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Tomato crops are affected in Mediterranean cold-greenhouse agrosystems by soilborne diseases, such as root-knot nematodes (Meloidogyne spp.), which represent a serious problem leading to losses in production. Agroecological soil management based on biocontrol agents and natural compounds has had increased grower interest in order to reduce chemical residues in the produce and to adopt environmentally friendly farming methods. In this frame, we evaluate and validate soil biofumigation by the use of glucosinolate (GLS) compounds. Among them, sinigrin showed biocontrol activities against several pests and diseases via nematotoxic action. Among the Brassicaceae species rich in sinigrin, we chose Brassica macrocarpa Guss. (BM) because its leaves show 90% of all GLSs, and we could better estimate the action of this single GLS. Different dosages of BM leaf flour, containing 200 to 300, 350, 400, 450, and 650 &mu, mol m&minus, 2 of sinigrin, were inserted into soil already infected by Meloidogyne spp. for evaluating their effects on tomatoes grown in cold greenhouses in comparison to absolute control (CTRL) and to the chemical one, Vydate 5G&reg, (CCTRL). The root disease index, caused by nematode attack, was the highest in CTRL, and a reduction of about 50% was observed with the 300 to 650 &mu, 2 sinigrin dosage. The CCTRL showed twofold marketable yield increase, and a fourfold increase was found in 650 &mu, 2 of sinigrin dosage, in comparison to the CTRL. Biofumigant applications improved tomato plant growth and development, and fruit quality, significantly for dry matter and soluble sugars (&deg, Brix). BM leaf flour inserted into the soil, at a dose of 300 &mu, 2 of sinigrin, showed similar effects to the CCTRL on root disease index, root weight, and marketable yield. Data showed the nematotoxic effect of sinigrin for the biocontrol of Meloydogine spp. by the use of B. macrocarpa leaves, very rich in this GLS compound, which represents a new tool for agroecological soil management and for organic farming.

Chinese cabbage (Brassica rapa) is a perennial crucifer vegetable that has long been used for forage. Crucifers are rich sources of glucosinolates (GSLs), which are anti-carcinogenic in humans and involved in plant defense responses. Myeloblastosis (MYB) proteins are a large family of transcription factors (TFs) in plants and play major regulatory roles in many biological processes. We identified 14 functional R2R3-MYB genes involved in glucosinolate biosynthesis in B. rapa ssp. pekinensis. Bioinformatic analysis of their phylogeny, protein motifs, gene interaction network, and molecular characteristics showed that Chinese cabbage MYB genes are comparable to those of Arabidopsis thaliana. The expression levels of the 14 BrMYB genes under fluorescent lamp, blue, and red light were quantitated using qRT-PCR analysis. Almost all of the R2R3-BrMYBs were upregulated and expressed more under red light than under fluorescent lamp or blue light, except BrMYB34s. We also calculated the total GSLs under each light condition. The total GSL content was higher under red light than under fluorescent lamp or blue light. Furthermore, the individual glucosinolates, comprised of four aliphatic GSLs (progoitrin, sinigrin, gluconapin, and glucobrassicanapin) and one indolic GSL (glucobrassicin), were higher under red light than the other light conditions. The relationships between light quality and glucosinolate biosynthesis require further investigation.

Nutritional value and disease-preventive effects of cabbage are well-known. Levels of the antioxidant compounds ascorbic acid (AA) and glucosinolates (GSL) in new Czech cabbage cultivars were determined in the context of different production systems. The contents of AA and GSLs in cabbage biomass were determined by HPLC. Individual GSLs were identified according to their exact masses with sinigrin used as the external standard. Artificial infection with A. brassicicola generally raised the AA levels. The major GSLs (&ge, 10 mg kg&minus, 1) were glucobrassicin, sinigrin, and glucoiberin. Indole and aliphatic GSLs were present, but no aromatic ones were detected. Ecological growth conditions and the artificial fungal infection increased the total content of GSLs and, also, of the methoxylated indole GSLs. Sulforaphane, iberin, indole-3-carbinol, and ascorbigen resulting from the hydrolysis of GSLs were found in both cultivars. The amounts and profiles of GSLs present in the two Czech cultivars demonstrated their good nutritional value. The decomposition products sulforaphane, iberin, indole-3-carbinol, and ascorbigen detected improve its health-promoting qualities and represent a suitable component of the human diet.

Glucosinolates (GSLs) and phenolic compounds (PCs) are biologically active and involved in the defense reaction of plants; these compounds have a beneficial effect on human health. In this study, we described the influence of biologically synthesized silver nanoparticles (Ag NPs) to enhance the phytochemicals (GSLs and PCs), their transcription levels, and their biological activities in genetically transformed root cultures (hairy root cultures) of Brassica rapa. The concentrations of silver and reactive oxygen species (malondialdehyde and hydrogen peroxide) were highly elevated in the Ag NP-elicited hairy roots (HRs). Glucosinolates (glucoallysin, glucobrassicanapin, sinigrin, progoitrin, gluconapin, 4-methoxyglucobrassicin, 4-hydroxyglucobrassicin, glucobrassicin, neoglucobrassicin, and gluconasturtiin) and their transcripts (MYB34, MYB51, MYB28, and MYB29) were significantly enhanced in the Ag NP-elicited HRs. Moreover, the phenolic compounds (flavonols, hydroxybenzoic, and hydroxycinnamic acids) were significantly enriched in the Ag NP-elicited HRs. Total phenolic and flavonoid concentrations and their transcripts (PAL, CHI, and FLS) were higher in the Ag NP-elicited HRs than in the non-elicited HRs. Additionally, biological (antioxidant, antimicrobial, and anticancer) activities were significantly higher in the Ag NP-elicited HRs than in the non-elicited HRs. The Ag NP-elicited HR cultures offered an efficient and promising in vitro method to increase the production of health-promoting bioactive compounds, which may be useful in nutraceutical and pharmaceutical industries.

Biofumigation is an integrated pest-management method involving the mulching of a glucosinolate-containing cover crop into a field in order to generate toxic isothiocyanates (ITCs), which are effective soil-borne-pest-control compounds. Variation in biofumigation efficacy demonstrates a need to better understand the factors affecting pest-control outcomes and develop best practices for choosing biofumigants, growth conditions, and mulching methods that allow the greatest potential isothiocyanate release. We measured the glucosinolate concentrations of six different commercial varieties of three biofumigant plant species: Brassica juncea (ISCI99, Vitasso, and Scala) Raphanus sativus (Diablo and Bento), and Sinapis alba (Ida Gold). The plants were grown in the range of commercially appropriate seeding rates and sampled at three growth stages (early development, mature, and 50% flowering). Within biofumigant species, the highest ITC-release potentials were achieved with B. juncea cv. ISCI99 and R. sativus cv. Bento. The highest ITC-release potential occurred at the 50% flowering growth stage across the species. The seeding rate had a minor impact on the ITC-release potential of R. sativus but had no significant effects on the ITC-release potentials of the B. juncea or S. alba cultivars.

A plant factory is a closed cultivation system that provides a consistent and modified environment for plant growth. We speculated that treatment of kale (Brassica oleracea) grown in a plant factory with NaCl, Na2SeO3, or both would increase the bioactive phytochemical levels including glucosinolates (GLSs) and isothiocyanates (ITCs), the key molecules in cancer prevention. The kale was harvested and analysed after treatment with NaCl and Na2SeO3 alone or in combination for 1 or 2 weeks. Exposure to NaCl alone but not Na2SeO3 increased plant root growth. Levels of sinigrin were increased by a 2-week exposure to Na2SeO3 alone or in combination with NaCl, whereas no changes were observed in glucoraphanin and gluconasturtiin gluconasturtiin levels. Importantly, the ITC concentration was affected by 2-week treatment with both compounds. To evaluate the bioactivity of kale, HepG2 human hepatoma cells were treated with plant extract for 6 h. Only the extract of kale roots exposed to a combination NaCl and Na2SeO3 for 2 weeks showed an increased expression of nuclear factor erythroid 2-related factor (Nrf2), which regulates genes encoding antioxidant proteins. These data suggest that co-treatment with NaCl and Na2SeO3 increased the ITC content and chemopreventive effects of kale root.

