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The summer climate in Maritime Antarctica is characterised by high humidity and cloudiness with slightly above zero temperatures. Under such conditions, photosynthetic activity is temperature-limited and plant communities are formed by a few species. These conditions could prevent the operation of the photoprotective xanthophyll (VAZ) cycle as low irradiance reduces the excess of energy and low temperatures limit enzyme activity. The VAZ cycle regulates the dissipation of the excess of absorbed light as heat, which is the main mechanism of photoprotection in plants. To test whether this mechanism operates dynamically in Antarctic plant communities, we characterised pigment dynamics under natural field conditions in two representative species: the moss Polytrichum juniperinum and the grass Deschampsia antarctica. Pigment analyses revealed that the total VAZ pool was in the upper range of the values reported for most plant species, suggesting that they are exposed to a high degree of environmental stress. Despite cloudiness, there was a strong conversion of violaxanthin (V) to zeaxanthin (Z) during daytime. Conversely, the dark-induced enzymatic epoxidation back to V was not limited by nocturnal temperatures. In contrast with plants from other cold ecosystems, we did not find any evidence of overnight retention of Z or sustained reductions in photochemical efficiency. These results are of interest for modelling, remote sensing and upscaling of the responses of Antarctic vegetation to environmental challenges. The Spanish Ministry of Science, Innovation and Universities (MICIU/FEDER, EU) and the Basque Government funded this research through the projects CTM2014-53902-C2-2-P, PGC2018-093824-B-C44 and UPV/EHU IT-1018-16. Open Access funding provided thanks to the CRUE-CSIC agreement with Springer Nature.

The aim of this study was to evaluate the effects of melatonin on photoinhibition under chilling stress in cucumber seedlings and to inquire into any mechanisms of mitigation. Under chilling stress, the net photosynthetic rate declined dramatically but the decline was significantly mitigated by irrigation with a melatonin solution. Possible mechanisms for this mitigation are that melatonin accelerates xanthophyll de-epoxidation by upregulating the transcription of the violaxanthin de-epoxidase gene (CsVDE) and down-regulating that of the zeaxanthin cyclase gene (CsZE) during chilling. There was also a rise in non-photochemical quenching (NPQ) if seedlings were pretreated with melatonin before chilling. The efficient operation of the xanthophyll cycle helped consume excessive excitation energy in photosystem (PS) Ⅱ and so protected the photosynthetic system. Melatonin also modulated the partitioning of absorbed excitation energy in PS Ⅱ as evidenced by alleviation of the decrease in quantum yield of photochemical energy conversion in PS Ⅱ under chilling stress, by alleviation of the rise in quantum yield of non-regulated, non-photochemical energy loss in PS II and by increasing the regulated non-photochemical energy loss in PS II. This study presents a new understanding of the mechanisms through which melatonin mitigates photoinhibition by modulating the partitioning of absorption energy in PS Ⅱ based on the xanthophyll cycle.

A valorization strategy for an aggravating type of plant waste is put to the test herein. It envisions the use of Japanese knotweed green leaves as a sustainable source of free lutein, from which bioactive diesters could be prepared as potential value-added products with improved properties. To this end, 13 structurally distinct model lutein diesters were synthesized and the relationships between their structure and stability were systematically determined. The forced degradation data show that the stability of a particular lutein diester may depend to a large extent on the type of exposure (elevated temperature, light, oxidant, or acidic environment) and, more importantly, not every esterification attempt necessarily leads to an enhancement of lutein's chemical stability. However, three branched and bulky products-lutein di(2,2-dimethylpropanoate), lutein di(2-methylpropanoate), and lutein di(3-methylbutanoate)-proved to be particularly relevant, as they consistently exhibited 1.5-21-fold higher stability compared to free lutein, regardless of the stress conditions used. Finally, we show that the Japanese knotweed plant matrix had a significant negative or positive effect on pigment degradation kinetics that could not be easily predicted. Thus, the proposed valorization strategy is quite feasible, but the esterification approach should be tailored to the intended use of a lutein diester.

