Salinity poses a serious threat to global agriculture and human food security. A better understanding of plant adaptation to salt stress is, therefore, mandatory. In the non-photosynthetic cells of the root, salinity perturbs oxidative balance in mitochondria, leading to cell death. In parallel, plastids accumulate the jasmonate precursor cis (+)12-Oxo-Phyto-Dienoic Acid (OPDA) that is then translocated to peroxisomes and has been identified as promoting factor for salt-induced cell death as well. In the current study, we probed for a potential interaction between these three organelles that are primarily dealing with oxidative metabolism. We made use of two tools: (i) Rice OPDA Reductase 7 (OsOPR7), an enzyme localised in peroxisomes converting OPDA into the precursors of the stress hormone JA-Ile. (ii) A Trojan Peptoid, Plant PeptoQ, which can specifically target to mitochondria and scavenge excessive superoxide accumulating in response to salt stress. We show that overexpression of OsOPR7 as GFP fusion in tobacco (Nicotiana tabacum L. cv. Bright Yellow 2, BY-2) cells, as well as a pretreatment with Plant PeptoQ can mitigate salt stress with respect to numerous aspects including proliferation, expansion, ionic balance, redox homeostasis, and mortality. This mitigation correlates with a more robust oxidative balance, evident from a higher activity of superoxide dismutase (SOD), lower levels of superoxide and lipid peroxidation damage, and a conspicuous and specific upregulation of mitochondrial SOD transcripts. Although both, Plant PeptoQ and ectopic OsOPR7, were acting in parallel and mostly additive, there are two specific differences: (i) OsOPR7 is strictly localised to the peroxisomes, while Plant PeptoQ found in mitochondria. (ii) Plant PeptoQ activates transcripts of NAC, a factor involved in retrograde signalling from mitochondria to the nucleus, while these transcripts are suppressed significantly in the cells overexpressing OsOPR7. The fact that overexpression of a peroxisomal enzyme shifting the jasmonate pathway from the cell-death signal OPDA towards JA-Ile, a hormone linked with salt adaptation, is accompanied by more robust redox homeostasis in a different organelle, the mitochondrion, indicates that cross-talk between peroxisome and mitochondrion is a crucial factor for efficient adaptation to salt stress.

Novel unimolecular bivalent glycoconjugates were assembled combining several functionalized capsular polysaccharides of Streptococcus pneumoniae and Neisseria meningitidis to a carrier protein by using an effective strategy based on the Ugi 4-component reaction. The development of multivalent glycoconjugates opens new opportunities in the field of vaccine design, but their high structural complexity involves new analytical challenges. Nuclear Magnetic Resonance has found wide applications in the characterization and impurity profiling of carbohydrate-based vaccines. Eight bivalent conjugates were studied by quantitative NMR analyzing the structural identity, the content of each capsular polysaccharide, the ratios between polysaccharides, the polysaccharide to protein ratios and undesirable contaminants. The qNMR technique involves experiments with several modified parameters for obtaining spectra with quantifiable signals. In addition, the achieved NMR results were combined with the results of colorimetric assay and Size Exclusion HPLC for assessing the protein content and free protein percentage, respectively. The application of quantitative NMR showed to be efficient to clear up the new structural complexities while allowing the quantitative assessment of the components.

Mannan is a class of cell wall polysaccharides widespread in the plant kingdom. Mannan structure and properties vary according to species and organ. The cell walls of cereal grains have been extensively studied due to their role in cereal processing and to their beneficial effect on human health as dietary fiber. Recently, we showed that mannan in wheat (Triticum aestivum) grain endosperm has a linear structure of β-1,4-linked mannose residues. The aim of this work was to study the biosynthesis and function of wheat grain mannan. We showed that mannan is deposited in the endosperm early during grain development, and we identified candidate mannan biosynthetic genes expressed in the endosperm. The functional study in wheat was unsuccessful therefore our best candidate genes were expressed in heterologous systems. The endosperm-specificTaCslA12 gene expressed in Pichia pastoris and in an Arabidopsis thaliana mutant depleted in glucomannan led to the production of wheat-like linear mannan lacking glucose residues and with moderate acetylation. Therefore, this gene encodes a mannan synthase and is likely responsible for the synthesis of wheat endosperm mannan.

