Decomposition of (1–3,1–4)-β-Glucan and Expression of the (1–3,1–4) -β-Glucanase Gene in Rice Stems during Ripening

Abstract

To elucidate the possible participation of hemicellulose decomposition in lodging resistance, we studied the change of hemicellulose and cellulose content in the stems of rice during the ripening stage by methylation analysis and the expression of related genes by Northern blotting. In the rice stem in ripening stage, content of (1-3,1-4)-β-glucan, a component of hemicellulose, decreased markedly although the content of arabinoxylan, a major component of hemicellulose, and cellulose showed little change during the same growth period. On the other hand, expression of the Gns 1 gene, which may encode (1-3,1-4) -β-glucanase that catalyzes the degradation of (1-3,14) -β-glucan, increased sharply in the stem. The mechanism of decomposition of (1-3,1-4) -β-glucan in rice stem and the possible association with lodging resistance is discussed.     

Varietal Differences in Cell Wall β-(1→3),(1→4)-Glucan and Nonstructural Carbohydrate in Rice Stems during the Grain Filling Stage

Abstract

The contribution of cell wall components and nonstructural carbohydrate (NSC) to grain filling in rice (Oryza sativa L.) was clarified by investigating the differences in the dynamics of hemicellulose, sugar composition of hemicellulose, β-(1→3),(1→4)-glucan, and NSC among cultivars with different grain filling capacities. This investigation was performed using the stems of standard, high yield and low harvest index (HI) cultivars. Hemicellulose concentration in stems tended to decrease slightly during the grain filling stage. This decrease was attributed to a decrease in β-(1→3),(1→4)-glucan concentration, which was detected as a decrease in glucose composition of hemicellulose in the stems during the grain filling stage. The rate of decrease and decrease in the amount of β-(1→3),(1→4)-glucan in the stems differed among the cultivars. These were higher in high yield and high HI cultivars than in relatively low yield and low HI cultivars. Moreover, a positive correlation was observed between the rate of decrease in β-(1→3),(1→4)-glucan and NSC, indicating similarities in the dynamics of β-(1→3),(1→4)-glucan and NSC among the cultivars. When the top half of panicle was removed, β-(1→3),(1→4)-glucan and NSC concentrations in the culm and leaf sheath did not decrease during the grain filling stage. Therefore, the β-(1→3),(1→4)-glucan in stems might be one of the sources that supply substrate to panicle as well as NSC.     

Molecular weight and structure units of (1→3, 1→4)-β-glucans in dough and bread made from hull-less barley milling fractions

Milling fractions of hull-less barley, and dough and bread with hull-less barley flour (40%) and wheat flour (60%) were analysed in an investigation of how the properties of (1→3, 1→4)-β-glucan were affected by milling, dough formation and bread making. Calcofluor average molecular weight $({\overline{M}}_{\text{cf}})$ and molecular weight distribution and the cellotriosyl/cellotetraosyl ratio of the (1→3, 1→4)-β-glucan were determined. Four different hull-less barley samples were milled to produce straight-run white flours, shorts, bran and whole-meal flours. The molecular weight distributions were unimodal for all fractions, and the $({\overline{M}}_{\text{cf}})$ range was between 117×10⁴ and 188×10⁴. These parameters were similar for all barleys, although $({\overline{M}}_{\text{cf}})$ was somewhat lower in white flour and bran fractions and somewhat higher in shorts and whole-meal. The cellotriosyl/cellotetraosyl ratio (1.5–1.8) was also similar in all fractions. Doughs and breads were made to study how flour type (sifted or whole-meal barley flour), water content, yeast, mixing time and fermentation time affect (1→3, 1→4)-β-glucan. The molecular weight distribution of (1→3, 1→4)-β-glucan was polymodal with two or three populations for all doughs and breads, and the $({\overline{M}}_{\text{cf}})$ decreased with increasing mixing and fermentation time. These results indicated that (1→3, 1→4)-β-glucan was degraded by endogenous β-glucanases in the barley and/or wheat flour. The molecular weight was not significantly affected by bread-baking and other factors. After mixing and fermentation the cellotriosyl/cellotetraosyl ratio was about 1.7–1.8 and was thus not significantly different from that of the flour blends. Thus to retain high molecular weight (1→3, 1→4)-β-glucan, which is important for its cholesterol-lowering effect, it is thus important to keep the mixing and fermentation time as short as possible when baking hull-less barley bread.     

