Henri W. Groeneveld

Photosynthetic acclimation to high light conditions in mature leaves of Coffea arabica L. : role of xanthophylls, quenching mechanisms and nitrogen nutrition

Young coffee plants (Coffea arabica L. cv. Catuai), originally from a shaded habitat, were separated in three groups to be grown under different levels of N fertilization: 0.3 mmol N supplements were given to the soil every 7 days (high N treatment, 2N), every 15 days (medium N treatment, 1N) and every 45 days (low N treatment, 0N). These plants were later exposed to a high sunlight irradiance (noon PPFD up to 1500 µmol m–2 s–1 ) for a period of 12 or 15 days. Among others, the values of electron trans-port capacity, maximum carboxylation activity, photosynthetic capacity (Amax) and several fluorescence parameters (Fv/Fm, Fv´/Fm´, qP, pe) first showed a reduction (until the 4th–7th day) in all N treatments, followed by an N-dependent recovery. The 2N plants were less affected in the first few days and, at the end of the stress period, showed a better recovery for most of the studied parameters and the highest increase in the saturating PPFD for net photosynthesis and A max . The present work shows that the ability to acclimate displayed by the mature leaves of 2N plants was accompanied by an increase in energy dis-sipation mechanisms. These include an increase in the ‘high energy’ quenching and, mostly, the presence of higher contents of some xanthophylls (zeaxanthin and lutein) and carotenes, which helped to decrease the energetic overcharge in the photosystems. Pigment changes in mature leaves suggest that N can promote specific mechanisms of acclimation others than those that might be expected from a preferential partition of the element N into photosynthetic components.

Submergence-induced petiole elongation in Rumex palustris is controlled by developmental stage and storage compounds

Submergence stimulates elongation of the leaves of Rumex palustris and under laboratory conditions the maximum final leaf length (of plants up to 7 weeks old) was obtained within a 9 day period. This elongation response, mainly determined by petiole elongation, depends on the availability of storage compounds and developmental stage of a leaf. A starch accumulating tap root and mature leaves and petioles were found to supply elongating leaves with substrates for polysaccharide synthesis in expanding cell walls. Changes in the composition of cell wall polysaccharides of elongated petioles suggest a substantial cell wall metabolism during cell extension. Reduced starch levels or removal of mature leaves caused a substantial limitation of submerged leaf growth. From the 5th leaf onward enough reserves were available to perform submerged leaf growth from early developmental stages. Very young petioles had a limited capacity to elongate. In slightly older petioles submergence resulted in the longest final leaf lengths and these values gradually decreased when submergence was started at more mature developmental stages. Submerged leaf growth is mainly a matter of petiole elongation in which cell elongation has a concurrent synthesis of xylem elements in the vascular tissue. Mature petioles still elongated (when submerged) by cell and tissue elongation only: the annular tracheary elements stretched enabling up to 70% petiole elongation.

Rapid, quantitative HPLC analysis ofAsclepias fruticosa L. andDanaus plexippus L. cardenolides.

The cardenolide extracts from latex and aerial parts ofAsclepias fruticosa and ofDanaus plexippus reared onA. fruticosa orA. curassavica were purified by adsorption chromatography on silica gel. HPLC analysis on a C18 reverse-phase column with an acetonitrile-water gradient as mobile phase, separated 28 compounds with a UV spectrum typical forcardenolides. Afroside and gomphoside (major components), as well as calotropagenin, calotoxin, calotropin, calactin, uscharidin, uscharin, and voruscharin, occurred as single peaks in the profiles of latex and aerial plant parts ofA. fruticosa. Calactin and calotropin were the major cardenolides inDanaus plexippus reared onA. fruticosa orA. curassavica. Quantitative data obtained with digitoxin as internal standard showed that 1.3–1.5% of the leaf cardenolides were sequestered byDanaus plexippus in which levels of 70–80μg cardenolide per butterfly were measured. The calotropin from the leaves was almost completely sequestered, and 10–13% of the calactin was stored by the butterfly, assuming that no conversion occurred in larval tissues.

The composition of phytosterols, latex triterpenols and wax triterpenoids in the seedling of Euphorbia lathyris L.

Abstract In the etiolated seedling of Euphorbia lathyris L., three groups of triterpenoids were found, located at different sites. The extracted triterpenoids were purified by TLC and HPLC and identified by GC/MS. Latex consisted mainly of triterpene alcohols. From these, guimarenol, 24-CH3-lanosterol and hopenol-B were not earlier identified in E. lathyris. The epicuticular wax contained mainly triterpene ketones, which were identified as taraxeron, hopenone-I, hopenone-II, hopenone-B, 3-oxo-22-hydroxyhopane, simiarenone and fernenone. Other components of the epicuticular wax were triterpenols, n-alkanes and aliphatic alcohols. The phytosterols were identified as cholesterol, campesterol, stigmasterol, sitosterol and isofucosterol.

