Examination of the translocation of sulfonylurea herbicides in sunflower plants by matrix-assisted laser desorption/ionisation mass spectrometry imaging.

Pesticides are widely used in agriculture to control weeds, pests and diseases. Successful control is dependent on the compound reaching the target site within the organism after spray or soil application. Conventional methods for determining uptake and movement of herbicides and pesticides include autoradiography, liquid scintillation and chromatographic techniques such as high-performance liquid chromatography (HPLC). Autoradiography using radiolabelled compounds provides the best indication of a compound's movement within the plant system. Autoradiography is an established technique but it relies on the synthesis of radiolabelled compounds. The distribution of four sulfonylurea herbicides in sunflower plants has been studied 24 h after foliar application. The use of matrix-assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI) images of protonated molecules and fragment ions (resulting from fragmentation at the urea bond within the sulfonylurea herbicides) has provided evidence for translocation above and below the application point. The translocation of nicosulfuron and azoxystrobin within the same plant system has also been demonstrated following their application to the plant stem. This study provides evidence that MALDI-MSI has great potential as an analytical technique to detect and assess the foliar, root and stem uptake of agrochemicals, and to reveal their distribution through the plant once absorbed and translocated.

Examination of the distribution of nicosulfuron in sunflower plants by matrix-assisted laser desorption/ionisation mass spectrometry imaging.

Matrix-assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI) has been used to image the distribution of the pesticide nicosulfuron (2-[[(4,6-dimethoxypyrimidin-2-yl)aminocarbonyl]aminosulfonyl]-N,N-dimethyl-3-pyridinecarboxamide) in plant tissue using direct tissue imaging following root and foliar uptake. Sunflower plants inoculated with nicosulfuron were horizontally sectioned at varying distances along the stem in order to asses the extent of translocation; uptake via the leaves following foliar application to the leaves and uptake via the roots from a hydroponics system were compared. An improved sample preparation methodology, encasing samples in ice, allowed sections from along the whole of the plant stem from the root bundle to the growing tip to be taken. Images of fragment ions and alkali metal adducts have been generated that show the distribution of the parent compound and a phase 1 metabolite in the plant. Positive and negative controls have been included in the images to confirm ion origin and prevent false-positive results which could originate from endogenous compounds present within the plant tissue.

Carboxylesterase activities toward pesticide esters in crops and weeds.

Proteins were extracted from maize, rice, sorghum, soybean, flax and lucerne; the weeds Abutilon theophrasti, Echinochloa crus-galli, Phalaris canariensis, Setaria faberii, Setaria viridis, Sorghum halepense and the model plant Arabidopsis thaliana and assayed for carboxylesterase activity toward a range of xenobiotics. These included the pro-herbicidal esters clodinafop-propargyl, fenoxaprop-ethyl, fenthioprop-ethyl, methyl-2,4-dichlorophenoxyacetic acid (2,4-d-methyl), bromoxynil-octanoate, the herbicide-safener cloquintocet-mexyl and the pyrethroid insecticide permethrin. Highest activities were recorded with alpha-naphthyl acetate and methylumbelliferyl acetate. Esters of p-nitrophenol were also readily hydrolysed, with turnover declining as the chain length of the acyl component increased. Activities determined with model substrates were much higher than those observed with pesticide esters and were of limited value in predicting the relative rates of hydrolysis of the crop protection agents. Substrate preferences with the herbicides were typically 2,4-d-methyl>clodinafop-propargyl>fenthioprop-ethyl, fenoxaprop-ethyl and bromoxynil-octanoate. Isoelectric focussing in conjunction with staining for esterase activity using alpha-naphthyl acetate as substrate confirmed the presence of multiple carboxylesterase isoenzymes in each plant, with major qualitative differences observed between species. The presence of serine hydrolases among the resolved isoenzymes was confirmed through their selective inhibition by the organophosphate insecticide paraoxon. Our studies identify potentially exploitable differences between crops and weeds in their ability to bioactivate herbicides by enzymic hydrolysis and also highlight the usefulness of Arabidopsis as a plant model to study xenobiotic biotransformation.

Determination of agrochemical compounds in soya plants by imaging matrix-assisted laser desorption/ionisation mass spectrometry.

Detection and imaging of the herbicide mesotrione (2-(4-mesyl-2-nitrobenzoyl)cyclohexane-1,3-dione) and the fungicide azoxystrobin (methyl (E)-2-{2-[6-(2-cyanophenoxy)pyrimidin-4-yloxy]phenyl}-3-methoxyacrylate), on the surface of the soya leaf, and the detection and imaging of azoxystrobin inside the stem of the soya plant, have been achieved using matrix-assisted laser desorption/ionisation quadrupole time-of-flight mass spectrometry. In leaf analysis experiments, the two pesticides were deposited onto the surface of individual soya leaves on growing plants. The soya leaves were removed and prepared for direct and indirect (following blotting onto matrix-coated cellulose membranes) imaging analysis at different periods after initial pesticide application. In stem analysis experiments, azoxystrobin was added to the nutrient solution of a soya plant growing in a hydroponics system. The plant was left for 48 h, and then horizontal and vertical stem sections were prepared for direct imaging analysis. The images obtained demonstrate the applicability of MALDI imaging to the detection and imaging of small organic compounds in plant tissue and further extend the analytical repertoire of the versatile MALDI technique.