The combined use of hull-less barley flour and xylanase as a strategy for wheat/hull-less barley flour breads with increased arabinoxylan and (1→3,1→4)-β-D-glucan levels

Bread-making with a composite flour (CF) consisting of 60% wheat flour (WF) and 40% hull-less barley flour, increased the total and soluble (1→3,1→4)-β-D-glucan and total arabinoxylan (AX) contents of dough and bread samples, but decreased the specific bread loaf volume. A xylanase insensitive to inhibition by Triticum aestivum L. xylanase inhibitor (TAXI) and xylanase inhibiting protein (XIP), increased loaf volume by 8.8 and 20.1% for WF and CF breads, respectively. Xylanase addition not only markedly improved loaf volume of CF bread, but also increased the soluble AX content of the WF and CF dough and bread samples because of conversion of water-unextractable AX into soluble AX. The xylanase had no impact on the extractability and molecular weight of (1→3,1→4)-β-D-glucan, but (1→3,1→4)-β-D-glucan was degraded during bread-making probably because of endogenous β-glucanase activity. Taken together, the results clearly show that the combined use of hull-less barley flour and a xylanase active during bread making, lead to palatable breads with high total and soluble AX and (1→3,1→4)-β-D-glucan contents. The sum of total AX and (1→3,1→4)-β-D-glucan was 1.70% for WF bread and 3.06% for CF bread, while the sum of soluble AX and (1→3,1→4)-β-D-glucan was 0.49 and 1.41% for control WF and CF xylanase supplemented breads, respectively.     

Effect of High β-Glucan Barley on Serum Cholesterol Concentrations and Visceral Fat Area in Japanese Men—A Randomized,Double-blinded,Placebo-controlled Trial

This study investigated whether the consumption of a diet in which high-β-glucan barley replaced rice would reduce the visceral fat area as well as the serum low-density lipoprotein cholesterol (LDL-C) and total cholesterol (TC) in hypercholesterolemic Japanese men. A randomized, double-blinded, placebo-controlled intervention study was conducted in 44 hypercholesterolemic Japanese men with a body mass index (BMI) >22 kg/m². The subjects were randomly assigned to groups consuming either rice (placebo group) or a mixture of rice and pearl barley with a high β-glucan content (test group, 7.0 g β-glucan per day) for 12 weeks. Blood samples were taken, and CT scan obtained before the trial and every four weeks during the trial. The pearl barley intake significantly reduced serum concentrations of LDL-C (P = 0.041) and TC (P = 0.037) during the trial. Significant differences between the test and placebo groups were found for the visceral fat (P = 0.039), BMI (P = 0.015), and waist circumference (P = 0.011) at the end point. The consumption of pearl barley with a high β-glucan content reduces not only LDL-C but also visceral fat area.     

Determination of (1→3),(1→4)-β-D-Glucanase Activity by a Calcofluor-Flow Injection Analysis Method

A new methodology for the determination of (1→3),(1→4)-β-d-glucanase activity has been developed. The concentration decay curves corresponding to the depolymerisation of high molecular weight barley (1→3),(1→4)-β-d-glucan by pure (1→3),(1→4)-β-d-glucanase (E.C. 3.2.1.73) fromBacillus subtilisand by crude (1→3),(1→4)-β-d-glucanase from different malts were monitored by the Calcofluor-FIA method. In all cases, the high molecular weight (17Instituto de Agroquı́mica y Tecnologı́a de Alimentos (CSIC)rarr;3),(1→4)-β-d-glucan decay curves fitted very well to an empirical formula describing the change in substrate concentration with time. The curves possess an inflexion point at which the depolymerisation rate of the substrate reaches a maximum. This maximum depolymerisation rate correlates with the initial concentrations of enzyme and substrate,E_oandS_o, and the enzyme kinetic constantsV_mandK_mthrough a hyperbola similar to that of Michaelis-Menten. TheK_mdetermined forB. subtilisβ-glucanase was rather low, about 0·99 g β-glucan/l, when compared with those corresponding to (1→3),(1→4)-β-d-glucanase from different malts, which were, in turn, practically identical at about 2·92 g β-glucan/l. Experiments with barley (1→3),(1→4)-β-d-glucans of different high initial molecular weights showed that initial molecular weight had no influence on the kinetics. Thus, this new methodology permits the determination of (1→3),(1→4)-β-d-glucanase activity in a direct way, i.e. the amount of (1→3),(1→4)-β-d-glucan degraded per amount of enzyme (or malt) per unit of time. Moreover, since it is insensitive to the initial molecular weight of the substrate, it seems to be well-suited for inter-laboratory comparisons of (1→3),(1→4)-β-d-glucanase activities.     

Distribution of (1-3)(1-4)-β-d-glucans in kernels of selected cultivars of naked and hulled barley

The objective of this study was to determine the distribution of (1-3)(1-4)-β-d-glucans in the kernels of three high-glucan lines of naked barley (STH 4561, STH 4671 and STH 4676), a low-glucan line (STH 4572), and two low-glucan comparative cultivars (naked – Rastik, and hulled – Stratus). The content of (1-3)(1-4)-β-d-glucans assayed with the enzymatic method varied within the range from 4.04 to 5.71% dry matter basis. Microspectrophotometric analysis revealed significant differentiation in the structure of the kernels and in the distribution of (1-3)(1-4)-β-d-glucans; it also indicated a necessity of applying such processing methods as are appropriate to the distribution of functional components. The distribution of functional components in the high-glucan lines of naked barley makes them ideal for utilization both in the milling technology and in the production of grits and flakes.     

