Germination Increases the Glucomoringin Content in Moringa Sprouts via Transforming Tyrosine.

Moringa seeds are an excellent dietary source of phytochemicals (i.e., glucosinolates, GSLs; isothiocyanates, ITCs) with health-beneficial effects. Although numerous studies have been conducted on moringa seeds, the effect of germination on the regulation of GSLs remains scarcely explored. The present study investigated the dynamic changes of GSLs in moringa seeds during germination (at 25, 30, and 35 °C for 6 days in the dark) through an untargeted metabolomics approach and compared the antioxidant capacity of ungerminated and germinated moringa seeds. Our results showed that germination significantly increased the total GSL content from 150 (day 0) to 323 µmol/g (35 °C, day 6) on a dry weight (DW) basis, especially glucomoringin (GMG), the unique glucosinolate in moringa seeds, which was significantly upregulated from 61 (day 0) to 149 µmol/g DW (35 °C, day 4). The upregulation of GMG corresponded to the metabolism of tyrosine, which might be the initial precursor for the formation of GMG. In addition, germination enhanced the total ITC content from 85 (day 0) to 239 µmol SE/g DW (35 °C, day 6), indicating that germination may have also increased the activity of myrosinase. Furthermore, germination remarkably increased the total phenolic content (109-507 mg GAE/100 g DW) and antioxidant capacity of moringa seeds. Our findings suggest that moringa sprouts could be promoted as a novel food and/or ingredient rich in GMG.

Natural H2S-donors: A new pharmacological opportunity for the management of overweight and obesity.

The prevalence of overweight and obesity has progressively increased in the last few years, becoming a real threat to healthcare systems. To date, the clinical management of body weight gain is an unmet medical need, as there are few approved anti-obesity drugs and most require an extensive monitoring and vigilance due to risk of adverse effects and poor patient adherence/persistence. Growing evidence has shown that the gasotransmitter hydrogen sulfide (H2S) and, therefore, H2S-donors could have a central role in the prevention and treatment of overweight/obesity. The main natural sources of H2S-donors are plants from the Alliaceae (garlic and onion), Brassicaceae (e.g., broccoli, cabbage, and wasabi), and Moringaceae botanical families. In particular, polysulfides and isothiocyanates, which slowly release H2S, derive from the hydrolysis of alliin from Alliaceae and glucosinolates from Brassicaceae/Moringaceae, respectively. In this review, we describe the emerging role of endogenous H2S in regulating adipose tissue function and the potential efficacy of natural H2S-donors in animal models of overweight/obesity, with a final focus on the preliminary results from clinical trials. We conclude that organosulfur-containing plants and their extracts could be used before or in combination with conventional anti-obesity agents to improve treatment efficacy and reduce inflammation in obesogenic conditions. However, further high-quality studies are needed to firmly establish their clinical efficacy.

Broccoli, Kale, and Radish Sprouts: Key Phytochemical Constituents and DPPH Free Radical Scavenging Activity.

Our research group previously found that broccoli sprouts possess neuroprotective effects during pregnancy. The active compound has been identified as sulforaphane (SFA), obtained from glucosinolate and glucoraphanin, which are also present in other crucifers, including kale. Sulforaphene (SFE), obtained from glucoraphenin in radish, also has numerous biological benefits, some of which supersede those of sulforaphane. It is likely that other components, such as phenolics, contribute to the biological activity of cruciferous vegetables. Notwithstanding their beneficial phytochemicals, crucifers are known to contain erucic acid, an antinutritional fatty acid. The aim of this research was to phytochemically examine broccoli, kale, and radish sprouts to determine good sources of SFA and SFE to inform future studies of the neuroprotective activity of cruciferous sprouts on the fetal brain, as well as product development. Three broccoli: Johnny's Sprouting Broccoli (JSB), Gypsy F1 (GYP), and Mumm's Sprouting Broccoli (MUM), one kale: Johnny's Toscano Kale (JTK), and three radish cultivars: Black Spanish Round (BSR), Miyashige (MIY), and Nero Tunda (NT), were analyzed. We first quantified the glucosinolate, isothiocyanate, phenolics, and DPPH free radical scavenging activity (AOC) of one-day-old dark- and light-grown sprouts by HPLC. Radish cultivars generally had the highest glucosinolate and isothiocyanate contents, and kale had higher glucoraphanin and significantly higher sulforaphane content than the broccoli cultivars. Lighting conditions did not significantly affect the phytochemistry of the one-day-old sprouts. Based on phytochemistry and economic factors, JSB, JTK, and BSR were chosen for further sprouting for three, five, and seven days and subsequently analyzed. The three-day-old JTK and radish cultivars were identified to be the best sources of SFA and SFE, respectively, both yielding the highest levels of the respective compound while retaining high levels of phenolics and AOC and markedly lower erucic acid levels compared to one-day-old sprouts.

