Volume 64, Issue 11-12

Calcium oxalate druses were isolated from the stems and leaves of six Pereskioideae family members and investigated by infrared spectroscopy, showing that in all samples the biomineral was present in the form of whewellite, CaC 2 O 4 · H 2 O. As Pereskia is thought to represent the “ancestral” condition of the leafl ess stem-succulent cacti, these results suggest that the biomineralization of calcium oxalate in Cactaceae represents a primitive characteristic of the group and also support a close genetic relationship between Pereskia and Opuntia.

A number of N-substituted pyrimidine acyclic nucleosides were synthesized by coupling reaction of 2-(2-chloroethoxy)ethyl acetate or (2,2-dimethyl-1,3-dioxolan-4-yl)methyl 4-methylbenzenesulfonate with the corresponding base followed by deprotection. The synthesized compounds were tested for their antiviral activity against hepatitis B virus (HBV). The plaque reduction infectivity assay was used to determine virus count reduction as a result of treatment with the synthesized compounds which showed moderate to high antiviral activities

A number of new 5-[(naphthalen-5-yloxy)methyl]-1,3,4-oxadiazole derivatives, 2 - 5 and 8 - 11, were synthesized. The 2-{5-[(naphthalen-5-yloxy)methyl]-1,3,4-oxadiazol-2-ylthio}acetohydrazones 6a and 6b were synthesized by the reaction of the hydrazide 4 with the corresponding monosaccharides. Cyclization of the sugar hydrazones 6a and 6b with acetic anhydride afforded the substituted oxadiazoline derivatives 7a and 7b. The synthesized compounds were evaluated for their antiviral activity against, the human immunodefi ciency virus (HIV-1) and some of these compounds showed moderate to high antiviral activity.

The antimycobacterial activity of (-)-cubebin (1), hinokinin (2), and some of their semisynthetic derivatives, namely (-)-O-acetyl-cubebin (3), (-)-O-methyl-cubebin (4), (-)-O-(N,Ndimethylamine- ethyl)-cubebin (5) and (-)-6,6´-dinitrohinokinin (6), was evaluated against Mycobacterium tuberculosis (ATCC 27294), M. kansasii (ATCC 12478), and M. avium (ATCC 15769). The MIC values ranged from 31.25 to 2000 μg/mL. Among the evaluated compounds, 2 displayed a MIC value of 62.5 μg/mL against M. tuberculosis, while 3 and 4 displayed MIC values of 62.5 and 31.25 μg/mL, respectively, against M. avium. All compounds were inactive against M. kansasii. These are promising results concerning the search for biologically active natural products, highlighting that new approaches to the prevention, treatment, and cure of tuberculosis are extremely important.

A number of new disubstituted 2,5-thiazolidinone derivatives were synthesized and tested for their antimicrobial activity against Bacillus subtilis (Gram-positive), Pseudomonas aeruginosa (Gram-negative), and Streptomyces species (Actinomycetes). They displayed different degrees of antimicrobial activities or inhibitory actions

The methanolic extract of the fruiting bodies of the mushroom Armillariella tabescens was found to show antifungal activity against Gibberella zeae. The active compound was isolated from the fruiting bodies of A. tabescens by bioassay-guided fractionation of the extract and identifi ed as armillarisin B. Armillarisin B eventually corresponds to 2-hydroxy-2- phenylpropanediamide and its structure was confi rmed on the basis of spectroscopic studies including 2D NMR experiments.

Chemical investigation of the EtOAc fraction (EF) obtained from the ethanolic extract of Zanthoxylum naranjillo (Rutaceae) leaves (EE) by preparative HPLC resulted in the isolation of protocatechuic acid (1), gallic acid (2), p-hydroxybenzoic acid (3), and 5-Ocaffeoylshikimic acid (4). This is the fi rst time that the presence of compounds 1 - 4 in Z. naranjillo has been reported. Compounds 1 - 4, the EE, and EF were tested in vitro against Schistosoma mansoni adult worms. The results showed that the S. mansoni daily egg production decreased by 29.8%, 13.5%, 28.4%, 17.7%, 16.3%, and 6.4%, respectively. Compounds 1 and 3 were also able to separate adult worm pairs into male and female. This activity may be correlated with the reduction in egg production, since 1 and 3 showed better inhibitory properties compared with 2 and 4.

