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Macrolides, as a class of natural or semisynthetic products, express their antibacterial activity primarily by reversible binding to the bacterial 50S ribosomal subunits and by blocking nascent proteins’ progression through their exit tunnel in bacterial protein biosynthesis. Generally considered to be bacteriostatic, they may also be bactericidal at higher doses. The discovery of azithromycin from the class of macrolides, as one of the most important new drugs of the 20th century, is presented as an example of a rational medicinal chemistry approach to drug design, applying classical structure-activity relationship that will illustrate an impressive drug discovery success story. However, the microorganisms have developed several mechanisms to acquire resistance to antibiotics, including macrolide antibiotics. The primary mechanism for acquiring bacterial resistance to macrolides is a mutation of one or more nucleotides from the binding site. Although azithromycin is reported to show different, two-step process of the inhibition of ribosome function of some species, more detailed elaboration of that specific mode of action is needed. New macrocyclic derivatives, which could be more potent and less prone to escape bacterial resistance mechanisms, are also continuously evaluated. A novel class of antibiotic compounds—macrolones, which are derived from macrolides and comprise macrocyclic moiety, linker, and either free or esterified quinolone 3-carboxylic group, show excellent antibacterial potency towards key erythromycin-resistant Gram-positive and Gram-negative bacterial strains, with possibly decreased potential of bacterial resistance to macrolides.

In the search for endogenous cardiac glycosides, two different inhibitors of the sodium pump have been isolated from bovine adrenals. Inhibitor A with a molecular mass of 600 Da and a UV maximum at 250 nm was purified from 16 kg of bovine adrenals. The pure substance (<1 ng) inhibited the sodium pump of human red blood cells with an affinity similar to that of ouabain, yet it cross-reacted with antibodies against the bufadienolide proscillaridin A but not against the cardenolide ouabain. Inhibitor A was slightly more hydrophilic than ouabain on RP-C 18 high pressure liquid chromatography. Hence, it showed properties similar to the proscillaridin A immunoreactivity (Sich, B., Kirch, U., Tepel, M., Zideck, W., and Schoner, W. (1996) Hypertension 27, 1073-1078) that increased in humans with systolic blood pressure and pulse pressure.Inhibitor B of the sodium pump with a molecular mass of 584 Da was purified 10 6 -fold from 20 kg of bovine adrenals. It cross-reacted with antibodies against ouabain but not with antibodies against proscillaridin A and inhibited the sodium pump of human and rat red blood cells with the same affinity as ouabain. All other properties, such as the retention time in a C 18 -reversed phase chromatography, molecular mass determination by electrospray mass spectrometry and fragmentation pattern, and UV and 1 H NMR spectroscopic data, were identical to ouabain. Hence, sodium pump inhibitor B from bovine adrenals is the cardenolide ouabain.

Right-side-out vesicles of pig kidney microsomes and amino-acid-sequence-specific antibodies were used to probe the sidedness of the C-terminus and the N-terminus of the catalytic CI subunit of Na+/K+-ATPase. Polyclonal antibodies were raised in rabbits against the peptide corresponding to the N-terminal sequence GRDKYEPAAVSE (peptide 1 -12) and against peptides corresponding to the C-terminal sequences IFVYDEVRKLIIRRR (peptide 991 -1005) and RPGGWVEKETYY (peptide 1005-1016). These antibodies were purified by affinity chromatography on the respective peptide-Sepharose columns. Moreover, antibodies against the N-terminal dodecapeptide GRDKYEPAAVSE were obtained by affinity purification from heteroclonal antibodies against the CI subunit of pork kidney Na+/K+-ATPase. These antibodies reacted with native as well as SDS-denaturated Na+/K+-ATPase. When the antibodies were used to probe the sidedness of the sequences in right-side-out vesicles of pig kidney microsomes, the N-terminal peptide 1 -12 as well as the C-terminal peptides 991 -1005 and 1005 -101 6 were found on the cytosolic side. Concanavalin A, however, which interacts with the j3 subunit, a glycoprotein, reacted with the outside of right-side-out vesicles.

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