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Background: Wasabi, a Brassicaceae member, is well-known for its unique pungent and hot flavor which is produced from glucosinolate (GSL) degradation. Myrosinase (MYR) is a principle enzyme catalyzing the primary conversion of GSLs to GSL hydrolysis products (GHPs) which is responsible for plant defense system and food quality. Due to the limited information in relation to MYRs present in wasabi (Wasabia japonica M.), this study aimed to identify the MYR isogenes in W. japonica and analyze their roles in relation to GSL metabolism., Results: In results, WjMYRI-1 was abundantly expressed in all organs, whereas WjMYRI-2 showed only trace expression levels. WjMYRII was highly expressed in the aboveground tissues. Interestingly, WjMYRII expression was significantly upregulated by certain abiotic factors, such as methyl jasmonate (more than 40-fold in petioles and 15-fold in leaves) and salt (tenfold in leaves). Young leaves and roots contained 97.89 and 91.17 µmol‧g -1 of GSL, whereas less GSL was produced in mature leaves and petioles (38.36 and 44.79 µmol‧g -1 , respectively). Similar pattern was observed in the accumulation of GHPs in various plant organs. Notably, despite the non-significant changes in GSL production, abiotic factors treated samples enhanced significantly GHP content. Pearson's correlation analysis revealed that WjMYRI-1 expression significantly correlated with GSL accumulation and GHP formation, suggesting the primary role of WjMYRI-1-encoding putative protein in GSL degradation. In contrast, WjMYRII expression level showed no correlation with GSL or GHP content, suggesting another physiological role of WjMYRII in stress-induced response., Conclusions: In conclusions, three potential isogenes (WjMYRI-1, WjMYRI-2, and WjMYRII) encoding for different MYR isoforms in W. japonica were identified. Our results provided new insights related to MYR and GSL metabolism which are important for the implications of wasabi in agriculture, food and pharmaceutical industry. Particularly, WjMYRI-1 may be primarily responsible for GSL degradation, whereas WjMYRII (clade II) may be involved in other regulatory pathways induced by abiotic factors., (© 2024. The Author(s).)

We previously found that feeding rats with broccoli or cauliflower leads to the formation of characteristic DNA adducts in the liver, intestine and various other tissues. We identified the critical substances in the plants as 1-methoxy-3-indolylmethyl (1-MIM) glucosinolate and its degradation product 1-MIM-OH. DNA adduct formation and the mutagenicity of 1-MIM-OH in cell models were drastically enhanced when human sulfotransferase (SULT) 1A1 was expressed. The aim of this study was to clarify the role of SULT1A1 in DNA adduct formation by 1-MIM-OH in mouse tissues in vivo. Furthermore, we compared the endogenous mouse Sult1a1 and transgenic human SULT1A1 in the activation of 1-MIM-OH using genetically modified mouse strains. We orally treated male wild-type (wt) and Sult1a1-knockout (ko) mice, as well as corresponding lines carrying the human SULT1A1-SULT1A2 gene cluster (tg and ko-tg), with 1-MIM-OH. N 2 -(1-MIM)-dG and N 6 -(1-MIM)-dA adducts in DNA were analysed using isotope-dilution UPLC-MS/MS. In the liver, caecum and colon adducts were abundant in mice expressing mouse and/or human SULT1A1, but were drastically reduced in ko mice (1.2-10.6% of wt). In the kidney and small intestine, adduct levels were high in mice carrying human SULT1A1-SULT1A2 genes, but low in wt and ko mice (1.8-6.3% of tg-ko). In bone marrow, adduct levels were very low, independently of the SULT1A1 status. In the stomach, they were high in all four lines. Thus, adduct formation was primarily controlled by SULT1A1 in five out of seven tissues studied, with a strong impact of differences in the tissue distribution of mouse and human SULT1A1. The behaviour of 1-MIM-OH in these models (levels and tissue distribution of DNA adducts; impact of SULTs) was similar to that of methyleugenol, classified as "probably carcinogenic to humans". Thus, there is a need to test 1-MIM-OH for carcinogenicity in animal models and to study its adduct formation in humans consuming brassicaceous foodstuff.

Cerebral ischaemia-reperfusion (CIR) injury occurs in stroke patients after the restoration of cerebral perfusion. Sinigrin, a phytochemical found in cruciferous vegetables, exhibits strong antioxidant activity. This study investigated the role of sinigrin in oxidative stress using a CIR injury model. The effects of sinigrin were studied in middle cerebral artery occlusion (MCAO) rats and oxygen-glucose deprivation/reoxygenation (OGD/R)-injured SH-SY5Y cells. Sinigrin treatment improved brain injury and neurological deficits induced by MCAO surgery in rats. Sinigrin inhibited apoptosis in brain tissues and SH-SY5Y cells following OGD/R induction. Additionally, sinigrin elevated the levels of superoxide dismutase (SOD), glutathione (GSH) and glutathione peroxidase (GSH-Px) while reducing malondialdehyde (MDA) levels. Furthermore, sinigrin inhibited the toll-like receptor 4 (TLR4)/myeloid differentiation factor 88 (MyD88) signalling pathway. The anti-apoptotic and antioxidant activities of sinigrin in OGD/R-injured SH-SY5Y cells were reversed by TLR4 overexpression. In conclusion, sinigrin inhibits oxidative stress in CIR injury by suppressing the TLR4/MyD88 signalling pathway., (© 2024 John Wiley & Sons Ltd.)

Inflammatory diseases are strongly associated with global morbidity and mortality. Several mediators are involved in this process, including proinflammatory interleukins and cytokines produced by damaged tissues that, somehow, act as initiators of the autoreactive immune response. Bioactive compounds present in plant-based foods and byproducts have been largely considered active agents with the potential to treat or prevent inflammatory diseases, being a valuable alternative to traditional therapeutic agents used nowadays, which present several side effects. In this regard, the present research uncovers the anti-inflammatory activity of the bioaccessible fraction of broccoli stalks processed, by applying different conditions that render specific concentrations of bioactive sulforaphane (SFN). The raw materials' extracts exhibited significantly different contents of total glucosinolates (GSLs) that ranged between 3993.29 and 12,296.48 mg/kg dry weight (dw), with glucoraphanin as the most abundant one, followed by GI and GE. The indolic GSLs were represented by hydroxy-glucobrassicin, glucobrassicin, methoxy-glucobrassicin, and neo-glucobrassicin, with the two latter as the most abundant. Additionally, SFN and indole-3-carbinol were found in lower concentrations than the corresponding GSL precursors in the raw materials. When exploring the bioaccessibility of these organosulfur compounds, the GSL of all matrices remained at levels lower than the limit of detection, while SFN was the only breakdown product that remained stable and at quantifiable concentrations. The highest concentration of bioaccessible SFN was provided by the high-ITC materials (~4.00 mg/kg dw). The results retrieved on the cytotoxicity of the referred extracts evidenced that the range of supplementation of growth media tested (0.002-430.400 µg of organosulfur compounds/mL) did not display cytotoxic effects on Caco-2 cells. The obtained extracts were assessed based on their capacity to reduce the production of key proinflammatory cytokines (interleukin 6 (IL-6), IL-8, and TNF-α) by the intestinal epithelium. Most of the tested processing conditions provided plant material with significant anti-inflammatory activity and the absence of cytotoxic effects. These data confirm that SFN from broccoli stalks, processed to optimize the bioaccessible concentration of SFN, may be potential therapeutic leads to treat or prevent human intestinal inflammation.