Phytoalexins of the crucifer Barbarea vulgaris: Structural profile and correlation with glucosinolate turnover.

Phytoalexins are antimicrobial plant metabolites elicited by microbial attack or abiotic stress. We investigated phytoalexin profiles after foliar abiotic elicitation in the crucifer Barbarea vulgaris and interactions with the glucosinolate-myrosinase system. The treatment for abiotic elicitation was a foliar spray with CuCl2 solution, a usual eliciting agent, and three independent experiments were carried out. Two genotypes of B. vulgaris (G-type and P-type) accumulated the same three major phytoalexins in rosette leaves after treatment: phenyl-containing nasturlexin D and indole-containing cyclonasturlexin and cyclobrassinin. Phytoalexin levels were investigated daily by UHPLC-QToF MS and tended to differ among plant types and individual phytoalexins. In roots, phytoalexins were low or not detected. In treated leaves, typical total phytoalexin levels were in the range 1-10 nmol/g fresh wt. during three days after treatment while typical total glucosinolate (GSL) levels were three orders of magnitude higher. Levels of some minor GSLs responded to the treatment: phenethylGSL (PE) and 4-substituted indole GSLs. Levels of PE, a suggested nasturlexin D precursor, were lower in treated plants than controls. Another suggested precursor GSL, 3-hydroxyPE, was not detected, suggesting PE hydrolysis to be a key biosynthetic step. Levels of 4-substituted indole GSLs differed markedly between treated and control plants in most experiments, but not in a consistent way. The dominant GSLs, glucobarbarins, are not believed to be phytoalexin precursors. We observed statistically significant linear correlations between total major phytoalexins and the glucobarbarin products barbarin and resedine, suggesting that GSL turnover for phytoalexin biosynthesis was unspecific. In contrast, we did not find correlations between total major phytoalexins and raphanusamic acid or total glucobarbarins and barbarin. In conclusion, two groups of phytoalexins were detected in B. vulgaris, apparently derived from the GSLs PE and indol-3-ylmethylGSL. Phytoalexin biosynthesis was accompanied by depletion of the precursor PE and by turnover of major non-precursor GSLs to resedine. This work paves the way for identifying and characterizing genes and enzymes in the biosyntheses of phytoalexins and resedine.

Synthesis and characterization of an artificial glucosinolate bearing a chlorthalonil-based aglycon as a potent inhibitor of glucosinolate transporters.

