The carotenoid content of plants may be impacted by stress with major consequences for photosynthesis and photoprotection. Most carotenoid stress research, however, has concentrated on abiotic stresses, and we know little about how biological stresses, such as herbivory, alter profiles of plant carotenoids and their degradation products. For example, carotenoid derivatives such as β-cyclocitral and β-ionone have been recently shown to act as signals in plant growth and protection against oxidative stress and herbivory. To understand how carotenoid composition is influenced by herbivory, changes in biosynthesis and degradation should be investigated. This chapter describes methods to simulate herbivory in a simple reproducible fashion and to assess carotenoid biosynthesis and degradation. Carotenoid biosynthesis depends on precursors provided by the methylerythritol 4-phosphate (MEP) pathway, which converts pyruvate and glyceraldehyde-3-phosphate to the five-carbon units used for construction of larger isoprenoids. We present protocols to quantify the activity of the first enzyme of the MEP pathway, deoxy-xylulose 5-phosphate synthase (DXS), usually assumed to be rate-controlling, and to estimate the concentration of the first intermediate of the pathway, deoxy-xylulose 5-phosphate (DXP). We also discuss procedures to measure the formation of volatile carotenoid breakdown products after herbivory. To monitor the activity of carotenoid-specific biosynthetic enzymes, such as phytoene synthase, protocols are available elsewhere in this volume (Wurtzel, 2022b).
Keywords: Apocarotenoids; GC–MS; Herbivory; LC-MS; Oxidative stress; Simulated herbivory; Singlet oxygen; Wounding.
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