The organic sulfur-containing compounds glucosinolates (GSLs) and the novel gasotransmitter H 2 S are known to have cardioprotective effects. This study investigated the antioxidant effects and H 2 S-releasing potential of three GSLs ((3 E )-4-(methylsulfanyl)but-3-enyl GSL or glucoraphasatin, 4-hydroxybenzyl GSL or glucosinalbin, and ( R S )-6-(methylsulfinyl)hexyl GSL or glucohesperin) in rat cardiac cells. It was found that all three GSLs had no effect on cardiac cell viability but were able to protect against H 2 O 2 -induced oxidative stress and cell death. NaHS, a H 2 S donor, also protected the cells from H 2 O 2 -stimulated oxidative stress and cell death. The GSLs alone or mixed with cysteine, N -acetylcysteine, glutathione, H 2 O 2 , iron and pyridoxal-5'-phosphate, or mouse liver lysates did not induce H 2 S release. The addition of GSLs also did not alter endogenous H 2 S levels in cardiac cells. H 2 O 2 significantly induced cysteine oxidation in the cystathionine gamma-lyase (CSE) protein and inhibited the H 2 S production rate. In conclusion, this study found that the three tested GSLs protect cardiomyocytes from oxidative stress and cell death but independently of H 2 S signaling.

Blindness is a physiopathy characterized by apical abortion that particularly affects the Brassica family. The occurrence of blindness has been related to exposure to low temperatures during early developmental stages. However, the causes of this selective sensitivity and how they affect the correct development remain unknown. In this study, we investigated the mechanisms involved in the occurrence of blindness in broccoli plants. The analysis of RNAseq, focused on membrane transporters and the synthesis pathways of glucosinolates and phenolics, was related with physiological changes in nutrient and water uptake, gas exchange, and metabolism. Comparative gene expression analysis between control and blindness-affected broccoli plants revealed distinct regulation patterns in roots and shoots, leading to reduced synthesis of glucosinolates and phenolics. Additionally, the expression levels of aquaporins and potassium transporters were found to be associated with mineral and water transport. In this way, our results revealed the causes of blindness by identifying differentially expressed genes, highlighting those related to secondary metabolism, as well as genes involved in water and nutrient uptake and transport as the crucial involved in the physiopathy appearance., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2023 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Cruciferous sprouts and microgreens are a good source of bioactive compounds for human health as they are rich in glucosinolates, polyphenols, carotenoids, and vitamins. Glucosinolates - sulfur-containing bioactive phytochemicals - have anti-cancer effects. They mainly exist in cruciferous vegetables. Sulfur is one of the essential elements for plants and is an indispensable component of glucosinolates. This paper summarizes the nutritional value of cruciferous spouts and microgreens, along with the effects of sulfur nutrition on bioactive phytochemical compounds of cruciferous sprouts and microgreens, especially glucosinolates, with the aim of providing information about the dietary effects of cruciferous sprouts and microgreens. © 2023 Society of Chemical Industry., (© 2023 Society of Chemical Industry.)

Widely targeted metabolomics were performed to explore the differences in glucosinolate and odor-active compound levels between flowering Chinese cabbage powder (FCCP) under vacuum-drying and oven-drying conditions. Twenty-three aliphatic, five indole, and three aromatic glucosinolates were identified in two pretreated FCCP. Higher aliphatic glucosinolate levels were retained in vacuum-dried cabbage powder compared to oven-dried samples, and they were negatively correlated with treated temperatures. A total of 36 major odor contributing compounds were detected, including 5 sulfur compounds, 10 aldehydes, 9 heterocyclic compounds, 7 nitriles, 3 acids, and 2 others. 5-Hexenenitrile and (methyldisulfanyl)methan, provide typical pungent, sulfous, and vegetable notes in FCCP. Four major GSLs, namely 2(R)-hydroxy-3-butenyl glucosinolate, (2S)-2-hydroxy-4-pentenyl glucosinolate, 5-(methylthio)pentyl glucosinolate and 2-phenylethyl glucosinolate were the key precursors to form odor-active compounds. Higher temperatures in thermal effects promotes the formation of sulfur-containing and nitrile compounds compared to the vacuum-dried ones. This work can provide a guide for flavor and nutrition retention in FCCP process., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Protein glycation can result in the formation of advanced glycation end products (AGEs), which pose a potential health risk due to their association with diabetic complications. Natural products are a source of drugs discovery and the search for potential natural inhibitors of AGEs is of great significance. Glucosinolates (GSLs) mainly from cruciferous plants have potential antioxidant, anti-inflammatory, and anti-glycation activities. In this study, the inhibitory activity of GSLs on bovine serum albumin (BSA) along with its mechanism was investigated by virtual screening and various computational simulation techniques. Virtual screening revealed that 174 GSLs were screened using Maestro based on the glide score and 89% of the compounds were found to have potential anti-glycation ability with the docking scores less than -5 kcal/mol. Molecular docking showed that the top 10 GSLs were bound to the IIA structural domain of BSA. Among them, glucohesperin (1) and 2-hydroxyethyl glucosinolate (2) had the lowest docking scores of -9.428 and -9.333 kcal/mol, respectively, reflecting their good binding affinity. Molecular dynamics simulations of 1 (ΔG = -43.46 kcal/mol) and 2 (ΔG = -43.71 kcal/mol) revealed that the complexes of these two compounds with proteins had good stability. Further binding site analysis suggested that the mechanism of inhibition of protein glycation by these two active ingredients might be through competitive hydrogen bonding to maintain the structural integrity of the protein, thus inhibiting glycation reaction. Moreover, the ADMET values and CYP450 metabolism prediction data were within the recommended values. Therefore, it can be concluded that 1 and 2 may act as potential anti-glycation agents., (© 2023 Wiley Periodicals LLC.)

The Optimal Defense Theory (ODT) predicts that the distribution of defenses within a plant should mirror the value and vulnerability of each tissue. Although the ODT has received much experimental support, very few studies have examined defense allocation among reproductive tissues and none assessed simultaneously how these defenses evolve with age. We quantified glucosinolates in perianths, anthers and pistils at different bud maturity stages (i.e., intermediate flower buds, old flower buds and flowers) of undamaged and mechanically damaged plants of an annual brassicaceous species. The youngest leaf was used as a reference for vegetative organs, since it is predicted to be one of the most defended. In line with ODT predictions, reproductive tissues were more defended than vegetative tissues constitutively, and within the former, pistils and anthers more defended than perianths. No change in the overall defense level was found between bud maturity stages, but a significant temporal shift was observed between pistils and anthers. Contrary to ODT predictions, mechanical damage did not induce systemic defenses in leaves but only in pistils. Our results show that defense allocation in plant reproductive tissues occurs at fine spatial and temporal scales, extending the application framework of the ODT. They also demonstrate interactions between space and time in fine-scale defense allocation., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Glucosinolates are precursors of bioactive and health-promoting isothiocyanates (ITCs). Upon enzymatic hydrolysis, Brassica vegetables, such as cabbage, also often yield nitriles and epithionitriles as main products next to ITCs. Here, we show that amines can be additional main enzymatic hydrolysis products of glucosinolates in Brassica vegetables. We propose that a plant endogenous ITC hydrolase (ITCase) is responsible for the enzymatic-like conversion of ITCs to amines in cabbage samples. This ITCase seems to have high activity towards alkenyl ITCs like allyl ITC and lower activity towards methylthioalkyl ITCs, and not to converting methylsulfinylalkyl ITCs like sulforaphane. In contrast, during heat treatment of homogenized cabbage material, methylsulfinylalkylamine levels increased by 400 % after 2 h of heating, which is likely due to thermal decomposition of ITCs, whereas alkenyl amine levels did not change due to heat treatment. The results show that amines from glucosinolates are part of the human diet., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Franziska S. Hanschen reports financial support was provided by Leibniz Association. Lars Andernach reports financial support was provided by German Research Foundation., (Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The flavor of Guizhou suancai fermented by the emerging varieties Zhuchang-2 (ZC-2) and Zhuchang Red (ZC Red) is superior to that of ZC-1, a typical local variety of leaf mustard in Guizhou Province, China. Here, changes in phytochemical qualities during the fermentation of three Guizhou suancai were characterized to identify the causes underlying differences in flavor. The content of pigments, antioxidants, gluconapin, gluconasturtiin, and total isothiocyanates, and antioxidant capacity were highest in fresh ZC Red. The content of sinigrin, indolic glucosinolates, and total glucosinolate breakdown products was highest in fresh ZC-2. The content of phytochemicals, with the exception of carotenoids, was significantly decreased after fermentation. Odor was correlated with gluconapin, while taste was correlated with sinigrin. These findings provide detailed insights into the sensory and phytochemical properties of three Guizhou suancai that could facilitate the selection of raw material varieties., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