This study aimed to optimize the key parameters of extraction methods and to increase the recovery yields of intact xanthophylls (violaxanthin, zeaxanthin, astaxanthin) from microalgae (Chlorella luteoviridis). An effective, simple, and fast extraction protocol is described. It consists of a grinding pretreatment followed by a microwave-assisted extraction, using ethanol 90% as an environmentally preferable extraction solvent. Xanthopylls were quantified using high performance liquid chromatography. Irradiation time of 6 s only resulted in the extraction of violaxanthin (4.479 ± 0.009 mg/g), astaxanthin (4.154 ± 0.013 mg/g), and zeaxanthin (4.776 ± 0.120 mg/g). The described protocol seems to be the fastest extraction method of xantophylls compared to the literature and could be an advantage for industrial scale, while saving time and energy.

The toxic potential of Chattonella is associated with a high production of reactive oxygen species (ROS). Chattonella species can tolerate high irradiance levels but seems not to be efficient in the induction of nonphotochemical chl a fluorescence (NPQ) under light stress conditions. Therefore, we postulated that high ROS production of this microalgal group is related to the lack of effective photoprotection mechanisms. We compared the NPQ induction, xanthophyll cycle interconversion (XC), and the production of the ion superoxide (O2 - ) in Chattonella marina var. antiqua, Chattonella sp., and C. marina acclimated to 43 (LL) and 300 µmol photons · m2 · s-1 (HL). We also evaluated the photosynthetic characteristics of the three strains. Photosynthesis saturated at relative high irradiances (above 500 µmol photons · m2 · s-1 ) in LL and HL Chattonella strains. For the first time, we documented the conversion of diadinoxanthin into diatoxanthin in microalgae that have violaxanthin as the major XC carotenoid. The slow NPQ induction indicated that qE (fast component of NPQ) was not present, and this process was related to the interconversion of XC pigments. However, the quenching efficiency (QE) of deepoxidated xanthophylls was low in the three Chattonella strains. The strain with the lowest QE produced the highest amount of a O2 - . Therefore, ROS production in Chattonella seems to be related to a low expression of XC-related thermal PSII dissipation.

Xanthophyll cycles are broadly important in photoprotection, and the reversible de-epoxidation of xanthophylls typically occurs in excess light conditions. However, as presented in this review, compiling evidence in a wide range of photosynthetic eukaryotes shows that xanthophyll de-epoxidation also occurs under diverse abiotic stress conditions in darkness. Light-driven photochemistry usually leads to the pH changes that activate de-epoxidases (e.g. violaxanthin de-epoxidase), but in darkness alternative electron transport pathways and luminal domains enriched in monogalactosyl diacyl glycerol (which enhance de-epoxidase activity) likely enable de-epoxidation. Another 'dark side' to sustaining xanthophyll de-epoxidation is inactivation and/or degradation of epoxidases (e.g. zeaxanthin epoxidase). There are obvious benefits of such activity regarding stress tolerance, and indeed this phenomenon has only been reported in stressful conditions. However, more research is required to unravel the mechanisms and understand the physiological roles of dark-induced formation of zeaxanthin. Notably, the de-epoxidation of violaxanthin to antheraxanthin and zeaxanthin in darkness is still a frequently ignored process, perhaps because it questions a previous paradigm. With that in mind, this review seeks to shed some light on the dark side of xanthophyll de-epoxidation, and point out areas for future work.

