Ionic thiocyanate (SCN-) production from 4-hydroxybenzyl glucosinolate contained in Sinapis alba seed meal

Meal produced from Sinapis alba seed by crushing to remove oil contains a glucosinolate that when hydrolyzed produces phytotoxic allelochemicals; however, the responsible compounds and pathways for their production have not been elucidated. S. alba seed meal and partially purified extracts containing 4-hydroxybenzyl glucosinolate were included in experiments to identify and monitor enzymatically released products using GC-MS and HPLC-MS. The initial product, 4-hydroxybenzyl isothiocyanate, was unstable in aqueous media, showing a half-life of 321 min at pH 3.0, decreasing to 6 min at pH 6.5. More alkaline pH values decrease the stability of 4-hydroxybenzyl isothiocyanate by promoting the formation of a proposed quinone that hydrolyzes to SCN-. Measurement of SCN- showed stoichiometric release from S. alba meal at 48 h when buffered at pH values as low as 4.0, demonstrating that SCN- production in soil is not only probable but likely responsible for observed phytoxicity of the meal.     

Quantitative detection of allergenic protein Sin a 1 from yellow mustard (Sinapis alba L.) seeds using enzyme-linked immunosorbent assay

Allergy to yellow mustard (YM; Sinapis alba L.) seed proteins has been reported and is currently seen as a constraint that hampers expansion of YM protein utilization. The most predominant allergenic protein of YM seed has been recognized as Sin a 1. In this study, Sin a 1 was purified ( S. alba var. Andante), rabbit polyclonal antibodies (pAb) specific to Sin a 1 were generated, and a sandwich enzyme-linked immunosorbent assay (S-ELISA) was developed to detect and quantify Sin a 1 from YM. The S-ELISA method using Sin a 1-pAb and its horseradish peroxidase conjugate resulted in a detection limit of 0.3 microg/mL for purified Sin a 1. The Sin a 1 contents of six YM lines were in the range of 0.82-2.94 mg/g when assayed by the developed S-ELISA method. The results showed that S-ELISA could distinguish Sin a 1 in YM seed-derived extracts rapidly and could be applied in controlling and/or monitoring of YM allergenic proteins.     

Genetic diversity analysis of yellow mustard (Sinapis alba L.) germplasm based on genotyping by sequencing

Recent advances in next generation sequencing technologies make genotyping by sequencing (GBS) more feasible for molecular characterization of plant germplasm with complex and unsequenced genomes. We used a GBS protocol consisting of Roche 454 pyrosequencing, genomic reduction and advanced bioinformatics tools to analyze genetic diversity of 24 diverse yellow mustard accessions. One and one half 454 pyrosequencing runs generated roughly 1.2 million sequence reads totaling about 392 million nucleotides. Application of the computational pipeline DIAL identified 512 contigs and 828 SNPs. The BLAST algorithm revealed alignments of 214 contigs with the sequences reported in NCBI nr/nt database. Sanger sequencing confirmed 95 % of 41 selected contigs and 94 % of 240 putative SNPs. The 454 scored SNPs were highly imbalanced among assayed samples. Diversity analysis of these SNPs revealed that 26.1 % of the total variation resided among landrace, cultivar and breeding lines and 24.7 % between yellow- and black-seeded germplasm. Cluster analysis showed that the black-seeded accessions were largely clustered together and the breeding lines were grouped with known origin. Computer simulation was performed to assess the impact of 454 SNPs missing and revealed considerable changes in allelic count, bias in detection of genetic structure, and large deviations from the expected genetic-distance matrix. These findings are useful for parental selection consideration in yellow mustard breeding, and our detailed analyses help illustrate the utility of GBS in genetic-diversity analysis of plant germplasm, particularly for genetic-relationship assessment.     

Optimizing the NPK application in white mustard (Sinapis alba L.) under an arid climate in Punjab,Pakistan

The production of oil from the conventional oilseed seed crops (viz. mustard, rapeseed) is not sufficient to meet the demands of Pakistani people. Due to this reason, a surplus quantity of oil is imported every year from different countries to fulfill the domestic demands of the cooking oil. This 2-year study was aimed to optimize the dose of NPK for profitable production of white mustard. Nitrogen (0, 60, 90?kg ha^?1), phosphorous (0, 30, 60?kg ha^?1) and potassium (0, 75, 100?kg ha^?1) were applied in various combinations. The results indicated that highest seed yield and profitability was recorded with NPK at 90–60–100?kg ha^?1, which was attributed to improvement in crop growth. In crux, application of NPK at pre-optimized rate might be a viable option to improve the growth, yield, oil quality, and economics of white mustard production under arid climates in Pakistan. According to the results, a balanced use of NPK fertilization is a key management strategy for white mustard growing farmers of arid region.     