In the last decade, most of the evidence on the clinical benefits of including cruciferous foods in the diet has been focused on the content of glucosinolates (GSL) and their corresponding isothiocyanates (ITC), and mercapturic acid pathway metabolites, based on their capacity to modulate clinical, biochemical, and molecular parameters. The present systematic review summarizes findings of human studies regarding the metabolism and bioavailability of GSL and ITC, providing a comprehensive analysis that will help guide future research studies and facilitate the consultation of the latest advances in this booming and less profusely researched area of GSL for food and health. The literature search was carried out in Scopus, PubMed and the Web of Science, under the criteria of including publications centered on human subjects and the use of Brassicaceae foods in different formulations (including extracts, beverages, and tablets), as significant sources of bioactive compounds, in different types of subjects, and against certain diseases. Twenty-eight human intervention studies met inclusion criteria, which were classified into three groups depending on the dietary source. This review summarizes recent studies that provided interesting contributions, but also uncovered the many potential venues for future research on the benefits of consuming cruciferous foods in our health and well-being. The research will continue to support the inclusion of GSL-rich foods and products for multiple preventive and active programs in nutrition and well-being.

Baseline levels of glucosinolates-important defensive phytochemicals in brassicaceous plants-are determined by both genotype and environment. However, the ecological causes of glucosinolate plasticity are not well characterized. Fertilization is known to alter glucosinolate content of Brassica crops, but the effect of naturally occurring soil variation on glucosinolate content of wild plants is unknown. Here, we conducted greenhouse experiments using Boechera stricta to ask (i) whether soil variation among natural habitats shapes leaf and root glucosinolate profiles; (ii) whether such changes are caused by abiotic soil properties, soil microbes, or both; and (iii) whether soil-induced glucosinolate plasticity is genetically variable. Total glucosinolate quantity differed up to 2-fold between soils from different natural habitats, while the relative amounts of different compounds were less responsive. This effect was due to physico-chemical soil properties rather than microbial communities. We detected modest genetic variation for glucosinolate plasticity in response to soil. In addition, glucosinolate composition, but not quantity, of field-grown plants could be accurately predicted from measurements from greenhouse-grown plants. In summary, soil alone is sufficient to cause plasticity of baseline glucosinolate levels in natural plant populations, which may have implications for the evolution of this important trait across complex landscapes., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Background: Chinese cabbage is one of the most widely grown leafy vegetables in China. Cytoplasmic male sterility (CMS) is a maternally inherited trait that produces abnormal pollen during anther development, which is commonly seen in cruciferous vegetables. However, the molecular mechanism of Chinese cabbage CMS is not clear. In this study, the metabolome and hormone profiles of Chinese cabbage male sterile line (CCR20000) and sterile maintainer line (CCR20001) were analyzed in flower buds during normal stamen development and abnormal stamen development, respectively., Results: A total of 556 metabolites were detected based on UPLC-MS/MS detection platform and database search, and the changes of hormones such as auxin, cytokinins, abscisic acid, jasmonates, salicylic acid, gibberellin acid and ethylene were analyzed. The results showed that compared with the male fertile line (MF), the male sterile line (MS) significantly decreased the content of flavonoids and phenolamides metabolites in the stamen dysplasia stage, accompanied by a large accumulation of glucosinolate metabolites. Meanwhile, the contents of GA9, GA20, IBA, tZ and other hormones in MS were significantly lower than those in MF strains. Further, by comparing the metabolome changes of MF and MS during stamen dysplasia, it was found that flavonoid metabolites and amino acid metabolites were distinctly different., Conclusions: These results suggest that flavonoids, phenolamides and glucosinolate metabolites may be closely related to the sterility of MS strains. This study provides an effective basis for further research on the molecular mechanism of CMS in Chinese cabbage., (© 2023. The Author(s).)

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.

Phenolic compounds and glucosinolates are secondary plant metabolites that play fundamental roles in plant resistance to abiotic stress. These compounds have been found to increase in stress situations related to plant adaptive capacity. This review assesses the functions of phenolic compounds and glucosinolates in plant interactions involving abiotic stresses such as drought, salinity, high temperature, metals toxicity, and mineral deficiency or excess. Furthermore, their relation with water uptake and transport mediated through aquaporins is reviewed. In this way, the increases of phenolic compounds and glucosinolate synthesis have been related to primary responses to abiotic stress and induction of resistance. Thus, their metabolic pathways, root exudation, and external application are related to internal cell and tissue movement, with a lack of information in this latter aspect.

Glucosinolates and their metabolites from Brassicaceae plants have received widespread attention due to their anti-inflammatory effects. Glucosinolates occurs an "enterohepatic circulation" in the body, and the glucosinolates metabolism mainly happens in the intestine. Glucosinolates can be converted into isothiocyanates by intestinal bacteria, which are active substances with remarkable anti-inflammatory, anti-cancer, anti-obesity and neuroprotective properties. This biotransformation can greatly improve the bioactivities of glucosinolates. However, multiple factors in the environment can affect the biotransformation to isothiocyanates, including acidic pH, ferrous ions and thiocyanate-forming protein. The derivatives of glucosinolates under those conditions are usually nitriles and thiocyanates, which may impair the potential health benefits. In addition, isothiocyanates are extremely unstable because of an active sulfhydryl group, which limits their applications. This review mainly summarizes the classification, synthesis, absorption, metabolism, physiological functions and potential application strategies of glucosinolates and their metabolites.

