A 24β-ethyl-Δ7-steryl glucopyranoside from Cucurbita pepo seeds

In the seeds of Cucurbita pepo three closely related 24-ethyl-Δ⁷-steryl glucosides were identified by hydrolytic studies and spectral analysis as spinasteryl-β-D-glucopyranoside, the new 3-O-(β-D-glucopyranosyl)-24β-ethyl-5α-cholesta-7,25(27)-dien-3β-ol and the corresponding Δ^22E,25,(27)-trienol. Except for its occurrence in cucumber seeds the latter is so far unknown as a natural product.     

Malonylation is a key reaction in the metabolism of xenobiotic phenolic glucosides in Arabidopsis and tobacco

Tobacco cells (Nicotiana tabacum L.) accumulate harmful naphthols in the form of malonylated glucosides ( Taguchi et al., 2005 ). Here, we showed that the malonylation of glucosides is a system to metabolize xenobiotics and is common to higher plants. Moreover, some plantlets including Arabidopsis thaliana excreted some of the incorporated naphthols into the culture media as their glucosides. In order to analyze the function of malonylation in the metabolism of these xenobiotics, we identified a malonyltransferase gene (At5g39050) responsible for the malonylation of these compounds in A. thaliana. The recombinant enzyme had malonyltransferase activity toward several phenolic glucosides including naphthol glucosides. A knockout mutant of At5g39050 (pmat1) exposed to naphthols accumulated only a few malonylglucosides in the cell, and released larger amounts of simple glucosides into the culture medium. In contrast, forced expression of At5g39050 in the pmat1 mutant resulted in increased malonylglucoside accumulation and decreased glucoside excretion to the media. The results provided clear evidence of whether the release of glucosides or the storage of malonylglucosides was determined by the At5g39050 expression level. A similar event in naphthol metabolism was observed in the tobacco mutant with a suppressed malonyltransferase gene (NtMaT1). These results suggested that malonylation could be a key reaction to separate the way of xenobiotics disposition, that is, release from cell surface or storage in vacuoles.     

Effects of toxic β‐glucosides on carbohydrate metabolism in cotton bollworm,Helicoverpa armigera (Hübner)

The purpose of this study was to evaluate the effects of three toxic β‐glucosides, phlorizin, santonin, and amygdalin, on carbohydrate metabolism in the cotton bollworm, Helicoverpa armigera (Hübner), when diets mixed with β‐glucosides were fed to third‐instar larvae. The growth of the larvae was significantly inhibited by exposure to santonin after 96 hr but not obviously affected by phlorizin and amygdalin. The midgut trehalase activities were only 51.7%, 32%, and 42.5% of that of the control after treatment with phlorizin, santonin and amygdalin at 2 mg/ml, respectively. In the hemolymph and fat body, the amount of trehalose decreased in all cases. However, the effects of santonin on the alteration of the glycogen and glucose levels as well as the activities of glycogen phosphorylase, were different than those of the other two β‐glucosides. It appears that the three β‐glucosides have different influences on the carbohydrate metabolism of cotton bollworm.     

Beta-glucosidase: substrate, solvent, and viscosity variation as probes of the rate-limiting steps

The second-order rate constants (kcat/Km) for the beta-glucosidase-catalyzed hydrolysis of aryl beta-D-glucopyranosides show a bell-shaped dependence of pH. The pKas that characterize this dependence are 4.4 (delta Hion approximately equal to 0) and 6.7 (delta Hion approximately equal to 0). In D2O these pKas are increased by 0.5 (+/- 0.1) unit, but there is no solvent isotope effect on the pH-independent second-order rate constant. Nath and Rydon [Nath, R. L., & Rydon, H. N. (1954) Biochem. J. 57, 1-10] examined the kinetics of the beta-glucosidase-catalyzed hydrolysis of a series of substituted phenyl glucosides. We have extended this study to include glucosides with phenol leaving groups of pKa less than 7. Br?nsted plots for this extended series were nonlinear for both kcat/Km and kcat. Br?nsted coefficients for those compounds with leaving groups of pKa greater than 7 (for kcat/Km) or pKa greater than 8.5 (for kcat) were nearly equal to -1.0, indicating substantial negative charge buildup on the leaving group in the transition state. The nonlinearity indicates an intermediate in the reaction. This was confirmed by partitioning experiments in the presence of methanol as a competing glucose acceptor. A constant product ratio, [methyl glucoside]/[glucose], was found with aryl glucoside substrates varying over 16,000-fold in reactivity (V/K), indicative of a common intermediate. Viscosity variation (in sucrose-containing buffers) was used to probe the extent to which the beta-glucosidase reactions are diffusion-controlled.(ABSTRACT TRUNCATED AT 250 WORDS)     

