title: Kinetic microplate bioassays for relative potency of antibiotics improved by partial Least Square (PLS) regression.
abstract: Microbiological assays are widely used to estimate the relative potencies of antibiotics in order to guarantee the efficacy, safety, and quality of drug products. Despite of the advantages of turbidimetric bioassays when compared to other methods, it has limitations concerning the linearity and range of the dose-response curve determination. Here, we proposed to use partial least squares (PLS) regression to solve these limitations and to improve the prediction of relative potencies of antibiotics. Kinetic-reading microplate turbidimetric bioassays for apramacyin and vancomycin were performed using Escherichia coli (ATCC 8739) and Bacillus subtilis (ATCC 6633), respectively. Microbial growths were measured as absorbance up to 180 and 300min for apramycin and vancomycin turbidimetric bioassays, respectively. Conventional dose-response curves (absorbances or area under the microbial growth curve vs. log of antibiotic concentration) showed significant regression, however there were significant deviation of linearity. Thus, they could not be used for relative potency estimations. PLS regression allowed us to construct a predictive model for estimating the relative potencies of apramycin and vancomycin without over-fitting and it improved the linear range of turbidimetric bioassay. In addition, PLS regression provided predictions of relative potencies equivalent to those obtained from agar diffusion official methods. Therefore, we conclude that PLS regression may be used to estimate the relative potencies of antibiotics with significant advantages when compared to conventional dose-response curve determination.
authors.affiliation: Departamento de Farmácia, Faculdade de Ciências Farmacêuticas, Universidade de São Paulo, Av. Prof. Lineu Prestes, 580 - Bloco 13, CEP 05508-000 São Paulo, SP, Brazil.
authors.forename: Fabiane Lacerda
authors.initials: FL
authors.lastname: Francisco
authors.affiliation: Departamento de Farmácia, Faculdade de Ciências Farmacêuticas, Universidade de São Paulo, Av. Prof. Lineu Prestes, 580 - Bloco 13, CEP 05508-000 São Paulo, SP, Brazil.
authors.forename: Alessandro Morais
authors.initials: AM
authors.lastname: Saviano
authors.affiliation: Departamento de Farmácia, Faculdade de Ciências Farmacêuticas, Universidade de São Paulo, Av. Prof. Lineu Prestes, 580 - Bloco 13, CEP 05508-000 São Paulo, SP, Brazil.
authors.forename: Túlia de Souza Botelho
authors.initials: Tde S
authors.lastname: Almeida
authors.affiliation: Departamento de Farmácia, Faculdade de Ciências Farmacêuticas, Universidade de São Paulo, Av. Prof. Lineu Prestes, 580 - Bloco 13, CEP 05508-000 São Paulo, SP, Brazil. Electronic address: feliperl@usp.br.
authors.forename: Felipe Rebello
authors.initials: FR
authors.lastname: Lourenço
mesh_terms: D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000494:pharmacology*; D001412:Bacillus subtilis / Q000737:chemistry / Q000187:drug effects / Q000254:growth & development; D001681:Biological Assay / Q000379:methods*; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000737:chemistry / Q000187:drug effects / Q000254:growth & development; D007700:Kinetics; D016018:Least-Squares Analysis; D008826:Microbial Sensitivity Tests / Q000379:methods*
chemical_list: D000900:Anti-Bacterial Agents
keywords: Aminoglycosides; Antimicrobial activity; Antimicrobial agents; Glycopeptides; Partial least squares (PLS) regression