A new triterpene saponin, named as 23-hydroxy-3α-[O-α-L- 1 C 4 -rhamnopyranosyl-(1´´4´)- O-α-L- 4 C 1 -arabinopyranosyl-oxy]olean-12-en-28-oic acid O-α-L- 1 C 4 -rhamnopyranosyl- (1´´´´´→4´´´´)-O-β-D-4C1-glucopyranosyl-(1´´´´→6´´´)-O-β-D-4C1-glucopyranosyl ester (9), was isolated from the leaves of Bauhinia variegata Linn. In addition, six flavonoid compounds along with two cinnamic acid derivatives were isolated and identified based on their chromatographic properties, and chemical and spectral data (ESI-high resolution-MSn, 1 H NMR, 13 C NMR, 1 H- 1 H COSY, HSQC, and HMBC). Compound 9 was found to be nontoxic (LD 50 ) and to have significant anti-inflammatory and antinociceptive activities. It also showed a slight antischistosomal activity.

Menthalactone, a new long-chain alkene with a bicyclic lactone moiety, was isolated as an analgesic constituent from the leaves of Mentha cordifolia Opiz. At a dosage of 100 mg/ kg mouse, it decreased the number of squirms induced by acetic acid by 67.3%. Statistical analysis using Kruskall Wallis one-way analysis of variance by ranks showed that menthalactone is different from the solvent control at α = 0.01 and approximates the analgesic activity of mefenamic acid at 0.001 level of significance.

Solanum species are traditionally employed as antiherpes and anticancer agents in different countries. S. paniculatum has widespread ethnomedical uses in Brazil, including the treatment of viral infections. This paper reports on the isolation of neotigogenin (1) and the new compound Δ 25 ( 27 )-tigogenin-3-O-β-D-glucopyranoside (2), obtained as a mixture of R and S diastereoisomers at C22 from an ethanol extract of S. paniculatum leaves, along with the determination of their cytotoxicity against Vero cells and antiviral effect against human herpes virus type 1 (HHV-1), murine encephalomyocarditis virus (EMCv), and vaccinia virus strain Western Reserve (VACV-WR). The extract of S. paniculatum inhibited HHV-1 replication [EC 50 = (298.0 ± 11.2) μg/ml] and showed no effect on EMCv and VACV-WR. On its turn, 1 was inactive against the assayed strains but presented high cytotoxicity [CC 50 = (2.03 ± 0.03) μg/ml], whereas 2 exhibited signifi cant antiherpes [EC 50 = (170.8 ± 1.7) μg/ml] and antivaccinia virus effects [EC 50 = (177.0 ± 3.3) μg/ml], with low cytotoxicity (CC 50 > 400 μg/ml). The results corroborate Solanum paniculatum as a source of cytotoxic and antiviral compounds.

Three labdane diterpenes, namely labda-8(17),14-diene, 13-epicupressic acid, and 13-Oacetyl- 13-epicupressic acid, were isolated from the resin collected from stem exudates of Araucaria heterophylla Salisb. (Araucariaceae). The isolated compounds were identified using different spectroscopic methods ( 1 H NMR, 13 C NMR, HMQC, HMBC and COSY). The resin extract showed antiulcerogenic activity against ethanol-induced stomach ulcers in Sprauge Dawely rats using ranitidine as standard. In addition, the resin and the isolated compounds showed variable cytotoxic activities against breast (MCF7) and colon (HCT116) cancer cell lines.