Field-grown grapevines are often exposed to multiple environmental stresses, which challenges wine-growers to develop sustainable measures to sustain vine growth, yield, and quality. Under field conditions this task is demanding, due to differences in the magnitudes of stresses and associated plant responses. In this study we explored the hypothesis that kaolin-particle film application improves grapevine photoprotection through the regulation of xanthophyll cycle genes, limiting the thermal dissipation of excess energy under harsh environmental conditions. Hence, we selected two grapevine varieties, Touriga-Nacional (TN) and Touriga-Franca (TF), grown in the Douro Demarcated Region, and evaluated changes in light dissipation mechanisms, xanthophyll cycle components, and the expression of xanthophyll cycle genes during the 2017 summer season. The results showed that, from veraison to ripening, kaolin triggered the up-regulation of violaxanthin de-epoxidase (VvVDE1) and zeaxanthin epoxidase (VvZEP1) genes, indicating optimised regulation of the xanthophyll cycle. Kaolin treatment also decreased chlorophyll (Chla, Chlb, Chl(a+b)) and carotenoid (Car) accumulation under increasing summer stress conditions in both varieties and lowered the non-photochemical quenching (NPQ) of grapevines on ripening, suggesting a long-term response to summer stress. In addition, kaolin-treated grapevines showed increased Chla/Chlb and lower Chl(a+b)/Car ratios, displaying some features of high light adapted leaves. Overall, this study suggests that kaolin application enabled grapevines to benefit from fluctuating periods of summer stress by managing chlorophyll and carotenoid content and limiting down-regulation of both photochemistry and photoinhibition processes. Under Mediterranean field conditions, kaolin application can be considered an efficient method of minimising summer stress impact on grapevines., This work was supported by National Funds by FCT - Portuguese Foundation for Science and Technology, under the project UIDB/04033/2020, the Clim4Vitis project – “Climate change impact mitigation for European viticulture: knowledge transfer for an integrated approach”, funded by the European Union's Horizon 2020 Research and Innovation programme, under grant agreement no. 810176, and by the project I&D&I AgriFood XXI, operation NORTE-01-0145-FEDER-000041, co-funded by the European Regional Development Fund (FEDER) through NORTE 2020 (Programa Operacional Regional do Norte, 2014/2020). Sara Bernardo acknowledges the financial support provided by the FCT-Portuguese Foundation for Science and Technology (PD/BD/128273/2017) under the Doctoral Programme “Agricultural Production Chains – from Fork to Farm”.

Chickpea is sensitive to heat stress and generally experiences high temperature during reproductive phase. Therefore, terminal heat stress is key constraints for its production. Exogenous application of bioregulators is an option to improve heat tolerance and yield. Therefore, an experiment was carried out using two contrasting Desi type (JG 14) and Kabuli type (Pusa 1108) chickpea varieties with an objective to interlink bioregulators induced heat tolerance with zeaxanthin pigments cycle. Experiment was comprising two main treatments i.e. Normal (18th November) and late (20th December) sown conditions for exposing the crop to high temperature and four sub-treatments (Foliar application of bioregulators viz ABA (10 ppm), BA (40 ppm), SA (100 ppm), thiourea (1000 ppm) and water spray (as control) prior to flowering. Under high temperature condition, reduction in leaf photosynthesis, photosynthetic pigments, chlorophyll fluorescence parameters, yield and its attributes was recorded. Foliar application of bioregulators augmented the grain yield by improving the photosynthetic rate (PN), photosynthetic pigments including zeaxanthin, maximum quantum yield of PS II (Fv/Fm), non-photochemical quenching, heat tolerance index, yield attributes and reducing the level of violaxanthin and heat susceptibility index over the control (water) under late sown condition which in turn suggested that bioregulators induced the heat tolerance by activating operation of zeaxanthin pigments cycle in chickpea. Moreover, Kabuli chickpea variety responded better to bioregulators than Desi type particularly under heat stress condition.

Marigold (Tagetes erecta L.) is an important ornamental plant with a wide variety of flower colors. Despite its economic value, few biochemical and molecular studies have explored the generation of flower color in this species. To study the mechanism underlying marigold petal color, we performed a metabolite analysis and de novo cDNA sequencing on the inbred line ‘V-01’ and its petal color mutant ‘V-01M’ at four flower developmental stages. A total of 49,217 unigenes were identified from 24 cDNA libraries. Based on our metabolites and transcriptomic analyses, we present an overview of carotenoid biosynthesis, degradation, and accumulation in marigold flowers. The carotenoid content of the yellow mutant ‘V-01M’ was higher than that of the orange inbred line ‘V-01’, and the abundances of the yellow compounds lutein, neoxanthin, violaxanthin, zeaxanthin, and antheraxanthin were significantly higher in the mutant. During flower development, the carotenoid biosynthesis genes were upregulated in both ‘V-01’ and ‘V-01M’, with no significant differences between the two lines. By contrast, the carotenoid degradation genes were dramatically downregulated in the yellow mutant ‘V-01M’. We therefore speculate that the carotenoid degradation genes are the key factors regulating the carotenoid content of marigold flowers. Our research provides a large amount of transcriptomic data and insights into the marigold color metabolome.

