Macrolide antibiotics assays

Data and chromatograms for four antibiotics will be used to help illustrate and characterize representative approaches to real situations. The work on cefadroxil, cefmenoxime, cefsulodin, and clarithromycin are all HPLC assays. The three cephalosporins used a UV finish, while the clarithromycin being a macrolide antibiotic and having a low chro-mophoric response, required an electrochemical detector for quantitation. 

Erythromycin, a macrolide antibiotic, lacks a significant chromophore. Detection sensitivity was enhanced by using a wavelength of 200 nm and selecting an injection solvent of lower conductivity than the BGE. In order to facilitate the separation of erythromycin and its related substances, 35% (v/v) ethanol was incorporated into a 150 mM phosphate buffer pH 7.5. Resolution of all of the compounds was achieved in approximately 45 min. The method was employed as an assay method for erythromycin and for impurity determination. Peptide antibiotics, such as colistin and polymyxin, are mixtures of many closely related compounds. A validated CZE method for impurity analysis of polymyxin B was described, employing 130 mM triethanolamine-phosphate buffer at pH 2.5 to reduce the adsorption of analyte onto the capillary wall. Methyl-/l-cyclodextrin (M-/1-CD) and 2-propanol were found to be necessary for selectivity enhancement. Using similar buffer additives, the same group developed and validated a method for colistin analysis. ... 

A Merck group was interested in the introduction of a basic amino group into the avermectin molecule [228]. This should change its physical properties, make it more polar, and result in a different tissue distribution. In addition, most of the anti-bacterially active macrolide antibiotics contain an aminosugar. After suitable protection and deprotection, reductive amination of a 4"-oxo intermediate gave the 4"-amino-4"-deoxy analogue as the major reaction product (Scheme 19). Since they also had observed better activity in the southern armyworm assay for certain monosaccharides, they prepared their 4 -amino derivatives as well as the... 

Nakajima, Y., Takeda, R., Tani, K., Endou, K., Matsuoka, M., and Yamagishi, S. (1990). Greatly improved activity of staphylococcal ribosomes in polyadenylate directed polylysine synthesis As an assay system for investigating their sensitivity to macrolide antibiotics. J. Pharmacobio. Dyn. 13, 378-383. 

Bekele, L.K., Gebeyehu, G.G., 2012. Application of different analytical techniques and microbiological assays for the analysis of macrolide antibiotics from pharmaceutical dosage forms and biological matrices. ISRN Anal. Chem. 2012, 1—17. Available from http //dx.doi.org/10.5402/2012/859473. 

More versatile than the growth-inhibition assays and potentially applicable to determining the presence of different antibiotic residues in different matrices are the microbial receptor CHARM I and II test assays (19, 20). The Charm I test, developed exclusively for -lactams in milk, constitutes the first rapid test recognized by The Association of Official Analytical Chemists (AOAC) with a test time of 15 min (19). The speed and sensitivity of this test permitted testing of milk tankers before they unloaded at the processing plant (21). In 1984-1985, the CHARM I test was further developed to test for antibiotics beyond -lactams to include tetracyclines, sulfonamides, aminoglycosides, chloramphenicol, novobiocin, and macrolides. The extended version has been referred to as CHARM II test. 

The HPLC-receptorgram assay combined the advantages of HPLC separation with the multiresidue detection of the Charm II tests. The procedure was tested for identification and quantitation of the most common veterinary drugs at regulatory levels or lower. It was validated for 40 individual drugs from seven antibiotic families 10 /3-lactams, 13 sulphonamides, 8 tetracyclines, 4 macrolides, 3 amphenicols, and other miscellaneous antimicrobials. This procedure combined a simple aqueous extraction and SPE with HPLC fractionation of individual drugs. Final identification and quantitation was achieved with the Charm II test. A drug contaminant could be identified in less then 3 hours (50). 

BogiaU S, Di Corcia A, Lagana A, et al., A simple and rapid confirmatory assay for analysing antibiotic residues of the macrolide class and lincomycin in bovine milk... 

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