Response of white mustard (Sinapis alba) and the soil microbial biomass to P and Zn addition in a greenhouse pot experiment

A 45‐d pot experiment was carried out to investigate the response of white mustard and the soil microbial biomass after Zn and P addition to a P deficient silt loam. The underlying hypothesis was that P application reduces the Zn availability to crops and microbial biomass. White mustard was supplied with different levels of P (0, 50, and 100 µg g^?1 soil) and Zn (0, 10, and 20 µg g^?1 soil). Amendments of P generally reduced extractable Zn, shoot Zn and soil microbial biomass Zn. Amendments of P generally decreased the microbial biomass C/P ratio. At 20 µg Zn g^?1 soil, a negative effect on the microbial biomass C/P ratio was observed, suggesting that high contents of extractable Zn have a negative impact on the microbial P uptake. However, the minimum Zn requirements of soil microorganisms and the consequences of microbial Zn deficiency for soil microbiological processes are completely unknown.     

Characterization of phytoecdysteroid glycosides in Meadowfoam (Limnanthes alba) seed meal by positive and negative ion LC-MS/MS

Meadowfoam ( Limnanthes alba) is an oilseed crop grown in western Oregon. The seed meal has potential value as a biopesticide due to glucosinolate degradation products and phytoecdysteroids, a group of polyhydroxylated triterpenoids with potent activities as arthropod molting hormones. Liquid chromatography in combination with tandem mass spectrometry operated in the precursor ion mode revealed the presence of four ecdysteroid glycosides in meadowfoam seed meal. The carbohydrate sequence and the identity of the ecdysteroid aglycones, ponasterone A and 20-hydroxyecdysone, were determined by product ion scanning. Ecdysteroids were detected in the negative ion mode as [M + formate] (-) ions, which yielded [M - H] (-) and alpha-cleavage fragments with retention of hydroxyl groups in MS/MS experiments (not seen in the positive ion mode), allowing the determination of the number of hydroxyl groups in the side chain and in the steroid ring system. MS/MS of glycoside ions ([MH] (+) or [M + formate] (-)) provided carbohydrate sequence information.     

Activity of meadowfoam (Limnanthes alba) seed meal glucolimnanthin degradation products against soilborne pathogens

Meadowfoam (Limnanthes alba L.) is a herbaceous winter-spring annual grown as a commercial oilseed crop. The meal remaining after oil extraction from the seed contains up to 4% of the glucosinolate glucolimnanthin. Degradation of glucolimnanthin yields toxic breakdown products, and therefore the meal may have potential in the management of soilborne pathogens. To maximize the pest-suppressive potential of meadowfoam seed meal, it would be beneficial to know the toxicity of individual glucolimnanthin degradation products against specific soilborne pathogens. Meloidogyne hapla second-stage juveniles (J2) and Pythium irregulare and Verticillium dahliae mycelial cultures were exposed to glucolimnanthin as well as its degradation products. Glucolimnanthin and its degradation product, 2-(3-methoxyphenyl)acetamide, were not toxic to any of the soilborne pathogens at concentrations up to 1.0 mg/mL. Two other degradation products, 2-(3-methoxymethyl)ethanethioamide and 3-methoxyphenylacetonitrile, were toxic to M. hapla and P. irregulare but not V. dahliae. The predominant enzyme degradation product, 3-methoxybenzyl isothiocyanate, was the most toxic compound against all of the soilborne pathogens, with M. hapla being the most sensitive with EC(50) values (0.0025 ± 0.0001 to 0.0027 ± 0.0001 mg/mL) 20-40 times lower than estimated EC(50) mortality values generated for P. irregulare and V. dahliae (0.05 and 0.1 mg/mL, respectively). The potential exists to manipulate meadowfoam seed meal to promote the production of specific degradation products. The conversion of glucolimnanthin into its corresponding isothiocyanate should optimize the biopesticidal properties of meadowfoam seed meal against M. hapla, P. irregulare, and V. dahliae.     