Chickpea being a winter season crop often experiences heat stress during reproductive phase. For chickpea production, terminal heat stress is one of the major constraints. Plants have built up numerous mechanisms to combat the heat stress. We considered the photosynthetic pigments for heat tolerance. Therefore, in order to investigate the heat tolerance in relation to photosynthetic pigments, a field trial was carried out having 4 contrasting genotypes namely BG 240 and JG 14 (relatively heat tolerant), SBD 377 (moderately tolerant) and ICC 1882 (relatively heat sensitive). Heat stress was imposed by altering the sowing date i.e. normal (18th November) and late sown (18th December). Under delayed sown condition, heat stress was faced by crop starting from flowering stage to crop maturity. Under heat stress condition, heat tolerant genotypes BG 240 and JG 14 maintained higher level of membrane stability, RWC (%), osmolytes, dry matter partitioning, grain yield, heat tolerance index and had higher values of zeaxanthin, quantum yield of PS II (Fv/Fm ratio), non-photochemical quenching (NPQ), photosynthetic rate, level of photosynthetic pigments (chlorophylls and carotenoids) and lower level of violaxanthin, and lipid peroxidation as compared to heat sensitive one (ICC 1882). In addition to this, Fv/Fm ratio and NPQ exhibited positive relationship with heat tolerance which suggested the involvement of xanthophyll cycle pigments in chickpea heat tolerance.

The accumulation of carotenoids (carotenogenesis) in the cells of Haematococcus pluvialis (strain IBCE H-17) upon cultivation in nutrient media deficient in nutrient elements (N and K + P) is shown. K and P deficiency in nutrient medium led to accumulation of astaxanthin, β-carotene, anteraxanthin, zeaxanthin and violaxanthin after 18 days of cultivation, while the amount of neoxanthin and lutein was at the control level. Carotenogenesis under such conditions was accompanied by high levels of chlorophyll (a + b). The use of nutrient medium deficient in N led to the accumulation of astaxanthin and, to a lesser extent, β-carotene in haematococcus cells, while lutein, anteraxanthin, zeaxanthin, violaxanthin and neoxanthin levels were lower or the same as control. N deficiency caused the decrease in amount of chlorophyll (a + b). Nitrogen deficiency was 4.3 times more effective than control medium and 1.8 times more effective than combined potassium and phosphorus deficiency in promoting astaxanthin accumulation.

Microbial production of carotenoids has mainly focused towards a few products, such as β-carotene, lycopene and astaxanthin. However, other less explored carotenoids, like violaxanthin, have also shown unique properties and promissory applications. Violaxanthin is a plant-derived epoxidated carotenoid with strong antioxidant activity and a key precursor of valuable compounds, such as fucoxanthin and β-damascenone. In this study, we report for the first time the heterologous production of epoxycarotenoids in yeast. We engineered the yeast Saccharomyces cerevisiae following multi-level strategies for the efficient accumulation of violaxanthin. Starting from a β-carotenogenic yeast strain, we first evaluated the performance of several β-carotene hydroxylases (CrtZ), and zeaxanthin epoxidases (ZEP) from different species, together with their respective N-terminal truncated variants. The combined expression of CrtZ from Pantoea ananatis and truncated ZEP of Haematococcus lacustris showed the best performance and led to a yield of 1.6 mg/gDCW of violaxanthin. Further improvement of the epoxidase activity was achieved by promoting the transfer of reducing equivalents to ZEP by expressing several redox partner systems. The co-expression of the plant truncated ferredoxin-3, and truncated root ferredoxin oxidoreductase-1 resulted in a 2.2-fold increase in violaxanthin yield (3.2 mg/gDCW). Finally, increasing gene copy number of carotenogenic genes enabled reaching a final production of 7.3 mg/gDCW in shake flask cultures and batch bioreactors, which is the highest yield of microbially produced violaxanthin reported to date.

The violaxanthin (V)-antheraxanthin (A)-zeaxanthin (Z) (VAZ) cycle was deemed a non-second-scale process of photoprotection in higher plants and microalgae, but the validity of this view has not been confirmed. To test this view, we explored responses of the VAZ cycle and the relationship between the VAZ cycle and non-photochemical quenching (NPQ) under highlight at second and minute scales in Heterosigma akashiwo and Platymonas sp. Both A and Z were generated in H. akashiwo during 15 s of light exposure, whereas only A rapidly accumulated within 15 s of exposure in Platymonas sp. The above results, together with a time-dependent sigmoidal relationship between the VAZ cycle (de-epoxidation state, A/Chl a, and Z/Chl a) and NPQ, proved that the VAZ cycle was a second-scale process related to NPQ. In addition, we found that not all NPQ was dependent on the VAZ cycle and suggested that NPQ model should be carefully modified due to the species-specific proportions of de-epoxidation-dependent NPQ.

