Permeation and toxicity of ethylene glycol and methanol in larvae of Anopheles gambiae

Abstract

In this study, we applied proton NMR to measure the permeation of two cryoprotective agents (CPAs), ethylene glycol (EG) and methanol, into 1st instar Anopheles larvae. Calibration with standard solutions of EG or methanol (0-10 mol l-1) confirmed the reliability of the NMR measurements for determining the concentration of these solutes. To assess permeation, larvae were immersed in 1.5 mol l-1 EG or 1.5 mol l-1 methanol for different periods of time at 22°C. The concentration of both CPAs in the larvae was then measured as a function of exposure time using 1H-NMR spectroscopy. Results show that after a 6 h exposure to 1.5 mol l-1 EG, the larval concentration of EG reaches a maximum value of 1.44 mol l-1, which is 96% of the theoretical maximum. By contrast, after just 1 h exposure to 1.5 mol l-1 methanol, the larval methanol concentration reaches its maximum, which, however, is only 75% of the theoretical maximum. Toxicity data show that larval survival remains 91% and 95% after 4 h and 1 h exposure to 1.5 mol l-1 EG and 1.5 mol l-1 methanol, respectively, at which time the larval concentration of EG and methanol has risen to 1.21 mol l-1 and 1.13 mol l-1, respectively. These results suggest that CPAs such as EG and methanol do permeate Anopheles larvae to up to 81% and 75% of equilibrium, respectively, before the exposure becomes toxic.