Glucosinolates (GSLs) are specialized metabolites in plants of the order Brassicales. GSL transporters (GTRs) are essential for the redistribution of GSLs and also play a role in controlling the GSL content of seeds. However, specific inhibitors of these transporters have not been reported. In the current study, we described the design and synthesis of 2,3,4,6-tetrachloro-5-cyanophenyl GSL (TCPG), an artificial GSL bearing a chlorothalonil moiety as a potent inhibitor of GTRs, and evaluated its inhibitory effect on the substrate uptake mediated through GTR1 and GTR2. Molecular docking showed that the position of the ß-D-glucose group of TCPG was significantly different from that of the natural substrate in GTRs and the chlorothalonil moiety forms halogen bonds with GTRs. Functional assays and kinetic analysis of the transport activity revealed that TCPG could significantly inhibit the transport activity of GTR1 and GTR2 (IC50 values (mean ± SD) being 79 ± 16 µM and 192 ± 14 µM, respectively). Similarly, TCPG could inhibit the uptake and phloem transport of exogenous sinigrin by Arabidopsis thaliana (L.) Heynh leaf tissues, while not affecting that of esculin (a fluorescent surrogate for sucrose). TCPG could also reduce the content of endogenous GSLs in phloem exudates. Together, TCPG was discovered as an undescribed inhibitor of the uptake and phloem transport of GSLs, which brings novel insights into the ligand recognition of GTRs and provides a new strategy to control the GSL level. Further tests on the ecotoxicological and environmental safety of TCPG are needed before using it as an agricultural or horticultural chemical in the future.

Glucosinolates in Wild-Growing Reseda spp. from Croatia.

Glucosinolates (GSLs) are a unique class of thioglucosides that evolved as defense mechanisms in the 16 families of the Brassicales order and present molecular tags which can be placed in a robust phylogenetic framework through investigations into their evolution and diversity. The GSL profiles of three Resedaceae species, Reseda alba, R. lutea, and R. phyteuma, were examined qualitatively and quantitatively with respect to their desulfo-counterparts utilizing UHPLC-DAD-MS/MS. In addition, NMR analysis of isolated 2-hydroxy-2-methylpropyl desulfoGSL (d31) was performed. Three Phe-derived GSLs were found in R. lutea, including glucotropaeolin (11) (0.6-106.69 mol g-1 DW), 2-(α-L-ramnopyranosyloxy)benzyl GSL (109) (8.10-57.89 µmol g-1 DW), glucolepigramin (22) (8.66 µmol g-1 DW in flower), and Trp-derived glucobrassicin (43) (0.76-5.92 µmol g-1 DW). The Phe-derived GSLs 109 (50.79-164.37 µmol g-1 DW), gluconasturtiin (105) (1.97 µmol g-1 DW), and 11 (tr), as well as the Trp-derived GSL glucobrassicin (43) (3.13-11.26 µmol g-1 DW), were all present in R. phyteuma. R. alba also contained Phe-derived 105 (0.10-107.77 µmol g-1 DW), followed by Trp-derived 43 (0.85-3.50 µmol g-1 DW) and neoglucobrassicin (47) (0.23-2.74 µmol g-1 DW). However, regarding the GSLs in R. alba, which originated from Leu biosynthesis, 31 was the major GSL (6.48 to 52.72 µmol g-1 DW) and isobutyl GSL (62) was the minor GSL (0.13 to 1.13 µmol g-1 DW). The discovered Reseda profiles, along with new evidence provided by GSL characterizations, were studied in the context of the current knowledge on GLSs in the Resedaceae family. With the exception of R. alba, the aliphatic GSLs of which were outliers among the Resedaceae species studied, this family typically contains GSLs derived primarily from Trp and Phe biosynthesis, which modifications resulted in GSLs unique to this family, implying presence of the specific genes. responsible for this diversification.

Metabolomic approach of azole fungicides in radish (Raphanus sativus): Perspective of functional metabolites.

Azole fungicides is one of the major fungicides in agricultural field. In this study, toxic effects of diniconazole (DIN), metconazole (MET), and tebuconazole (TEB) to radish leaves and roots were investigated using targeted metabolomics with gas chromatography-mass spectrometry (GC-MS/MS). Especially, the changes of functional chemicals, including phytosterols and glucosinolates evaluated. Radish leaves and roots were harvested after 7 days and 14 days from last exposure. In multivariate analysis, the experimental groups showed clear separation in PCA and PLS-DA score plots. Phytosterols and glucosinolates were significantly changed by azole fungicide. Six metabolic pathways which are affected by fungicides were selected and showed similar patterns regardless of the type of azole fungicide used. As a result, azole fungicide induces the defense mechanisms of plants and affects both primary and secondary metabolism.