The myrosinase-glucosinolate system, physicochemical properties, and bacterial community were profiled during fermentation of high hydrostatic pressure (HHP) pretreated brine-pickled radishes; traditionally brine-pickled radishes were utilised as the control. Scanning electron microscopy (SEM) analysis revealed that 300 MPa pretreatment promoted brine infiltration in radish cells and damaged cellular microstructures, which activated the myrosinase-glucosinolate system. The conversion of glucosinolate (GLs) to isothiocyanates (ITC) increased and significantly enhanced the raphasatin and sulforaphene contents of pickled radish. However, 600 MPa pretreatment suppressed myrosinase activity. HHP pretreatment altered the natural radish bacterial communities by reducing the total bacterial and lactic acid bacteria counts. Lactobacillus spp. became the dominant bacterial genus within 15 d of fermentation. However, the destruction of cellular structures by HHP pretreatment also significantly decreased hardness and caused the dissolution of amino acids and TTA into brine. This caused reduced amino acid and TTA contents compared to the control group, as well as decreases in pH. HHP pretreatment suppressed the growth of spoilage bacteria (e.g. Pseudomonas, Staphylococcus, and Shewanella genera). This study provides new insight into the potential applications of HHP treatment in pickling, as it demonstrates that HHP can increase the ITC conversion rate of pickled radish, modify its physiochemical characteristics, and decrease microbial risk. Therefore, HHP is a promising preprocessing technique to be used for pickle manufacturing industry., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The effects of supplemental UV-A (385 nm) period and UV-A intensity for 5 days before harvest (DBH) on growth, antioxidants, antioxidant capacity, and glucosinolates contents in Chinese kale ( Brassica oleracea var. alboglabra Bailey) were studied in plant factory. In the experiment of the UV-A period, three treatments were designed with 10 W·m -2 UV-A supplement, T1(5 DBH), T2 (10 DBH), and no supplemental UV-A as control. In the experiment of UV-A intensity, four treatments were designed with 5 DBH, control (0 W·m -2 ), 5 w (5 W·m -2 ), 10 w (10 W·m -2 ), and 15 w (15 W·m -2 ). The growth light is as follows: 250 μmol·m -2 ·s -1 ; red light: white light = 2:3; photoperiod: 12/12. The growth and quality of Chinese kale were improved by supplemental UV-A LED. The plant height, stem diameter, and biomass of Chinese kale were the highest in the 5 W·m -2 treatment for 5 DBH. The contents of chlorophyll a, chlorophyll b, and total chlorophyll were only highly increased by 5 W·m -2 UV-A for 5 DBH, while there was no significant difference in the content of carotenoid among all treatments. The contents of soluble sugar and free amino acid were higher only under 10 DBH treatments than in control. The contents of total phenolic and total antioxidant capacity were the highest in 5 W·m -2 treatment for 5 DBH. There was a significant positive correlation between total phenolic content and DPPH and FRAP value. After 5 DBH treatments, the percentages and contents of total aliphatic glucosinolates, sinigrin (SIN), gluconapin (GNA), and glucobrassicanapin (GBN) were highly increased, while the percentages and contents of glucobrassicin (GBS), 4-methoxyglucobrassicin (4-MGBS), and Progoitrin (PRO) were significantly decreased, especially under 10 W·m -2 treatment. Our results show that UV-A LED supplements could improve the growth and quality of Chinese kale, and 5 W·m -2 UV-A LED with 5 DBH might be feasible for Chinese kale growth, and 10 W·m -2 UV-A LED with 5 DBH was better for aliphatic glucosinolates accumulation in Chinese kale.

The Brassica oleracea industry generates large amounts of by-products to which value could be added because of the characteristics of their composition. The aim was to extract different fibre fractions from broccoli stalks to obtain potential new added-value ingredients. Using an ethanol and water extraction procedure, two fibre-rich fractions (total fibre fraction, TF B , and insoluble fibre fraction, IF B ) were obtained. These fractions were analysed to determine the nutritional, (poly)phenols and glucosinolates composition and physicochemical properties, comparing the results with those of freeze-dried broccoli stalks (DBS). Although TF B showed a higher content of total dietary fibre, IF B had the same content of insoluble dietary fibre as TF B (54%), better hydration properties, higher content of glucosinolates (100 mg/100 g d.w.) and (poly)phenols (74.7 mg/100 g d.w.). The prebiotic effect was evaluated in IF B and compared with DBS by in vitro fermentation with human faecal slurries. After 48 h, the short-chain fatty acid (SCFA) production was higher with IF B than with DBS because of the greater presence of both uronic acids, the main component of pectin, and (poly)phenols. These results reveal that novel fibre-rich ingredients-with antioxidant, technological and physiological effects-could be obtained from broccoli stalks by using green extraction methods.

The glucosinolates (GLs) and myrosinase defensive systems in cruciferous plants were circumvented by Plutella xylostella using glucosinolate sulfatases (PxGSSs) during pest-plant interaction. Despite identifying three duplicated GSS-encoding genes in P. xylostella, limited information regarding their spatiotemporal and induced expression is available. Here, we investigated the tissue- and stage-specific expression and induction in response to GLs of PxGSS1 and PxGSS2 (PxGSS1/2) at the protein level, which shares a high degree of similarity in protein sequences. Western blotting (WB) analysis showed that PxGSS1/2 exhibited a higher protein level in mature larvae, their guts, and gut content. A significantly high protein and transcript levels of PxGSS1/2 were also detected in the salivary glands using WB and qRT-PCR. The immunofluorescence (IF) and immunohistochemistry (IHC) results confirmed that PxGSS1/2 is widely expressed in the larval body. The IHC was more appropriate than IF when autofluorescence interference was present in collected samples. Furthermore, the content of PxGSS1/2 did not change significantly under treatments of GL mixture from Arabidopsis thaliana ecotype Col-0, or commercial ally (sinigrin), 4-(methylsulfinyl)butyl, 3-(methylsulfinyl)propyl, and indol-3-ylmethyl GLs indicating that the major GLs from leaves of A. thaliana Col-0 failed to induce the expression of proteins for both PxGSS1 and PxGSS2. Our study systemically characterized the expression properties of PxGSS1/2 at the protein level, which improves our understanding of PxGSS1/2-center adaptation in P. xylostella during long-term insect-plant interaction., (© The Author(s) 2022. Published by Oxford University Press on behalf of Entomological Society of America.)

Pickled tuber mustard is a traditional fermented pickle widely consumed in China, and it is characterized by the presence of glucosinolates (GSLs). To understand the biotransformation of GSLs in tuber mustard during pickling, the dynamics of the glucosinolate-myrosinase (GSL-MYR) system and its potential associations with bacterial communities and fermentation characteristics (i.e., salinity, titratable acidity [TAA], and pH) were investigated. In total, 18 GSLs were identified in fresh tuber mustard; 12 were aliphatic, 4 were indolic, and 2 were aromatic, with aliphatic sinigrin and aromatic gluconasturtiin being the dominating components. The pickling process resulted in complete degradation of GSLs, with isothiocyanates (ITCs) and nitriles being the main breakdown products. Total ITCs reached maximum concentrations on day 21-28, while total nitriles peaked at the end of pickling. Based on Spearman's correlation analysis, our study showed that lactic acid bacteria (LAB) species might contribute to GSL transformation in pickled tuber mustard. Specifically, Weissella paramesenteroides, Pediococcus pentosaceus, and unclassified Lactococcus exhibited positive correlations with GSL contents (p < 0.01), suggesting that they might contribute to the increasing amounts of GSLs in the initial pickling, while the Lactobacillus-related populations that dominated in the later stages (i.e., Companilactobacillus alimentarius and Lactiplantibacillus Plantarum) were positively correlated with nitrile product concentrations. Moreover, redundancy analysis showed that pH and TAA had strong effect on myrosinase activity during tuber mustard pickling, which was dictated via the organic acids produced by microorganisms. This study provided a perspective for understanding the effect of fermentation on the transformation of tuber mustard GSLs., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Premise: Invasive plant species often escape from specialist herbivores and are more likely to be attacked by generalist herbivores in the exotic range. Consequently, the shifting defense hypothesis predicts that invasive plants will produce higher concentrations of qualitative defense compounds to deter dominant generalist herbivores in the exotic range. Here, I additionally propose a reduced chemical diversity hypothesis (RCDH), which predicts that reduced herbivory pressure will select for invasive plant genotypes that produce lower diversities of chemical defense compounds in the exotic range., Methods: I tested whether (1) invasive Brassica nigra populations express a lower diversity and an overall higher concentration of glucosinolate compounds than native-range B. nigra; (2) Brassica nigra individuals that express high diversities and concentrations of glucosinolate compounds are more attractive to specialist and deterrent to generalist herbivores; and (3) tissues of invasive B. nigra are less palatable than tissues of native-range B. nigra to the generalist herbivores Theba pisana and Helix aspersa., Results: Invasive B. nigra populations produced a significantly lower diversity of glucosinolate compounds, a marginally higher concentration of total glucosinolates, and a significantly higher concentration of sinigrin (the dominant glucosinolate). Leaf tissues of invasive B. nigra were significantly less palatable to T. pisana and marginally less so to H. aspersa. Brassica nigra individuals that expressed high concentrations of total glucosinolate compounds were visited by a low diversity of generalist herbivore species in the field., Conclusions: In line with the RCDH, the lower diversity of glucosinolate compounds produced by invasive B. nigra populations likely resulted from selection imposed by reduced herbivory pressure in the exotic range., (© 2022 Botanical Society of America.)

