplasts of Bacillus licheniformis which was able to synthesize bacitracin from its constituent amino acids in the presence of an energy-generating system (Ishihara, Sasaki & Shimura, 1968), whereas an extract prepared by ultrasonication was less effective (Shimura, P. A. F A W C E T T A N D O T H E R SSasaki & Sugawara, 1964). This paper describes the preparation of protoplasts from strains of Penicillium chrysogenum and Cephalosporium acremonium and some of the properties of these protoplasts which are relevant to the biosynt...