The mysterious non-arbuscular mycorrhizal status of Brassicaceae species.

The Brassicaceae family is unique in not fostering functional symbiosis with arbuscular mycorrhiza (AM). The family is also special in possessing glucosinolates, a class of secondary metabolites predominantly functioning for plant defence. We have reviewed what effect the glucosinolates of this non-symbiotic host have on AM or vice versa. Isothiocyanates, the toxic degradation product of the glucosinolates, particularly the indolic and benzenic glucosinolates, are known to be involved in the inhibition of AM. Interestingly, AM colonization enhances glucosinolate production in two AM-host in the Brassicales family- Moringa oleifera and Tropaeolum spp. PHOSPHATE STARVATION RESPONSE 1 (PHR1), a central transcription factor that controls phosphate starvation response also activates the glucosinolate biosynthesis in AM non-host Arabidopsis thaliana. Recently, the advances in whole-genome sequencing, enabling extensive ecological microbiome studies have helped unravel the Brassicaceae microbiome, identifying new mutualists that compensate for the loss of AM symbiosis, and reporting cues for some influence of glucosinolates on the microbiome structure. We advocate that glucosinolate is an important candidate in determining the mycorrhizal status of Brassicaceae and has played a major role in its symbiosis-defence trade-off. We also identify key open questions in this area that remain to be addressed in the future.

The Impact of Nitrile-Specifier Proteins on Indolic Carbinol and Nitrile Formation in Homogenates of Arabidopsis thaliana.

Glucosinolates, specialized metabolites of the Brassicales including Brassica crops and Arabidopsis thaliana, have attracted considerable interest as chemical defenses and health-promoting compounds. Their biological activities are mostly due to breakdown products formed upon mixing with co-occurring myrosinases and specifier proteins, which can result in multiple products with differing properties, even from a single glucosinolate. Whereas product profiles of aliphatic glucosinolates have frequently been reported, indole glucosinolate breakdown may result in complex mixtures, the analysis of which challenging. The aim of this study was to assess the breakdown of indole glucosinolates in A. thaliana root and rosette homogenates and to test the impact of nitrile-specifier proteins (NSPs) on product profiles. To develop a GC-MS-method for quantification of carbinols and nitriles derived from three prominent indole glucosinolates, we synthesized standards, established derivatization conditions, determined relative response factors and evaluated applicability of the method to plant homogenates. We show that carbinols are more dominant among the detected products in rosette than in root homogenates of wild-type and NSP1- or NSP3-deficient mutants. NSP1 is solely responsible for nitrile formation in rosette homogenates and is the major NSP for indolic nitrile formation in root homogenates, with no contribution from NSP3. These results will contribute to the understanding of the roles of NSPs in plants.

All Ion Fragmentation Analysis Enhances the Untargeted Profiling of Glucosinolates in Brassica Microgreens by Liquid Chromatography and High-Resolution Mass Spectrometry.

An analytical approach based on reversed-phase liquid chromatography coupled to electrospray ionization Fourier-transform mass spectrometry in negative ion mode (RPLC-ESI-(-)-FTMS) was developed for the untargeted characterization of glucosinolates (GSL) in the polar extracts of four Brassica microgreen crops, namely, garden cress, rapeseed, kale, and broccoli raab. Specifically, the all ion fragmentation (AIF) operation mode enabled by a quadrupole-Orbitrap mass spectrometer, i.e., the systematic fragmentation of all ions generated in the electrospray source, followed by the acquisition of an FTMS spectrum, was exploited. First, the best qualifying product ions for GSL were recognized from higher-energy collisional dissociation (HCD)-FTMS2 spectra of representative standard GSL. Extracted ion chromatograms (EIC) were subsequently obtained for those ions from RPLC-ESI(-)-AIF-FTMS data referred to microgreen extracts, by plotting the intensity of their signals as a function of retention time. The alignment of peaks detected in the EIC traces was finally exploited for the recognition of peaks potentially related to GSL, with the EIC obtained for the sulfate radical anion [SO4]•- (exact m/z 95.9523) providing the highest selectivity. Each putative GSL was subsequently characterized by HCD-FTMS2 analyses and by collisionally induced dissociation (CID) multistage MSn (n = 2, 3) acquisitions based on a linear ion trap mass spectrometer. As a result, up to 27 different GSLs were identified in the four Brassica microgreens. The general method described in this work appears as a promising approach for the study of GSL, known and novel, in plant extracts.

