Lipid hydroperoxide levels in plant tissues

Hydroperoxides are the primary oxygenated products of polyunsaturated fatty acids and are key intermediates in the octadecanoid signalling pathway in plants. Lipid hydroperoxides (LHPO) were determined spectrophotometrically based on their reaction with an excess of Fe2+ at low pH in the presence of the dye xylenol orange. Triphenylphosphine-mediated hydroxide formation wa s used to authenticate the signal generated by the hydroperoxides. The meth od readily detected lipid peroxidation in Phaseolus microsomes, senescing p otato leaves and in a range of other plant tissues including Phaseolus hypo cotyls (26 +/- 5 nmol g(-1) FW), Alstroemeria floral tissues (sepals 66+/-1 3 nmol g(-1) FW; petals 49+/-6 nmol g(-1) (1) FW), potato leaves (334 +/- 7 5 nmol g(-1) FW), broccoli florets (568+/-68 nmo g(-1) FW) and Chlamydomona s cells (602 +/- 40 nmol g(-1) FW). Relative to the total fatty acid conten t of the tissues, the % LHPO was within the range of 0.6-1.7% for all tissu e types (photosynthetic and non-photosynthetic) and represents the basal ox idation level of membrane fatty acids in plant cells. In order to relate th e levels of LHPO to specific signalling pathways, transgenic potato plant l ines were used in which lipoxygenase (LOX) (responsible for hydroperoxide b iosynthesis) and hydroperoxide lyase (a route of hydroperoxide degradation) activities were largely reduced by an antisense-mediated approach. While t he LHPO levels were similar to wild type in the individual LOX antisensed p lants, basal LHPO levels, by contrast, were elevated by 38% in transgenic p otato leaves antisensed in hydroperoxide lyase, indicating a role for this enzyme in the maintenance of cellular levers of LHPOs.