Sugar Composition Analysis Research Articles

The effect of MSMEG_6402 gene disruption on the cell wall structure of Mycobacterium smegmatis

Arabinogalactan (AG) of mycobacterial cell wall consists of arabinan region, galactan region and disaccharide linker. The arabinan is composed of d-arabinofuranose residues, and decaprenyphosphoryl- d-arabinose (DPA) is the donor of the d-arabinofuranose residues. DPA is formed from phosphoribose diphosphate (PRPP) in a four-step process catalyzed by transferase, phosphatase and epimerase, respectively. Mycobacterium tuberculosis Rv3806c has been identified as PRPP: decaprenyl-phosphate 5-phosphoribosyltransferase，and heteromeric Rv3790/Rv3791 has epimerase activity. Rv3807c is putative phospholipid phosphatase. However, there is no direct biochemical evidence since expression of Rv3807c has been unsuccessful. Mycobacterium smegmatis MSMEG_6402 is ortholog of Rv3807c. To investigate the function of MSMEG_6402 on AG biosynthesis, a conditional MSMEG_6402 gene knock out ( M. sm-Δ M_6402) strain was constructed through homologous recombination technique. The morphological and compositional changes of cell wall were examined in the M. sm-Δ M_6402 strain. The M. sm-Δ M_6402 strain grew at non-permissive temperature slower than that at permissive temperature, indicating that MSMEG_6402 is non-essential for growth of M. smegmatis. The change of cell shape and detectable bulging on the cell surface of M. sm-Δ M_6402 strain were observed by scanning electron microscopy, and curled as well as deformed cell wall of M. sm-Δ M_6402 strain was revealed by transmission electron microscopy. Analysis of sugar composition in the cell wall by HPLC indicated that the ratio of arabinofuran to galacto furan in M. sm-Δ M_6402 strain was changed to 1.7:1 comparing with 2:1 in the wild type. It demonstrates that the lacking MSMEG_6402 interferes the biosynthesis of arabinan. Analyzing 5′ P-DPR and DPR from both M. sm-Δ M_6402 strain and wild type M. smegmatis is undergoing in this lab.

Molecular Characterization of Arabinoxylans from Hull-Less Barley Milling Fractions

Arabinoxylans were prepared from different hull-less barley milling fractions (bran, shorts and flour). The yields of hull-less bran arabinoxylan (HBB-AX), shorts arabinoxylan (HBS-AX) and flour arabinoxylan (HBF-AX) were 8.42%, 4.08% and 2.13% respectively. Sugar composition analysis showed that arabinose and xylose were the main sugars. HBF-AX had the highest Ara/Xyl ratio, followed by HBS-AX and HBB-AX. Size exclusion chromatography analysis (HPSEC) showed that HBF-AX had the highest molecular weight, followed by HBS-AX and HBB-AX, which had the lowest molecular weight. Aqueous solutions of HBB-AX, HBS-AX and HBF-AX had higher molecular weight(Mw) than in 0.5 mol/L NaOH and 1.0 mol/L NaOH. Dynamic light scattering analysis (DLS) showed that HBB-AX, HBS-AX and HBF-AX had existed in two states in distilled water, 0.5 mol/LNaOH, 1.0 mol/L NaOH, and DMSO/H2O (90/10), an unaggregated state and an aggregated state, with the latter predominating.

Chemical compositions and some physical properties of the water and alkali-extracted mucilage from the young fronds of Asplenium australasicum (J. Sm.) Hook