Utilizing genetically engineered bacteria to produce plant-specific glucosides

Plant-derived glucosides have attracted much attention due to their widespread applications. This class of products is difficult to isolate or to synthesize in pure form because of the resulting low yields. Thus, simple approaches for the generation of such glucosides would be highly beneficial. We purified and characterized a novel glucosyltransferase from plant cell suspension cultures of Rauvolfia serpentina, which showed rather low substrate specificity. We obtained its cDNA and expressed the active recombinant protein in bacteria (Escherichia coli) with excellent plant-specific glucosylation efficiencies. Compared with the plant system, the bacteria delivered the new enzyme, which was in the form of a soluble or matrix-bound enzyme, approximately 1800 times more efficiently for the synthesis of a wide range of glucosides. More importantly, the engineered E. coli strain allowed for in vivo glucosylation and release of the product into the culture medium, as shown by the formation of arbutin, which is a potent inhibitor of human melanin biosynthesis with commercial value.     

A new flavone xyloside and two new flavan-3-ol glucosides from Juniperus communis var. depressa

A new flavone xyloside, 1, and two new flavan-3-ol glucosides, 3 and 4, were isolated together with three known flavones, 2, 11, and 12, five known flavans, 5-9, and a known dihydrochalcone, 10, from the stems and leaves of Juniperus communis var. depressa (Cupressaceae) collected in Oregon, U.S.A., and their structures were determined on the basis of spectral evidence. A novel flavone nucleus such as that in 1 is seldom found in nature today, and new methylcatechin glucosides 3 and 4 are also rare in nature. In addition, we investigated the inhibitory activity of individual components, i.e., 8-11, and others, that were abundantly isolated from the same plant material for the Maillard reaction.     

Analysis of Rhizoma Polygoni Cuspidati by HPLC and HPLC-ESI/MS

An HPLC method with photodiode array detection (PAD) and ESI/MS detection was developed for the qualitative and quantitative analysis of the major chemical constituents of the dried rhizome of Polygonum cuspidatum Sieb. et Zucc. (Rhizoma Polygoni Cuspidati; Chinese name Hu-Zhang). Based on the chromatographic separation on an Altima C(18) column using 0.5% aqueous acetic acid and acetonitrile as the mobile phase, nine compounds, including stilbenes, stilbene glucosides, anthraquinones and anthraquinone glucosides, were identified by online ESI/MS analysis and seven were quantified by HPLC-PAD. A full validation of the method including sensitivity, linearity, repeatability and recovery was conducted. Linear calibration was achieved over the concentration range 1-200 mg/L with R(2) > 0.999, whilst the limits of detection ranged from 0.51 to 1.57 ng. Repeatability was evaluated by intra- and inter-day assays and the RSD value was within 1.79%. Recoveries of the quantified compounds were within the range 96.0-100.1% with RSD values of less than 2.2%. Five samples of Rhizoma Polygoni Cuspidati from different regions were analysed using the developed method. The major constituents piceid, resveratrol, emodin-8-O-beta-D-glucoside and emodin were selected to provide an index for the quality assessment of the herbal drug.     

Diversification of an ancient theme: hydroxynitrile glucosides

Many plants produce cyanogenic glucosides as part of their chemical defense. They are alpha-hydroxynitrile glucosides, which release toxic hydrogen cyanide (HCN) upon cleavage by endogenous plant beta-glucosidases. In addition to cyanogenic glucosides, several plant species produce beta- and gamma-hydroxynitrile glucosides. These do not release HCN upon hydrolysis by beta-glucosidases and little is known about their biosynthesis and biological significance. We have isolated three beta-hydroxynitrile glucosides, namely (2Z)-2-(beta-D-glucopyranosyloxy)but-2-enenitrile and (2R,3R)- and (2R,3S)-2-methyl-3-(beta-D-glucopyranosyloxy)butanenitrile, from leaves of Ribesuva-crispa. These compounds have not been identified previously. We show that in several species of the genera Ribes, Rhodiola and Lotus, these beta-hydroxynitrile glucosides co-occur with the L-isoleucine-derived hydroxynitrile glucosides, lotaustralin (alpha-hydroxynitrile glucoside), rhodiocyanosides A (gamma-hydroxynitrile glucoside) and D (beta-hydroxynitrile glucoside) and in some cases with sarmentosin (a hydroxylated rhodiocyanoside A). Radiolabelling experiments demonstrated that the hydroxynitrile glucosides in R. uva-crispa and Hordeum vulgare are derived from L-isoleucine and L-leucine, respectively. Metabolite profiling of the natural variation in the content of cyanogenic glucosides and beta- and gamma-hydroxynitrile glucosides in wild accessions of Lotus japonicus in combination with genetic crosses and analyses of the metabolite profile of the F2 population provided evidence that a single recessive genetic trait is most likely responsible for the presence or absence of beta- and gamma-hydroxynitrile glucosides in L. japonicus. Our findings strongly support the notion that the beta- and gamma-hydroxynitrile glucosides are produced by diversification of the cyanogenic glucoside biosynthetic pathway at the level of the nitrile intermediate.     