Metabolism-based resistance of a wild mustard (Sinapis arvensis L.) biotype to ethametsulfuron-methyl

Under controlled-environment conditions, ethametsulfuron-methyl doses that inhibited growth by 50% (ED(50)) were >100 and <1 g of active ingredient (ai) ha(-)(1) for ethametsulfuron-methyl-resistant (R) and -susceptible (S) wild mustard, respectively. There were no differences between the two biotypes with regard to absorption and translocation of the herbicide. Three days after treatment, approximately 90, 5, and 2% of the applied [(14)C]ethametsulfuron-methyl was found in the treated leaf, foliage, and roots of each biotype, respectively. Acetolactate synthase extracted from the two biotypes was equally sensitive to both ethametsulfuron-methyl and chlorsulfuron. These results indicate that resistance was not due to differences in the target site, absorption, or translocation. However, ethametsulfuron-methyl was metabolized more rapidly in the R than the S biotype. Approximately 82, 73, 42, 30, and 17% of the recovered radioactivity remained as ethametsulfuron-methyl in R wild mustard 3, 6, 18, 48, and 72 h after treatment, respectively. Conversely, 84, 79, 85, and 73% of the (14)C was ethametsulfuron-methyl in the S biotype 12, 24, 48, and 72 h after treatment, respectively. On the basis of these results, it is proposed that resistance is due to enhanced metabolism of ethametsulfuron-methyl in the R biotype.     

Dynamics of 13C‐labeled mustard litter (Sinapis alba) in particle‐size and aggregate fractions in an agricultural cropland with high‐ and low‐yield areas

The application of ¹³C‐labeled litter enables to study decomposition processes as well as the allocation of litter‐derived carbon to different soil C pools. ¹³Carbon‐labeled mustard litter was used in order to compare decomposition processes in an agricultural cropland with high‐yield (HY) and low‐yield (LY) areas, the latter being characterized by a finer texture and a lower organic‐C (OC) content. After tracer application, ¹³C concentrations were monitored in topsoil samples in particulate organic matter (POM) and in fine mineral fractions (silt‐ and clay‐sized fractions). After 568 d, approximately 5% and 10% of the initial ¹³C amount were found in POM fractions of LY and HY areas, respectively. Higher amounts were found in POM occluded in aggregates than in free POM. Medium‐term (0.5–2 y) storage of the initial ¹³C in fine silt‐ and clay‐sized fractions amounts to 10% in HY and LY soils, with faster enrichment but also faster disappearance of the ¹³C signal from LY soils. Amounts of 80%–90% of the added ¹³C were mineralized or leached in the observed period. Decomposition of free POM was faster in HY than in LY areas during the first year, but the remaining ¹³C amounts in occluded‐POM fractions were higher in HY soils after 568 d. High‐yield and low‐yield areas showed different ¹³C dynamics in fine mineral fractions. In LY soils, ¹³C amounts and concentrations in mineral‐associated fractions increased within 160 d after application and decreased in the following time period. In HY areas, a significant increase in ¹³C amounts did not occur until after 568 d. The results indicate initially faster decomposition processes in HY than in LY areas due to different soil conditions, such as soil texture and water regime. The higher silt and clay contents of LY areas seem to promote a faster aggregate formation and turnover, leading to a closer contact between POM and mineral surfaces in this area. This favors the OC storage in fine mineral fractions in the medium term. Lower aggregate formation and turnover in the coarser textured HY soil leads to a delayed C stabilization in silt‐ and clay‐sized fractions.     

Response of mustard to seed treatment with pyridioxine and basal and foliar application of nitrogen and phosphorus

In a simple randomised field trial conducted during 1985–86/ 1986–87, the effect of basal nitrogen (45 and 60 kg N/ha) and basal phosphorus (15 and 20 kg P/ha) applications together with the soaking of seeds in 0.025% aqueous pyridoxine hydrochloride solution for 4h and foliar applications of 15 kg N/ha and 5 kg P/ha in two installments at 70 and 90 days after sowing (DAS) was studied on the performance of mustard (Brassica juncea L.). Recommended basal applications of 90 kg N/ha and 30 kg P/ha (BN₉₀P₃₀) was used as the control. The parameters studied included leaf area index (LAI) at 60, 80, and 100 DAS, net assimilation rate at 60–80 and 80–100 day intervals, and pods/plant, seeds/pod, seed yield, oil content, and oil yield at harvest. In general, the pyridoxine treatment proved superior over water soaking. The higher basal fertilizer dose was effective and foliar application of N and P gave higher values as compared to the water foliar application alone. The combination of pyridoxine + BN₆₀P₂₀+ FN₁₅FP₅ significantly enhanced the performance of the crop and enhanced seed yield and oil yield by 15.8 and 13.5%, respectively, over the check BN₉₀P₃₀ treatment.     