An LC-MS/MS assay for enzymatic characterization of methylthioalkylmalate synthase (MAMS) involved in glucosinolate biosynthesis.

Brassicaceae are blessed with specialized metabolites called glucosinolates (GSLs), which along with their degradation products, are beneficial in agriculture and human health. To date, more than 130 GSL structures have been identified, mostly derived from the amino acid methionine. The biosynthesis of methionine-derived aliphatic GSLs starts with a side-chain elongation step involving a recursive three-step cyclic process that incorporates a new methylene group into the 2-oxo acid to form a series of elongated 2-oxo acids. Methylthioalkylmalate synthase (MAMS) catalyzes the first committed step in the side-chain elongation of methionine-derived GSLs. The substrate specificity of MAMS with different 2-oxo acids determines whether reaction products of a given cycle enter for an additional round of chain elongation or enter into core GSLs structure formation. Multiple MAMS encoding genes are present in the Brassicaceae species and are known to play a central role in shaping the diverse profile of aliphatic GSLs. We recently established a highly sensitive LC-MS/MS-based methodology that quantifies the MAMS activity by estimating the amount of the next intermediate of the pathway, the 2-malate derivatives. Overall, this chapter describes the protocol for the expression, purification, and steady-state kinetic analysis of the recombinant MAMS protein.

Inhibitor of Glucosinolate Sulfatases as a Potential Friendly Insecticide to Control Plutella xylostella.

The glucosinolate-myrosinase system is a two-component defense system characteristic of cruciferous plants. To evade the glucosinolate-myrosinase system, the crucifer specialist insect, Plutella xylostella, promptly desulfates the glucosinolates into harmless compounds by glucosinolate sulfatases (GSSs) in the gut. In this study, we identified an effective inhibitor of GSSs by virtual screening, molecular docking analysis, and in vitro enzyme inhibition assay. The combined effect of the GSS inhibitor with the plant glucosinolate-myrosinase system was assessed by the bioassay of P. xylostella. We show that irosustat is a GSS inhibitor and the inhibition of GSSs impairs the ability of P. xylostella to detoxify the glucosinolate-myrosinase system, leading to the systematic accumulation of toxic isothiocyanates in larvae, thereby severely affecting feeding, growth, survival, and reproduction of P. xylostella. While fed on the Arabidopsis mutants deficient in myrosinase or glucosinolates, irosustat had no significant negative effect on P. xylostella. These findings reveal that the GSS inhibitor is a novel friendly insecticide to control P. xylostella utilizing the plant glucosinolate-myrosinase system and promote the development of insecticide-plant chemical defense combination strategies.

A new source of bacterial myrosinase isolated from endophytic Bacillus sp. NGB-B10, and its relevance in biological control activity.