The increasing demand for food and the need for a sustainability vision in the agri-food sector have boosted novel approaches for food management, enhancing the valorization of wastes and by-products belonging to the food industry. Herein, we present a novel paper-based origami device to assess the amount of both glucosinolate and glucose in a food waste product belonging to Brassicaceae plants, to evaluate the quality value and the correct management of waste samples. The device has been designed as an origami paper-based platform constituted of two paper-based biosensors to work synergistically in a multiplexed detection. In detail, a monoenzymatic biosensor and a bienzymatic biosensor were configured for the detection of glucose and glucosinolates, respectively, using filter paper pads preloaded with glucose oxidase and/or myrosinase. To complete the paper-based platform, the enzyme-preloaded pads were combined with office paper-based electrodes modified with Carbon black/Prussian Blue nanoparticles for the measurement of enzymatic by-product at a low applied potential (i.e., 0 V versus Ag/AgCl). Overall, this paper-based platform measured glucose and glucosinolate (i.e., sinigrin) with a linear range up to 2.5 and 1.5 mM, and detection limits of 0.05 and 0.07 mM, respectively. The repeatability corresponded to an RSD% equal to 5% by testing 10 mM of glucose, and 10% by testing 1 mM of sinigrin. The accuracy of the developed multiplex device was evaluated by recovery studies at two different levels of sinigrin, i.e., 0.25 and 0.5 mM, obtaining recoveries values equal to (111 ± 3) % and (86 ± 1) %, respectively. The multiplex detection of both glucose and glucosinolate in Brassicaceae samples evaluates the quality values of the waste sample, ensuring the quality of the re-used food product waste by using an eco-designed analytical tool. The combination of paper-based devices for quality control of food waste with the re-use of these food products represents a sustainable approach that perfectly matches sustainable agrifood practices as well as the overall approach of the circular economy., (© 2022. The Author(s).)

The glucosinolate pathway, which is present in the order Brassicales, is one of the most researched defensive natural product biosynthesis pathways. Its core molecules, the glucosinolates are broken down upon pathogen challenge or tissue damage to yield an array of natural products that may help plants defend against the stressor. Though the most widely known glucosinolate decomposition products are the antimicrobial isothiocyanates, there is a wide range of other volatile and non-volatile natural products that arise from this biosynthetic pathway. This review summarizes our current knowledge on the interaction of these much less examined, non-isothiocyanate products with fungi. It deals with compounds including (1) glucosinolates and their biosynthesis precursors; (2) glucosinolate-derived nitriles (e.g. derivatives of 1H-indole-3-acetonitrile), thiocyanates, epithionitriles and oxazolidine-2-thiones; (3) putative isothiocyanate downstream products such as raphanusamic acid, 1H-indole-3-methanol (= indole-3-carbinol) and its oligomers, 1H-indol-3-ylmethanamine and ascorbigen; (4) 1H-indole-3-acetonitrile downstream products such as 1H-indole-3-carbaldehyde (indole-3-carboxaldehyde), 1H-indole-3-carboxylic acid and their derivatives; and (5) indole phytoalexins including brassinin, cyclobrassinin and brassilexin. Herein, a literature review on the following aspects is provided: their direct antifungal activity and the proposed mechanisms of antifungal action, increased biosynthesis after fungal challenge, as well as data on their biotransformation/detoxification by fungi, including but not limited to fungal myrosinase activity., (Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The utilization of rapeseed meal in food is limited due to its abundant glucosinolates (GLs). In this study, an LC-MS/MS method for GLs determination in rapeseed meal was developed. Then, the degradation of GLs using rapeseed sprouts derived myrosinase (MYR) was investigated. Results showed that 11 kinds of GLs were identified in rapeseed meal. The LC-MS/MS method had a high linearity (R 2  greater than 0.9999), repeatability (RSD < 5%) and recovery rate (92%-102%). The optimum condition for hydrolyzing GLs in rapeseed meal was reacting for 4 h with the addition of 2236.35 U/g MYR, 9.63 μg/g ascorbic acid and 26.68 μg/g EDTA. Under this condition, more than 80% of GLs were degraded and the yields of isothiocyanates and oxazolidinone-2-thione were 859.30 μg/g and 685.59 μg/g, respectively. To conclude, this study reported a reliable method for GLs determination and an effective way to degrade GLs in rapeseed meal., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Introduction: The high mortality rate in severe cases of COVID-19 is mainly due to the strong upregulation of cytokines, called a cytokine storm. Hyperinflammation and multiple organ failure comprise the main clinical features of a cytokine storm. Nrf2 is a transcription factor which regulates the expression of genes involved in immune and inflammatory processes. Furthermore, Nrf2, as a master regulator, controls the activity of NF-κB which binds to the promoter of many pro-inflammatory genes inducible of various inflammatory factors. Inhibition of Nrf2 response was recently demonstrated in biopsies from patients with COVID-19, and Nrf2 agonists inhibited SARS-CoV-2 replication across cell lines in vitro. Glucosinolates and their hydrolysis products have excellent anti-inflammatory and antioxidant effects via the Nrf2 activation pathway, reduction in the NF-κB activation, and subsequent reduced cytokines levels., Conclusion: Accordingly, these compounds can be helpful in combating the cytokine storm associated with COVID-19., (© 2022. Springer Nature Switzerland AG.)

Mitochondrial retrograde signaling (MRS) plays an essential role in sensing and responding to internal and external stimuli to optimize growth to adapt to the prevailing environmental conditions. Previously studies showed alterations on MRS in cytoplasmic male sterile (CMS) plant. However, the regulators involved in MRS in CMS plants remain largely unknown. In this study, we used alternative oxidase 1a (AOX1a) as an indicator of MRS and found that the expression of AOX1a was significantly downregulated in a CMS line comparing to its revertant line, thus indicating an alteration in MRS in the CMS line. By performing a BLAST search of known regulatory components involved in MRS in yeast, we identified general regulatory factor 3 (GRF3), an orthologue of Bmh1/2 in yeast, and demonstrated an association between this gene and MRS in plants, as evidenced by change in AOX1a expression. GRF3 protein was found to be located in the nucleus and the plasma membrane. Further studies showed that GRF3 interacted with MYB29, and regulated the biosynthesis of glucosinolates in Brassica juncea. These findings revealed that GRF3, a negative regulator of AOX1a, is involved in MRS, and also plays a vital role in the accumulation of glucosinolates in CMS crops., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Glucoraphanin is a plant specialized metabolite found in cruciferous vegetables that has long been a target for production in a heterologous host because it can subsequently be hydrolyzed to form the chemopreventive compound sulforaphane before and during consumption. However, previous studies have only been able to produce small amounts of glucoraphanin in heterologous plant and microbial systems compared to the levels found in glucoraphanin-producing plants, suggesting that there may be missing auxiliary genes that play a role in improving production in planta . In an effort to identify auxiliary genes required for high glucoraphanin production, we leveraged transient expression in Nicotiana benthamiana to screen a combination of previously uncharacterized coexpressed genes and rationally selected genes alongside the glucoraphanin biosynthetic pathway. This strategy alleviated metabolic bottlenecks, which improved glucoraphanin production by 4.74-fold. Our optimized glucoraphanin biosynthetic pathway provides a pathway amenable for high glucoraphanin production.

