Reversed phase HPLC antibiotics

A selective, sensitive and stability indicating reversed phase-HPLC method was developed for the determination of clarithromycin antibiotic in human plasma. 

FIGURE 7 Effect of mobile phase composition on the resolution of enantiomers of different racemates in reversed-phase HPLC on antibiotic CSPs. (a) First ( , O) and second ( , ) enantiomers of 5-methyl-5-phenylhydantoin on a Chirobiotic T column using an acetonitrile-triethylammonium acetate buffer (—) and a methanol-triethylammo-... 

K. R. Bagon, The assay of antibiotics in pharmaceutical preparations using reversed-phase HPLC, HRC CC. J. High Resol. Chromatogr. Chromatogr. Commun., 2 211 (1979). 

H. S. Huang, H. R. Wu, and M. L. Chen, Reversed phase HPLC of amphoteric /3-lactam antibiotics effects of columns ion-pairing reagents and mobile phase pH on their retention times, J. Chromatogr., 564 195(1991). 

Reversed-phase HPLC will find increased application in the analysis of purine antimetabolites and nucleoside antibiotics, both in the chemical laboratory for monitoring serum levels in chemotherapeutic treatment and in quality control in the pharmaceutical industry. In addition, RPLC will be used as a clinical diagnostic tool and aid the clinician in the detection of disease, in confirming a diagnosis, and in monitoring the causes of disease or effectiveness of therapy. 

Kedarcidin, isolated by Zein et al. (Bristol-Myers), was first reported in 1991 as the fermentation product of a novel actinomycete strain obtained from soils collected in India [129]. It exhibits potent in-vivo antitumor activity similar to that of the other enediyne antibiotics, and pronounced activity against Gram-positive bacteria. Kedarcidin was separated relatively easily by reversed-phase HPLC into the apoprotein and chromophore components, and is found to be a varying complex depending upon fermentation conditions [28],... 

In 1991, Ayer et al. reported on the isolation and structure elucidation of a second phenanthridine type alkaloid antibiotic called jadomycin (118), which is a product of the culture of S. venezuelae ISP5230 when the medium is fed with galactose and the temperature is raised to 37°C. When the medium contains glucose-isoleucine at 28°C, the product is chloramphenicol (120) (47). The filtrate of the culture broth was acidified with 6 M HCl and extracted with EtOAc. After evaporation of the solvent, the product was subjected to a reverse-phase flash column chromatography, which was first washed with H2O and then eluted with a MeOH/H20 step gradient. The fraction eluted with 90% MeOH gave jadomycin, which was further purified by reverse-phase HPLC. 

The aim of the work is investigate possibilities of application of Cartridges Packed DIAPAK for concentrating antibiotics Cefazoline and Levomycetine and analyze them by Reversed Phase High Performance Liquid Chromatography (RP HPLC). 

Colistin (COL) is a multicomponent antibiotic (polymyxins E) that is produced by strains of inverse Bacillus polymyxa. It consists of a mixture of several closely related decapeptides with a general structure composed of a cyclic heptapeptide moiety and a side chain acetylated at the N-terminus by a fatty acid. Up to 13 different components have been identified. The two main components of colistin are polymyxins El and E2 they include the same amino acids but a different fatty acid (216). A selective and sensitive HPLC method was developed for the determination of COL residues in milk and four bovine tissues (muscle, liver, kidney, and fat). The sample pretreatment consists of protein precipitation with trichloracetic acid (TCA), solid-phase purification on Cl 8 SPE cartridges, and precolumn derivatization of colistin with o-phthalaldehyde and 2-mercaptoethanol in borate buffer (pH 10.5). The last step was performed automatically, and the resulting reaction mixture was injected into a switching HPLC system including a precolumn and the reversed-phase analytical column. Fluorescence detection was used. The structural study of El and E2 derivatives was carried out by HPLC coupled with an electrospray MS. Recoveries from the preseparation procedure were higher than 60%. 

Chiral separations can be considered as a special subset of HPLC. The FDA suggests that for drugs developed as a single enantiomer, the stereoisomeric composition should be evaluated in terms of identity and purity [6]. The undesired enantiomer should be treated as a structurally related impurity, and its level should be assessed by an enantioselective means. The interpretation is that methods should be in place that resolve the drug substance from its enantiomer and should have the ability to quantitate the enantiomer at the 0.1% level. Chiral separations can be performed in reversed phase, normal phase, and polar organic phase modes. Chiral stationary phases (CSP) range from small bonded synthetic selectors to large biopolymers. The classes of CSP that are most commonly utilized in the pharmaceutical industry include Pirkle type, crown ether, protein, polysaccharide, and antibiotic phases [7]. 

Tesarova and Bosakova [58] proposed an HPLC method for the enantio-selective separation of some phenothiazine and benzodiazepine derivatives on six different chiral stationary phases (CSPs). These selected CSPs, with respect to the structure of the separated compounds, were either based on b-CD chiral selectors (underivatized (J>-CD and hydroxypropyl ether (3-CD) or on macrocyclic antibiotics (vancomycin, teicoplanin, teicoplanin aglycon and ristocetin A). Measurements were carried out in a reversed-phase separation mode. The influence of mobile phase composition on retention and enantio-selective separation was studied. Enantioselective separation of phenothiazine derivatives, including levopromazine (LPZ), promethazine and thioridazine, was relatively difficult to achieve, but it was at least partly successful with both types of CSPs used in this work (CD-based and glycopeptide-based CSP), except for levomepromazine for which only the [CCD-based CSP was suitable. 

Gentamycin is a widely used antibiotic but presents a number of undesirable side-effects when administered in high doses. Thus, it is vital to be able to monitor serum levels accurately. In the past this has been achieved using a number of assay methods other than HPLC. However, recently an HPLC method which compares favourably with microbiological assay methods has been described (Marples and Oates, 1982). This system utilises pre-column derivatisation with o-phthal-aldehyde to facilitate fluorimetric detection with subsequent resolution on a Spherisorb 5-ODS reversed phase support using isocratic conditions with methanol-water-EDTA (Fig. 11.9.4). Baseline resolution of tobramycin and four gentamycin components was achieved by this method. 

In previous papers/ we reported the effect of column temperature on resolution in reversed-phase high-performance liquid chromatography (RP-HPLC) to separate various (3-lactams (penicillins and cephalosporins) from a single sample. In this work we describe the effect of column temperature and volume fraction of an organic solvent on resolution in the isocratic elution conditions of some (3-lactam antibiotics. 

Recently, Aboul-Enein and Ali reviewed the chiral resolution on antibiotic CSPs by HPLC [3,47]. It was observed that chiral resolution on antibiotic CSPs is governed by various HPLC parameters. The antibiotic CSPs may be used in normal, reversed, and new modified polar organic phase modes. The most important parameters which control the chiral resolution on antibiotic CSPs by HPLC are mobile phase composition, pH of the mobile phase, flow rate, temperature, structures of solutes, structures of antibiotics, and other parameters. These parameters are discussed herein. 

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