In the course of our continuous search for bioactive metabolites from endophytic fungi living in plants from the Brazilian fl ora, leaves of Alibertia macrophylla (Rubiaceae) were submitted to isolation of endophytes, and two species of Penicillium were isolated. The acetonitrile fraction obtained in corn from a culture of Penicillium sp.1 afforded orcinol (1). On the other hand, Penicillium sp.1 cultivated in potato-dextrose-broth furnished two different compounds, cyclo-(L-Pro-L-Val) (2) and uracil (3). The chromatographic fractionation of the acetonitrile fraction obtained from Penicillium sp.2 led to three dihydroisocoumarins, 4-hydroxymellein (4), 8-methoxymellein (5) and 5-hydroxymellein (6). Compounds 5 and 6 were obtained from the Penicillium genus for the fi rst time. Additionally, metabolites 1 - 6 were evaluated for their antifungal and acetylcholinesterase (AChE) inhibitory activities. The most active compounds 1 and 4 exhibited detection limits of 5.00 and 10.0 μg against Cladosporium cladosporioides and C. sphaerospermum, respectively. Compound 2 showed a detection limit of 10.0 μg, displaying potent AChE inhibitory activity.

Phytochemical studies of an ethanolic extract of the aerial parts of Salvia disermas resulted in the isolation of seven known compounds, rosmarinic (1) and caffeic (2) acids, salvigenin (3), luteolin (4), luteolin 7-O-β-arabinoside (5), luteolin 7-O-β-glucoside (6), and ocotillol II (7). The initiation stage of carcinogenesis is triggered by activation of procarcinogens by phase I enzymes, such as cytochrome P-450 1A, and oxidative stress that leads to DNA damage. The initiation stage is countered by phase II detoxifi cation enzymes such as glutathione S-transferases (GST), quinine reductase (QR), epoxide hydrolase (mEH) besides conjugation with thiols. We aimed to investigate the cancer chemopreventive and tumour anti-initiating activity of the ethanolic extract of the aerial parts of Salvia disermas and its constituents. The S. disermas extract was a promising inhibitor of CYP1A activity, inducer of GST, QR, and mEH activities, enhancer of thiol content, radical scavenger, and inhibitor of DNA damage. On the other hand, 3 was an enhancer of thiol content and QR activity, while 4 was an inhibitor of CYP1A activity, inducer of QR activity, and radical scavenger of ROO • , and 5 was an inducer of GST activity and inhibitor of DNA damage. The present study indicated that the ethanolic extract of S. disermas and 4 are promising anti-initiating and multipotent blocking agents.

Aromadendranes belong to a class of sesquiterpenes present in higher plant essential oils and marine animals. Although the biological activities include antifungal, antibacterial, antiviral, plant growth regulatory, antifeedant, repellent and cytotoxic, there is only one precedent for spasmolytic effects. In a previous report we have shown that the aromadendrene molecule known as spathulenol, isolated from Lepechinia caulescens, efficiently relaxes rat uterus rings and therefore in the present work we describe structure-activity relationships of thirteen aromadendranes, most of them having the trans-fused perhydroazulene skeleton, with spasmolytic activity.

The zoospores of the phytopathogenic Aphanomyces cochlioides are chemotactically attracted by a host-specific flavone, cochliophilin A (5-hydroxy-6,7-methylenedioxyflavone), and repelled from the mammalian estrogens or estrogenic compounds. This study further examined the responses of A. cochlioides zoospores to some flavonoids structurally related to cochliophilin A or compounds known as phytoestrogens. The bioassay revealed that some synthetic flavones (such as 6-methyl-4'-methoxyflavone, 3-hydroxy-4´methoxyfl avone, 7-hydroxy- 5-methylflavone, 3-hydroxy-4´methoxy-6-methylflavone) elicited attractant activity at concentrations as low as picomolar (10 pM), which was an 100-fold lower concentration than that of the threshold concentration of the host-specifi c attractant cochliophilin A. Apparently, a hydrophobic B-ring with an alkylated (methylated) A-ring or a methoxylated B-ring with an unsubstituted A-ring in the flavone skeleton played a significant role in higher attractant activity. On the other hand, all known estrogenic flavonoids (such as equol, 3´ or 8-prenylated naringenins) displayed potent repellent activity toward zoospores. Surprisingly, zoospores were attracted by non-estrogenic 6-prenylated naringenin indicating that repellent activity is linked to the estrogenic activity of the phytoestrogens.

