Stability of β-lactam antibiotics in bacterial growth media

• Published in: PloS one 2020-07, Vol.15 (7), p.e0236198-e0236198
• Main Authors: Brouwers, Rebecca, Vass, Hugh, Dawson, Angela, Squires, Tracey, Tavaddod, Sharareh, Allen, Rosalind J.
• Format: Article
• Language: English
• Subjects: Amdinocillin; Amides; Amino acids; Antibiotics; Aqueous solutions; Astronomy; Aztreonam; Bacteria; Bioassays; Biodegradation; Biology and Life Sciences; Cefotaxime; Cell division; Degradation; Delay time; E coli; Glucose; Gram-negative bacteria; Growth media; Half-life; Laboratories; Medicine and Health Sciences; Methods; Minimum inhibitory concentration; Mopping; Penicillin; pH effects; Physical Sciences; Physics; Research and Analysis Methods; Stability; β-Lactam antibiotics
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0236198

Laboratory assays such as MIC tests assume that antibiotic molecules are stable in the chosen growth medium—but rapid degradation has been observed for antibiotics including β-lactams under some conditions in aqueous solution. Degradation rates in bacterial growth medium are less well known. Here, we develop a ‘delay time bioassay’ that provides a simple way to estimate antibiotic stability in bacterial growth media, using only a plate reader and without the need to measure the antibiotic concentration directly. We use the bioassay to measure degradation half-lives of the β-lactam antibiotics mecillinam, aztreonam and cefotaxime in widely-used bacterial growth media based on MOPS and Luria-Bertani (LB) broth. We find that mecillinam degradation can occur rapidly, with a half-life as short as 2 hours in MOPS medium at 37°C and pH 7.4, and 4-5 hours in LB, but that adjusting the pH and temperature can increase its stability to a half-life around 6 hours without excessively perturbing growth. Aztreonam and cefotaxime were found to have half-lives longer than 6 hours in MOPS medium at 37°C and pH 7.4, but still shorter than the timescale of a typical minimum inhibitory concentration (MIC) assay. Taken together, our results suggest that care is needed in interpreting MIC tests and other laboratory growth assays for β-lactam antibiotics, since there may be significant degradation of the antibiotic during the assay.