5-O-glucosyldihydroflavones from the leaves of Helicia cochinchinensis

From the leaves of Helicia cochinchinensis, collected on Okinawa Island, seven phenolic glucosides and two terpenic glucosides were isolated. Five of the phenolic glucosides were previously known, being identified with p-coumaric and ferulic acids glucosyl esters, rhodioloside, helicidiol, and naringenin 5-O-beta-D-glucopyranoside. The structures of two other phenolic glucosides, named heliciosides A and B, were elucidated to be 5-O-beta-D-glucosides of 3-hydroxyflavanone, namely aromadendrin and taxifolin, by means of spectroscopic analyses. The two terpenic glucosides were identified with ampelopsisionoside and icariside C1.     

Purification and kinetic mechanism of the major glutathione S-transferase from bovine brain.

This paper addresses the similarities and differences in the topology of the catalytic centres of human liver cytosolic beta-glucosidase and placental lysosomal glucocerebrosidase, and utilizes well-documented reversible active-site-directed inhibitors. This comparative kinetic study was performed mainly to decipher the chemical and structural nature of the active site of the cytosolic beta-glucosidase, whose physiological function is unknown. Specifically, analysis of the effects of a family of alkyl beta-glucosides consistently displayed 100-250-fold lower inhibition constants with the cytosolic broad-specificity beta-glucosidase compared with the placental glucocerebrosidase; for example, with octyl beta-D-glucoside the Ki values were 10 microM and 1490 microM for the cytosolic and lysosomal beta-glucosidases respectively. Furthermore the higher affinity of the cytosolic beta-glucosidase than glucocerebrosidase for the amphipathic alkyl beta-D-glucosides was validated by the greater increase in the free energy of binding with increasing alkyl chain length [delta delta G0 (K,)/CH2: lysosomal enzyme, 2.01 kJ/mol (480 cal/mol); cytosolic enzyme, 3.05 kJ/mol (730 cal/mol)]. The implications of the presence of highly non-polar domains in the active site of the cytosolic beta-glucosidase are discussed with regard to its potential physiological substrates.     

Isolation and purification of closely related citrus limonoid glucosides by flash chromatography

Several citrus limonoid glycosides have proved to be particularly difficult to purify using conventional techniques. A reversed-phase flash chromatographic technique has been developed for the separation and isolation of the closely related limonoid glucosides, nomilin 17-beta-D-glucopyranoside and nomilinic acid 17-beta-D-glucopyranoside, with confirmation of their identities by electrospray ionisation mass spectrometry. Furthermore, the semi-purification of the mixture of glucosides enriched with flavanone glucosides such as naringin, narirutin and other limonoid glucosides was obtained. The closely eluting glucosides were successfully separated to achieve a good yield and purity of 93%.     

Iridoid glucosides from roots of Vietnamese Paederia scandens

Four iridoid glucosides, three of which are dimeric were isolated from the methanol extract of roots of Vietnamese Paederia scandens (Lour) Merrill together with the five known glucosides, paederoside, asperuloside, paederosidic acid, asperulosidic acid and geniposide. Seven sulfur-containing iridoid glucosides were also isolated. The structures of the iridoid glucosides were determined by a combination of high-resolution NMR, MS, IR and UV spectra, and chemical reaction such as acetylation.     

Polar constituents of celery seed

From the water-soluble portion of the methanol extract of celery seed (fruit of Apium graveolens L.) five sesquiterpenoid glucosides (celerioside A-E) and three phthalide glycosides (celephtalide A-C) were isolated together with six aromatic compound glucosides, two norcarotenoid glucosides and a lignan glucoside. Their structures were determined by spectral investigations.     