Effect of extraction conditions, including solvent types (water and sodium bicarbonate) and extraction temperatures (25, 50, 70 and 90 °C), on the physicochemical properties of the water and alkali-extracted mucilage from the young fronds of Asplenium australasicum (J. Sm.) Hook was evaluated. It was found that sodium bicarbonate extract generally showed higher yield than water extract, and its yield increased with increasing extraction temperature. Composition analysis revealed that A. australasicum mucilage was composed of mainly carbohydrates in conjunction with some glycoproteins. The weight average molecular size of the carbohydrate fraction of crude mucilage obtained from alkali and water extractions were about 6.30 × 10 7 and 2.63 × 10 7, respectively. Sugar composition analysis revealed that A. australasicum young fronds mucilage contained significant amount of uronic acid (14.3–56.6% based on total sugars). For the water extracted mucilage, GlcA is predominant. However, for the sodium bicarbonate extracted mucilage, GalA is predominant. Furthermore, Rha can only be detected in sodium bicarbonate extract. Other major neutral sugars include Gal, Man, Xyl and Fuc. The intrinsic viscosity of the water and alkali-extracted mucilage in deionized water were around 21.12 and 5.92 dL/g, respectively. This information would be useful when considering A. australasicum young fronds mucilage as a new Agro-ingredient.

Golgi nucleotide sugar transporter modulates cell wall biosynthesis and plant growth in rice

Golgi-localized nucleotide sugar transporters (NSTs) are considered essential for the biosynthesis of wall polysaccharides and glycoproteins based on their characteristic transport of a large number of nucleotide sugars to the Golgi lumen. The lack of NST mutants in plants has prevented evaluation of this hypothesis in plants. A previously undescribed Golgi NST mutant, brittle culm14 (bc14), displays reduced mechanical strength caused by decreased cellulose content and altered wall structure, and exhibits abnormalities in plant development. Map-based cloning revealed that all of the observed mutant phenotypes result from a missense mutation in a putative NST gene, Oryza sativa Nucleotide Sugar Transporter1 (OsNST1). OsNST1 was identified as a Golgi-localized transporter by analysis of a fluorescence-tagged OsNST1 expressed in rice protoplast cells and demonstration of UDP-glucose transport activity via uptake assays in yeast. Compositional sugar analyses in total and fractionated wall residues of wild-type and bc14 culms showed a deficiency in the synthesis of glucoconjugated polysaccharides in bc14, indicating that OsNST1 supplies the glucosyl substrate for the formation of matrix polysaccharides, and thereby modulates cellulose biosynthesis. OsNST1 is ubiquitously expressed, with high expression in mechanical tissues. The inferior mechanical strength and abnormal development of bc14 plants suggest that OsNST1 has pleiotropic effects on cell wall biosynthesis and plant growth. Identification of OsNST1 has improved our understanding of how cell wall polysaccharide synthesis is regulated by Golgi NSTs in plants.

Monosaccharide-NAIM Derivatives for D-, L-Configurational Analysis

The D-, L-enantiomeric pairs of common monosaccharides (xylose, ribose, rhamnose, arabinose, fucose, glucose, mannose, galactose, N-acetylgalactosamine, glucuronic acid and galacturonic acid) were derivatized with 2,3-naphthalenediamine to form the corresponding D-, L-aldo-NAIM derivatives. A simple and facile capillary electrophoretic method was established for sugar composition analysis by simultaneously determining the migration times of these aldo-NAIMs using borate buffer at high pH (100 mM, pH 9.0). The methodology is also applicable to sialic acid (ketose monosaccharides). The quantitation level of the proposed method was in the 10~500 ppm range and the LOD was 1 ppm. The enantioseparation of D, L pairs of aldo-NAIMs were also achieved by using modified sulfated−α−cyclodextrin as the chiral selector in phosphate buffer (300 mM, pH 3.0). In addition, the combination by reductive amination of amino-aldo-NAIM agent and D-, L-enantiomeric pairs of monosaccharides formed a diastereomeric pair for saccharide configuration analysis. Aldo-NAIM derivatives are thus shown to be rapid and efficient agents for analyzing saccharide compositions and configurations with good linearity and short analysis times via capillary electrophoresis.