Plant metabolism interacts strongly with the plant microbiome. Glucosinolates, secondary metabolites synthesized by Brassica plants, are hydrolyzed by myrosinase into bioactive compounds of great importance in human health and plant protection. Compared with myrosinase from plant sources, myrosinase enzymes of microbial origin have not been extensively investigated. Therefore, seven endophytic strains corresponding to Bacillus sp. were isolated from Eruca vesicaria ssp. sativa plants that could hydrolyse glucosinolates (sinigrin) in the culture medium and showed myrosinase activity (0.08-19.92 U mL-1). The bglA myrosinase-related gene encoding the 6-phospho-ß-glucosidase (GH 1) from Bacillus sp. NGB-B10, the most active myrosinase-producing bacterium, was successfully identified. Response surface methodology (RSM) was applied to statistically optimize culture conditions for myrosinase production from Bacillus sp. strain NGB-B10. The Plackett-Burman design indicated that nitrogen concentration, incubation period, and agitation speed were the significant parameters in myrosinase production. The application of the Box-Behnken design of RSM resulted in a 10.03-fold increase in enzyme activity as compared to the non-optimized culture conditions. The myrosinase was partially purified by 40% fractionation followed by SDS-PAGE analysis which yielded two subunits that had a molecular weight of 38.6 and 35.0 KDa. The purified enzyme was stable under a broad range of pH (5.5-10) and temperatures (10-65 °C). The hydrolysis products released by bacterial myrosinase from some glucosinolate extracts had higher and/or equivalent in vitro antagonistic activity against several phytopathogenic fungi compared to the nystatin (a broad-spectrum antifungal agent). This study provides original information about a new source of bacterial myrosinase and affords an optimized method to enhance myrosinase production.

Effects of Post-Harvest Elicitor Treatments with Ultrasound, UV- and Photosynthetic Active Radiation on Polyphenols, Glucosinolates and Antioxidant Activity in a Waste Fraction of White Cabbage (Brassica oleracea var. capitata).

Biosynthesis of phytochemicals in leaves of Brassica can be initiated by abiotic factors. The aim of the study was to investigate elicitor treatments to add value to waste of cabbage. A leaf waste fraction from industrial trimming of head cabbage was exposed to UV radiation (250-400 nm, 59 and 99 kJ∙m-2, respectively), photosynthetic active radiation (PAR, 400-700 nm, 497 kJ∙m-2), and ultrasound in water bath (35 kHz, at 15, 30 and 61 kJ∙l-1 water), in order to improve nutraceutical concentration. UV was more effective than PAR to increase the level of flavonols (2 to 3-fold higher) and hydroxycinnamate monosaccharides (1 to 10-fold higher). PAR was three times as effective as UV to increase anthocyanins. Interaction of PAR + UV increased antioxidant activity (30%), the content of five phenolics (1.4 to 10-fold higher), and hydroxycinnamic monosaccharides (compared with PAR or UV alone). Indoles were reduced (40-52%) by UV, but the other glucosinolates (GLS) were unaffected. Ultrasound did not influence any parameters. The results are important for white cabbage by-products by demonstrating that UV + PAR can be successfully used as an effectual tool to increase important phenolics and antioxidant activity of waste fraction leaves without an adverse effect on the main GLS.

Characterization of anti-proliferative and anti-oxidant effects of nano-sized vesicles from Brassica oleracea L. (Broccoli).

In this in vitro study, we test our hypothesis that Broccoli-derived vesicles (BDVs), combining the anti-oxidant properties of their components and the advantages of their structure, can influence the metabolic activity of different cancer cell lines. BDVs were isolated from homogenized fresh broccoli (Brassica oleracea L.) using a sucrose gradient ultracentrifugation method and were characterized in terms of physical properties, such as particle size, morphology, and surface charge by transmission electron microscopy (TEM) and laser doppler electrophoresis (LDE). Glucosinolates content was assessed by RPLC-ESI-MS analysis. Three different human cancer cell lines (colorectal adenocarcinoma Caco-2, lung adenocarcinoma NCI-H441 and neuroblastoma SHSY5Y) were evaluated for metabolic activity by the MTT assay, uptake by fluorescence and confocal microscopy, and anti-oxidant activity by a fluorimetric assay detecting intracellular reactive oxygen species (ROS). Three bands were obtained with average size measured by TEM based size distribution analysis of 52 nm (Band 1), 70 nm (Band 2), and 82 nm (Band 3). Glucobrassicin, glucoraphanin and neoglucobrassicin were found mostly concentrated in Band 1. BDVs affected the metabolic activity of different cancer cell lines in a dose dependent manner compared with untreated cells. Overall, Band 2 and 3 were more toxic than Band 1 irrespective of the cell lines. BDVs were taken up by cells in a dose- and time-dependent manner. Pre-incubation of cells with BDVs resulted in a significant decrease in ROS production in Caco-2 and NCI-H441 stimulated with hydrogen peroxide and SHSY5Y treated with 6-hydroxydopamine, with all three Bands. Our findings open to the possibility to find a novel "green" approach for cancer treatment, focused on using vesicles from broccoli, although a more in-depth characterization of bioactive molecules is warranted.