The involvement of sucrose, glucose and other metabolites in the synthesis of triterpenes and dopa in the laticifers of Euphorbia lathyris

Abstract A quantitative triterpene analysis was made of latex stem tissue of Euphorbia lathyris . Young plants seedlings of E. lathyris were incubated with various labelled precursors. Incorporation into triterpenes was obtained from [2- 14 C]mevalonic acid, [1- 14 C]acetate, [3- 14 C]pyruvate, [U- 14 C]sucrose, [U- 14 C]glucose, [U- 14 C]xylose, [U- 14 C]glyoxylate, [2,3- 14 C]succinic acid, [1- 14 C]glycerol [U- 14 C]serine. Both sugars tyrosine appeared to be effective precursors in DOPA synthesis inside the laticifers. Exogenously supplied mevalonic acid was only involved in triterpene synthesis outside the laticifers. GC-RC of triterpenes synthesized from [U- 14 C]glucose revealed the origin of these compounds in the latex. The labelled triterpenes obtained after incorporation of the other mentioned labelled precursors were only partly synthesized in the laticifers. For quantitative data on latex triterpene synthesis seedlings were incubated with [U- 14 C]sucrose, [U- 14 C]glucose, [U- 14 C]xylose [1- 14 C]acetate in the presence of increasing amounts of unlabelled substrate. From the amount of 14 C incorporated into the triterpenes the amount of substrate directly involved in triterpene synthesis was calculated, as was the absolute triterpene yield. Sucrose showed the highest triterpene yield, equivalent to the daily increase of the triterpene content of growing seedlings. The possible significance of the other precursors in triterpene synthesis in the laticifers is discussed.

Cardenolide biosynthesis from malonate in Asclepias curassavica.

Abstract Cardenolides, free sterols, triacylglycerols, triterpenyl acetates, steryl- and triterpene esters were the major lipid classes in the stem of five- to eight-week-old Asclepias curassavica plants, in which 30% of the cardenolides occurred in the latex. Excised defoliated stems incorporated up to 6.7% of radioactive acetate into these lipids in a three day incorporation period. The label from [2- 14 C]acetate proceeded mainly to the fatty acid moiety of the triterpene and steryl esters, the fatty acids of the triacylglycerols, the free sterols and the cardenolides. Optimal results were obtained with 10 cm stems from six-week-old plants. An increased supply of acetate (2.5–10 μmol stem −1 ) showed a saturated synthesis of cardenolides and triacylglycerols and caused an enhanced production of triterpenyl acetates. The highest yield of [ 14 C]cardenolides was obtained after uptake of 5 μmol of acetate per stem. The cardenolide composition of the incubated stem tissue increased on the absorption of 10 μmol of acetate and major changes in organic acid composition were measured after uptake of 20 μmol of acetate. [ 13 C]Uscharidin was biosynthesized from 0.5 mmol of [1,2- 13 C]acetate by 100 stem tips. 13 C NMR spectroscopy showed an unequal relative enrichment in both the genin and the dideoxyhexosulose moiety. The construction of the butenolide ring by the condensation of a pregnane derivative with one molecule acetate as observed for 5β-cardenolides is not confirmed by these-NMR data.

Triterpenoid biosynthesis in the seedling of Euphorbia lathyris L. from sucrose and amino acids

Abstract The triterpenoid biosynthesis from sucrose and several amino acids was investigated with 9-day-old Euphorbia lathyris seedlings. Tracer experiments with [U-14C]sucrose demonstrated that sucrose was mainly incorporated into latex triterpenols. Several [U-14C]amino acids were found to be efficient precursors in the synthesis of sterols and the wax triterpene ketones. The incorporation data of sucrose and amino acids into triterpenoids together with the daily uptake of these substrates from the endosperm were used to estimate the daily triterpenoid production from each substrate species. Sucrose was quantitatively the most important precursor in triterpenoid biosynthesis. About 10% of the sterols and wax triterpenoids were produced from the amino acids.

Triterpenoid biosynthesis in the etiolated seedling of Euphorbia lathyris L.. Developmental changes and the regulation of local triterpenoid production

Summary Intact seedlings of Euphorbia lathyris were used to measure the daily production of sterols, latex triter-penols and wax triterpene ketones, and to estimate the participation of sucrose and threonine in the synthesis of these lipids. In the early stage of seedling development, 3–4 days after seed imbibition (DAI), the biosynthesis of these triterpenoids is limited by enzymatic production capacity. From 5 DAI onwards, the triterpenoid production seemed to be restricted by the availability of sugars. Threonine played only a marginal role in the synthesis of triterpenoids. From 5-8 DAI, most of the triterpenoids are synthesized directly from sucrose taken up from the endosperm by the cotyledons. From 8–12 DAI, sugars from the hypocotyl and root became another source of carbon for triterpenoid synthesis. In 9-day-old seedlings, the production of latex triterpenols and triterpene ketones was most prominent in the rapidly elongating parts of the hypocotyl. The activity of sterol synthesis was most pronounced in the root tip.

The involvement of amino acids in latex lipid synthesis in Euphorbia lathyris seedlings

The breakdown of triglycerides and proteins in the endosperm of Euphorbia lathyris was assayed in a 14 day germination period. Six days after germination, the average daily production was 2.7 μmol of amino acids. Arginine, glutamine, asparagine and glutamic acid accounted for 53% of the total amino acids. Excised cotyledons with 1 cm hypocotyls were used for amino acid uptake and their involvement in terpenoid synthesis was studied. Glutamine and aspartate were hardly involved in apolar lipid synthesis. Leucine, isoleucine, valine and threonine were mainly incorporated into the triterpenes in the laticifers. Alanine and serine were also involved in phytosterol synthesis in the adjacent tissue. In the 14 day germination period, ca3% of the daily yield of latex triterpenes may be synthesized from a variety of amino acids.

Serine in sterol synthesis in euphorbia lathyris seedlings

Abstract During germination serine was shown to be taken up from the endosperm by the cotyledons and partly translocated to the hypocotyl. Up to 1.7% of the [ 14 C]serine taken up by the seedling was involved in phytosterol synthesis in which the hypocotyl was the most effective plant part. Compared with [ 14 C]sucrose, serine was found to be 12 times more effective in this synthesis. From the data obtained it could be calculated that this amino acid may yield about 7% of all the free sterols in an etiolated seedling of Euphorbia lathyris , making this substrate a suitable marker in sterol synthesis in endospermous seedlings.