Recombinant Escherichia coli cells, which express the higher-plant CaZEP (zeaxanthin epoxidase) and AtCYO1 (molecular chaperone) genes and an NADPH regeneration system in addition to the IDI, crtE, crtB, crtI, crtY, and crtZ genes for zeaxanthin biosynthesis, were cultured in 2 × YT medium for the efficient production of antheraxanthin and violaxanthin. The carotenoids accumulated in the cells were purified by two-phase partition, silica gel column chromatography, and preparative silica gel HPLC. Mass spectrometry and nuclear magnetic resonance analyses of these carotenoids revealed that a novel carotenoid, determined to be (3R,6′S)-3,6′-dihydroxy-β,e-caroten-3′-one and named as 6′-hydroxy-3′-didehydrolutein, was produced in addition to zeaxanthin, antheraxanthin and violaxanthin. It is likely that the new carotenoid was generated from an unstable epoxy-carotenoid antheraxanthin that was synthesized from zeaxanthin with CaZEP. Its antioxidant activities were also examined.

Heatwaves are becoming more frequent with climate warming and can impact tree growth and reproduction. Eucalyptus parramattensis can cope with an extreme heatwave in the field via transpiratory cooling and enhanced leaf thermal tolerance that protected foliar tissues from photo-inhibition and photo-oxidation during natural midday irradiance. Here, we explored whether changes in foliar carotenoids and/or the xanthophyll cycle state can facilitate leaf acclimation to long-term warming and/or an extreme heatwave event. We found that leaves had similar carotenoid levels when grown for one year under ambient and experimental long-term warming (+ 3 °C) conditions in whole tree chambers. Exposure to a 4-day heatwave (> 43 °C) significantly altered the xanthophyll de-epoxidation state of carotenoids revealing one mechanism by which trees could minimise foliar photo-oxidative damage. The levels of zeaxanthin were significantly higher in both young and old leaves during the heatwave, revealing that violaxanthin de-epoxidation and perhaps de novo zeaxanthin synthesis contributed to enhancement of the xanthophyll cycle state. In a future climate of long-term warming and increased heatwave events, leaves of E. parramattensis will be able to utilise biochemical strategies to alter the xanthophyll cycle state and cope with extreme temperatures under natural solar irradiation.

Carotenoids are a class of natural product compound that are currently being used as colouring agents and widely used in food industry. Carotenoids are bioactive pigments obtained mainly from plants through dietary intake. They possess good features in terms of dietary supplement, food colourant, and polymer stabiliser. The presence of 4 main carotenoids, which are β-carotene, zeaxanthin, lutein, and violaxanthin, were determined in 4 classes of allelopathic plant groups namely trees, ferns, grasses and herbaceous plants. This research aims to explore the carotenoid’s content and composition in 11 allelopathic species by HPLC analysis. A. auriculiformis (tree) was found to have the highest total carotenoid concentration (146.36 µg/g DW) that was substantially higher than all other species tested whereas the lowest total carotenoid concentration was found in S. palustris (fern) (3.76 µg/g DW). Lutein and β-carotene were detected highest in A. auriculiformis (tree), with 1024 ± 25.5 µg/g DW and 37.55 ± 3.16 µg/g DW, respectively. Violaxanthin and zeaxanthin were found substantially highest in M. cajuputi (tree) (5.02 ± 0.5 µg/g DW) and S. palustris (fern) (5.88 ± 0.19µg/g DW), respectively.