Phytochemical and functional analysis of horseradish (Armoracia rusticana) fermented and non-fermented root extracts.

BACKGROUND: The roots of horseradish (Armoracia rusticana) are used for infections of respiratory airway and for urinary tract infections due to isothiocyanates (ITC), enzymatically formed during fermentation of glucosinolates by myrosinase. HYPOTHESIS/PURPOSE: The present study aims to present a comprehensive overview on the phytochemical composition of A. rusticana roots, especially concerning isothiocyanates and respective glucosinolates. The complex flavonoid spectrum of the herbal material is reviewed. Published data on in vitro activity of horseradish extracts and isolated compounds are summarized. These data indicate well-established use of horseradish as an antibacterial remedy against bacterial infections of the airway and urinary tract. STUDY DESIGN: To answer the question if other compounds from A. rusticana beside ITC contribute to the antibacterial activity, non-targeted LC-MS studies were performed with fermented and non-fermented horseradish extracts, and detailed phytochemical profiles were established. RESULTS: Comparative investigations on the antibacterial activity indicated that only ITC-containing extracts and fractions exert antibacterial activity. The huge variety of non-ITC compounds do not significantly contribute to the antibacterial activity, but can be used for analytical characterisation and quality control of the herbal material. Detailed phytochemical analysis additionally revealed a variety of compounds, not described until now for horseradish roots: the flavonol glycosides kaempferol-3-O-ß-d-xylopyranosyl-(1''' â†’ 2'')-ß-d-galactopyranoside, kaempferol-3-O-α-l-rhamnopyranosyl-(1''' â†’ 6'')-ß-d-glucopyranoside, kaempferol-3-O-ß-d-glucopyranoside, Kaempferol-3-O-ß-d-xylopyranosyl-7-O-ß-d-glucopyranoside, Kaempferol-3-O-ß-d-xylopyranosyl-(1'''' â†’ 2''')-ß-d-galactopyranoside-7-O-ß-d-glucopyranoside, the oxo-indole derivative spirobrassinin, the phenylthiazole 2-methylsulfanyl-4-phenyl-4,5-dihydro-1,3-thiazole, a series of lysophophatidylethanolamine and 13 different N-phenylpropenoyl-L-amino acids. CONCLUSION: The antibacterial effects of horseradish are only due to the presence of glucosinolates resp. the corresponding ITC, and the detailed overall composition of horseradish extracts has been reported.

Development of a quantification method for routine analysis of glucosinolates and camalexin in brassicaceous small-sized samples by simultaneous extraction prior to liquid chromatography determination.

Glucosinolates and camalexin are secondary metabolites that, as phytoanticipins and phytoalexins, play a crucial role in plant defence. The present work proposes an improved analytical method for routine analysis and quantification of glucosinolates and camalexin in brassicaceous small-sized samples by using the very specific desulfation process of glucosinolates analysis and the specificity of fluorescence detection for camalexin analysis. The approach is based on a simultaneous ultrasound-assisted extraction followed by a purification on an anion-exchange column. Final analyses are conducted by HPLC-UV-MS for desulfo-glucosinolates and HPLC coupled to a fluorescence detector (HPLC-FLD) for camalexin. The method is linear for glucosinolates (50-3500 µM) and camalexin (0.025-5 µg.mL-1) with an LOD/LOQ of 3.8/12.6 µM and 0.014/0.046 µg.mL-1 respectively. The method demonstrated adequate precision, accuracy and trueness on certified reference rapeseed. A practical application of our approach was conducted on different Brassicaceae genera (Barbarea vulgaris, Brassica nigra, Capsella bursa-pastoris, Cardamine hirsuta, Coincya monensis, Sinapis arvensis, and Sisymbrium officinale) and Arabidopsis thaliana genotypes (Columbia and Wassilewskija). Futhermore, different plant organs (seeds and leaves) were analysed, previously inoculated or not with the pathogenic fungus Alternaria brassicicola.

