Browsing by Author "Du Toit, Etienne Andre"
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- ItemThe development of an antibacterial assay and its use in the investigation of the combined effect of tetracycline and synthetic antimicrobial peptides on strains of Escherichia coli(Stellenbosch : Stellenbosch University, 1999-11) Du Toit, Etienne Andre; Rautenbach, Marina; Stellenbosch University. Faculty of Science. Dept. of Biochemistry.ENGLISH SUMMARY: Magainin 2 is a 23-residue antimicrobial peptide secreted on the skin of Xenopus laevis. In our study on antimicrobial peptides, this cationic, α-helical peptide and two N-terminal deletion analogues were synthesised. The first of the deletion peptides differed from the full length peptide by the omission of two amino acid residues from the N-terminus of rnagainin 2, while the second deletion peptide had four residues omitted from the N-terminus (termed magainin 2 N21 , and magainin 2 N19 respectively). Peptides were purified by gel permeation chromatography and high performance liquid chromatography (HPLC). The synthetic peptides were of high chemical purity as verified with' electro spray ionisation mass spectrometry (ESI-MS) and analytical HPLC. To further evaluate the antimicrobial activity of the peptides in this study, a highly sensitive micro-gel well diffusion assay was developed. This assay was compared to a radial diffusion assay and a microtiter broth dilution method, using gramicidin S as model antibiotic, and Micrococcus lute us as the indicator organism. The micro-gel well diffusion assay was as sensitive as the microtiter broth dilution method, and approximately twice as sensitive as the radial diffusion method. Data analysis to calculate minimum inhibitory concentration (MIC), 50% microbial growth inhibition (IC50), and maximum bactericidal concentration (MBC) was refined by generating dose-response curves with Prism® 2.01 (Graphpad Software Inc.). The MICs, determined by the three methods, were significantly different (P