Vegetable crops of Brassica oleracea contain significant amounts of antioxidant compounds, such as glucosinolates (GLS) which, upon disruption of the tissue, are broken down by the endogenous enzyme myrosinase (MYR), resulting in production of isothiocyanates, thiocyanates, nitriles, goitrin and epithionitriles. In particular, isothiocyanates are known to exert beneficial effects for human health. Several studies describe isothiocyanates (ITCs) as inhibitors of mitosis and stimulators of apoptosis in human tumor cells. The Sicilian black broccoli is a neglected crop of Mt. Etna towns showing a particular GLS profile mainly represented by GRA and NGBS and their derivatives. We investigated the effects of different concentrations (0-25 μM) of various GLS: gluconasturtiin (GNT), glucoraphanin (GRA), progoitrin (PRO), sinigrin (SIN), glucoiberin (GIB), gluconapin (GNA), sinalbin (GSI), glucobrassicin (GBS), glucobrassicanapin (GBN) and glucoerucin (GER) on the growth of HT29 (colon cancer) and A2058 (melanoma cancer) cells, after 24 h of treatment, in presence or not of MYR (0.018 U, in situ method). The same cell lines were also exposed to the Sicilian black broccoli juice extract (from leaves and stems) at different amount (0.05, 0.1, 0.5, 1, and 5%), with and without MYR. Cellular viability was evaluated by MTT assay. GLS, alone and in presence of MYR, elicited different grades of toxicity in both cell lines models. GRA and GNT showed less toxicity than their breakdown products and opposite results were obtained with GBS and GBN. Black broccoli juice showed to be less toxic in presence of MYR; this effect is particular evident at higher juice percentages (1 and 5%), corresponding to 5 and 25 µM, respectively of GRA and NGBS. Certainly, juice extract activity on in vitro cell lines cannot only be explained by the peculiar GLS profile; probably, the interaction of others compounds, such as polyphenols, have to be taken into account. In conclusion, not all breakdown GLS products show more toxicity than their precursors, at least in the in vitro cell cancer models we utilized.

The effect of different doses of supplemental ultraviolet-B (UV-B) illumination on yield, plant growth, biochemical changes and antioxidant activity of broccoli florets was evaluated. The broccoli plants were grown under three different supplemental UV-B illumination doses (2.2, 8.8 and 16.4 kJ m−2 d−1) in the glasshouse. Plant height decreased with increasing supplementary UV-B illumination dose. However, leaf thickness increased with increasing UV-B dose. Chlorophyll content in the leaves also increased during the growing period. The lowest chlorophyll content was found at 16.4 kJ m−2 d−1 UV-B dose. Total yield decreased with supplemental illumination, especially at 16.4 kJ m−2 d−1 illuminated dose. Total dry matter, total soluble solids, carotenoids, chlorophyll a and b content in broccoli florets decreased with increasing UV-B illumination dose. Conversley, ascorbic acid, sinigrin, total phenolic, and flavonoid content and antioxidant activity increased in UV-B illuminated florets. Surprisingly, glucotropaeolin content, one of the forms of glucosinolate in broccoli, was not affected significantly by UV-B illumination or enhanced by UV-B illumination doses.

Soilless cultivation systems (SCS) are increasingly used to produce high quality baby-leaf arugula, appreciated by consumers for its pungent taste, due to the content of glucosinolates (GLS). Given all of the health benefits attributed to GLSs, there is great interest in understanding whether and how soilless growing systems may affect the GLS profile of arugula. For this purpose, a study was conducted to compare the GLS profile of Diplotaxis erucoides, D. tenuifolia, and Eruca sativa grown side-by-side under protected environment in a conventional soil-system (CSS) and in a SCS. Genotype and growing-system resulted in a significant effect on GLS content, while the resulting qualitative GLS profile was species-specific. Despite the growing-system, six different GLSs were identified from plants of D. tenuifolia and E. sativa, while only sinigrin was detected in plants of D. erucoides. Total GLS content was on average 9.85, 8.23, and 7.96 mg g−1 of dry weight in D. erucoides, D. tenuifolia, and E. sativa, grown in CSS, respectively. The same three species grown in SCS synthesized 36%, 51%, and 41% more GLSs than plants grown in CSS, respectively. Soilless cultivation may be adopted to increase the GLS content and enhance the nutritional quality of brassicas leafy vegetables.

International audience; Glucosinolates are sulfur-containing secondary metabolites found in plants of the Brassicale order. They are precursors of isothiocyanate species, resulting from C–S hydrolysis catalysed by the thioglucohydrolase myrosinase. We describe the synthesis of bifunctional glucosinolate–mannoside glycoconjugates combining both the structural features of a substrate of myrosinase and a ligand of the lectin FimH. We show that these glycoconjugates serve as enzyme substrates and that myrosinase can indeed hydrolyze the glucosinolate moiety with affinities (KM, Vmax) comparable to the natural substrates glucomoringin and sinigrin. This enzymatic hydrolysis of the thioglycosidic bond led to the efficient formation of an isothiocyanate which was assessed by the formation of the corresponding dithiocarbamate derivatives. Finally, we show that our synthetic bifunctional glycoconjugates also serve as FimH ligands where the glucosinolate moiety does not hamper the interaction with the lectin. Our findings set the stage for an original bioconjugation tool, allowing for myrosinase-triggered specific labelling of lectins using glucosinolate glycoconjugates as non-toxic, water soluble isothiocyanate precursors.

Adipocyte differentiation is a critical adaptive response to nutritional overload and affects the metabolic outcome of obesity. Sinigrin (2-propenyl glucosinolate) is a glucosinolate belong to the glucoside contained in broccoli, brussels sprouts, and black mustard seeds. We investigated the effects of sinigrin on adipogenesis in 3T3-L1 preadipocytes and its underlying mechanisms. Sinigrin remarkably inhibited the accumulation of lipid droplets and adipogenesis by downregulating the expression of CCAAT-enhancer-binding protein α (C/EBPα), peroxisome proliferator-activated receptor gamma (PPARγ), leptin and aP2. Sinigrin arrested cells in the G0/G1 phase of the cell cycle and increased the expression of p21 and p27. CDK2 expression was suppressed by sinigirn in MDI-induced adipocytes. Sinigrin increased the phosphorylation of adenosine monophosphate-activated protein kinase (AMPK), mitogen-activated protein kinase (MAPK) and acetyl-CoA carboxylase (ACC) in the early stage of adipocyte differentiation, suggesting that sinigrin has anti-adipogenic effects through AMPK, MAPK and ACC activation. Sinigrin also inhibited the production of pro-inflammatory cytokines including tumor necrosis factor -alpha (TNF-α) and interleukin (IL)-6, IL-1β and IL-18. Taken together, these data suggest that sinigrin inhibits early-stage adipogenesis of 3T3-L1 adipocytes through the AMPK and MAPK signaling pathways.

