Macrophylloside, a flavone glucoside from Primula macrophylla

A new flavone glucoside macrophylloside has been isolated from the whole plant of Primula macrophylla and its structure was determined by spectroscopic methods as 2′-hydroxy-7-O-β- -glucopyranosyloxyflavone. Sitosterol glucoside was also isolated for the first time from this plant.     

A divergent synthesis of [1-14C]-mono-E isomers of fatty acids

A convenient synthesis of [1-14C]-mono-trans fatty acid using olefin inversion as a key-step is described. This methodology allows for a facile synthesis of [1-14C]-labelled mono-trans analogues of oleic, linoleic and linolenic acids. As an example, only eleven steps were necessary to obtain the [1-14C]-mono-E isomers of linolenic acid from its commercial all-Z form. In the first step, Barton's decarboxylation procedure yielded a bromo intermediate. Epoxidation of this compound resulted in the formation of three monoepoxides, which could be separated by HPLC. After identification by 1H NMR and MS, the pure monoepoxides were then subjected to inversion consisting of a stereospecific deoxygenation followed by a beta-elimination step. Finally, the labelling was introduced by substitution of the bromine by a [14C]-cyano group followed by hydrolysis.     

Side reactions in the synthesis of phosphotyrosine-containing peptides

Summary A series of phosphopeptides Tyr(PO₃H₂)-Val-Pro-Xxx-Leu (Xxx=Met, Met(O), Nle, Dab or Cys), derived from the native platelet-derived growth factor- receptor (PDGF-) sequence, has been prepared to study their interaction with the src-homology 2 (SH2) domains of the p85 subunit of PI3 kinase. The phosphopeptides were synthesized using Fmoc methodology incorporating N-Boc dibenzyl-protected phosphotyrosine (Boc-Tyr[PO₃(Bzl)₂]) as the N-terminal amino acid, since the benzyl groups can be removed during resin cleavage with TFA. Only peptides containing methionine were found to exist partially as S-benzyl sulfonium salts after TFA cleavage from the resin. The desired peptide could be obtained from the S-benzyl sulfonium salt by hydrogenolysis.     

Solid-phase synthesis of human salivary mucin-derived O-linked glycopeptides

Summary Short glycopeptides derived from salivary mucin have been synthesized in order to delineate the O-glycosylation pattern that is important in the biological activity of mucin. Two glycopoptides, APPETT*AAP-OMe and PAPPSS*SAP-OMe (*=-d-GalNAc), were prepared by solid-phase peptide synthesis integrating the Fmoc and Boc strategies. Since these peptides contain a C-terminal proline, we devised an efficient strategy using facile Boc chemistry, where the glycosylation at the desired position in the sequence was achieved using corresponding Fmoc-glycoamino acid esters A and B as building blocks. The transformation of the 2-azido group into the acetamido derivative was performed with thioacetic acid on the polymer-bound glycopeptides. Corresponding nonglycosylated peptides were also synthesized to study the influence of -d-GalNAc on peptide backbone conformation.     

DNA repair and replication in human fibroblasts treated with (±)-r-7,t-8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene

DNA repair and replication were examined in diploid human fibroblasts after treatment with (±)-r-7,t-8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE-I). Unscheduled DNA synthesis exhibited a linear response to BPDE-I concentrations up to 1.5 μM and a saturation plateau after higher concentrations. Maximal unscheduled DNA synthesis was observed in the first hour after treatment with synthesis diminishing progressively thereafter. Half-maximal unscheduled DNA synthesis was seen within 4–6 h after treatment with 0.7 μM BPDE-I. DNA replication was inhibited by BPDE-I in a dose- and time-dependent fashion. The mechanisms of this inhibition were characterized by velocity sedimentation of pulse-labeled nascent DNA in alkaline sucrose gradients. Very low concentrations of BPDE-I (0.03 and 0.07 μM) were found to inhibit replicon initiation by up to 50% within 30–60 min after treatment. Recovery of initiation following these low concentrations was evident within 3 h after treatment. Higher concentrations of carcinogen inhibited DNA synthesis in active replicons. This effect was manifested by a reduction in incorporation of precursor into replication intermediates of greater than 1·10⁷ Da with the concurrent production of abnormally small nascent DNA. When viewed 45 min after treatment with 0.17 μM BPDE-I the combination of these two effects partially masked the inhibition of replicon initiation. However, even after treatment with 0.33 μM BPDE-I an effect on initiation was evident. These results reveal a pattern of response to BPDE-I that is quite similar to that produced by 254 nm radiation.     