Structures of acidic polysaccharides from Basella rubra L. and their antiviral effects

In this study four acidic polysaccharides (BRP-1, BRP-2, BPR-3 and BRP-4) were isolated from the aerial part of Basella rubra L. Sugar composition and methylation analyses indicated that both BRP-2 and BRP-4 were native pectins containing dominant homogalacturonan regions and minor ramified rhamnogalacturonan type I regions branched with arabinogalactan type II neutral side chains. They exhibited antiviral activity against herpes simplex virus type 2 by interfering with absorption and penetration of virus to host cells. The most abundant component, BRP-4, showed a high therapeutic efficacy in the mouse model infected intravaginally with herpes simplex virus type 2. Moreover, orally administered BRP-4 resulted in moderate therapeutic efficacy in the mice infected intranasally with influenza A virus, and was shown to stimulate the production of neutralizing antibody and the secretion of mucosal IgA in influenza virus-infected mice.

Altered glycosylation profile of purified plasma ACT from Alzheimer's disease.

BackgroundAlzheimer’s disease (AD) is one of the most frequent cause of neurodegenerative disorder in the elderly. Inflammation has been implicated in brain degenerative processes and peripheral markers of brain AD related impairment would be useful. Plasma levels of alpha-1-antichymotrypsin (ACT), an acute phase protein and a secondary component of amyloid plaques, are often increased in AD patients and high blood ACT levels correlate with progressive cognitive deterioration. During inflammatory responses changes in the micro-heterogeneity of ACT sugar chains have been described.MethodsN-Glycanase digestion from Flavobacterium meningosepticum (PNGase F) was performed on both native and denatured purified ACT condition and resolved to Western blot with the purpose to revealed the ACT de-glycosylation pattern.Further characterization of the ACT glycan profile was obtained by a glycoarray; each lectin group in the assay specifically recognizes one or two glycans/epitopes. Lectin-bound ACT produced a glyco-fingerprint and mayor differences between AD and controls samples were assessed by a specific algorithms.ResultsWestern blot analysis of purified ACT after PNGase F treatment and analysis of sugar composition of ACT showed significantly difference in “glyco-fingerprints” patterns from controls (CTR) and AD; ACT from AD showing significantly reduced levels of sialic acid. A difference in terminal GlcNac residues appeared to be related with progressive cognitive deterioration.ConclusionsLow content of terminal GlcNac and sialic acid in peripheral ACT in AD patients suggests that a different pattern of glycosylation might be a marker of brain inflammation. Moreover ACT glycosylation analysis could be used to predict AD clinical progression and used in clinical trials as surrogate marker of clinical efficacy.

An Insect Herbivore Microbiome with High Plant Biomass-Degrading Capacity

Herbivores can gain indirect access to recalcitrant carbon present in plant cell walls through symbiotic associations with lignocellulolytic microbes. A paradigmatic example is the leaf-cutter ant (Tribe: Attini), which uses fresh leaves to cultivate a fungus for food in specialized gardens. Using a combination of sugar composition analyses, metagenomics, and whole-genome sequencing, we reveal that the fungus garden microbiome of leaf-cutter ants is composed of a diverse community of bacteria with high plant biomass-degrading capacity. Comparison of this microbiome's predicted carbohydrate-degrading enzyme profile with other metagenomes shows closest similarity to the bovine rumen, indicating evolutionary convergence of plant biomass degrading potential between two important herbivorous animals. Genomic and physiological characterization of two dominant bacteria in the fungus garden microbiome provides evidence of their capacity to degrade cellulose. Given the recent interest in cellulosic biofuels, understanding how large-scale and rapid plant biomass degradation occurs in a highly evolved insect herbivore is of particular relevance for bioenergy.