Chinese cabbage (Brassica rapa ssp. pekinensis) is an economically important vegetable crop throughout the world, especially in Asia. High-quality genome sequences are available for Chinese cabbage, but gene functional studies remain challenging. To promote functional genomic studies of Chinese cabbage, we generated an ethyl methane sulfonate (EMS) mutant population of ∼8000 M 2 plants using the double haploid inbred line A03 as the parent. The genome of A03 was sequenced and used as a reference for high-throughput functional characterization of gene mutations at the whole-genome level. A total of 300 M 2 to M 5 EMS mutants were phenotypically screened and then sequenced, revealing 750 629 SNPs and 46 272 InDel mutations that cover 98.27% of all predicted genes in the A03 genome. A forward-genetics approach was successfully used to identify two genes with chloroplast-related functions that are responsible for the yellow leaf mutant trait. A reverse-genetics approach was also used to identify associations between mutations in five genes of the glucosinolate biosynthetic pathway and variations in glucosinolate content of the mutant plants. In addition, we built the Chinese cabbage EMS mutation database (CCEMD, www.bioinformaticslab.cn/EMSmutation/home) to increase the usability of this mutant population resource. In summary, we performed large-scale screening of a heading Chinese cabbage EMS mutant collection at the phenotypic and genotypic levels, which will facilitate gene mining of Chinese cabbage and might also be useful for the study of other Brassica crops., (Copyright © 2022 The Author. Published by Elsevier Inc. All rights reserved.)

Radish ( Raphanus sativus L.) is rich in specific glucosinolates (GSLs), which benefit human health and special flavor formation. Although the basic GSLs metabolic pathway in Brassicaceae plants is clear, the regulating mechanism for specific glucosinolates content in radish fleshy taproots is not well understood. In this study, we discovered that there was a significant difference in the GSLs profiles and the content of various GSLs components. Glucoraphasatin (GRH) is the most predominant GSL in radish taproots of different genotypes as assessed by HPLC analysis. Further, we compared the taproot transcriptomes of three radish genotypes with high and low GSLs content by employing RNA-seq. Totally, we identified forty-one differentially expressed genes related to GSLs metabolism. Among them, thirteen genes ( RsBCAT4 , RsIPMDH1 , RsMAM1a , RsMAM1b , RsCYP79F1 , RsGSTF9 , RsGGP1 , RsSUR1 , RsUGT74C1 , RsST5b , RsAPK1 , RsGSL-OH , and RsMYB28 ) were significantly higher co-expressed in the high content genotypes than in low content genotype. Notably, correlation analysis indicated that the expression level of RsMYB28 , as an R2R3 transcription factor directly regulating aliphatic glucosinolate biosynthesis, was positively correlated with the GRH content. Co-expression network showed that RsMYB28 probably positively regulated the expression of the above genes, particularly RsSUR1 , and consequently the synthesis of GRH. Moreover, the molecular mechanism of the accumulation of this 4-carbon (4C) GSL in radish taproots was explored. This study provides new perspectives on the GSLs accumulation mechanism and genetic improvements in radish taproots.

Juices of Brassica vegetables are mutagenic and form characteristic DNA adducts in bacteria and mammalian cells. In this study, we examined whether such adducts are also formed in vivo in animal models. Rats fed raw broccoli ad libitum in addition to normal laboratory chow for 5 weeks showed one major adduct spot and sometimes an additional minor adduct spot in liver, kidney, lung, blood and the gastrointestinal tract, as determined by 32 P-postlabelling/thin-layer chromatography. Adducts with the same chromatographic properties were formed when herring sperm DNA (or dG-3'-phosphate) was incubated with 1-methoxy-3-indolylmethyl glucosinolate (phytochemical present in Brassica plants), in the presence of myrosinase (plant enzyme that hydrolyses glucosinolates to bioactive breakdown products). UPLC-MS/MS analysis corroborated this finding: 1-Methoxy-3-indolylmethyl-substituted purine nucleosides were detected in the hepatic DNA of broccoli-fed animals, but not in control animals. Feeding raw cauliflower led to the formation of the same adducts. When steamed rather than raw broccoli was used, the adduct levels were essentially unchanged in liver and jejunum, but elevated in large intestine. Due to inactivation of myrosinase by the steaming, higher levels of the glucosinolates may have reached the large bowl to be activated by glucosidases from intestinal bacteria. In conclusion, the consumption of common Brassica vegetables can lead to the formation of substantial levels of DNA adducts in animal models. The adducts can be attributed to a specific phytochemical, neoglucobrassicin (1-methoxy-3-indolylmethyl glucosinolate)., (© 2022. The Author(s).)

The glucosinolate (GSL) profiles of four Limnanthaceae species, including the oil crop Limnanthes alba (meadowfoam), were investigated by an ultrahigh-performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry (UHPLC-QToF-MS/MS) analysis of desulfoGSLs after desulfation of native GSLs, supplemented by NMR of desulfated 2-hydroxy-2-methylpropylGSL and 3-methoxybenzylGSL. Leaves, roots, and seeds were investigated, providing an overview of biosynthetic capabilities in the genera Floerkea and Limnanthes . Methoxyl groups on benzylGSLs were in meta but not para positions; two 3,5-disubstituted benzylGSLs are tentatively proposed. 2-Hydroxy-2-methylpropylGSL was accompanied by an isomer that was not a previously reported GSL. The combined GSL profile of the family included GSLs derived from valine, leucine, isoleucine, phenylalanine, and tyrosine, and possibly methionine and tryptophan. Substituted indole GSLs and GSLs derived from chain-elongated amino acids or alanine were searched for but not detected. Hypothetic glycosides of GSLs were detected at low levels. Based on biochemical interpretation, we suggest biosynthetic schemes and gene families ( CYP79C , GSOH ) relevant for tailoring GSL profiles in Limnanthes crops.

Glucosinolate transporters (GTRs) are part of the nitrate/peptide transporter (NPF) family, members of which also transport specialized secondary metabolites as substrates. Glucosinolates are defense compounds derived from amino acids. We selected 4-methylthiobutyl (4MTB) and indol-3-ylmethyl (I3M) glucosinolates to study how GTR1 from Arabidopsis thaliana transports these substrates in computational simulation approaches. The designed pipeline reported here includes massive docking of 4MTB and I3M in an ensemble of GTR1 conformations (in both inward and outward conformations) extracted from molecular dynamics simulations, followed by clustered and substrate-protein interactions profiling. The identified key residues were mutated, and their role in substrate transport was tested. We were able to identify key residues that integrate a major binding site of these substrates, which is critical for transport activity. In silico approaches employed here represent a breakthrough in the plant transportomics field, as the identification of key residues usually takes a long time if performed from a purely wet-lab experimental perspective. The inclusion of structural bioinformatics in the analyses of plant transporters significantly speeds up the knowledge-gaining process and optimizes valuable time and resources.