The goal of our present research was to develop a simple and rapid method for the quantitation of desulfoglucosinolates (desulfoGLS) without using column chromatography. The proposed method involves extraction, concentration, incubation of glucosinolates with a sulfatase enzyme, and HPLC analysis. Identification of desulfoGLS in green kohlrabi was performed by LC-HR-ESI-QTOF-MS in positive-ionization mode. A total of 11 desulfoGLS were identified with neoglucobrassicin (3.32 ± 0.05 μmol/g DW) as the predominant indolyl, whereas progoitrin and sinigrin were the major aliphatic desulfoGLS. The levels of the aliphatic desulfoGLS glucoiberin, progoitrin, and glucoerucin at 7 h were found to be 3.6-, 1.9-, and 1.6-fold higher, respectively, than those produced through the conventional method. This technique was successfully applied in the identification of desulfoGLS from cabbage. The developed method has fewer unit operations, has maximum recovery, and is reproducible in the determination of desulfoGLS in a large number of Brassicaceae samples in a short time.

Department of Bio-Environmental Chemistry, Chungnam National University, Daejeon 305-764, KoreaReceived on 8 May 2015, revised on 10 August 2015, accepted on 19 August 2015Abstract : The present study aimed to investigate the effects of drought stress on the accumulation of glucosinolates (GSLs) in the leaves of Kale cultivated in autumn and spring. HPLC analysis guided to identify seven GSLs including progoitrin, glucoraphanin, sinigrin, gluconapin, glucobrassicin, 4-methoxyglucobrassicin and neoglucobrasscin. Quantification of GSLs revealed that the contents of sigirin was the highest (45%) followed by the level of progoitrin (24%) in terms of total GSLs. The ranges of total GSL contents was 1.16 (84)-15.88 (89 DAS, µmol/g dry wt. (DW)) in treatment plot and 1.23 (84)-7.05 (74 DAS, µmol/g dry wt.) in control plot showed the enhancement in the contents of GSLs in treatment than in the control plot. The present results evidenced that the variation of total GSL contents were depending on the harvest period. In 105 DAS, comparatively no differences in the GSL contents on each sample in autumn season, whereas in spring season, although there was decrease in the GSLs tendency from 74 DAS to 84 DAS in both control and treatment plot, the GSL contents of treatment plot was dramatically increased in 89 DAS. In treatment plot, the GSL contents on 89 DAS (1.16) was 15 fold higher to 84 DAS (15.88 µmol/g DW). The variation in the contents of GSL in spring and autumn did not documented significant differences because of their differences in the growth time and cultivation conditions. In conclusion, the GSL contents in kale was likely to be affected by drought stress treatment. Scrutiny and further research for exact relation between drought stress and GSL contents in kale should be needed.Key words : Kale, Brassica crops, Glucosinolates, Drought-stress, HPLC analysis*Corresponding author: Tel: +82-42-821-6738E-mail address: kimsunju@cnu.ac.kr

In some plant-insect interactions, specialist herbivores exploit the chemical defenses of their food plant to their own advantage. Brassica plants produce glucosinolates that are broken down into defensive toxins when tissue is damaged, but the specialist aphid, Brevicoryne brassicae, uses these chemicals against its own natural enemies by becoming a "walking mustard-oil bomb". Analysis of glucosinolate concentrations in plant tissue and associated aphid colonies reveals that not only do aphids sequester glucosinolates, but they do so selectively. Aphids specifically accumulate sinigrin to high concentrations while preferentially excreting a structurally similar glucosinolate, progoitrin. Surveys of aphid infestation in wild populations of Brassica oleracea show that this pattern of sequestration and excretion maps onto host plant use. The probability of aphid infestation decreases with increasing concentrations of progoitrin in plants. Brassica brassicae, therefore, appear to select among food plants according to plant secondary metabolite profiles, and selectively store only some compounds that are used against their own enemies. The results demonstrate chemical and behavioral mechanisms that help to explain evidence of geographic patterns and evolutionary dynamics in Brassica-aphid interactions.

The aims of this study was to optimize the extraction conditions of sinigrin from Dolsan leaf mustard. Dolsan leaf mustard (Dolsan-eup, Yeosu-si) harvested during at May 2014 was used for sinigrin extraction. After the extraction of sinigrin using 50% CH3CN, 10% NH4Cl, 60% CH2OH, and 70% CH3OH, the sinigrin content was measured by HPLC analysis. The results showed that sinigrin content was highest with 50% CH3CN solvent extraction and UV detector sensitivity was greater at 228 nm rather than at 242 nm. The sinigrin concentrations of leaf, stem and root with 50% CH3CN extraction were 345 ppm, 728 ppm, and 539 ppm, respectively. After extraction of the different parts of Dolsan leaf mustard, The standard retention time by HPLC analysis of sinigrin content was 2.054, 2.032, 2.059, and 2.035 min from the root, stalk, and leaf, respectively. On the other hand, HPLC analysis showed that the leaf extracts contained glucoraphanin, one of glucosinolates. The optimum time and extraction solvent for the sinigrin extraction from Dolsan leaf mustard was found to be 24 hr with 50% CH3CN solvent. In addition, opotimum UV detector k at 228 nm. These results showed that the optimum extraction conditions for Dolsan leaf mustard were 24 hr extraction with 50% CH3CN solvent. In addition, the optimum wavelength of UV detector was determined to be 228 nm for sinigrin analysis. Therefore, this study could provide a useful information for sinigrin extraction and its systematic analysis during the storage.

The grubs of Melolontha melolontha and Melolontha hippocastani damage the roots of many plants and are therefore considered serious pests in agriculture, horticulture and forestry in central and eastern Europe. Since the implementation of legislative regulations banning the use of synthetic insecticides against these pests in soil, alternative methods have gained high priority. The plants of the family Brassicaceae have recently received much attention due to the high content of glucosinolates (GSLs) in their tissues. The GSL breakdown products are highly biocidal in relation to many soil-borne pests. We tested Brassica juncea granulated seed meal against Melolontha grubs under semi-natural (the concentration-response experiments) and field conditions. The efficacy and the indirect lethal concentrations of GSLs that were necessary to obtain 50% and 95% grub mortality (ILC50 and ILC95, respectively) were estimated. The concentration-response experiments revealed that the ILC50 of GSLs dominated by sinigrin (97%) for the L1, L2 and L3 grubs was 118.4, 167.1 and 173.5 μmol/L, respectively, and the ILC95 was 293.3, 312.7 and 401.7 μmol/L, respectively. Under field conditions, the mortality of the L3 grubs in the plots that were treated with granulated B. juncea seed meal at ILC99.8 was almost twice as high (82.2%) as that of the grubs in the untreated plots (45.4%), and the changes in the grub density with time in these plots were significantly different. The results of our experiments demonstrate the high potential of soil biofumigation using B. juncea granulated seed meal to control Melolontha grubs.