Exopolysaccharide from Lactobacillus helveticus: identification of chemical structure and effect on biscuit duality

Exopolysaccharide (LB-gum) was produced from Lactobacillus helveticus by ethanol precipitation and gel-permeation chromatography. The structures of LB1 and LB2 were estimated by sugar composition analysis, methylation, and FT-IR analysis. The results proved the contents of glucose and galactose in molar ratio of 2:1 and 2.3:1, and molecular weights ~5.4 × 10<sup>5</sup> Da and ~20.3 × 10<sup>5</sup> Da, respectively. Xanthan and LB-gum were added to wheat flour during biscuits making at levels of 0, 0.5, 1.0, 1.5, and 2.0%. Rheological properties and chemical quality attributes of the biscuits during storage for six months were evaluated. The data revealed slight increases in water absorption, dough development time, and dough stability but weakening decreased about 50 %. Also, height, weight, volume, and specific volume were found to increase as xanthan and LB-gum level increased. The colour was slightly affected. There were no significant differences between the samples containing xanthan and LB-gum at different levels with respect to all parameters tested except for the taste. Shelf – life of biscuit prolonged as a result of xanthan or LB-gum addition. The best addition level to improve the biscuits quality was 1.0% of xanthan or 1.5% of LB-gum.  

Cell wall polysaccharides from cell suspension cultures of the Atlantic Forest tree Rudgea jasminoides (Rubiaceae)

Rudgea jasminoides (Rubiaceae) is a tropical tree species native of the Atlantic Forest in the south of Brazil. Previous studies with leaf cell walls of R. jasminoides showed a different proportion of cross-linked glycans compared to what is usually reported for eudicots. However, due to the difficulties of working with whole plant organs, cell suspensions of R. jasminoides, consisting of predominantly undifferentiated cells with mainly primary cell walls, were used to examine cell walls and extracellular soluble polysaccharides (EP) released into the culture medium. Sugar composition and linkage analysis showed homogalacturonans, xylogalacturonans and arabinogalactans to be the predominant EP. In the cell wall, homogalacturonans and arabinogalactans are the major pectins, and xyloglucans and xylans are the major cross-linking glycans. The presence of xylogalacturonans in the R. jasminoides cell cultures seems to be related to the occurrence of a homogeneous cell suspension with loosely attached cells. Although all alkali extractions from the cell walls yielded amounts of xyloglucan that exceed those of the xylans, the latter was found in a proportion that is higher than what has been usually reported for primary cell walls of most eudicots. The xyloglucan from cell walls of cell suspension cultures of R. jasminoides has low fucosylation levels and high proportion of galactosyl residues, a branching pattern commonly found in storage cell-wall xyloglucans.

Sugar treatment inhibits IAA-induced expression of endo-1,3:1,4-β-glucanase EI transcripts in barley coleoptile segments

The degradation of 1,3:1,4-beta-glucan by glucanases is believed to be critical for auxin-induced elongation in Gramineae coleoptile. In the present study, we reinvestigated the relationship between auxin-induced elongation and gene expression of glucanases upon treatment of coleoptile segments with sugars. Gene expression of exo-beta-1,3:1,4-glucanase ExoII was not affected by treatment with IAA and/or sucrose. In contrast, levels of endo-beta-1,3:1,4-glucanase EI transcripts increased in response to IAA treatment, which was negated by the addition of glucose or sucrose, although the addition of sucrose or glucose did not suppress IAA-induced elongation. Sugar composition analysis of the hemicellulosic fraction revealed that the addition of glucose suppressed the IAA-induced reduction of beta-glucan. In the coleoptile segments that were starved by pre-incubation in water, the IAA-induced accumulation of EI mRNA was accelerated, as compared with the non-starved segments, which suggests that the level of carbon source in the cytoplasm regulates EI expression. Moreover, in the basal region of coleoptiles, where IAA treatment does not induce elongation growth, high levels of EI transcripts were observed in the presence and absence of IAA treatment. These results strongly demonstrated that the expressions of exo- and endo-beta-glucanase genes are not directly involved in the IAA-induced loosening of cell walls associated with elongation and also suggests that cell walls may degrade 1,3:1,4-beta-glucan to provide glucose as an energy source for cell elongation.