Glucosinolates are an important class of secondary metabolites in Brassicales plants with a critical role in chemical defense. Glucosinolates are chemically inactive but can be hydrolyzed by myrosinases to produce a range of chemically active compounds toxic to herbivores and pathogens, thereby constituting the glucosinolate-myrosinase defense system or the mustard oil bomb. During the evolution, Brassicales plants have developed not only complex biosynthetic pathways for production of a large number of glucosinolate structures but also different classes of myrosinases that differ in catalytic mechanisms and substrate specificity. Studies over the past several decades have made important progress in the understanding of the cellular and subcellular organization of the glucosinolate-myrosinase system for rapid and timely detonation of the mustard oil bomb upon tissue damage after herbivore feeding and pathogen infection. Progress has also been made in understanding the mechanisms that herbivores and pathogens have evolved to counter the mustard oil bomb. In this review, we summarize our current understanding of the function and organization of the glucosinolate-myrosinase system in Brassicales plants and discuss both the progresses and future challenges in addressing this complex defense system as an excellent model for analyzing plant chemical defense.

Glucosinolate (GSL) not only has highly physiological function for plants but also has considerable human interest. We analyzed the GSL compositions and levels in four organs of 111 radish accessions. Seven major GSLs were detected (approximately 5-245 μmol g -1 DW), among which 4-(methylsulfinyl)but-3-enyl GSL and 4-methylsulfanyl-3-butenyl GSL were the dominant GSLs. GSL levels varied substantially among species and groups, and some genotypes/groups with special GSL profiles were identified. The total GSL level was higher in seeds than in sprouts, taproots, and leaves. Additionally, a correlation analysis revealed that seed 4-(methylsulfinyl)but-3-enyl GSL levels were highly correlated with sprout GSL levels. Moreover, a candidate gene ( RsCYP81F2.3 ) encoding an enzyme that catalyzes the conversion of indol-3-ylmethyl GSL to 4-hydroxyindol-3-ylmethyl GSL was identified based on the detection and analysis of three radish accessions with relatively high indol-3-ylmethyl GSL, low 4-hydroxyindol-3-ylmethyl GSL, and 4-methoxyindol-3-ylmethyl GSL levels in their seeds. Our results provide some insights for finding materials and genes relevant for breeding new varieties with ideal GSL compositions and levels.

Glucosinolate-producing plants have long been recognized for both their distinctive benefits to human nutrition and their resistance traits against pathogens and herbivores. Despite the accumulation of glucosinolates (GLS) in plants is associated with their resistance to various biotic and abiotic stresses, the defensive and biological activities of GLS are commonly conveyed by their metabolic products. In view of this, metabolism is considered the driving factor upon the interactions of GLS-producing plants with other organisms, also influenced by plant and plant attacking or digesting organism characteristics. Several microbial pathogens and insects have evolved the capacity to detoxify GLS-hydrolysis products or inhibit their formation via different means, highlighting the relevance of their metabolic abilities for the plants' defense system activation and target organism detoxification. Strikingly, some bacteria, fungi and insects can likewise produce their own myrosinase (MYR)-like enzymes in one of the most important adaptation strategies against the GLS-MYR plant defense system. Knowledge of GLS metabolic pathways in herbivores and pathogens can impact plant protection efforts and may be harnessed upon for genetically modified plants that are more resistant to predators. In humans, the interest in the implementation of GLS in diets for the prevention of chronic diseases has grown substantially. However, the efficiency of such approaches is dependent on GLS bioavailability and metabolism, which largely involves the human gut microbiome. Among GLS-hydrolytic products, isothiocyanates (ITC) have shown exceptional properties as chemical plant defense agents against herbivores and pathogens, along with their health-promoting benefits in humans, at least if consumed in reasonable amounts. Deciphering GLS metabolic pathways provides critical information for catalyzing all types of GLS towards the generation of ITCs as the biologically most active metabolites. This review provides an overview on contrasting metabolic pathways in plants, bacteria, fungi, insects and humans towards GLS activation or detoxification. Further, suggestions for the preparation of GLS containing plants with improved health benefits are presented., (Copyright © 2021 Elsevier Inc. All rights reserved.)

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., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

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., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Sauerkraut juice has not gained much scientific attention to date. Therefore, this study aimed to track changes in glucosinolates (GLS) during fermentation of white cabbage and the formation of corresponding breakdown products in sauerkraut and sauerkraut juice separately and to evaluate their stability during prolonged storage of the final products. The results obtained indicate that both sauerkraut and sauerkraut juice are a good source of bioactive compounds: ascorbigen and isothiocyanates. The stability of individual compounds during storage varied, and the absolute content of phytochemicals depended on the content of native GLS in the raw material and its bacterial composition. The dominant compound was ascorbigen, stable in acidic pH in both sauerkraut and sauerkraut juice, even after prolonged storage. Sauerkraut juice was also found as a rich source of bioactive isothiocyanates. One 250 mL glass of sauerkraut juice (after two weeks) can deliver approx. 75 μmol of bioactive ascorbien and isothiocyanates, hence it can be considered as functional food, delivering beneficial health effects., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Plant-derived antimalarials are indispensable for malaria treatment and a platform for new drugs. The present study explores sinigrin, for malaria using in vitro, in silico and in vivo strategies and the immune response generated after administration. The compound exhibited promising activity against chloroquine (CQ)-resistant (RKL-9) IC 50 5.14 μg/mL and CQ-sensitive (3D7) IC 50 5.47 μg/mL strains of P. falciparum and was safe in both in vitro (CC 50  > 640 μg/mL) and in vivo (LD 50  > 2 g/kg) toxicity studies. In addition, virtual screening showed hydrogen bonding, hydrophobic and van der Waals interactions with amino acid residues of 3BPM (falcipain-3). In vivo studies revealed promising antimalarial activity of sinigrin (200 mg/kg) with 87.44% chemo-suppression on day 5 and significantly (p < 0.0001) enhanced the mean survival time (21 ± 4.74 days) in contrast to the infected control (5.4 ± 1.14 days). In combination therapy, sinigrin (100 mg/kg and 200 mg/kg) augmented the efficacy of artesunate (AS 50 mg/kg) with 100% survival and no recrudescence. These observations are further corresponded and supported by DLC, NO production, cytokine analysis, biochemical and histopathological studies. Treatment with the combination resulted in a regulated interplay of immune cells and cytokines aiding in parasite clearance in addition to its specific inhibitory activity. We report the antimalarial activity of sinigrin first time with best D-score against falcipain-3. These findings highlight sinigrin as a HIT molecule, which may potentially be used in drug and vaccine development approaches., (Copyright © 2021 Elsevier B.V. All rights reserved.)