The glucosinolates content of brassica plants is a distinctive characteristic, representing a healthy advan- tage as many of these compounds are associated to antioxidant and anti-carcinogenic properties. Brassica sprouts are still an underutilized source of these bioactive compounds. In this work, four varieties of bras- sica sprouts (red cabbage, broccoli, Galega kale and Penca cabbage), including two local varieties from the North of Portugal, were grown to evaluate the glucosinolate profile and myrosinase activity during the sprouting. Also the influence of light/darkness exposure during sprouting on the glucosinolate content was assessed. Glucosinolate content and myrosinase activity of the sprouts was evaluated by HPLC meth- ods. All sprouts revealed a higher content of aliphatic glucosinolates than of indole glucosinolates, con- trary to the profile described for most of brassica mature plants. Galega kale sprouts had the highest glucosinolate content, mainly sinigrin and glucoiberin, which are recognized for their beneficial health effects. Penca cabbage sprouts were particularly richer in glucoraphanin, who was also one of the major compounds in broccoli sprouts. Red cabbage showed a higher content of progoitrin. Regarding myrosi- nase activity, Galega kale sprouts showed the highest values, revealing that the use of light/dark cycles and a sprouting phase of 7?9 days could be beneficial to preserve the glucosinolate content of this variety. B514-8F2F-A1FB | Ana Vale N/A

The purpose of this study was to determine the possibility of using Wasabi japonica Matsum leaves as natural health food source. To accomplish this purpose, the contents of general and antioxidative nutrients of Wasabi japonica Matsum leaves were measured. The contents of carbohydrate, crude protein, crude lipid and ash are 53.41%, 25.00%, 7.95% and 13.64%. And the calories of Wasabi leaves was 385.23 Kcal. Total dietary fiber was 52.27%. The K was the largest mineral followed by Ca, P, Mg which means Wasabi leaves is alkali material. The contents of sinigrin and allyisothiocyanate in the wasabi leaves were 69.2 mg/g and 241.0 mg/g, respectively. Total phenol contents of the hot water extract and the 70% ethanolic extract were and GAE/g, respectively. The total flavonoids content of the hot water extract and the 70% ethanol extract were and QE/g, espectively. The general nutrients and other antioxidant bioactive materials in Wasabi japonica Matsum leaves were also potential materials for good health food.

The antifungal properties of isothiocyanates released from the hydrolysis of the glucosinolate sinigrin were assessed on an Armillaria mellea strain in Petri dishes. The fumigation with sinigrin at 100 µM showed a fungicidal effect, while lower rates determined a temporary fungistatic effect. The application of increasing rates of solid and liquid biofumigant formulations on A. mellea inoculated potted peach plants showed an increase of soil nitrate concentration, basal respiration, microbial biomass, leaf nitrogen and chlorophyll concentration. These effects indicated a stimulating effect of Brassicaceae derivatives on soil biological activity and plant growth. The application of a rate of meal higher than 6.42 g kg−1 soil caused a stunted growth and the death of some plants showing a clear phytotoxic effect of the treatments. The inhibition of A. mellea growth observed in in vitro trials was not evidenced in in vivo experiments due to the lack of infection symptoms in experimentally inoculated potted trees.

Control of Salmonella in poultry is a public health concern as salmonellosis is one of the most common foodborne diseases worldwide. This study aimed to screen the ability of 5 Salmonella serovars to degrade the mustard glucosinolate, sinigrin (by bacterial myrosinase) in Mueller-Hinton broth at 25 °C for 21 d and to reduce Salmonella on fresh chicken breasts by developing an edible 0.2% (w/v) κ-carrageenan/2% (w/v) chitosan-based coating containing Oriental mustard extract, allyl isothiocyanate (AITC), EDTA or their combinations. Individual Salmonella serovars degraded 50.2%-55.9% of the sinigrin present in 21 d. κ-Carrageenan/chitosan-based coatings containing 250 mg Oriental mustard extract/g or 50 μl AITC/g reduced the numbers of Salmonella on chicken breasts 2.3 log10 CFU/g at 21 d at 4 °C. However, when either mustard extract or AITC was combined with 15 mg/g EDTA in κ-carrageenan/chitosan-based coatings, Salmonella numbers were reduced 2.3 log10 CFU/g at 5 d and 3.0 log10 CFU/g at 21 d. Moreover, these treatments reduced numbers of lactic acid bacteria and aerobic bacteria by 2.5-3.3 log10 CFU/g at 21 d. κ-Carrageenan/chitosan coatings containing either 50 μl AITC/g or 250 mg Oriental mustard extract/g plus 15 mg EDTA/g have the potential to reduce Salmonella on raw chicken.

UNLABELLED Nanocomposite carboxymethyl cellulose films containing sinigrin (SNG) were prepared by stirring 2% (w/v) carboxymethyl cellulose (CMC) and 2% (w/v) glycerol (as a plasticizer) in distilled water with or without SNG (an antimicrobial precursor) as a 99% pure reagent (pSNG) or as a crude extract (cSNG). These films plus normal CMC film with or without SNG were tested on Escherichia coli O157:H7- inoculated beef for antimicrobial activity. Beef pieces measuring 6 × 5 × 2 cm(3) (L × W × H) were dipped in an E. coli O157:H7 broth suspension containing >8 log10 CFU ml(-1) and were drained for 3 min over a sterile cloth. They were wrapped in CMC or NCMC films, placed in a high oxygen barrier film (Deli *1), vacuum-packaged and stored at 8°C for 5, 8, 12 and 18 days. The CMC and NCMC films without SNG were not antimicrobial against E. coli O157:H7; however, NCMC and CMC films with SNG were highly antimicrobial. After 5 days at 8°C, E. coli O157:H7 was reduced more than 4 log10 by the NCMC•pSNG film and this reduction remained almost the same until 18 days at 8°C when E. coli O157:H7 was reduced >5 log10 CFU g(-1) meat. SIGNIFICANCE AND IMPACT OF THE STUDY Transparent nanoparticulate carboxymethyl cellulose (CMC) films containing sinigrin (SNG), an antimicrobial precursor, controlled surface contamination of packaged fresh beef by the pathogen Escherichia coli O157:H7 when stored at 8°C. Films with nanoparticulation that carried pure SNG or the naturally occurring SNG in Oriental mustard were significantly more antimicrobial than similar films without nanoparticulation. As films without sinigrin were not antimicrobial, the combinations studied showed that nanoparticulation of the packaging film enhanced delivery of the antimicrobial incorporated within the film.

Aims: This study explores the biofumigation effects of glucosinolate (GSL) containing Brassica oleracea plant material on beneficial, non-target soil organisms, and aims to relate those effects to differences in GSL profiles. Methods: Leaf material of purple sprouting broccoli 'Santee', Savoy cabbage 'Wintessa', and the wild B. oleracea accession Winspit was analysed for GSL production and used for biofumigation experiments on the beneficial soil invertebrates, Folsomia candida (springtail) and Eisenia andrei (earthworm) and the soil bacterial community. Results: When mixed into soil, the Winspit plant material exerted the highest toxic effects on beneficial soil invertebrates by reducing survival and reproduction. Total GSL levels varied substantially between genotypes, in particular the aliphatic GSL (AGSL) sinigrin and gluconapin being highly abundant or exclusively present in Winspit. Differences between the genotypes regarding biofumigation effects on the soil microbial community were only observed on a temporal basis with the largest difference in bacterial community structure after 1 week. Conclusions: The high total GSL content in biofumigated soil could explain the toxicity of Winspit for soil invertebrates. These effects are likely to be the results of high AGSL levels in Winspit. The use of wild B. oleracea crops, such as Winspit, for biofumigation practices would need a proper assessment of the overall impact on soil biota before being applied on a wide scale.

The identification of genetic factors influencing the accumulation of individual glucosinolates in broccoli florets provides novel insight into the regulation of glucosinolate levels in Brassica vegetables and will accelerate the development of vegetables with glucosinolate profiles tailored to promote human health. Quantitative trait loci analysis of glucosinolate (GSL) variability was conducted with a B. oleracea (broccoli) mapping population, saturated with single nucleotide polymorphism markers from a high-density array designed for rapeseed (Brassica napus). In 4 years of analysis, 14 QTLs were associated with the accumulation of aliphatic, indolic, or aromatic GSLs in floret tissue. The accumulation of 3-carbon aliphatic GSLs (2-propenyl and 3-methylsulfinylpropyl) was primarily associated with a single QTL on C05, but common regulation of 4-carbon aliphatic GSLs was not observed. A single locus on C09, associated with up to 40 % of the phenotypic variability of 2-hydroxy-3-butenyl GSL over multiple years, was not associated with the variability of precursor compounds. Similarly, QTLs on C02, C04, and C09 were associated with 4-methylsulfinylbutyl GSL concentration over multiple years but were not significantly associated with downstream compounds. Genome-specific SNP markers were used to identify candidate genes that co-localized to marker intervals and previously sequenced Brassica oleracea BAC clones containing known GSL genes (GSL-ALK, GSL-PRO, and GSL-ELONG) were aligned to the genomic sequence, providing support that at least three of our 14 QTLs likely correspond to previously identified GSL loci. The results demonstrate that previously identified loci do not fully explain GSL variation in broccoli. The identification of additional genetic factors influencing the accumulation of GSL in broccoli florets provides novel insight into the regulation of GSL levels in Brassicaceae and will accelerate development of vegetables with modified or enhanced GSL profiles.