Fractionation of Alkali-Solubilized Hemicelluloses from Delignified Populus gansuensis: Structure and Properties

The dewaxed cell walls of Populus gansuensis were delignified with NaClO(2) and then sequentially extracted with 0.25, 0.5, and 1.0 M KOH under a solid to liquid ratio of 1: 25 (g mL(-1)) at 25 degrees C for 10 h. The successive treatments together resulted in the dissolution of 83.7% of original hemicelluloses. The solubilized hemicellulosic fractions were further fractionated into six hemicellulosic subfractions by an iodine-complex precipitation technique. Their chemical and physical characteristics were determined by HPAEC, GPC, FT-IR, and (1)H and (13)C NMR spectroscopy. Neutral sugar composition and molecular weight analysis showed that, for each extract, the hemicellulosic subfractions that precipitated with aqueous potassium iodide-iodine had lower overall uronic acid/xylose (Uro/Xyl) ratios and higher molecular weights (M(w)) than those remaining in the solution. FT-IR, (1)H, and (13)C NMR spectroscopy analysis indicated that the alkali-soluble hemicelluloses of Populus gansuensis had a structure composed of the (1 --> 4)-linked beta-D-xylopyranosyl backbone with 4-O-methyl-alpha-D-glucuronic acid attached to O-2 of the xylose residues.

Fractional Study of Alkali-Soluble Hemicelluloses Obtained by Graded Ethanol Precipitation from Sugar Cane Bagasse

The two hemicellulosic fractions were subsequentially extracted with 5% and 8% NaOH aqueous solution at a solid to liquid ratio of 1:25 (g mL(-1)) at 50 degrees C for 3 h from the water, 1 and 3% NaOH-treated sugar cane bagasse, and subfractionated into six preparations by a graded ethanol precipitation method at concentrations of 15%, 30% and 60% (v/v). Sugar composition and molecular weight analysis showed that, with an increasing concentration of ethanol, hemicellulosic subfractions with both higher Ara/Xyl ratios and higher molecular weights were obtained. In other words, with an increasing ethanol concentration from 15% to 60%, the Ara/Xyl ratios increased from 0.043 in H(1) to 0.088 in H(3) and from 0.040 in H(4) to 0.088 in H(6), and the weight-average molecular weights of hemicellulosic subfractions increased from 42 430 (H(1)) to 85 510 (H(3)) g mol(-1) and from 46 130 (H(4)) to 64 070 (H(6)) g mol(-1), respectively. The results obtained by the analysis of Fourier transform infrared, sugar composition, and (1)H and (13)C nuclear magnetic spectroscopy showed that the alkali-soluble hemicelluloses had a backbone of xylose residues with a beta-(1-->4)-linkage and were branched mainly through arabinofuranosyl units at C-2 and/or C-3 of the main chain, whereas the differences may occur in the distribution of branches along the xylan backbone.

Permeabilization of Microbacterium oxylans shifts the conversion of puerarin from puerarin-7-O-glucoside to puerarin-7-O-fructoside

The main product of the conversion of puerarin by unpermeabilized cells of bacterium Microbacterium oxydans CGMCC 1788 was puerarin-7-O-glucoside (241 +/- 31.9 microM). Permeabilization with 40% ethanol could not increase conversion yield, whereas it resulted in change of main product; a previous trace product became a main product (213 +/- 48.0 microM) which was identified as a novel puerarin-7-O-fructoside by electrospray ionization time-of-flight MS, (13)C NMR, (1)H NMR, and GC-MS analysis of sugar composition, and puerarin-7-O-glucoside became a trace product (14.8 +/- 5.4 microM). However, the extract from cells of M. oxydans CGMCC 1788 permeabilized with ethanol converted puerarin to form 113.9 +/- 27.7 microM puerarin-7-O-glucoside and 187.8 +/- 29.5 microM puerarin-7-O-fructoside under the same conditions. When unpermeabilized intact cells were recovered and used repeatedly for the conversion of puerarin, with increase of reuse times, the yield of puerarin-7-O-glucoside gradually decreased, whereas the yield of puerarin-7-O-fructoside increased gradually in the conversion mixture. The main product of the conversion of puerarin by the tenth recycled unpremerbilized cells was puerarin-7-O-fructoside (288.4 +/- 24.0 microM). Therefore, the change of permeability of cell membrane of bacterium M. oxydans CGMCC 1788 contributed to the change of conversion of the product's composition.