Effect of sequential or ternary starters-assisted fermentation on the phenolic and glucosinolate profiles of sauerkraut in comparison with spontaneous fermentation.

This study investigated the effectiveness of two novel started-assisted sauerkraut fermentations in comparison with spontaneous fermentation. Three lactic acid bacteria strains were selected as best starters for sauerkraut processing, based on the complementarity of pro-technological (kinetics of growth and acidification) traits, phenotypic fingerprints through OmniLog Phenotype MicroArray, and phenolics metabolism. The selected strains were applied according to two different fermentation methods based on steering sequential and temporally deferred inoculum of three strains, and ternary simultaneous inoculum. Sequential and ternary starters-assisted fermentations lasted 9 and 7 days, respectively, and were compared to conventional spontaneous fermentation lasting 35 days. Sequential and ternary fermentations resulted in a higher and constant number of lactic acid bacteria compared to spontaneous fermentation, which reflected on the acidification and sugar utilization. Ternary fermentation enhanced phenolic compounds conversion (hydrocaffeic acid, hydroferulic acid, 4-ethyl catechol), ensuring at the same time higher level of aliphatic (glucobrassicanapin) and indole glucosinolates (glucobrassicin) and derivatives(ascorbigen). Short fermentation of sequential and ternary starters-assisted processing caused only slight changes in the sensory profile compared to the spontaneous process, preserved the structural integrity of sauerkraut, and did not affect the colour lightness.

Manipulation of the Phytochemical Profile of Tenderstem® Broccoli Florets by Short Duration, Pre-Harvest LED Lighting.

Light quality has been reported to influence the phytochemical profile of broccoli sprouts/microgreens; however, few studies have researched the influence on mature broccoli. This is the first study to investigate how exposing a mature glasshouse grown vegetable brassica, Tenderstem® broccoli, to different light wavelengths before harvest influences the phytochemical content. Sixty broccoli plants were grown in a controlled environment glasshouse under ambient light until axial meristems reached >1 cm diameter, whereupon a third were placed under either green/red/far-red LED, blue LED, or remained in the original compartment. Primary and secondary spears were harvested after one and three weeks, respectively. Plant morphology, glucosinolate, carotenoid, tocopherol, and total polyphenol content were determined for each sample. Exposure to green/red/far-red light increased the total polyphenol content by up to 13% and maintained a comparable total glucosinolate content to the control. Blue light increased three of the four indole glucosinolates studied. The effect of light treatments on carotenoid and tocopherol content was inconclusive due to inconsistencies between trials, indicating that they are more sensitive to other environmental factors. These results have shown that by carefully selecting the wavelength, the nutritional content of mature broccoli prior to harvest could be manipulated according to demand.

Untargeted Metabolomic Analysis of Nonvolatile and Volatile Glucosinolates in Brassicaceae.

Glucosinolates (GLSs) are a group of plant secondary metabolites mainly found in Cruciferous plants. The main hydrolysis products of GLSs, isothiocyanates (ITCs), are the bioactive metabolites that have shown plant defense and human cancer prevention properties. Untargeted metabolomic analysis of plant metabolites can uncover the profiles of these bioactive phytochemicals in specific plants and discover potential human health promoting products. We have developed an integrated metabolomic analysis method for plant samples, with specific focus on nonvolatile GLSs and volatile ITCs. In this chapter, we describe in detail the protocols, including metabolite extraction, high resolution LC-MS and GC-MS analysis, and data processing. The method is readily applicable to untargeted analysis of other nonvolatile and volatile metabolites in plants.