We evaluated the effects of three common cabbage cooking methods (blanching, steaming and microwaving) on glucosinolate and S-methylmethionine (SMM) content and total antioxidant capacity of cabbage leaves. We detected four glucosinolates, including glucoraphanin, sinigrin, glucobrassicin, and 4-methoxyglucobrassicin, by high-pressure liquid chromatography (HPLC). Cabbage contained high levels of SMM (192.85 mg/100 g dry weight), compared to other cruciferous vegetables. Blanching cabbage leaves for one to ten minutes decreased glucosinolate and SMM levels, whereas microwaving or steaming cabbage for 5-10 min preserved glucosinolate and SMM levels. 2,2-Diphenyl-1-picrylhydrazyl (DPPH) and 2-2"-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) radical scavenging activities of cooked cabbage generally decreased as cooking time increased, but microwave cooking had a smaller negative effect on antioxidant activities than blanching or steaming. This study demonstrates that some domestic cooking methods, such as microwaving and steaming, can increase the bioaccessibility of glucosinolates and SMM, highlighting the positive role of cooking on the nutritional qualities of cabbage.

This study was aimed to identify sinigrin-induced bacterial proteins potentially involved in the metabolism of glucosinolate in two glucosinolate-metabolising bacteria Lactobacillus agilis R16 and Escherichia coli VL8. Sinigrin (2 mM) was used to induce the proteins in both bacteria under anaerobic incubation for 8 h at 30 °C for L. agilis R16 and 37 °C for E. coli VL8 and the controls without sinigrin were performed. Allyl isothiocyanate and allyl nitrile as two degradation products of sinigrin were detected in sinigrin-induced cultures of L. agilis R16 (27 % total products) and E. coli VL8 (38 % total products) from a complete sinigrin degradation in 8 h for both bacteria. 2D gel electrophoresis was conducted to identify induced proteins with at least twofold increased abundance. Sinigrin-induced L. agilis R16 and the control produced 1561 and 1543 protein spots, respectively. For E. coli VL8, 1363 spots were detected in sinigrin-induced and 1354 spots in the control. A combination of distinct proteins and upregulated proteins of 32 and 35 spots in L. agilis R16 and E. coli VL8, respectively were detected upon sinigrin induction. Of these, 12 and 16 spots from each bacterium respectively were identified by LC–MS/MS. In both bacteria most of the identified proteins are involved in carbohydrate metabolism, oxidoreduction system and sugar transport while the minority belong to purine metabolism, hydrolysis, and proteolysis. This indicated that sinigrin induction led to the expressions of proteins with similar functions in both bacteria and these proteins may play a role in bacterial glucosinolate metabolism.

Glucosinolates may deter generalist insect feeding as their toxicity causes fitness damage, whereas insects specialized in brassicaceous plants may circumvent the toxic effect. By using no-choice leaf tests, we investigated whether larval development time, body weight, mortality and feeding rate of the generalist Mamestra brassicae (Lepidoptera, Noctuidae) and the specialist Pieris rapae (Lepidoptera, Pieridae), were affected by six genotypes of Brassica oleracea var. acephala, selected for having high or low concentration of sinigrin, glucoiberin (aliphatics) and glucobrassicin (indole). Two phenological plant stages were used. On young plants, M. brassicae most consumed the high sinigrin and low glucoiberin genotypes. Larvae weighed more on the high sinigrin plants. Development time took longer on the low glucoiberin genotype. On mature plants, consumption rate decreased on the high glucoiberin genotype. Larval weight decreased on the high sinigrin, glucoiberin and glucobrassicin genotypes, and development time increased with high glucobrassicin concentration. Pupal weight and mortality rate increased on mature plants, irrespective of the genotype. Pieris rapae fed most on young plants with high sinigrin, and larval weight increased on the high glucoiberin genotype. Mortality increased with low glucoiberin and low glucobrassicin. On mature plants, larval weight decreased with high sinigrin and glucoiberin. The high glucoiberin genotype was the less consumed and also induced a longer development time. High content of aliphatic glucosinolates offered mature plants significant antibiosis defence against both the lepidopterans, whereas the indole glucosinolate was marginally effective. Young plants were more consumed and increased larval weight likely because glucosinolate concentration was still not optimal.

Horseradish (Armoracia rusticana) is a perennial crop and its ground root tissue is used in condiments because of the pungency of the glucosinolate (GS)-hydrolysis products allyl isothiocyanate (AITC) and phenethyl isothiocyanate (PEITC) derived from sinigrin and gluconasturtiin, respectively. Horseradish roots are sold in three grades: U.S. Fancy, U.S. No. 1, and U.S. No. 2 according to the USDA standards. These grading standards are primarily based on root diameter and length. There is little information on whether root grades vary in their phytochemical content or potential health promoting properties. This study measured GS, GS-hydrolysis products, potential anticancer activity (as quinone reductase inducing activity), total phenolic content, and antioxidant activities from different grades of horseradish accessions. U.S. Fancy showed significantly higher sinigrin and AITC concentrations than U.S. No. 1 ,whereas U.S. No. 1 showed significantly higher concentrations of 1-cyano 2,3-epithiopropane, the epithionitrile hydrolysis product of sinigrin, and significantly higher total phenolic concentrations than U.S. Fancy.

4-methylthio-3-butenyl isothiocyanate (4-MTBI) is released from 4-methylthio-3-butenyl glucosinolate (4-MTBG) in radish roots by endogenous myrosinase hydrolysis. In this research, factors influencing the extraction of 4-MTBI from radish roots by an endogenous myrosinase method, such as the time for endogenous myrosinase hydrolysis under 37 °C, the storage time after hydrolysis, were optimized. A new GC method was developed to determine 4-MTBI in radish roots, which was identified by GC-MS and compared with the authentic 4-MTBI. Through optimization of the starting point and increasement of column temperature, both the internal standard allyl isothiocyanate (AITC) and 4-MTBI were clearly separated with a capillary gas chromatography column HP-5. Coefficient of variation (CV) of GC analysis was 3.883%. Recovery percentage of AITC remained 90.3–105.4%. CV of determination and recovery percentage of sinigrin were 5.35% and 91.3–101.9%, respectively.