Kaolin-particle film has been considered a low-cost technology to mitigate the adverse effects of high light and temperature, and drought in several crops. However, the underlying excess energy absorption and dissipation mechanisms, and related components associated with kaolin photoprotective effects in grapevines are poorly explored. This study aims to understand the interactions between kaolin foliar treatment and photosynthetic pigments accumulation, carotenoids metabolism, xanthophyll cycle regulation, and its putative role on the non-photochemical quenching (NPQ) processes in Touriga-Franca (TF) and Touriga-Nacional (TN) varieties. The experiments were conducted during the 2017 summer season in a commercial vineyard, and measurements were performed at pre-dawn and midday in each sampling date (EL35 – veraison; EL38 – full mature). Overall, TF variety showed higher accumulation of chlorophylls, xanthophylls, and de-epoxidation state (DPS) than TN. Kaolin treatment enhanced TN chlorophyll accumulation up to 114 % at EL35 (veraison) and 123 % at EL38 (full mature), highlighting its protective role on chlorophyll degradation, while no changes were found in TF, which might indicate a lower need for particle-film technology in this variety under the current environmental conditions. Individual carotenoids were mainly higher in the treated leaves of both varieties, as well as the xanthophyll cycle pigments zeaxanthin (Zx) and violaxanthin (Vx). Simultaneously, the DPS and NPQ values were lower in TN and TF treated leaves (1.92 – 2.36) compared to untreated vines (3.19 – 3.24), suggesting that there might be other components influencing NPQ levels beyond Zx, with an indirect role in long-lasting NPQ processes. In addition, in the TF kaolin-treated leaves, violaxanthin de-epoxidase (VvVDE1) and zeaxanthin epoxidase (VvZEP1) gene expression were respectively 3-fold and 4-fold upregulated at stage EL35, while VvZEP1 gene expression decreased at stage EL38 in TN kaolin-treated leaves, indicating an optimised regulation of the xanthophyll cycle. These findings suggest that kaolin treatment promoted a fine-tuning of grapevine summer stress responses under sustained summer stress factors, by managing xanthophyll cycle dynamics, and pigments accumulation., This work was supported by National Funds by FCT - Portuguese Foundation for Science and Technology, under the project UIDB/04033/2020, the Clim4Vitis project – "Climate change impact mitigation for European viticulture: knowledge transfer for an integrated approach", funded by European Union's Horizon 2020 research and innovation programme, under grant agreement no. 810176. Authors acknowledge Rui Flores from “Herdade do Esporão”, for the collaboration and efforts in making the vineyard facilities available for this study, as well as BASF collaboration, namely Paulo Matos. The technical assistance of Inmaculada Carbonell (IATA-CSIC) in pigment analysis is also gratefully acknowledged. Sara Bernardo acknowledges the financial support provided by the FCT-Portuguese Foundation for Science and Technology (PD/BD/ 128273/2017), under the Doctoral Programme "Agricultural Production Chains – from fork to farm", and Dinis L.-T. thanks the FCT and UTAD for the research contract (D.L. Law no. 57/2017).

Carotenoids are naturally synthesized in some species of bacteria, archaea, and fungi (including yeasts) as well as all photosynthetic organisms. Escherichia coli has been the most popular bacterial host for the heterologous production of a variety of carotenoids, including even xanthophylls unique to photosynthetic eukaryotes such as lutein, antheraxanthin, and violaxanthin. However, conversion efficiency of these epoxy-xanthophylls (antheraxanthin and violaxanthin) from zeaxanthin remained substantially low. We here examined several factors affecting their productivity in E. coli. Two sorts of plasmids were introduced into the bacterial host, i.e., a plasmid to produce zeaxanthin due to the presence of the Pantoea ananatis crtE, crtB, crtI, crtY, and crtZ genes in addition to the Haematococcus pluvialis IDI gene, and one containing each of zeaxanthin epoxidase (ZEP) genes originated from nine photosynthetic eukaryotes. It was consequently found that paprika (Capsicum annuum) ZEP (CaZEP) showed the highest conversion activity. Next, using the CaZEP gene, we performed optimization experiments in relation to E. coli strains as the production hosts, expression vectors, and ribosome-binding site (RBS) sequences. As a result, the highest productivity of violaxanthin (231 μg/g dry weight) was observed, when the pUC18 vector was used with CaZEP preceded by a RBS sequence of score 5000 in strain JM101(DE3).