Barley (1→3; 1→4)-β-glucan and arabinoxylan content are related to kernel hardness and water uptake

Harder kernels in barley are thought to be a factor affecting the modification of the endosperm during malting by restricting water and enzyme movement within the endosperm. The objective of this study was to investigate the relationship between kernel hardness, water uptake and the endosperm composition in barley. A range of barley samples from 2003 and 2004 crops were analyzed for kernel hardness by the Single Kernel Characterization System, water uptake during steeping and chemical composition of the endosperm including (1→3; 1→4)-β-glucan, arabinoxylan and total protein. Both (1→3; 1→4)-β-glucan and arabinoxylan content of the endosperm were correlated significantly with kernel hardness in barley samples from both 2003 (r=0.873 and 0.601, respectively, p<0.01) and 2004 seasons (r=0.764 and 0.501, respectively, p<0.01). Hardness of the kernel was highly correlated with its water uptake in both 2003 and 2004 samples (r=−0.853 and −0.752, respectively, p<0.01). β-Glucan content of the endosperm was also correlated significantly with the kernel water uptake for both years (2003: r=−0.752, p<0.01; 2004: r=−0.551, p<0.01). Arabinoxylan content of the endosperm was correlated significantly with the kernel water uptake for the 2003 barley but not for 2004 barley (2003: r=−0.523, p<0.01; 2004: r=−0.151, p>0.01). Protein content of the endosperm was not correlated with the kernel hardness in either year. These results demonstrate that endosperm cell wall components may have significant impact on kernel hardness as well as water uptake of barley.     

Effects of 1-methylcyclopropene (1-MCP) Treatment on Postharvest Physiology and Quality of ‘Younai' Fruits

研究1.2 μL/L的1-甲基环丙烯(1-methylcyclopropene,1-MCP)处理对在(25±1)℃下贮藏的‘油檬’(Prunus salicina Lindl.cv.Younai)果实采后生理和品质的影响.结果表明,与对照果实相比,1-MCP处理可有效降低‘油(檫)’果实呼吸强度、呼吸峰值和乙烯释放量,延缓果实细胞膜相对渗透率升高和果实表面色调角h.值的下降,保持较高的果实硬度、可滴定酸和果皮叶绿素含量,延迟果实外观颜色转变,减少果实失重和腐烂.经1-MCP处理的果实在(25±1)℃下贮藏15d时的好果率为80％,而对照果实只有58％.因此认为,1-MCP处理可以延缓采后‘油榇’果实后熟衰老和保持果实品质,延长果实保鲜期.     

The Effect of Sulfated (1→3)-α-L-Fucan from the Brown Alga Saccharina cichorioides Miyabe on Resveratrol-Induced Apoptosis in Colon Carcinoma Cells

Accumulating data clearly indicate that the induction of apoptosis by nontoxic natural compounds is a potent defense against the development and progression of many malignancies, including colon cancer. Resveratrol and the fucoidans have been shown to possess potent anti-tumor activity in vitro and in vivo. The aim of the present study was to examine whether the combination of a fucoidan from the brown alga Saccharina cichorioides Miyabe and resveratrol would be an effective preventive and/or therapeutic strategy against colon cancer. Based on NMR spectroscopy and MALDI-TOF analysis, the fucoidan isolated and purified from Saccharina cichorioides Miyabe was (1→3)-α-L-fucan with sulfate groups at C2 and C4 of the α-L-fucopyranose residues. The fucoidan enhanced the antiproliferative activity of resveratrol at nontoxic doses and facilitated resveratrol-induced apoptosis in the HCT 116 human colon cancer cell line. Apoptosis was realized by the activation of initiator caspase-9 and effector caspase-7 and -3, followed by the cleavage of PARP. Furthermore, significant inhibition of HCT 116 colony formation was associated with the sensitization of cells to resveratrol by the fucoidan. Taken together, these results demonstrate that the combination of the algal fucoidan with resveratrol may provide a potential therapy against human colon cancer.     