Isolation and identification of molecular species of phosphatidylcholine and lysophosphatidylcholine from jojoba seed meal (Simmondsia chinensis)

A mixture of lysophosphatidylcholine (LPC) and phosphatidylcholine (PC) has been isolated by column chromatography from a jojoba meal (Simmondsia chinensis) extract. The molecular species of both classes could be separated and isolated by C18 reversed phase HPLC. The two major compounds were identified by 1D and 2D (1)H and (13)C NMR, by MS, and by GC-MS as 1-oleoyl-3-lysophosphatidylcholine and 1,2-dioleoyl-3-phosphatidylcholine. Eight other molecular species of LPC and four other molecular species of PC could be assigned by comparison of the mass spectra of the isolated compounds with the spectra of the two major compounds. Complete characterization of the individual molecular species was achieved by GC and GC-MS analysis of the fatty acyl composition from the isolated compounds. The PC/LPC proportion in the phospholipid mixture from three different samples is 1.6 +/- 0.1. LPC is considered to be an important bioactive compound; the results of this study suggest further research for the evaluation of potential health benefits of jojoba meal phospholipids.     

Comparative Efficacy of White Mustard (Sinapis alba L.) and Soybean (Glycine max L. Merr.) Seed Meals as Bioherbicides in Organic Broccoli (Brassica oleracea Var. Botrytis) and Spinach (Spinacea oleracea) Production

White mustard and soybean seed meals were compared for weed control and yield of organically grown broccoli and spinach. The meals were incorporated into the soil 2 weeks before crop planting at two rates (1.24 and 4.48 t ha^?1). Weed densities and hand-weeding time were recorded twice during the growing seasons and weed biomass was measured at crop harvest. Compared to the 1.24 t ha^?1 soybean treatment, weed densities were 52 to 95% and 41 to 45% less at 3 and 6 weeks after planting, respectively, in both crops with the 4.48 t ha^?1 white mustard seed meal treatment. Time required for hand weeding at these times was also reduced by up to 82% and 48%, respectively. Broccoli yield was similar in all the treatments, but spinach yield was greatest in the 4.48 t ha^?1 treatments for both seed meals. Petiole nitrate and nutrient concentrations in both crops were generally similar in all the treatments.     

Nutritional and physiological responses of young growing rats to diets containing raw cowpea seed meal,protein isolate (globulins), or starch

The nutritional and physiological effects of raw cowpea (Vigna unguiculata (L) Walp.) seed meal, protein isolate (globulins), or starch on the metabolism of young growing rats have been evaluated in 14-day trials. Wet and dry weight gain, feed conversion efficiency, and lipid and protein accretion were significantly reduced as a result of inclusion of seed meal, globulins, or starch in the diet, with growth retardation being most marked with the seed meal. The proportional weights of the small intestine and pancreas were increased by meal diets, and serum cholesterol levels were slightly reduced. The globulins and raw starch also increased relative small intestine weights but had no effect on the pancreas or serum constituents. The effects of cowpeas on rats appeared to be due primarily to the combined actions of globulins, resistant starches, protease inhibitors, and possibly fiber and non-starch polysaccharides on intestinal and systemic metabolism.     

Development of yellow pigmentation in squid (Loligo peali) as a result of lipid oxidation

The impact of lipid oxidation on yellow pigment formation in squid lipids and proteins was studied. When the squid microsomes were oxidized with iron and ascorbate, thiobarbituric acid reactive substance were observed to increase simultaneously with b values (yellowness) and pyrrole compounds concomitantly with a decrease in free amines. Oxidized microsomes were not able to change the solubility, sulfhydryl content, or color of salt-soluble squid myofibrillar proteins. Aldehydic lipid oxidation products were able to decrease solubility and sulfhydryl content of salt-soluble squid myofibrillar proteins but had no impact on color. Aldehydic lipid oxidation products increased b values (yellowness) and pyrrole compounds and decreased free amines in both squid phospholipid and egg yolk lecithin liposomes. The ability of aldehydic lipid oxidation products to change the physical and chemical properties of egg yolk lecithin liposomes increased with increasing level of unsaturation and when the carbon number was increased from 6 to 7. These data suggest that off-color formation in squid muscle could be due to nonenzymatic browning reactions occurring between aldehydic lipid oxidation products and the amines on phospholipids headgroups.     