Plants have developed multiple self-regulatory mechanisms to efficiently function under varying sunlight conditions. At high light intensities, non-photochemical quenching (NPQ) is activated on a molecular level, safely dissipating an excess excitation as heat. The exact molecular mechanism for NPQ is still under debate, but it is widely agreed that the direct participation of the carotenoid pigments is involved, one of the proposed candidate being the zeaxanthin. In this work, we performed fluorescence measurements of violaxanthin- and zeaxanthin-enriched major light-harvesting complexes (LHCII), in ensemble and at the single pigment-protein complex level, where aggregation is prevented by immobilization of LHCIIs onto a surface. We show that a selective enrichment of LHCII with violaxanthin or zeaxanthin affects neither the ability of LHCII to switch into a dissipative conformation nor the maximal level of induced quenching. However, the kinetics of the fluorescence decrease due to aggregation on the timescale of seconds are different, prompting towards a modulatory effect of zeaxanthin in the dynamics of quenching.

We studied the effects of different radiation treatments on the physiology and UV-absorbing compounds of the model liverwort Marchantia polymorpha subsp. ruderalis. Starting from gemmae, samples were exposed to five radiation treatments: low photosynthetically active radiation (PAR), low PAR+ UV-A, low PAR + UV-B, low PAR + UV-A + UV-B, and high PAR. After 35 days, the maximum quantum yield of photosystem II was similar between treatments, which suggested comparable photoinhibition and physiological vitality, also supported by results showing an unchanged chlorophyll a/b ratio and only slight changes in growth. However, the total contents of both chlorophylls and carotenoids decreased in the UV radiation treatments and, more strongly, in the high-PAR samples, suggesting mainly PAR-dependent damage to the photosynthetic pigments. The xanthophyll index (antheraxanthin + zeaxanthin)/(violaxanthin + antheraxanthin + zeaxanthin) was only increased in the high-PAR samples, indicating an increase in photoprotection through nonphotochemical dissipation of the excess energy. The sclerophylly index (the ratio between the thallus dry mass and surface area) was increased in the UV-B-exposed samples, suggesting a UV-induced structural protection. Only the UV-B-exposed samples showed DNA damage. Several apigenin and luteolin derivatives were found in the methanol-soluble vacuolar fraction of the liverwort and p-coumaric and ferulic acids in the methanol-insoluble cell wall-bound fraction. Most individual soluble compounds, the bulk level of soluble compounds, and chalcone synthase expression increased in UV-B-exposed samples, whereas individual insoluble compounds increased in the samples exposed to only PAR. Principal components analysis summarized these responses, showing the strong influence of both UV-B and PAR levels on the physiology and UV protection of the samples.

An analytical method was validated for the quantitative determination of isoprenoids compounds in faecal samples, based on liquid-liquid extraction from a small aliquot (0.3-0.5 g of sample) and subsequent analysis by Rapid Resolution Liquid Chromatography (RRLC) on a C30 column. An excellent linear response was observed over the range specified for all dietary isoprenoids, as confirmed by the correlation coefficient, which ranged from 0.9977 to 0.9999. LODs ranged from 0.002 μg to 0.036 μg for lutein and α-tocopherol, respectively. Depending on the compound, LOQs ranged from 0.001 μg (lutein) to 0.120 μg (α-tocopherol). For accuracy testing, spiking of faeces samples with trans-β-apo-8′-carotenal, α-tocopherol and chlorophyll a were performed (three concentration levels). Excellent recoveries were obtained in all levels (>90%). The intra-day RSD% ranged from 0.86 to 9.78%. The inter-day RSD% was not higher than 10%, except to α-tocopherol (11.34%). In order to assess the applicability of the method faecal samples from a baby fed with different purees formulated from various vegetables were analysed during a six month period. α-carotene, β-carotene, capsanthin, lycopene, lutein, phytoene, phytofluene, violaxanthin, zeaxanthin and ζ-carotene), and their isomers were identified and quantified using this method. Besides, 2 tocopherols and 9 chlorophylls and derivatives were identified and quantified in the faecal samples analysed. This method is suitable to determine dietary isoprenoids from complex matrices such as human faeces within 28 min.