The objective of this study was to determine the effectiveness of rapeseed meal in controlling soil nematode. Two different rapeseed meals (Jeju local rape varieties and ‘Sunmang’ variety) were mixed with the soil to control nematodes environmentally. When soil physical properties in the rapeseed meal-mixed soils were analy zed, OM (organic matter), P 2 0 5 , Ca, Mg, CEC (Cation Exchange Capacity) value increased. Especia lly, the level of OM was 3-fold higher than control soil. Glucosinolate content of rapeseed meal was higher in Jeju local rape varieties than ‘Sunmang’ variety. The major components of glucosinolates were consisted of progoitrin, gluconapin, glucobrassiaca napin, and sinigrin. These components were likely to be involved in reducing nematode density. Key words - Rapeseed meal, Nematode, Continuous cultivation, Glucosinolate, Soil *교신저자(E-mail): j570510@korea.kr ⓒ 본 학회지의 저작권은 (사)한국자원식물학회지에 있으며, 이의 무단전재나 복제를 금합니다.This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Cauliflower ( Brassica oleraceae L. var. Botrytis ) is a good source of bioactive compounds, such as glucosinolates, phenolic compounds and vitamins. In this study, the effects of some processes ( i.e. boiling, steaming) on the sinigrin bioaccessibility as a major glucosinolate found in cruciferous vegetables after in vitro digestion, also in vitro antimutagenic activities, total phenols and total antioxidant capacities of cauliflower were determined. The sinigrin content was reduced by approximately 9.6% and 29.1% in steamed and boiled cauliflower ( p > 0.05), respectively. After in vitro simulated digestion, sinigrin content was decreased by 26.4% in raw samples, increased by 29.5% and 114.7% in steamed and boiled samples, respectively. In all samples, mutagenic effect to Salmonella typhimurium TA 100 was not seen. When samples were steamed, phenol content was increased by 14.83%. After boiling total phenol content of cauliflower was decreased by 1.8%. Total antioxidant capacities (TAC) measured by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azino-bis(3 ethylbenzothiazoline-6-sulfonic acid) (ABTS) methods were increased by 47% and 39%, respectively ( p p > 0.05). TAC in raw sample of cauliflower, which was investigated in phosphomolybdenum assays, was determined as 18.7 mg ascorbic acid equivalents (AAE)/100 g. In all cases, the highest antioxidant activity was determined in the steamed samples, while the lowest antioxidant activity was in boiled samples.

Field experiment was conducted on clay loam soil during winter season of 2010–2011 and 2011–2012 at the Research Farm, Bidhan Chandra Krishi Viswavidyalaya, West Bengal to study the influence of sulphur (S) levels and irrigation on quality and yield of mustard (cv. Varuna, T-59). Results revealed that yield attributes and yield of crop were highest with 60 kg S/ha, mostly at par with 45 kg S/ha. Double irrigation at flower initiation (30 days after sowing (DAS)) and siliqua development stages (60 DAS) was best with respect to growth, yield attributes, yield, S uptake and oil percent in seed. Effects of both S levels and irrigations on glucosinolate and fatty acid content were non-significant except on progoitrin. The erucic:oleic acid ratio was inversely related to the subsequent increase in S doses, thereby suggesting the qualitative improvement of oil with S application. Oil percent has a negative correlation with sinigrin and gluconapin content. The uptake of S was positively correlated with oleic acid content but showed lower or even negative correlation with other fatty acids. Therefore, irrigation (twice at 30 DAS and 60 DAS) in combination with 45 kg S/ha are recommended for improving yield attributes, yield, oil percent and S uptake of Indian mustard.

Glucosinolates (GSLs) are secondary metabolites found in Brassica vegetables that confer on them resistance against pests and diseases. Both GSLs and glucosinolate hydrolysis products (GHPs) have shown positive effects in reducing soil pathogens. Information about their in vitro biocide effects is scarce, but previous studies have shown sinigrin GSLs and their associated allyl isothiocyanate (AITC) to be soil biocides. The objective of this work was to evaluate the biocide effects of 17 GSLs and GHPs and of leaf methanolic extracts of different GSL-enriched Brassica crops on suppressing in vitro growth of two bacterial (Xanthomonas campestris pv. campestris and Pseudomonas syringae pv. maculicola) and two fungal (Alternaria brassicae and Sclerotinia scletoriorum) Brassica pathogens. GSLs, GHPs, and methanolic leaf extracts inhibited the development of the pathogens tested compared to the control, and the effect was dose dependent. Furthermore, the biocide effects of the different compounds studied were dependent on the species and race of the pathogen. These results indicate that GSLs and their GHPs, as well as extracts of different Brassica species, have potential to inhibit pathogen growth and offer new opportunities to study the use of Brassica crops in biofumigation for the control of multiple diseases.

Factors, including pH, temperature, glucose concentration, and iron compounds, affect the activity of plant myrosinase and, consequently, endogenous glucosinolate degradation. These factors also may affect glucosinolate degradation by bacterial myrosinase. Therefore, this study examined the effect of temperature (4 to 21°C), glucose (0.05 to 1.0%), and iron (10 mM ferrous or ferric) on sinigrin degradation by Salmonella or Listeria monocytogenes cocktails in Mueller-Hinton broth and the effect of sinigrin degradation on bacterial viability. The degradation of sinigrin by both pathogens increased with higher temperatures (21 > 10 > 4°C). Salmonella and L. monocytogenes cocktails hydrolyzed 59.1 and 53.2% of sinigrin, respectively, at 21°C up to 21 days. Both iron compounds significantly enhanced sinigrin degradation by the pathogens. On day 7, sinigrin was not detected when the Salmonella cocktail was cultured with ferrous iron or when the L. monocytogenes cocktail was cultured in Mueller-Hinton broth containing ferric iron. In contrast, ferric and ferrous iron inhibited the activity of 0.002 U/ml myrosinase from white mustard by 63 and 35%, respectively, on day 1. Salmonella and L. monocytogenes cocktails were able to degrade >80% of sinigrin at 0.05 and 0.1% glucose; however, 0.25 to 1.0% glucose significantly reduced sinigrin degradation. Although both pathogens significantly degraded sinigrin, the allyl isothiocyanate (AITC) recoverable was ≤6.2 ppm, which is not inhibitory to Salmonella or L. monocytogenes. It is probable that the gradual hydrolysis of sinigrin to form AITC either did not produce an inhibitory level of AITC or the AITC formed was unstable in the aqueous medium and rapidly decomposed to new compounds that were less bactericidal against the pathogens.

The aim of this study was to assess the feasibility of controlling the potato cyst nematode Globodera pallida through biofumigation with glucosinolate-rich Brassica juncea genotypes. The main glucosinolate of B. juncea is 2-propenyl glucosinolate which is the precursor of 2-propenyl isothiocyanate. Toxicity of 2-propenyl isothiocyanate to encysted G. pallida was tested in vitro . Fifty percent reduction in hatching was found within 2 h of exposure to 0.002% 2-propenyl isothiocyanate. Based on the in vitro results, we hypothesized that biofumigation with B. juncea would reduce hatching of G. pallida in vivo and higher 2-propenyl glucosinolate levels would have a stronger effect. Plant genotype, sulfur fertilization and insect herbivory affected 2-propenyl glucosinolate concentration of B. juncea . However, increasing 2-propenyl glucosinolate concentration of B. juncea did not affect G. pallida hatching after biofumigation. The absence of a biofumigation effect was most likely due to lower concentrations of 2-propenyl isothiocyanate in vivo compared to in vitro conditions. These results show that it is difficult to reach sufficiently high levels of toxicity to reduce hatching of G. pallida under realistic conditions.

Myrosinase (β-thioglucosidase glucohydrolase, EC 3.2.1.147) from broccoli (Brassica oleracea var. italica) was purified by ammonium sulfate precipitation followed by concanavalin A affinity chromatography, with an intermediate dialysis step, resulting in 88% recovery and 1318-fold purification. These are the highest values reported for the purification of any myrosinase. The subunits of broccoli myrosinase have a molecular mass of 50-55 kDa. The native molecular mass of myrosinase was 157 kDa, and accordingly, it is composed of three subunits. The maximum activity was observed at 40 °C and at pH below 5.0. Kinetic assays demonstrated that broccoli myrosinase is subjected to substrate (sinigrin) inhibition. The Michaelis-Menten model, considering substrate inhibition, gave Vmax equal to 0.246 μmol min(-1), Km equal to 0.086 mM, and K(I) equal to 0.368 mM. This is the first study about purification and characterization of broccoli myrosinase.

