Bioactive metabolites isolated from Penicillium sp. YY-20, the endophytic fungus from Ginkgo biloba.

Six known metabolites, adenosine (1), methyl ß-D-ribofuranoside (2), adenine (3), 2'-deoxyadenosine (4), 3-methylpiperazine-2,5-dione (5) and 2'-deoxyuridine (6), were isolated from the extracts of the endophytic fungus Penicillium sp. YY-20 isolated from the root of Ginkgo biloba, and their structures were elucidated by spectroscopic methods. The antioxidant and growth-promoting activities of these compounds were first evaluated. The results indicated that compounds 1, 3 and 4 exhibited potential DPPH-scavenging activities compared with positive control. In addition, all the compounds (except 5) stimulated seed germination of Raphanus sativus, Brassica napus and Brassica chinensis but had weak stimulating effect on their root and hypocotyl growth.

Deoxyuridines from the marine sponge associated actinomycete Streptomyces microflavus.

One new nucleoside derivative, named 3-acetyl-5-methyl-2'-deoxyuridine (1), along with two known compounds 3,5-dimethyl-2'-deoxyuridine (2) and 3-methyl-2'-deoxyuridine (3), were isolated from the cultures of Streptomyces microflavus. This strain was an associated actinomycete isolated from the marine sponge Hymeniacidon perlevis collected from the coast of Dalian (China). Their structures were elucidated by detailed NMR and MS spectroscopic analysis as well as comparison with literature data.

Identification and characterization of 2'-deoxyuridine from the supernatant of conidial suspensions of rice blast fungus as an infection-promoting factor in rice plants.

We previously detected infection-promoting activity in the supernatant of the conidial suspension (SCS) of the rice blast fungus. In the present study, a molecule carrying the activity was purified and identified as 2'-deoxyuridine (dU). The infection-promoting activity of dU was strictly dependent on its chemical structure and displayed characteristics consistent with those of the SCS. Notably, the activity of dU was exclusively detected during interactions between rice and virulent isolates of the fungus, the number of susceptible lesions in leaf blades was increased by dU, and nonhost resistance in rice plants was not affected by treatment with dU. In addition, the expression of pathogensis-related genes, accumulation of H(2)O(2), and production of phytoalexins in rice in response to inoculation with virulent fungal isolates was not suppressed by dU. The infection-promoting activity of dU was not accompanied by elevated levels of endogenous abscissic acid, which is known to modify plant-pathogen interactions, and was not detected in interactions between oat plants and a virulent oat blast fungus isolate. Taken together, these results demonstrate that dU is a novel infection-promoting factor that acts specifically during compatible interactions between rice plants and rice blast fungus in a mode distinct from that of toxins and suppressors.

Marine sesquiterpenoids that inhibit the lyase activity of DNA polymerase beta.

Bioassay-directed fractionation of an extract of the marine species Spongia sp. led to the discovery of the new sesquiterpenoid derivative 17-O-isoprenyldictyoceratin-C (1), the known sesquiterpenoid derivative dictyoceratin-C (2), and the sesquiterpenoid quinone ilimaquinone (3), in addition to the nucleoside 2'-deoxyuridine. The structure of the new compound 1 was determined on the basis of spectroscopic methods and by conversion of dictyoceratin-C (2) to 1.

[Development of methods for the chromatographic analysis of Hevizos]. / A hevizos kromatográfiás analitikai módszereinek kifejlesztése.

Normal phase HPLC and OPTLC methods are described for the analysis of Hevizos (5-isopropyl-2'-beta-deoxyuridine) and it's impurities. The eluent is 50% water saturated ethyl-acetate to which 0.5-1.5% methanol is added depending on the silica used. The water content is important as it practically eliminates tailing. The methods can also be used for the determination of 5-isopropyl-2'-beta-deoxyuridine and it's impurities in Hevizos ointment after a simple extraction procedure. The OPTLC method uses a CHROMPRES 10 apparatus, aluminum backed HPTLC plates and the same eluent-system as the HPLC method. The reproducibility of the HPLC method (rel. st. dev.) is 0.8%, the correlation coefficient is > 0.999 for the main component, while the impurities can be determined at the 0.1% level with a reproducibility of 20%, and a correlation coefficient of > 0.997. The reproducibility of the OPTLC method (rel. st. dev.) is 4% for the main component, while for the impurities at the 0.5% level the reproducibility is 20%.

Mass spectrometry of electrophore-labeled nucleosides. Pentafluorobenzyl and cinnamoyl derivatives.

Three structurally similar deoxynucleosides (thymidine, O4-ethylthymidine, and 2'-deoxyuridine) were studied by mass spectrometry as pentafluorobenzyl, cinnamyl, or mixed derivatives. The purpose of the work was to define the usefulness of such derivatives for structural elucidation of deoxynucleosides. The compounds were ionized in three ways: electron capture negative ion, positive ion chemical ionization, and electron impact. For each of the derivatives examined, the combined spectra were well suited for structural elucidation purposes.

Simple separation of tritiated water and [3H]deoxyuridine from [5-3H]deoxyuridine 5'-monophosphate in the thymidylate synthase assay.

A simple micromethod was developed for the accurate measurement of the activity of dTMP synthase in rat liver crude extracts. The reaction product of dTMP synthase activity assay, i.e., tritiated water, generated by the release of tritium from carbon-5 of [5-3H]deoxyuridine 5'-monophosphate (dUMP), was separated simply by 100% KOH absorption from [5-3H]deoxyuridine (dUrd), which is the side-product by dephosphorylation of [5-3H]deoxyuridine (dUrd), which is the side-product by dephosphorylation of [5-3H]dUMP during the enzyme reaction. Tritiated water was trapped in three droplets of 100% KOH deposited on the underside of the vessels' lids, while [3H]dUrd remained in the bottom of vessels after absorption of the substrate, [5-3H]dUMP, from the reaction mixture by charcoal treatment. Under standard assay conditions in the crude extract of rat liver, the specific activities of dTMP synthase and dUMP phosphatase were 0.092 +/- 0.002 and 0.351 +/- 0.013 nmol/h/mg protein, respectively. This method was also adapted for dTMP synthase assay in crude extracts of rat hepatoma 3924A. The major advantages of this procedure are the elimination of the phosphatase activity which interferes with the estimation of dTMP synthase activity in crude extracts, one-step separation of 3H2O, high sensitivity (with a limit of detection of 10 pmol of 3H2O production), high reproducibility (less than +/- 4.3%), and capability to measure activity in small amounts of sample (30-45 micrograms protein).