Comparative studies on the physico-chemical properties of hemicelluloses obtained by DEAE-cellulose-52 chromatography from sugarcane bagasse

Abstract Water- and alkali-soluble hemicelluloses isolated from dewaxed sugarcane bagasse were sub-fractionated on DEAE-cellulose-52 chromatography and obtained six hemicellulosic sub-fractions by eluting with water, 0.1 M and 0.3 M NaCl aqueous solution, respectively. Sugar composition and molecular weight analysis revealed that the lower molecular weight (14,180–43,590 g mol −1 ) and more branches of hemicelluloses could be extracted by the hot water, which are rich in glucose, galactose, and xylose, while the higher molecular weight (75,430–138,170 g mol −1 ) and more linear hemicelluloses were able to be dissolved into 1% NaOH aqueous solution, which are rich in xylose, principally resulting from l -arabino-(4- O -methyl-glucurono)- d -xylans. In addition, it was found that with increasing the concentration of NaCl (aqueous), the hemicellulosic sub-fractions with both higher arabinose to xylose ratio and higher molecular weight were eluted. Based on the FT-IR, sugar composition and 1 H and 13 C NMR comparative studies, the alkali-soluble hemicellulosic sub-fractions had a classical structure, with a backbone of β-(1→4)-linked xylosyl residue substituted with arabinose at C–2 and/or C–3 of main chain, whereas the difference may occur in the distribution of branches along the xylan backbone.

Rice BRITTLE CULM 5 (BRITTLE NODE) is Involved in Secondary Cell Wall Formation in the Sclerenchyma Tissue of Nodes

Several brittle culm (bc) mutants known in grasses are considered excellent materials to study the process of secondary cell wall formation. The brittle phenotype of the rice bc5 (brittle node) mutant appears exclusively in the developed nodes, which is distinct from other bc mutants (bc1, 2, 3, 4, 6 and 7) that show the brittle phenotype in culms and leaves. To address the defects of the rice bc5 mutant in node-specific cell wall formation, we analyzed tissue morphology and cell wall composition. The bc5 mutation was found to affect the cell wall deposition of node sclerenchyma tissues at 1 week after heading, the stage at which the cell wall sugar content is reduced, in the bc5 nodes, compared with wild-type nodes. Moreover, decreased accumulation of lignin and thickness of cell walls in the sclerenchyma tissues were also observed in the bc5 nodes. The amounts of cellulose and hemicellulose were reduced to 53 and 65% of those in the wild-type plants, respectively. Sugar composition and glycosidic linkage analyses of the hemicellulose showed that the accumulation of glucuronosyl arabinoxylan in bc5 nodes was perturbed by the mutation. The bc5 locus was narrowed to an approximately 3.1 Mb region of chromosome 2, where none of the other bc genes is located. The bc5 mutation appeared to reduce the expression levels of the OsCesA genes in the nodes after heading. The results indicate that the BC5 gene regulates the development of secondary cell walls of node sclerenchyma tissues.

Glycogen phosphorylase is involved in stress endurance and biofilm formation inAzospirillum brasilense Sp7

Here we report the identification of a glycogen phosphorylase (glgP) gene in the plant growth-promoting rhizobacterium Azospirillum brasilense, Sp7, and the characterization of a glgP marker exchange mutant of this strain. The glgP mutant showed a twofold reduction of glycogen phosphorylase activity and an increased glycogen accumulation as compared with wild-type Sp7, indicating that the identified gene indeed encodes a protein with glycogen phosphorylase activity. Interestingly, the glgP mutant had higher survival rates than the wild type after exposure to starvation, desiccation and osmotic pressure. The mutant was shown to be compromised in its biofilm formation ability. Analysis of the exopolysaccharide sugar composition of the glgP mutant revealed a decrease in the amount of glucose, accompanied by increases in rhamnose, fucose and ribose, as compared with the Sp7 exopolysaccharide. To the best of our knowledge, this is the first study that demonstrates GlgP activity in A. brasilense, and shows that glycogen accumulation may play an important role in the stress endurance of this bacterium.