The effects of oxygen supply on growth and carotenoids accumulation by Xanthophyllomyces dendrorhous were studied. Initial volumetric oxygen transfer coefficients (KLa) within the range 21.5 - 148.5 h -1 had significant effects on growth and carotenoids accumulation, and an increase of the initial KLa value led to higher carotenoids, astaxanthin and biomass yields by X. dendrorhous. At an initial KLa value of 148.5 h -1 , a maximal cell concentration of 19.37 g l -1 and optimal carotenoids and astaxanthin productions of 18.1 and 14.5 mg l -1 were obtained, as well as a maximal astaxanthin content of 0.8 mg g DCW -1 , respectively. A higher oxygen supply was advantageous to astaxanthin biosynthesis and the ratio of astaxanthin in the total carotenoids. An increasing initial KLa value gave stronger fluorescence intensities by X. dendrorhous, resulting in the maximal intensity of fluorescence at the KLa value 148.5 h -1 . The cell growth of X. dendrorhous was significantly inhibited when dissolved oxygen tension (DOT) was controlled at ~ 20% air saturation, which was due to the oxygen limitation in broth. The astaxanthin yield and content at ~ 50% DOT were higher than those at ~ 20% DOT.

Jatropha curcas embryos were grown in vitro to observe the effects of lead on cotyledon responses. The cotyledon biomass increased initially and then decreased with increasing lead concentration. The SOD activity increased gradually up to 200 μM and then decreased. The POD activity showed a similar trend. The CAT activity was increased at all lead concentrations, the highest activity being observed at 200 μM. However, the PAL activity was inhibited significantly except for 100 μM. Anaylsis by electrophoresis suggested a significant correlation between lead concentration and patterns of SOD, POD and CAT isoenzymes, and these results were consistent with changes of the antioxidant enzyme activities as assayed in solution.

This is the fi rst evidence on successful Agrobacterium rhizogenes-mediated genetic transformation of two species from the genus Hypericum, H. tomentosum L. and H. tetrapterum Fries. Hairy root cultures were induced from root segments of both Hypericum species by two agropine wild-type strains of A. rhizogenes, ATCC 15834 and A4. The transgenic character of the hairy root cultures was proved by PCR amplifi cation of the rolABCD genes. In some H. tetrapterum transgenic lines aux genes were detected as well.

Hairy root cultures of Nicotiana tabacum were set up by excised root tips with Agrobacterium rhizogenes. The successful transformation was confirmed by analyzing rolC and virC genes using polymerase chain reaction (PCR). Hairy root cultures were employed to study the formation of pyridine alkaloids, mainly nicotine. The transformed cultures were incubated with potential elicitors, such as methyljasmonate, quercetin and salicylic acid, in order to stimulate the biosynthesis of pyridine alkaloids. Profile and amounts of pyridine alkaloids were analyzed using capillary GLC-MS. Treatment of the cultures with methyljasmonate (50 μM) increased the alkaloid accumulation ca. 7-fold up to a level of 0.58 mg/g fresh weight as compared to untreated controls. Quercetin (200 μM) enhanced the alkaloid production ca. 4-fold (0.34 mg/g fresh weight) within 24 h. In contrast salicylic acid in all tested concentrations decreased the alkaloid level to 1 μg/g fresh weight. Also the inhibitory effect of salicylic acid on the elicitation effect of methyljasmonate and quercetin was investigated.

