Stability of β-lactam antibiotics in bacterial growth media

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

D, L-Sulforaphane loaded Fe3O4@ gold core shell nanoparticles: A potential sulforaphane delivery system

A novel design of gold-coated iron oxide nanoparticles was fabricated as a potential delivery system to improve the efficiency and stability of d, l-sulforaphane as an anticancer drug. To this purpose, the surface of gold-coated iron oxide nanoparticles was modified for sulforaphane delivery via furnishing its surface with thiolated polyethylene glycol-folic acid and thiolated polyethylene glycol-FITC. The synthesized nanoparticles were characterized by different techniques such as FTIR, energy dispersive X-ray spectroscopy, UV-visible spectroscopy, scanning and transmission electron microscopy. The average diameters of the synthesized nanoparticles before and after sulforaphane loading were obtained ∼ 33 nm and ∼ 38 nm, respectively, when ∼ 2.8 mmol/g of sulforaphane was loaded. The result of cell viability assay which was confirmed by apoptosis assay on the human breast cancer cells (MCF-7 line) as a model of in vitro-cancerous cells, proved that the bare nanoparticles showed little inherent cytotoxicity, whereas the sulforaphane-loaded nanoparticles were cytotoxic. The expression rate of the anti-apoptotic genes (bcl-2 and bcl-xL), and the pro-apoptotic genes (bax and bak) were quantified, and it was found that the expression rate of bcl-2 and bcl-xL genes significantly were decreased when MCF-7 cells were incubated by sulforaphane-loaded nanoparticles. The sulforaphane-loaded into the designed gold-coated iron oxide nanoparticles, acceptably induced apoptosis in MCF-7 cells

Magnetization hysteresis loop of NPs.

<p>The room-temperature magnetization (<i>M</i>) hysteresis loop curves of the same mass of [Fe₃O₄@Au] NPs and FITC/FA@[Fe₃O₄@Au] NPs as a function of applied magnetic field (<i>H</i>).</p

Release of FITC from NPs (A) and average hydrodynamic (B) of NPs.

<p>A: Release of FITC from FITC/FA@[Fe₃O₄@Au] NPs. B: The average hydrodynamic diameter of [Fe₃O₄@Au] and FITC/FA@[Fe₃O₄@Au] NPs at <i>p</i>H = 7.4.</p

Optical microscopy images of cells after 24 h.

<p>A: Bright-field image, B: Fluorescence image of DAPI detection, and C: Combined of A and B, after 24 h incubation of MCF-7 cells (control). D: Bright-field image, E: Fluorescence image of DAPI detection, and F: Combined of D and E, after 24 h incubation of MCF-7 cells with free SF. G: bright-field image, H: fluorescence image of DAPI detection, and I: fluorescence image of FITC detection of the same area after 24 h incubation of MCF-7 cells with FITC@[Fe₃O₄@Au] NPs.</p

Viability of cells at different conditions.

<p>Viability of MCF-7 cells which were assessed A: after 24 h, 48 h, and 72 h of incubation with different concentrations of SF-loaded FITC/FA@[Fe₃O₄@Au] NPs. B: at different concentrations of incubation with free SF, FITC/FA@[Fe₃O₄@Au] NPs, SF-loaded FITC@[Fe₃O₄@Au] NPs, and SF-loaded FITC/FA@[Fe₃O₄@Au] NPs. Data are presented as the mean ± standard deviation of replicates.</p

Optical microscopy images of incubated cells by SF-loaded FITC/FA@[Fe₃O₄@Au] NPs after 24 and 72 h.

<p>A: bright-field image, B: fluorescence image of FITC detection, and C: fluorescence image of DAPI detection of the same area after 24 h incubation of MCF-7 cells with SF-loaded FITC/FA@[Fe₃O₄@Au] NPs. D: The corresponding fluorescence intensity maps (colorbar shown) of FITC (in SF-loaded FITC/FA@[Fe₃O₄@Au] NPs) distribution of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0151344#pone.0151344.g009" target="_blank">Fig 9B</a>. E: Combined of A and B. F: Combined of A and C. G: bright-field image, H: fluorescence image of FITC detection, and I: fluorescence image of DAPI detection of the same area after 72 h incubation of MCF-7 cells with SF-loaded FITC/FA@[Fe₃O₄@Au] NPs. J: The corresponding fluorescence intensity maps (colorbar shown) of FITC (in SF-loaded FITC/FA@[Fe₃O₄@Au] NPs) distribution of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0151344#pone.0151344.g009" target="_blank">Fig 9B</a>. K: Combined of G and H. L: Combined of G and I.</p