TEMPO-mediated oxidation of softwood thermomechanical pulp

AbstractA softwood thermomechanical pulp (TMP) was suspended in water and oxidized with sodium hypochlorite and catalytic amounts of sodium bromide and 2,2,6,6-tetramethylpiperidine-1-oxyl radical (TEMPO) at pH 10. When the NaClO addition levels were 20–26 mmol g-1of the TMP, the yields of water-insoluble TEMPO-oxidized TMPs fractions were approximately 40%. Sugar composition and other analyses revealed that most of the lignin and hemicellulose components in the TMP were removed as water-soluble fractions by the oxidation. Thus, almost pure TEMPO-oxidized celluloses can also be prepared from TMP, although the addition levels of NaClO are much higher than for the TEMPO-mediated oxidation of pure celluloses. The water-soluble fractions were analyzed by NMR, showing that polyuronic acids formed from glucomannan and cellulose by oxidation were the main compounds. The water-insoluble fractions of TEMPO-oxidized TMPs prepared with NaClO of 20–26 mmol g-1had carboxylate contents of approximately 1.2 mmol g-1, and had the same cellulose I allomorph and the same crystal widths as in the original TMP. Transparent and highly viscous gels were obtained by disintegration of these TEMPO-oxidized TMPs in water, and the gels consisted of individual nanofibers 4–6 nm in width. Thus, TEMPO-oxidized cellulose nanofibers can also be prepared from TMP.

Diallel analysis of sugar composition of 10 vegetable soybean lines

AbstractFew studies have evaluated vegetable soybean for sugar content at the green pod stage. Information on combining ability and type of gene action that governs inheritance of seed traits can help breeders to select suitable parents and devise an appropriate breeding strategy. Ten vegetable soybean accessions were crossed in a complete diallel mating design. Parent lines and F2 and F3 progenies were evaluated for two nutritional components. In this study, both general and specific combining ability and reciprocal effects were significant for sucrose and total sugar. Cultivars ‘Kanrich’, ‘Pella’, ‘Verde’ and V81‐1603 had good general combining ability for high sucrose. In general, high sucrose content was observed in progeny of the early maturity group genotypes ‘Kanrich’, ‘Pella’ and ‘Verde’. The best combiners for high total sugar content were ‘Verde’, V81‐1603 and PI 399055. These genotypes could serve as genetic sources in a vegetable soybean breeding programme.

Purification and characterization of a novel high molecular weight exotoxin produced by red tide phytoplankton, Alexandrium tamarense

Our recent studies have demonstrated that the aqueous extract prepared from Alexandrium tamarense, a harmful red tide phytoplankton, showed cytotoxicity on Vero cells. In this study, the toxic substance was purified from the culture supernatant of A. tamarense. Based on the gel-filtration profile, the molecular mass of a purified toxin was estimated to be about 1,000 kDa. On sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis, a main band with molecular mass of 1,000 kDa was detected with periodic acid-Schiff (PAS) staining, but no protein bands were detected by Coomassie brilliant blue (CBB) protein staining. Sugar composition analysis of the toxin suggested that the toxin contains galactose, fucose, mannose, N-acetylglucosamine, xylose, and other minor saccharides, whereas no significant levels of amino acids were detected by amino acid analysis. These results suggest that the toxin is a polysaccharide-based compound. The toxin showed cytotoxic effects on various cell lines in a concentration-dependent manner. Among the cell lines tested, U937 cells were the most susceptible to the toxin. In U937 cells treated with the toxin, a typical apoptotic nuclear morphological change and DNA fragmentation were observed. This is the first report demonstrating that a polysaccharide-based toxin isolated from red tide phytoplankton can induce apoptotic cell death.