This study evaluated the effects of CaCl 2 -HCl electrolyzed water (CHEW) with different calcium chloride concentrations on broccoli sprouts. CHEW treatment reduced the malondialdehyde (MDA) and H 2 O 2 contents of broccoli sprouts. The results showed that 10 kinds of glucosinolates were detected, and glucoraphanin was the dominant component. After hydrolysis, three kinds of isothiocyanates and two kinds of nitriles were detected in broccoli sprouts; however, the corresponding nitrile 4-isothiocyanato-1-butene was not detectable. The sulforaphane content of broccoli sprouts in the 10CHEW (Electrolyte of 10 mM CaCl 2 acidic solution) treatment increased by 34.4%, and the content of sulforaphane nitrile decreased by 53.3% compared with that of the tap water treatment. CHEW changed the metabolism of glucosinolates in broccoli sprouts by promoting the synthesis of glucoraphanin, increasing the activity of myrosinase and decreasing the activity of epithiospecifier protein (ESP) for the generation of more bioactive isothiocyanates. In addition, compared to the tap water treatment, the calcium content in broccoli sprouts treated with 25CHEW (Electrolyte of 25 mM CaCl 2 acidic solution) was dramatically enhanced from 15.8 to 49.7 mg/g DW. CHEW can be a useful tool for enhancing the amount of secondary metabolites and calcium content in broccoli sprouts intended for fresh consumption as a functional food., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Glucosinolates (GSLs), secondary metabolites synthesized by cruciferous plants, can be hydrolyzed by myrosinase into compounds, such as isothiocyanates (ITCs), with various bioactivities. Thus, myrosinase plays an important role in the utilization of GSLs. We isolated a bacterial strain, which was identified as Leclercia adecarboxylata , from the rhizosphere soil of rape seedlings and identified two myrosinase genes and an ITC hydrolase gene. Both myrosinases are intracellular and have 658 amino acid residues. Via molecular docking and chemical modification assays investigating the active sites of the myrosinases, arginine was found to be essential for their catalytic activity. Transcriptomic analysis of the response to sinigrin revealed significant up-regulation of some genes involved in allyl-ITC detoxification, with metallo-β-lactamase 3836 having the highest fold change. Thus, we discovered two myrosinases from L. adecarboxylata and demonstrated that the mechanism of tolerance of the bacterium to allyl-ITC likely involved metallo-β-lactamase activity.

The invasive pest whitefly (Bemisia tabaci) is a complex species, of which Middle East-Minor Asia 1 (MEAM1) and Mediterranean (MED) are the two most damaging members. Previous research showed that cabbage is frequently infested with MEAM1 but seldomly with MED, and this difference in performance is associated with glucosinolate (GS) content. Some insects can modify GS using glucosinolate sulfatase (SULF), the activity of which is regulated by sulfatase modifying factor 1 (SUMF1); therefore, to increase our understanding of different performances of MEAM1 and MED on cabbage plants, we identified and compared nine putative SULFs and one SUMF in MEAM1 and MED. We found that the lengths of two genes, BtSulf2 and BtSulf4, differed between MEAM1 and MED. The messenger RNA levels of BtSulf4 increased more than 20-fold after MEAM1 and MED adults were exposed to GS, but BtSulf2 expression was only induced by GS in MEAM1. Knockdown of BtSulf2 and BtSulf4 in MEAM1 resulted in a substantial increase in the mortality of GS-treated adults but not in MED. These results indicate that differences in BtSulf2 and BtSulf4 sequences and/or expression may explain why MEAM1 performs better than MED on cabbage. Our results provide a basis for future functional research on SULF and SUMF in B. tabaci., (© 2020 Institute of Zoology, Chinese Academy of Sciences.)

Allopolyploid Brassica juncea crops in Brassicaceae are becoming increasingly revitalized as vegetables and oilseeds owing to wide adaptability and significant economic values. However, the genomic differentiation of diversified vegetables and oilseed B. juncea and the genetic basis underlying glucosinolates accumulation have yet to be elucidated. To address this knowledge gap, we report the sequencing of pairwise genomes of vegetable and oilseed B. juncea at chromosome scale. Comparative genomics analysis unveils panoramic structural variation footprints, particularly the genetic loci of HSP20 and TGA1 associated with abiotic and biotic stresses responses between oilseed and vegetable subgroups. We anchored two major loci of MYB28 (HAG1) orthologues caused by copy number variations on A02 and A09 chromosomes using scored genomic SNPs-based GWAS that are responsible for seed oil quality-determining glucosinolates biosynthesis. These findings will provide valuable repertories of polyploidy genomic information enabling polyploidy genome evolution studies and precise genomic selections for crucial traits like functional components of glucosinolates in B. juncea crops and beyond., (© 2021 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.)

Rapeseed contains high levels of glucosinolates (GSLs), playing pivotal roles in defense against herbivores and pests. As their presence in rapeseed reduces the value of the meal for animal feeding, intensive efforts to reduce them produced low-seed GSL cultivars. However, there is no such variety suitable for the south part of Japan. Here, we tested the effects of cold oxygen plasma (oxygen CP) on seed germination and GSL and lipid content, in 3 rapeseed cultivars. According to the cultivars, oxygen CP slightly stimulated seed germination and modified the GSL levels, and decreased GSL levels in Kizakinonatane but increased those in Nanashikibu. In contrast, it negligibly affected the lipid content and composition in the 3 cultivars. Thus, oxygen CP modulated seed GSL levels without affecting seed viability and lipid content. Future optimization of this technique may help optimize rapeseed GSL content without plant breeding., (© The Author(s) 2021. Published by Oxford University Press on behalf of Japan Society for Bioscience, Biotechnology, and Agrochemistry.)

Glucoraphanin (GRA) is found in the seeds and vegetative organs of broccoli ( Brassica oleracea L. var. italica Planch) as the precursor of anti-carcinogen sulforaphane (SF). The yield of GRA obtained from these materials is weak and the cost is high. In recent years, the production of plant secondary metabolites by large-scale hairy roots culture in vitro has succeeded in some species. Melatonin (MT) is a natural hormone which existed in numerous organisms. Studies have demonstrated that MT can improve the synthesis of secondary metabolites in plants. At present, it has not been reported that MT regulates the biosynthesis of glucoraphanin in broccoli hairy roots. In this study, the broccoli hairy roots that grew for 20 d were respectively treated by 500 µM MT for 0, 6, 12, 20 and 32. To explore the reason of changes in secondary metabolites and reveal the biosynthetic pathway of glucoraphanin at transcriptional level. Compared with 0 h, the yield of GRA under other treatments was increased, and the overall trend was firstly increased and then decreased. The total yield of GRA reached the highest at 12 h, which was 1.22-fold of 0 h. Then, the genome of broccoli as the reference, a total of 13234 differentially expressed genes (DEGs) were identified in broccoli hairy roots under treatment with 500 µM MT for 0, 6, 12, 20 and 32 h, respectively. Among these DEGs, 6266 (47.35%) were upregulated and 6968 (52.65%) were downregulated. It was found that the pathway of 'Glucosinolates biosynthesis (ko00966)' was enriched in the 16th place by Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis of the upregulated DEGs. The expression of key genes in the GRA biosynthesis pathway was upregulated at all time points, and a deduced GRA biosynthesis pathway map was constructed for reference.

Indole-3-acetaldoxime (IAOx) and phenylacetaldoxime (PAOx) are precursors for the growth hormones indole-3-acetic acid (IAA) and phenylacetic acid (PAA) and the defense compounds glucosinolates in Brassicales. Our recent work has shown that Arabidopsis transgenic lines overexpressing AtCYP79A2, a PAOx-production enzyme, accumulate the PAOx-derived compounds benzyl glucosinolate and PAA. Here we report that they also accumulate the benzyl glucosinolate hydrolysis products benzyl isothiocyanate and benzyl cyanide, which indicates that the turnover of benzyl glucosinolate can occur in intact tissues. Myrosinases or β-glucosidases are known to catalyze glucosinolate breakdown. However, transcriptomics analysis detected no substantial increase in expression of known myrosinases or putative β-glucosidases in AtCYP79A2 overexpressing lines. It was previously shown that accumulation of aldoximes or their derivatives represses the phenylpropanoid pathway. For instance, ref2 mutant having a defect in one of the aldoxime catabolic enzymes decreases phenylpropanoid production. Considering that AtCYP79A2 is not expressed in most organs under optimal growth condition, ref2 accumulates aliphatic aldoximes but not PAOx. Interestingly, overexpression of AtCYP79A2 in ref2 resulted in a further decrease in sinapoylmalate content compared to ref2 . This indicates that accumulation of PAOx has an additive effect on phenylpropanoid pathway suppression mediated by other aldoximes.