Yellow or green spots related to pigment changes found at the early stage of oleocellosis can cause severe economic damage. However, little information exists on pigment changes during oleocellosis development, so this study investigated the main changes in chlorophyll and carotenoid metabolites and related gene expression. Among the variations, the increased contents of chlorophyll a and b, and decreased concentrations of lutein, β-cryptoxanthin, zeaxanthin, violaxanthin, α-carotene and β-carotene were responsible for chlorophyll and carotenoid changes, respectively. Regarding gene expression, the up-regulated genes, magnesium chelatase subunit H (MgCh), magnesium-protoporphyrin IX monomethyl ester (oxidative) cyclase 1/2 (MPEC1/2), protochlorophyllide reductase a, chloroplastic 1/2 (PORA1/2) and chlorophyllide a oxygenase (CAO), regarding chlorophyll synthesis as well as the down-regulated genes, phytoene synthase (PSY), phytoene dehydrogenase (PDS), lycopene β-cyclase (LCYb), and zeaxanthin epoxidase 1/2 (ZEP 1/2) and the up-regulated genes (+)-abscisic acid 8'-hydroxylase 1/2 (ABA-HX 1/2), regarding carotenoid metabolism, constituted the major variations in oleocellosis peels.

Key message: The suppression of the HYD-1 gene by a TILLING approach increases the amount of ?-carotene in durum wheat kernel. Abstract: Vitamin A deficiency is a major public health problem that affects numerous countries in the world. As humans are not able to synthesize vitamin A, it must be daily assimilated along with other micro- and macronutrients through the diet. Durum wheat is an important crop for Mediterranean countries and provides a discrete amount of nutrients, such as carbohydrates and proteins, but it is deficient in some essential micronutrients, including provitamin A. In the present work, a targeting induced local lesions in genomes strategy has been undertaken to obtain durum wheat genotypes biofortified in provitamin A. In detail, we focused on the suppression of the ?-carotene hydroxylase 1 (HYD1) genes, encoding enzymes involved in the redirection of ?-carotene toward the synthesis of the downstream xanthophylls (neoxanthin, violaxanthin and zeaxanthin). Expression analysis of genes involved in carotenoid biosynthesis revealed a reduction of the abundance of HYD1 transcripts greater than 50% in mutant grain compared to the control. The biochemical profiling of carotenoid in the wheat mutant genotypes highlighted a significant increase of more than 70% of ?-carotene compared to the wild-type sibling lines, with no change in lutein, ?-carotene and zeaxanthin content. This study sheds new light on the molecular mechanism governing carotenoid biosynthesis in durum wheat and provides new genotypes that represent a good genetic resource for future breeding programs focused on the provitamin A biofortification through non-transgenic approaches.

Plastidial isoprenoids, such as carotenoids and tocopherols, are important anti-oxidant metabolites synthesized in plastids from precursors generated by the methylerythritol 4-phosphate (MEP) pathway. In this study, we found that irradiation of Arabidopsis thaliana plants with UV-B caused a strong increase in the accumulation of the photoprotective xanthophyll zeaxanthin but also resulted in slightly higher levels of γ-tocopherol. Plants deficient in the MEP enzymes 1-deoxy-D-xylulose 5-phosphate synthase and 1-hydroxy-2-methyl-2-butenyl 4-diphosphate synthase showed a general reduction in both carotenoids and tocopherols and this was associated with increased DNA damage and decreased photosynthesis after exposure to UV-B. Genetic blockage of tocopherol biosynthesis did not affect DNA damage accumulation. In contrast, lut2 mutants that accumulate β,β-xanthophylls showed decreased DNA damage when irradiated with UV-B. Analysis of aba2 mutants showed that UV-B protection was not mediated by ABA (a hormone derived from β,β-xanthophylls). Plants accumulating β,β-xanthophylls also showed decreased oxidative damage and increased expression of DNA-repair enzymes, suggesting that this may be a mechanism for these plants to decrease DNA damage. In addition, in vitro experiments also provided evidence that β,β-xanthophylls can directly protect against DNA damage by absorbing radiation. Together, our results suggest that xanthophyll-cycle carotenoids that protect against excess illumination may also contribute to protection against UV-B., This research was supported by Argentina FONCyT grants PICT 2016–141 and 2015–157 to PC, by Spanish grants from MINECO (BIO2015-71703-REDT and BIO2014-59092-P) and AGAUR (2014SGR-1434) to MRC, and by an EMBO Short-Term Fellowship to JE. MLFF, ER, and PC are members of the Researcher Career of the Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET). MLFF, ER, and PC are Professors of UNR; and JE and EM are postdoctoral fellows from CONICET.