Evaluation of mushroom dietary fiber (nonstarch polysaccharides) from sclerotia of Pleurotus tuber-regium (Fries) singer as a potential antitumor agent

Mushroom dietary fiber or nonstarch polysaccharides (NSPs) that were soluble in hot alkali and belonged to the beta-glucan type were isolated from the sclerotia of an edible mushroom, Pleurotus tuber-regium. The mushroom NSPs were further separated into a number of fractions [hot alkali extracts (HAEs)] with weight-average molecular weights (M(w)) ranging from 1 x 10(4) to 42.2 x 10(4). The HAE fractions [with M(w) of (5.8-17.1) x 10(4)] administered intraperitoneally at a dose of 20 mg/kg of body weight to BALB/c mice implanted with solid tumor Sarcoma 180 were found to be effective in inhibiting tumor proliferation with an inhibition ratio of > or =50%. In vitro experiments using human tumor cell lines HL-60 and HepG2 had shown that HAE fractions with M(w) of (5.8-42.2) x 10(4) also had antiproliferative activity at three different concentrations (50, 100, and 200 microg/mL) toward the tumor cell lines tested. All HAE fractions did not inhibit the growth of a normal kidney cell line (Vero) from monkey. It is therefore postulated that the antitumoral effect of NSPs from the sclerotia of P. tuber-regium is probably host-mediated and cytocidal.     

Characterization of polyphenol oxidase from litchi pericarp using (-)-epicatechin as substrate

Polyphenol oxidase (PPO) from litchi (Litchi chinensis Sonn.) pericarp was characterized using (-)-epicatechin, which was the major endogenous polyphenol in litchi pericarp as a substrate. The optimum pH for PPO activity with (-)-epicatechin was 7.5, and the enzyme was unstable below pH 4.5 and stable in the pH range of 6.0-8.0. Residual activities of PPO were 86.25, 86.31, and 80.17% after 67 days of incubation at 4 degrees C at pH 6.0, 7.5, and 8.0, respectively. From thermostability studies, the Ki value increased with temperature and the results suggested that the enzyme was unstable above 45 degrees C. Moreover, the results also provided strong evidence that the denaturalization temperature of PPO was near 70 degrees C. The inhibition studies indicated that l-cysteine and glutathione were strong inhibitors even at low concentrations while NaF inhibited moderately. In addition, the results also indicated that the inhibition mechanisms of thiol groups were different from those of halide salts.     

Characterization of protein changes associated with sugar beet (Beta vulgaris) resistance and susceptibility to Fusarium oxysporum

Fusarium oxysporum (F-19) is a serious threat to sugar beet. Resistance exists, but the basis for resistance and disease is unknown. Protein extracts from sugar beet genotypes C1200.XH024 (resistant, R) and Fus7 (susceptible, S) were analyzed by multidimensional liquid chromatography at 2 and 5 days postinoculation (dpi) and compared to mock-inoculated controls. One hundred twenty-one (R) and 73 (S) protein peaks were induced/repressed by F-19, approximately 12 (R) and 8% (S) of the total proteome detected. Temporal protein regulation occurred within and between each genotype, indicating that the timing of expression may be important for resistance. Thirty-one (R) and 48 (S) of the differentially expressed peaks were identified using matrix-assisted laser desorption-ionization with tandem time-of-flight mass spectrometry; others were below detection level. Comparison between the two genotypes uncovered R- and S-specific proteins with potential roles in resistance and disease development, respectively. Use of these proteins to select for new sources of resistance and to develop novel disease control strategies is discussed.     

New flavanol derivatives from grape (Vitis vinifera) byproducts. Antioxidant aminoethylthio-flavan-3-ol conjugates from a polymeric waste fraction used as a source of flavanols

A new family of antioxidants has been obtained from a residual fraction of polymeric polyphenols of grape origin. The integral exploitation of resources is important in any sustainable production scheme. Many byproducts and residues generated by the agroindustries contain polyphenols with potential application as preventative agents against cancer and cardiovascular diseases. Among these polyphenols oligomeric proanthocyanidins are particularly significant. The polymeric forms, considered of less interest because of their astringent properties, constitute the largest portion of the biologically active plant proanthocyanidins. The new compounds described here result from the breakdown of polymeric flavanols in the presence of cysteamine and bear an amino function, which facilitates their isolation from complex mixtures by cation-exchange gels or resins. In this way, valuable antioxidant molecules can be efficiently obtained from otherwise wasted polymers. The new molecules appear to be as effective as their underivatized counterparts (flavan-3-ols) as antioxidants.