Effects of Cultivar and Environment on β-(1,3)-(1,4)-D-glucan Content and Acid Extract Viscosity of Spanish Barleys

The acid extract viscosities and β-glucan contents of ten two- and six-rowed barley cultivars grown at seven locations in three consecutive years in Spain were studied in the present work. The viscosities varied from 2·4 to 24·8 centistokes (cSt) and the mean value was 6·4 cSt. The average β-glucan content of barleys determined by HPLC was 3·5% with a range of 1·9–5·5%. Significant differences were found in both β-glucan content and acid extract viscosity between different cultivars, locations and years. The β-glucan contents and viscosities of winter cultivars were higher than those of spring. Cvs. Barbarrosa and Hatif de Grignon were the genotypes with the highest values for both parameters, while cv. Beka had the lowest viscosity and β-glucan content. Environmental factors influenced both parameters. The acid extract viscosities of barleys were correlated negatively with the amount of precipitation (r=−0·754;P<0·05). Barleys grown in wet and rainy areas (Girona and La Coruña) had lower viscosity values.     

Characterization and online detection of surfactin isomers based on HPLC-MS(n) analyses and their inhibitory effects on the overproduction of nitric oxide and the release of TNF-α and IL-6 in LPS-induced macrophages

A rapid method for characterization and online detection of surfactin isomers was developed based on HPLC-MS(n) (n = 1, 2, 3) analyses, and many surfactin isomers were detected and characterized from the bioactive fraction of the mangrove bacterium Bacillus sp. Inhibitory activities of surfactin isomers on the overproduction of nitric oxide and the release of TNF-α and IL-6 in LPS-induced macrophages were systematically investigated. It was revealed that the surfactin isomers showed strong inhibitory properties on the overproduction of nitric oxide and the release of IL-6 on LPS-induced murine macrophage cell RAW264.7 with IC(50) values ranging from 1.0 to 7.0 μM. Structure-activity relationship (SAR) studies revealed that the existence of the free carboxyl group in the structure of surfactin isomers was crucial. These findings will be very helpful for the development of this novel kind of natural product as new anti-inflammatory agents.     

Effects of microbial,organically acceptable,and reduced risk insecticides on Anthonomus signatus (Curculionidae: Coleoptera) in strawberries (Fragaria × ananassa)

Anthonomus signatus, the strawberry bud weevil, is active in early spring coinciding with the bloom period of host plants and when managed and unmanaged pollinators are active. Female A. signatus cause injury to the host plant during egg laying when they deposit a single egg inside an unopened flower bud and then girdle or “clip” the bud at the pedicel. Past efforts to control A. signatus populations have relied on the use of broad spectrum insecticides. Fields are either treated at the first signs of damage, or precautionary treatments may be applied prophylactically in fields with a history of damage. Because A. signatus damage occurs during bloom, there is concern about the potential harm to pollinators caused by these treatments. In order to identify materials more compatible for use during bloom, the efficacy of reduced risk pesticides against A. signatus was tested in semi-field bioassays over two years. Beauveria bassiana, cyantraniliprole, novaluron, and pyrethrins were assessed in the first year, and none caused significant A. signatus mortality. Acetamiprid, flonicamid, and spinosad were compared in the second year, acetamiprid and spinosad had higher A. signatus mortality than an untreated control, and exposure to spinosad resulted in similar mortality to bifenthrin, an industry standard material.     

Rove beetles (Coleoptera Staphylinidae) – Their abundance and competition with other predatory groups in Bt maize expressing Cry34Ab1, Cry35Ab1, Cry1F and CP4 EPSPS proteins

This study investigated the effects of different GM maize crops on the abundances of rove beetles. It also investigated competition between rove beetles and other epigeic arthropod predatory groups (ground beetles and spiders). Altogether a total of 5926 rove beetle individuals belonging to 25 species were collected. Whilst the average number of rove beetle individuals per replicate per sampling date varied and, moreover, decreased each year, there were no observed differences in rove beetle abundance when compared between isogenic and GM maize plots within the study period. Furthermore, no clear trends on positive correlations between weed coverage and rove beetle abundances were detected. Altogether a total number of 76,070 ground beetle individuals and 11,126 spiders (including all Araneae) were collected with pitfall traps. Analyses revealed a strong negative relationship between rove beetles and the other two groups in each treatment and year. According to the results the effect of interspecific competition on rove beetle abundance in GM maize is much higher than the effects of treatments including both Bt toxins and extra glyphosate applications.     

Accumulation of mixed linkage (1 → 3) (1 → 4)-β-d-glucan during grain filling in barley: A vibrational spectroscopy study

The accumulation of mixed linkage barley (1 → 3) (1 → 4)-β-d-glucan (BG) during grain filling at eight stages was studied using standard reference methods and infrared spectroscopy. Two mutant barley genotypes having higher (starch mutant lys5f) and lower (high lysine mutant lys3a) BG content than the normal control Cork were studied. The Cork and lys3a genotypes showed a linear BG accumulation throughout the grain filling to reach a maximum of approximately 6 and 4% BG (w/w) dry matter, respectively. However, lys5f mutant exhibited an exponential increase in BG synthesis to a maximum of approximately 18% BG (w/w) dry matter 30 days after flowering (DAF), seemingly compensating for a decreased synthesis of starch.