A new full-length β-carbonic anhydrase cDNA was obtained from Brassica napus by homologous cloning. The cDNA has an open-reading frame of 996 nucleotides, encoding 331 amino acids with a calculated molecular weight of 35,692 Da and an estimated pI value of 5.459. The deduced amino acid sequence of β-carbonic anhydrase from Brassica napus shared significant identity with β-carbonic anhydrases from Brassica carinata, Arabidopsis thaliana, and Thlaspi caerulescens (97.9%, 94%, and 93.5% identity, respectively). This cDNA was expressed in Escherichia coli BL21 (DE3) using the expression vector pET-32a(+). The expression band corresponded to the calculated mass plus the N-terminal fusion protein derived from the vector

Genetic linkage analysis of 151 restriction fragment length polymorphism (RFLP) loci, that included eight new loci, detected by the six probes in the present study, and four trait loci including seed colour, leaf pubescence, resistance to white rust caused by Albugo candida race-2 (AC-2) and race-7 (AC-7) employing the MAPMAKER/EXP 3.0 programme led to the development of 10 linkage groups (LGs) spanning over 44.4 centiMorgan (cM) to 130.4 cM containing 9 to 22 loci and two short LGs with two or three marker loci in Brassica rapa. The enriched map covers 993.1 cM of B. rapa genome with an average marker interval of 6.41. Eight new RFLP loci occupied new map positions on five linkage groups, LG 2, 3, 6, 8 and 9. Addition of these RFLP loci led to appreciable changes in the corresponding linkage groups and resulted in an increase of the total map length by 102.8 cM and of the marker interval by 0.35 cM. Interval mapping by using the computer programme MAPMAKER/ QTL 1.1 for scanning the genetic map led to the detection of one major quantitative trait locus (QTL) in LG 4 and one minor QTL in LG 8 governing resistance to AC-7. Both QTLs contributed 7.89 to the interaction phenotype (IP) score with 96.3% genetic variation. The multi-locus model suggested additive gene action with 96.8% genetic variation.

The pyridoxal-5’-phosphate (PLP)-dependent family of enzymes is a very diverse group of proteins that metabolize small molecules like amino acids and sugars, and synthesize cofactors for other metabolic pathways through transamination, decarboxylation, racemization, and substitution reactions. In this study we employed degenerated primer-based PCR amplification, using genomic DNA isolated from the soil bacterium Exiguobacterium acetylicum strain SN as template. We revealed the presence of a PLP-dependent family of enzymes, such as PLP-dependent acyltransferase, and similarity to 8-amino-7-oxononoate synthase. Sequencing analysis and multiple alignment of the thymidine-adenine-cloned PCR amplicon revealed PLP-dependent family enzymes with specific confering codes and consensus amino acid residues specific to this group of functional proteins. Amino acid residues common to the majority of PLP-dependent enzymes were also revealed by the Lasergene MegAlign software. A phylogenetic tree was constructed. Its analysis revealed a close relationship of E. acetylicum to other bacteria isolated from extreme environments suggesting similarities in anabolic adaptability and evolutionary development.

The vacuolar ATPase (V-ATPase) is a multifunctional enzyme that consists of several subunits. Subunit B is a part of the catalytic domain of the enzyme. The result of the RTPCR suggested that the V-ATPase B subunit is a ubiquitous gene. 24 h after the larvae were infected with the Bombyx mori nucleopolyhedrovirus (BmNPV), the expression level of the V-ATPase B subunit in the midgut of the resistant strain NB was about 3 times higher than in the susceptible strain 306, and then the expression level of the V-ATPase B subunit decreased rapidly to a very low level. This indicated that the virus may cause a lot of changes of physiological conditions in the midgut. Localization of the V-ATPase B subunit was attempted in midgut cells of Bombyx mori by immunohistochemistry. The immunohistochemical localization with the antibody against the B subunit revealed a positive staining in goblet cell apical membranes of Bombyx mori midgut cells as well as in the midgut of Manduca sexta. This sequence has been registered in GenBank under the accession number EU727173.