Mustard is a Brassica vegetable that provides a number of phytonutrients. However, the phytonutrient profile of mustard has been relatively limited. We analyzed the glucosinolates and their hydrolysis products, carotenoids, total anthocyanin and phenolic contents, and antioxidant capacity of the leaves of 11 mustard cultivars grown in a greenhouse at the baby leaf and mature stages. An aliphatic glucosinolate sinigrin and its hydrolysis products allyl isothiocyanate and 1-cyano-2,3-epithiopropane were the major phytonutrients in the mustard leaves. Carotenoids β-carotene, lutein, violaxanthin, and neoxanthin were detected. We found phytonutrient concentration and their change with plant growth were cultivar-dependent. The %RDA value for vitamin A calculated using β-carotene content and retinol activity equivalents suggests that mustard cultivars used in this study can be a good source of vitamin A. Phenolic contents and antioxidant capacity also varied among cultivars and between physiological stages. Our results suggest that mustard leaves are rich in various phytochemicals and their composition depends on cultivar and the physiological stage. This is the first report on phytochemical composition in various mustard cultivars at different physiological stages.

Drought can severely damage crops, resulting in major yield losses. During drought, vascular land plants conserve water via stomatal closure. Each stomate is bordered by a pair of guard cells that shrink in response to drought and the associated hormone abscisic acid (ABA). The activation of complex intracellular signaling networks underlies these responses. Therefore, analysis of guard cell metabolites is fundamental for elucidation of guard cell signaling pathways. Brassica napus is an important oilseed crop for human consumption and biodiesel production. Here, non-targeted metabolomics utilizing gas chromatography mass spectrometry (GC-MS/MS) and liquid chromatography mass spectrometry (LC-MS/MS) were employed for the first time to identify metabolic signatures in response to ABA in B. napus guard cell protoplasts. Metabolome profiling identified 390 distinct metabolites in B. napus guard cells, falling into diverse classes. Of these, 77 metabolites, comprising both primary and secondary metabolites were found to be significantly ABA responsive, including carbohydrates, fatty acids, glucosinolates, and flavonoids. Selected secondary metabolites, sinigrin, quercetin, campesterol, and sitosterol, were confirmed to regulate stomatal closure in Arabidopsis thaliana, B. napus or both species. Information derived from metabolite datasets can provide a blueprint for improvement of water use efficiency and drought tolerance in crops.

Vegetables of the Brassica oleracea group, such as broccoli, cauliflower, and cabbage, play an important role for glucosinolate consumption in the human diet. Upon maceration of the vegetable tissue, glucosinolates are degraded enzymatically to form volatile isothiocyanates, nitriles, and epithionitriles. However, only the uptake of isothiocyanates is linked to the cancer-preventive effects. Thus, it is of great interest to evaluate especially the isothiocyanate formation. Here, we studied the formation of glucosinolates and their respective hydrolysis products in sprouts and fully developed vegetable heads of different genotypes of the five B. oleracea varieties: broccoli, cauliflower as well as white, red, and savoy cabbages. Further, the effect of ontogeny (developmental stages) during the head development on the formation of glucosinolates and their respective hydrolysis products was evaluated at three different developmental stages (mini, fully developed, and over-mature head). Broccoli and red cabbage were mainly rich in 4-(methylsulfinyl)butyl glucosinolate (glucoraphanin), whereas cauliflower, savoy cabbage and white cabbage contained mainly 2-propenyl (sinigrin) and 3-(methylsulfinyl)propyl glucosinolate (glucoiberin). Upon hydrolysis, epithionitriles or nitriles were often observed to be the main hydrolysis products, with 1-cyano-2,3-epithiopropane being most abundant with up to 5.7 μmol/g fresh weight in white cabbage sprouts. Notably, sprouts often contained more than 10 times more glucosinolates or their hydrolysis products compared to fully developed vegetables. Moreover, during head development, both glucosinolate concentrations as well as hydrolysis product concentrations changed and mini heads contained the highest isothiocyanate concentrations. Thus, from a cancer-preventive point of view, consumption of mini heads of the B. oleracea varieties is recommended.

Background Glucosinolates, anionic sulfur rich secondary metabolites, have been extensively studied because of their occurrence in the agriculturally important brassicaceae and their impact on human and animal health. There is also increasing interest in the biofumigant properties of toxic glucosinolate hydrolysis products as a method to control agricultural pests. Evaluating biofumigation potential requires rapid and accurate quantification of glucosinolates, but current commonly used methods of extraction prior to analysis involve a number of time consuming and hazardous steps; this study aimed to develop an improved method for glucosinolate extraction. Results Three methods previously used to extract glucosinolates from brassicaceae tissues, namely extraction in cold methanol, extraction in boiling methanol, and extraction in boiling water were compared across tissue type (root, stem leaf) and four brassicaceae species (B. juncea, S. alba, R. sativus, and E. sativa). Cold methanol extraction was shown to perform as well or better than all other tested methods for extraction of glucosinolates with the exception of glucoraphasatin in R. sativus shoots. It was also demonstrated that lyophilisation methods, routinely used during extraction to allow tissue disruption, can reduce final glucosinolate concentrations and that extracting from frozen wet tissue samples in cold 80% methanol is more effective. Conclusions We present a simplified method for extracting glucosinolates from plant tissues which does not require the use of a freeze drier or boiling methanol, and is therefore less hazardous, and more time and cost effective. The presented method has been shown to have comparable or improved glucosinolate extraction efficiency relative to the commonly used ISO method for major glucosinolates in the Brassicaceae species studied: sinigrin and gluconasturtiin in B. juncea; sinalbin, glucotropaeolin, and gluconasturtiin in S. alba; glucoraphenin and glucoraphasatin in R. sativus; and glucosatavin, glucoerucin and glucoraphanin in E. sativa. Electronic supplementary material The online version of this article (doi:10.1186/s13007-017-0164-8) contains supplementary material, which is available to authorized users.

