AbstractMetabolomics is a technique that can be used to investigate the metabolic profiles of an organism by measuring a large number of the low molecular weight metabolites in the metabolic pool. 1H NMR spectroscopy is an analytical tool that is unbiased, and a non-destructive way of investigating metabolic perturbations in organisms exposed to environmental stress.
In this study two model organisms, Mytilus edulis and Chironomus riparius were used to investigate the effects of several pesticides: lindane and atrazine in the mussels, and fenitrothion, methiocarb and permethrin in the midge larvae. The impact of hypoxia and starvation were also investigated alongside the exposure to atrazine in the mussel. The pesticides used in the mussel study have different modes of action, but produce similar changes to behaviour and can cause starvation and mild hypoxia. Acetonitrile/2H2O (60/40 % v/v) extracts of foot muscle of mussels subjected to hypoxia, or starvation, or to low or high doses of pesticide were analysed using 1H NMR spectroscopy to produce metabolic profiles associated with these treatments. Discriminant analysis showed significant differences between treated and control animals and gave a clear separation between all treatment groups. Atrazine profiles were clearly separated from the starved and hypoxic animals and the animals exposed to high and low doses were also separated. Lindane treatment was separated from control animals in a dose-dependent way. This was associated with an increase in alanine concentrations and a decrease in all other identified metabolites. The study of midge larvae used the same approach, but using extracts of pooled whole body homogenates (10 animals per sample) instead of tissue from individuals. The animals were subjected to low, environmentally relevant dose levels of three pesticides; fenitrothion, methiocarb and permethrin. The first two of these have a similar mode of action, inhibition of acetylcholine esterase, whilst the latter is an axonal poison acting on cation channels. The metabolic profiles associated with these treatments showed a clear separation between all treatment groups and between treated and control animals. Fenitrothion treatment was associated with an increase in alanine (on average of 93.3 M relative to controls) and lactate concentrations compared with controls and other treatments. Methiocarb caused a reduction in arginine, leucine and lysine concentrations to half of the control level. Permethrin produced a reduction in tyrosine and phenylalanine concentrations to half that of the control group. In both these experiments the use of 1H NMR metabolomics enabled the separation the effects of all of the treatments and stressors from each other and from the controls. It demonstrates the potential of the metabolomic approach to provide separation of the effects of poisoning from those of environmental stress, and to distinguish between toxicants with similar modes of action.
|Date of Award||Sept 2010|
|Supervisor||Graham Mills (Supervisor) & Richard Greenwood (Supervisor)|