Contents SummaryPlants are subject to a wide range of abiotic stresses, and their cuticular wax layer provides a protective barrier, which consists predominantly of long-chain hydrocarbon compounds, including alkanes, primary alcohols, aldehydes, secondary alcohols, ketones, esters and other derived compounds. This article discusses current knowledge relating to the effects of stress on cuticular waxes and the ways in which the wax provides protection against the deleterious effects of light, temperature, osmotic stress, physical damage, altitude and pollution. Topics covered here include biosynthesis, morphology, composition and function of cuticular waxes in relation to the effects of stress, and some recent findings concerning the effects of stress on regulation of wax biosynthesis are described.New Phytologist (2006) 171: 469-499

SummaryMetabolic profiling was carried out in the forage grass Lolium perenne L. (perennial ryegrass) to uncover mechanisms involved in the plants response to water stress. When leaf and root materials from two genotypes, with a contrasting water stress response, were analysed by GC-MS, a clear difference in the metabolic profiles of the leaf tissue under water stress was observed. Differences were principally due to a reduction in fatty acid levels in the more susceptible Cashel genotype and an increase in sugars and compatible solutes in the more tolerant PI 462336 genotype. Sugars with a significant increase included: raffinose, trehalose, glucose, fructose and maltose. Increasing the ability of perennial ryegrass to accumulate these sugars in response to a water deficit may lead to more tolerant varieties. The metabolomics approach was combined with a transcriptomics approach in the water stress tolerant genotype PI 462336, which has identified perennial ryegrass genes regulated under water stress.

Phytochemical diversity with respect to a range of polar (including amino acids, organic acids, sugars, and sugar alcohols) and nonpolar (including fatty acids, alkanols, and sterols) metabolites was examined within tubers from a total of 29 genetically diverse potato cultivars and Chilean landraces using a metabolomics approach by gas chromatography-mass spectrometry. From principal component analysis of the polar and nonpolar metabolite data there was insufficient variation to differentiate the majority of cultivars and landraces. Analysis of all polar metabolite profiles revealed separation of two cultivars (Glenna and Morag) from the other cultivars and landraces and a separate cluster of one landrace line, largely due to higher levels of sugars. Pentland Javelin was distinct in containing high levels of many amino acids. The two Solanum tuberosum group phureja cultivars (Inca Sun and Mayan Gold) were not particularly similar and were not separated from the S. tuberosum group tuberosum cultivars. Analysis of the nonpolar metabolite data revealed partial separation of two landrace lines and, on the basis of some minor fatty acids, Mayan Gold was distinct. The differences in metabolite profiles are considered in terms of the taxonomy and breeding history of the cultivars and possible influences from other factors such as developmental stage of the tuber. With a view to exploring biosynthetic links between metabolites, a pairwise correlation analysis was performed on all metabolites. The significance of high correlations between many amino acids and between several nonpolar metabolites is discussed.

Metabolic profiling methods are not ideally suited to the simultaneous analysis of all metabolite classes within a biological sample and must be optimized for maximum applicability. Several factors related to the optimization, validation and limitations of a GC-MS-based metabolic profiling method for potato were examined. A key step is conversion of reducing sugars to methyloximes, and optimum reaction conditions were 50°C for 4 h. Shorter times or lower temperatures resulted in incomplete oximation whereas longer times and higher temperatures caused hydrolysis of sucrose, the major tuber dissacharide. Metabolite concentration gradients were observed in tuber sections. Glucose, fructose, alanine, methionine, threonine and tyrosine were more concentrated in the interior, whereas asparagine, putrescine, and caffeic and chlorogenic acids were higher in the skin and citrate was concentrated at the tuberÕs bud end. These results impact upon choice of sampling strategy, consequently the use of freeze-dried (FD) material from a sampling protocol developed to avoid gradient-induced bias was examined. Using FD material, the method was highly linear and there was little qualitative or quantitative difference in the metabolite composition between fresh and FD material. The short-and long-term repeatability of the method was studied, and the use of reference materials to monitor and to improve data quality is discussed. Ascorbate is an important tuber metabolite that is readily measured by targeted approaches, but can be a problem in metabolic profiling. It was shown for standards and FD potato that ascorbate was largely degraded during oximation, although some survived in FD material.

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