The large cabbage moth, Crocidolomia pavonana (F.) (Lepidoptera: Crambidae), is one of the most significant pests of Brassica crops and has re-emerged as a serious pest of crucifer crops in tropical and sub-tropical Asia because of the control used against the diamondback moth, Plutella xylostella (L.) (Lepidoptera: Plutellidae). The damage caused by this pest can be significant because larvae devour young tissues of the plant at different stages and bore into the centre of the cabbage head at maturity and can damage the whole cabbage plant. Despite its economic importance the interaction of this pest with its host plant is not well understood. The current study showed that larval feeding by C. pavonana was influenced by the presence of cabbage in artificial diet, in laboratory bioassays. Of the four diets developed and tested, neonates survived and completed developmental stages (81%) on diet containing cabbage (Brassica oleracea cv sugarloaf) more so than on base diet (diet without cabbage), diet containing wheat grass or spinach powder. Neonates were able to locate diet with cabbage powder but not when cabbage was absent (i.e. base diet). Similarly, in an olfactometer test, neonates were strongly attracted to the odour of diet with cabbage compared with diet without cabbage. More than twice the number of feeding events was observed on filter paper disks impregnated with diet containing cabbage powder than on paper disks with base diet. The behavioural responses of neonates to diet with cabbage are linked to the glucosinolates present on cabbage plants. Sinigrin elicited feeding when added to diet without cabbage and also when applied to a non-host plant, Gossypium hirsutum (Sicot 71RRF). Neonates’ response to allyl isothiocyanate in a two-arm olfactometer was stronger than to sinigrin, which would explain larval attraction to damaged cabbage plants. These results indicate that sinigrin and allyl isothiocyanate provide important feeding and olfactory cues for C. pavonana larvae. Larval feeding and moth oviposition behaviour of C. pavonana was further studied using whole cabbage plants of two different ages (6 and 8-leaf stage) and this was compared to the response of a well-studied crucifer specialist, P. xylostella. Plant age did not significantly affect oviposition and feeding preference by C. pavonana whereas P. xylostella preferred younger plants for oviposition and larval feeding. The intra-plant distribution of eggs and feeding sites was similar for both species, but egg distribution showed a pattern different from feeding sites. Neonates that hatched from eggs deposited on lower leaves moved towards the apical portion of the plant and established a majority of feeding sites on younger leaves. Preference for younger leaves was also observed on excised leaves, indicating architectural position was not the factor driving choice. Younger leaves are known to have a higher concentration of nitrogen content and proteins suitable for larval development, whereas lower leaves are considered not palatable because of fibrous content and low nutritional quality. Therefore, leaf age is an important factor affecting choice of feeding sites by the crucifer specialists. Sinigrin content, inferred from a separate study, did not explain feeding choice. The significant increase in the concentration of total aliphatic and indole glucosinolates with reduced leaf age does correspond with larval preference for younger leaves. Leaf wax and leaf toughness brought about by leaf age were studied as possible factors involved in leaf choice. Wax morphology, observed using scanning electron microscopy, differed between old and young leaves. The removal of leaf surface wax did affect C. pavonana neonates’ feeding choice, with greater preference for leaf disks with wax in situ, compared against leaf disks with wax removed, in a dual choice assay. Leaf wax extracted from either old or young leaves also induced feeding when added to artificial diet without cabbage. Leaf toughness, was not found to be a significant factor in feeding choice. The effect of sinigrin (allyl glucosinolate) and its hydrolysis product, allyl isothiocyanate on growth and survival of C. pavonana was investigated using artificial base diet and diet containing cabbage leaf powder, and compared with the response of P. xylostella. Consistent with other findings in this study, sinigrin promoted larval feeding for both crucifer specialists. Response to allyl isothiocyanate though, differed between C. pavonana and P. xylostella. Allyl isothiocyanate was found to be toxic and lethal for P. xylostella but C. pavonana neonates fed and survived, and when assessed at 24 hours, had gained weight. Similarly, varying the concentrations of allyl isothiocyanate (1 to 10 μmol/g diet) did not significantly affected survival of C. pavonana neonates whereas 1 μmol/g diet caused maximal mortality for P. xylostella neonates. The very high survival of C. pavonana neonates on diet containing allyl isothiocyanate indicates that C. pavonana handles glucosinolate hydrolysis products in a way that differs from P. xylostella, and this finding points to a new or different mechanism for glucosinolate metabolism.

Glucosinolates are a well-studied and highly diverse class of natural plant compounds. They play important roles in plant resistance, rapeseed oil quality, food flavoring, and human health. The biological activity of glucosinolates is released upon tissue damage, when they are mixed with the enzyme myrosinase. This results in the formation of pungent and toxic breakdown products, such as isothiocyanates and nitriles. Currently, more than 130 structurally different glucosinolates have been identified. The chemical structure of the glucosinolate is an important determinant of the product that is formed, which in turn determines its biological activity. The latter may range from detrimental (e.g., progoitrin) to beneficial (e.g., glucoraphanin). Each glucosinolate-containing plant species has its own specific glucosinolate profile. For this reason, it is important to correctly identify and reliably quantify the different glucosinolates present in brassicaceous leaf, seed, and root crops or, for ecological studies, in their wild relatives. Here, we present a well-validated, targeted, and robust method to analyze glucosinolate profiles in a wide range of plant species and plant organs. Intact glucosinolates are extracted from ground plant materials with a methanol-water mixture at high temperatures to disable myrosinase activity. Thereafter, the resulting extract is brought onto an ion-exchange column for purification. After sulfatase treatment, the desulfoglucosinolates are eluted with water and the eluate is freeze-dried. The residue is taken up in an exact volume of water, which is analyzed by high-pressure liquid chromatography (HPLC) with a photodiode array (PDA) or ultraviolet (UV) detector. Detection and quantification are achieved by conducting comparisons of the retention times and UV spectra of commercial reference standards. The concentrations are calculated based on a sinigrin reference curve and well-established response factors. The advantages and disadvantages of this straightforward method, when compared to faster and more technologically advanced methods, are discussed here.

International audience; Plants from the Brassicaceae family are known to contain secondary metabolites called glucosinolates. Our goal was to establish by LC/MS the glucosinolate profile of seeds of three Brassicaceae species known to hyperaccumulate heavy metals. We investigated Alyssum fallacinum auct. non Hausskn., Iberis intermedia Guers., and Noccaea caerulescens (J. Presl & C. Presl) F. K. Mey. Our results indicate that A. fallacinum seeds contain glucoiberin and glucoibervirin, which had not been previously identified in this plant. Furthermore, we report for the first time the presence of glucoiberin, glucoibervirin, glucotropaeolin, and sinigrin in I. intermedia. We have detected for the first time glucoconringiin in N. caerulescens. In addition, glucosinalbin, 4-hydroxyglucobrassicin, and glucomoringin were also detected.

Phytophthora cinnamomi causes a highly destructive root rot that seriously affects oak trees in semi-natural woodlands known as “dehesas”. Biofumigation with Brassica spp. is a promising tool for disease management. We demonstrated that ground seeds from B. carinata and B. juncea can inhibit mycelial growth and decrease chlamydospore viability of P. cinnamomi in soil. In contrast, B. napus seedmeals were ineffective. Reduction of root necrosis in Lupinus plants was also achieved when soils were biofumigated with B. carinata or B. juncea seedmeals. Seedmeal effectiveness was strongly correlated with high sinigrin (2-propenyl glucosinolate) content. We conclude that biofumigation with seedmeals rich in sinigrin could be effective as part of integrated management of oak disease caused by P. cinnamomi in “dehesas”., The authors thank the Andalusian Government (Project P10-359 AGR6501), the Spanish Ministry of Economy and Competitiveness (MINECO-INIA; Project RTA2014-00063-C04-03 and FEDER Funds), and the European Union (Project LIFE11 BIO/ ES/000726) for financial support.

Natural compounds isolated from vegetables have protective and disease-preventing potential for the development of functional food. This study investigates the in vitro antioxidant capacity, total polyphenol and total flavonoid content, glucosinolate and glucosinolate degradation products of broccoli and collard extracts. In vitro antitumor activities on two human tumour cell lines (MCF-7 and HeLa) were studied. Broccoli and collard extracts were extracted from fresh, blanched and boiled broccoli florets and collard leaves. The Oxygen Radical Absorbance Capacity (ORAC) assay revealed that collard extracts exhibited higher antioxidant capacity than broccoli extracts. Total polyphenol and total flavonoid contents were also higher in collard extracts. The extracts were compared for the contents of total glucosinolates, indol glucosinolates and aliphatic glucosinolates. Glucoraphanin was the predominant compound in total glucosinolate content, followed by glucobrassicin and neoglucobrassicin (95.97 μmol 100 g −1 fresh weight (fw), 85.42 μmol 100 g −1 fw and 82.12 μmol 100 g −1 fw, respectively) in broccoli extracts; while in collard extracts, the major glucosinolate was glucobrassicin (80.33 μmol 100 g −1 fw), followed by glucoiberin (67.50) μmol 100 g −1 fw and sinigrin (21.91 μmol 100 g −1 fw). The strongest antiproliferative effect was observed in extracts obtained by blanching. The present study established that the extracts that were examined possess antioxidant and promising antitumour activities.