Salvadora persica L. (toothbrush tree, Miswak) is well recognized in most Middle Eastern and African countries for its potential role in dental care, albeit the underlying mechanism for its effectiveness is still not fully understood. A comparative MS and NMR metabolomics approach was employed to investigate the major primary and secondary metabolites composition of S. persica in context of its organ type viz., root or stem to rationalize for its use as a tooth brush. NMR metabolomics revealed its enrichment in nitrogenous compounds including proline-betaines i.e., 4-hydroxy-stachydrine and stachydrine reported for the first time in S. persica. LC/MS metabolomics identified flavonoids (8), benzylurea derivatives (5), butanediamides (3), phenolic acids (8) and 5 sulfur compounds, with 21 constituents reported for the first time in S. persica. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) of either NMR or LC/MS dataset clearly separated stem from root specimens based on nitrogenous compounds abundance in roots and is justifying for its preference as toothbrush versus stems. The presence of betaines at high levels in S. persica (9−12 μg/mg dry weight) offers novel insights into its functioning as an osmoprotectant that maintains the hydration of oral mucosa. Additionally, the previously described anti-inflammatory activity of stachydrine along with the antimicrobial effects of sulfonated flavonoids, benzylisothiocynate and ellagic acid derivatives are likely contributors to S. persica oral hygiene health benefits. Among root samples, variation in sugars and organic acids levels were the main discriminatory criterion. This study provides the first standardization of S. persica extract using qNMR for further inclusion in nutraceuticals.

Ceramides (CERs) play a major role in skin barrier function and direct replacement of depleted skin CERs,due to skin disorder or aging, has beneficial effects in improving skin barrier function and skin hydration.Though, plants are reliable source of CERs, absence of economical and effective method of hydrolysis toconvert the dominant plant sphingolipid, glucosylceramides (GlcCERs), into CERs remains a challenge.This study aims at exploring alternative GlcCERs sources and chemical method of hydrolysis into CERsfor dermal application. GlcCERs isolated from lupin bean (Lupinus albus), mung bean (Vigna radiate) andnaked barley (Hordium vulgare) were identified using ultra high performance liquid chromatographyhyphenated with atmospheric pressure chemical ionization - high resolution tandem mass spectrometer(UHPLC/APCI-HRMS/MS) and quantified with validated automated multiple development-high perfor-mance thin layer chromatography (AMD-HPTLC) method. Plant GlcCERs were hydrolyzed into CERs withmild acid hydrolysis (0.1 N HCl) after treating them with oxidizing agent, NaIO4,and reducing agent,NaBH4. GlcCERs with 4,8-sphingadienine, 8-sphingenine and 4-hydroxy-8-sphingenine sphingoid baseslinked with C14 to C26 -hydroxylated fatty acids (FAs) were identified. Single GlcCER (m/z 714.5520)was dominant in lupin and mung beans while five major GlcCERs species (m/z 714.5520, m/z 742.5829,m/z 770.6144, m/z 842.6719 and m/z 844.56875) were obtained from naked barley. The GlcCERs con-tents of the three plants were comparable. However, lupin bean contains predominantly (> 98 %) a singleGlcCER (m/z 714.5520). Considering the affordability, GlcCER content and yield, lupin bean would bethe preferred alternative commercial source of GlcCERs. CER species bearing 4,8-sphingadienine and 8-sphingenine sphingoid bases attached to C14 to 24 FAs were found after mild acid hydrolysis. CER specieswith m/z 552.4992 was the main component in the beans while CER with m/z 608.5613 was dominantin the naked barley. However, CERs with 4-hydroxy-8-sphingenine sphingoid base were not detected inUHPLC-HRMS/MS study suggesting that the method works for mainly GlcCERs carrying dihydroxy sph-ingoid bases. The method is economical and effective which potentiates the commercialization of plantCERs for dermal application.

Knipholone (1) and knipholone anthrone (2), isolated from the Ethiopian medicinal plant Kniphofia foliosa Hochst. are two phenyl anthraquinone derivatives, a compound class known for biological activity. In the present study, we describe the activity of both 1 and 2 in several biological assays including cytotoxicity against four human cell lines (Jurkat, HEK293, SH-SY5Y and HT-29), antiplasmodial activity against Plasmodium falciparum 3D7 strain, anthelmintic activity against the model organism Caenorhabditis elegans, antibacterial activity against Aliivibrio fischeri and Mycobacterium tuberculosis and anti-HIV-1 activity in peripheral blood mononuclear cells (PBMCs) infected with HIV-1c. In parallel, we investigated the stability of knipholone (2) in solution and in culture media. Compound 1 displays strong cytotoxicity against Jurkat, HEK293 and SH-SY5Y cells with growth inhibition ranging from approximately 62–95% when added to cells at 50 μM, whereas KA (2) exhibits weak to strong activity with 26, 48 and 70% inhibition of cell growth, respectively. Both 1 and 2 possess significant antiplasmodial activity against Plasmodium falciparum 3D7 strain with IC50 values of 1.9 and 0.7 μM, respectively. These results complement previously reported data on the cytotoxicity and antiplasmodial activity of 1 and 2. Furthermore, compound 2 showed HIV-1c replication inhibition (growth inhibition higher than 60% at tested concentrations 0.5, 5, 15 and 50 μg/ml and an EC50 value of 4.3 μM) associated with cytotoxicity against uninfected PBMCs. The stability study based on preincubation, HPLC and APCI-MS (atmospheric-pressure chemical ionization mass spectrometry) analysis indicates that compound 2 is unstable in culture media and readily oxidizes to form compound 1. Therefore, the biological activity attributed to 2 might be influenced by its degradation products in media including 1 and other possible dimers. Hence, bioactivity results previously reported from this compound should be taken with caution and checked if they differ from those of its degradation products. To the best of our knowledge, this is the first report on the anti-HIV activity and stability analysis of compound 2.