Sexually dimorphic gall structures correspond to differential phytohormone contents in male and female wasp larvae

Sexually dimorphic galls are rare among gall‐inducing insects and the reason for their occurrence is unknown. The pteromalid wasp Trichilogaster acaciaelongifoliae, which induces galls on Acacia longifolia, is one such species. In the present study, the anatomical and physiological attributes of male and female galls of T. acaciaelongifoliae are examined and compared. Histological preparations are used to characterize anatomical differences between male and female gall chambers. Bioassays, high‐performance liquid chromatography‐mass spectrometry and an enzyme immunoassay are used to measure concentrations of auxin and cytokinin in normal buds, galled tissues, and larvae of both sexes. Female chambers are found to be 3.3‐fold larger, and are associated with 1.5‐fold more storage tissue and 3.5‐fold more vascular tissues than male chambers. Tissues from female chambers induce stronger cytokinin‐like bioactivity than tissues from male chambers. Female larvae have considerably higher concentrations of cytokinin free bases, ribosides, glucosides and monophosphates than male larvae; higher auxin‐like bioactivity than in normal or galled plant tissues; and almost twice the concentration of auxin than male larvae. Both male and female larvae contain much higher auxin concentrations than either galled or normal plant tissues. These findings suggest that differing levels of phytohormones are involved in the development of sexual dimorphism of gall structures in this species.     

Lanceocrepidiasides A-F, glucosides of guaiane-type sesquiterpene from Crepidiastrum lanceolatum

From the aerial parts of Crepidiastrum lanceolatum, six guaiane-type sesquiterpene glucosides, lanceocripidiasides A-F were isolated together with five known sesquiterpene glucosides, ixerin Y, crepidialanceosides A and B, and youngiasides A and D, two known megastigmane glucosides, icariside B1 and corchoionoside A, and benzyl 6'-O-beta-D-apiofuranosyl-beta-D-glucopyranoside. Structures were elucidated by spectroscopic analyses.     

Molecular characterization of flavonoid malonyltransferase from Oryza sativa

In this study, a flavonoid malonyltransferase (OsMaT-2) was cloned from Oryza sativa, and the recombinant protein OsMaT-2 was purified via affinity chromatography. OsMaT-2 utilized a variety of flavonoid glucosides, including flavanone glucosides, flavone glucosides, flavonol glucosides, and isoflavone glucosides as substrates, but did not utilize anthocyanin. As an acyl donor, OsMaT-2 utilized only malonyl-CoA. Based on reactions with various quercetin 3-O-sugars, we identified the probable position of malonylation as the 6″-hydroxyl group of the sugar. This is the first report, to the best of our knowledge, of the cloning of a flavonoid malonyltransferase from O. sativa.     

Triterpenoids and glycosides from Geum japonicum

Two new glucosides and two known ester glucosides have been isolated from Geum japonicum. The two new glucosides were isolated by formation of their acetates and were identified as glucosides of 2-isopropyl-5-methylhydroquinone by chemical and spectral studies. The two known ester glucosides were identified as niga-ichigoside F1 and suavissimoside F1 by direct comparison with authentic samples. 2α,19α-Dihydroxyursolic acid and the known glycoside, gein, were also isolated from the same plant, in addition to a mixture of 2α-hydroxyursolic acid and 2α-hydroxyoleanolic acid.     

Isolation and measurement of quercetin glucosides in flower buds of Japanese butterbur (Petasites japonicus subsp. gigantea Kitam.)

Three quercetin glucosides were isolated from flower buds of Japanese butterbur (Petasites japonicus subsp. gigantea Kitam.) together with caffeic acid as the ingredients that had DPPH radical scavenging activity, using the DPPH-HPLC method for measuring the radical scavenging activity. These quercetin glucosides were identified as quercetin 3-O-beta-D-glucoside, quercetin 3-O-beta-D-6'-O-acetylglucoside, and rutin, and the amounts of the glucosides in flower buds were also examined by HPLC. The flower buds were harvested from four different sites, the total amount of quercetin glucosides in each site was 100-170 mg/100 g fr. wt., and there were no great differences of the amounts between growing fields.     

Glucosides from the roots of Capparis tenera

Two new lignan glucosides, compounds 2 and 3, two new 1H-indole-alkaloid glucosides, 5 and 6, as well as two new phenolic glucosides, 7 and 10, were isolated from the roots of Capparis tenera, together with five known compounds. Their structures were characterized by chemical and spectroscopic methods. Most of these isolates were obtained for the first time from Capparidaceae. The antioxidant and anti-inflammatory activities of the new compounds were investigated.     

Iridoid glucosides from Satureja Vulgaris

Satureja vulgaris was shown to contain two new iridoid glucosides, 5-deoxylamiol and 4-methylantirrhinoside, as well as the known iridoid glucosides lamiol and 5-deoxylamioside. The structures of the new glucosides were established by spectroscopic studies and chemical evidence.