Inactivation of Escherichia coli O157:H7 and Salmonella on artificially or naturally contaminated mung beans (Vigna radiata L) using a stabilized oxychloro-based sanitizer

AIMS: To evaluate the efficacy of a stabilized oxychloro-based (SOC) sanitizer to decontaminate mung beans artificially or naturally contaminated with Escherichia coli O157:H7 or Salmonella. METHODS AND RESULTS: Naturally contaminated beans were produced by introducing a five-strain cocktail of E. coli O157:H7 or Salmonella onto the flowers of growing mung bean plants. Escherichia coli O157:H7 was only sporadically recovered from sprout lots (three testing positive from 10 tested) derived from harvested beans. In contrast, Salmonella was recovered from 18 of 20 lots screened. Pathogens present on naturally contaminated seed could be successfully inactivated with SOC applied at 200 ppm for 24 h at 28 degrees C. SOC treatment could also decontaminate artificially inoculated mung bean batches containing different levels of contaminated seed. SOC inactivated E. coli O157:H7, but not Salmonella introduced onto damaged (scarified) beans. CONCLUSIONS: SOC sanitizer could inactivate Salmonella or E. coli O157:H7 naturally or artificially introduced onto mung beans. However, the SOC treatment failed to inactivate Salmonella introduced onto damaged mung beans. SIGNIFICANCE AND IMPACT OF THE STUDY: SOC sanitizer represents an effective method for decontaminating undamaged mung beans.     

Genetical investigations into β-glucan content in barley

Summary Random inbred lines produced by doubled haploidy (DH) and single seed descent (SSD) have been used to investigate the genetics of -glucan (gum) content in barley (Hordeum vulgare). Genetical analyses indicated that gum content is controlled by a simple additive genetic system. Significant negative genetic correlations were observed between -glucan content, thousand grain weight and height in the DH samples. These correlations were much reduced in the SSD samples and would suggest linkage of the genes controlling these characters. The presence of repulsion linkages could be exploited in a barley breeding programme by producing F1 derived DH to generate recombinants with high thousand grain weight and low -glucan content. Genetical parameters estimated from DH and F3 samples have successfully been used to predict the number of inbred lines transgressing the parental range for -glucan content and bivariate combinations involving -glucan.     

N-ω-chloroacetyl-l-ornithine,a new competitive inhibitor of ornithine decarboxylase,induces selective growth inhibition and cytotoxicity on human cancer cells versus normal cells

Abstract

Many cancer cells have high expression of ornithine decarboxylase (ODC) and there is a concerted effort to seek new inhibitors of this enzyme. The aim of the study was to initially characterize the inhibition properties, then to evaluate the cytotoxicity/antiproliferative cell based activity of N-ω-chloroacetyl-l-ornithine (NCAO) on three human cancer cell lines. Results showed NCAO to be a reversible competitive ODC inhibitor (K_i?=?59?µM) with cytotoxic and antiproliferative effects, which were concentration- and time-dependent. The EC_50,72h of NCAO was 15.8, 17.5 and 10.1?µM for HeLa, MCF-7 and HepG2 cells, respectively. NCAO at 500?µM completely inhibited growth of all cancer cells at 48?h treatment, with almost no effect on normal cells. Putrescine reversed NCAO effects on MCF-7 and HeLa cells, indicating that this antiproliferative activity is due to ODC inhibition.     

The elimination ofO-linked glycans from glycoproteins under non-reducing conditions

A simple procedure is described for the elimination ofO-linked glycans from bovine submaxillary mucin under non-reducing conditions, using triethylamine in aqueous hydrazine. The glycans were isolated as the hydrazones, which were converted to the reducing glycans by exchange with acetone in neutral aqueous solution. The glycan alditols obtained after reduction corresponded to those obtained by the reductive -elimination ofO-glycans.