The contribution of carbon assimilation and allocation and of invertases to the stimulation of adventitious root formation in response to a dark pre-exposure of petunia cuttings was investigated, considering the rooting zone (stem base) and the shoot apex as competing sinks. Dark exposure had no effect on photosynthesis and dark respiration during the subsequent light period, but promoted dry matter partitioning to the roots. Under darkness, higher activities of cytosolic and vacuolar invertases were maintained in both tissues when compared to cuttings under light. This was partially associated with higher RNA levels of respective genes. However, activity of cell wall invertases and transcript levels of one cell wall invertase isogene increased specifically in the stem base during the first two days after cutting excision under both light and darkness. During five days after excision, RNA accumulation of four invertase genes indicated preferential expression in the stem base compared to the apex. Darkness shifted the balance of expression of one cytosolic and two vacuolar invertase genes towards the stem base. The results indicate that dark exposure before planting enhances the carbon sink competitiveness of the rooting zone and that expression and activity of invertases contribute to the shift in carbon allocation.

Various cleavage products of C40 carotenoid substrates are formed preferentially or exclusively in roots. Such apocarotenoid signaling or regulatory compounds differentially induced in roots during environmental stress responses including root colonization by arbuscular mycorrhizal fungi include ABA, strigolactones and C13 α-ionol/C14 mycorradicin derivatives. The low carotenoid levels in roots raise the question of whether there is a regulated precursor supply channeled into apocarotenoid formation distinct from default carotenoid pathways. This review describes root-specific isogene components of carotenoid pathways toward apocarotenoid formation, highlighting a new PSY3 class of phytoene synthase genes in dicots. It is clearly distinct from the monocot PSY3 class co-regulated with ABA formation. At least two members of the exclusive dicot PSY3s are regulated by nutrient stress and mycorrhization. This newly recognized dicot PSY3 (dPSY3 vs. mPSY3 from monocots) class probably represents an ancestral branch in the evolution of the plant phytoene synthase family. The evolutionary history of PSY genes is compared with the evolution of MEP pathway isogenes encoding 1-deoxy-d-xylulose 5-phosphate synthases (DXS), particularly DXS2, which is co-regulated with dPSY3s in mycorrhizal roots. Such stress-inducible isoforms for rate-limiting steps in root carotenogenesis might be components of multi-enzyme complexes committed to apocarotenoid rather than to carotenoid formation.

Trigonelline (3-carboxy-1-methyl pyridinium) was identified as a relevant bioactivity and taste imparting component in Balanites aegyptiaca fruit, using 1H NMR of crude extracts without any fractionation or isolation step. The structural integrity of trigonelline was established within the extract matrix via1H NMR, 1H–1H COSY, HMQC and HMBC and by comparison with authentic standard. A quantitative 1H NMR method (qHNMR) was used to determine trigonelline concentrations in the peel and pulp of B. aegyptiaca fruit of 8 and 13 mg g−1, respectively. Trigonelline so far has not been reported from B. aegyptiaca or its genus as it easily escapes LC–MS based detection. Its discovery provides novel insight into the balanite fruits antidiabetic properties as the compound is known for a pronounced hypoglycemic effect. In addition, it is likely to impart the perceptible bitter taste portion to balanites sweet bitter taste. UPLC–MS of the crude extract additionally revealed the fruit flavonoid pattern showing quercetin/isorhamnetin flavonol conjugates in addition to epicatechin, the latter being present at much lower levels.

Apocarotenoids are a class of compounds that play important roles in nature. In recent years, a prominent role for these compounds in arbuscular mycorrhizal (AM) symbiosis has been shown. They are derived from carotenoids by the action of the carotenoid cleavage dioxygenase (CCD) enzyme family. In the present study, using tomato as a model, the spatio-temporal expression pattern of the CCD genes during AM symbiosis establishment and functioning was investigated. In addition, the levels of the apocarotenoids strigolactones (SLs), C13 α-ionol and C14 mycorradicin (C13/C14) derivatives were analyzed. The results suggest an increase in SLs promoted by the presence of the AM fungus at the early stages of the interaction, which correlated with an induction of the SL biosynthesis gene SlCCD7. At later stages, induction of SlCCD7 and SlCCD1 expression in arbusculated cells promoted the production of C13/C14 apocarotenoid derivatives. We show here that the biosynthesis of apocarotenoids during AM symbiosis is finely regulated throughout the entire process at the gene expression level, and that CCD7 constitutes a key player in this regulation. Once the symbiosis is established, apocarotenoid flux would be turned towards the production of C13/C14 derivatives, thus reducing SL biosynthesis and maintaining a functional symbiosis.