Volume 54, Issue 7-8

Strobilurin N (1), a new member of the strobilurin family of antibiotics, has been isolated from the toadstool Mycena crocata, together with dehydrochorismic acid lactone (5) and 3-[1-carboxyvinyl)oxy]benzoic acid (6), which are closely related to chorismic acid (10). Their structures were determined mainly by 1 H and 13 C NMR spectroscopy. Interestingly, strobi­lurin N is the first strobilurin without antifungal activity

6,9-Dihydroxy-3(15)-caryophyllen-4,8-dione (1), a new cytotoxic caryophyllane sesquiterpene, was isolated from fermentations of a tropical Marasmius species. The structure was established by spectroscopic methods. 1 exhibits strong cytotoxic effects on different cell lines, but only weak antimicrobial activities. 1 weakly inhibits the incorporation of leucine and thymidine into proteins and DNA of mammalian cells and interferes with the aggregation of hum an and bovine platelets

Hypoxysordarin (1), a new sordarin derivative, was isolated from the fermentation broth of the facultative marine Hypoxylon croceum together with a new y-lactone, hypoxylactone (2) and sordarin (3). The structures were determined by spectroscopic m ethods. Sordarin (3) has previously been isolated from the terrestrial Sordaria araneosa (Sordariaceae). Like the parent com pound hypoxysordarin exhibits high antifungal activities due to a specific inhibition of protein biosynthesis

22 ent-kaurane and ent-pimarane type diterpenoids, ten of which are new, ent- 19-acetoxykaur- 16-en-15-one, ent-19-oxo-kaur-16-en-15-one, ent-11α-acetoxy-20-oxo-kaur-16-en-15-one, enf-11α-acetoxy-15-oxo-kaur-16-en-20-acid,-11α-acetoxy-7β-hydroxy-kaur-16-en-15-one, ent-11α-acetoxy-19-hydroxy-kaur-16-en- 15-one, ent-19-acetoxy-11α-hydroxy-kaur-16-en-15- one, ent-11α-acetoxy-6β, 19-dihydroxy-kaur-16-en-15-one, ent-11α-acetoxy-19-hydroxykauran- 15-one and em -19-acetoxy-11α-hydroxy-kauran-15-one were isolated from the liverwort Blepharidophyllum densifolium. Furthermore three known triterpenoids cabraleadiol, cabraleadiolacetate and shoreic acid have been found. Their structures were established by NMR spectroscopy

Galium humifusum, R ubiaceae, Iridoid G lycosides Two new iridoid glucosides, hum ifusin A and hum ifusin B, with a 3-(4-hydroxyphenyl)-p ropionate ester unit were isolated from Galium hum ifusum to g eth er with 11 know n iridoid glucosides asperuloside, scandoside, desacetylasperulosidic acid, asperulosidic acid, genipo-sidic acid, desacetylasperuloside. m onotropein, V I, V2, V3 and daphylloside. The new iri-doids w ere identified on the basis of spectral d ata. 13C N M R d ata for V I and V2 iridoids are reported.

Sporopollenin was isolated and purified from pollen of the gymnospermae Torreya californica using an enzymatic treatment followed by extraction with organic solvents. 1 H -NMR (ID and 2D) was used for analysis of this biopolym er and reveals the presence of three phenolic com pounds. Comparison with the analysis of sporopollenin from the angiospermae Typha angustifolia L. shows high similarity between the aromatic compounds

Dioscorea esculenta. Peroxidase. Ion Exchange Chrom atography. Gel Filtration We isolated two types of peroxidase from the fresh tuber of Dioscorea esculenta using a combination of ion exchange and gel filtration chrom atography. The results showed one type to be neutral and the other to be strongly ionic. The strongly ionic type constitutes 70% of total peroxidase activity in the tissue. The apparent molecular weight of the neutral type is 38 kD a while the anionic type has an apparent molecular weight of 57 kD a. It was possible with the use of gel filtration on Sephadex G-200 followed by FPLC on phenyl superose to purify the lower size POD by a factor of 15, while the larger ionic peroxidase was purified 68 fold com pared to the crude with protein yields of 0.90% and 1.30% respectively. The ionic POD is m ore therm o-stable, has a higher optimum temperature for activity and has a higher ap parent activation energy com pared to the neutral POD from this source.

In the leaf extract of Trillium apetalon (Liliaceae) distributed in Japan, an enzyme was demonstrated which catalyzes a m ethyl group transfer from S-adenosyl-ʟ-m ethionine (SAM) to the 3′ position of quercetin and its glycosides. The enzyme ( Trillium F3′OMT ) was purified 433-fold with a yield of 0.2% by (NH 4 ) 2 SO 4 precipitation and chromatographies of DEAE - cellulose, SAH-EAH-Sepharose 4B , Sephacryl S-200 and additional chrom atofocusing. Trillium F3′OMT has a pH optimum of 7.0 and a pi of 5.3. The apparent m olecular weight was estimated by Sephacryl S-200 to be about 78 kD a; SD S-PAGE profile showed that the enzyme was a dim er com posed of MW 38 kDa 2 subunits. The enzyme activity was stimulated by EDTA and dithiothreitol (DTT), but strongly inhibited by p-chlorom ercuribenzoate (PCMB) and iodoacetate. The activity was m oderately inhibited by Mg 2+ and Zn 2+ , and strongly inhibited by Co 2+ , Mn 2+ and Hg 2+ . The apparent K m values for quercetin and SAM were 10 μm and 3.6 μm , respectively. Lower substrate specificity of the glycosides compared with quercetin indicates that methylation precedes glycosylation in flavonoid biosynthesis of T. apetalon

Cells of the green alga Micrasterias denticulata cultivated at 15 °C, 20 °C or 25 °C were exposed to heat shocks at different temperatures (30 -40 °C) for varying duration ( 5 - 90 m in). Cell pattern formation, division rate as well as photosynthesis and respiration by measuring oxygen production and consumption have been studied. The degree of cell shape malformations was found dependent on the preceding cultivation temperature along with the mode of the heat shock. Cells cultivated at 15 °C and 20 °C could counteract a 90 min heat shock at 35 °C much better than those cultivated at 25 °C, which was seen by a less reduced young semicell. Cells cultivated at 15 °C and 25 °C reveal a reduced division activity compared to those grown at 20 °C even with a marked retardation when affected by a preceding heat shock. Photosynthesis and the level of plastid pigments (carotenoids, chlorophylls, β-carotene, lutein) of controls determined by HPLC analysis reached a plateau after about 26 days when starting with 22-day old cultures. Photosynthesis and respiration were determined in a range between 15 °C and 40 °C in defined Micrasterias cell cultures of about this age (cultivation temperature 15 °C, 20 °C or 25 °C). Both processes rose steadily with increasing temperature starting with 15 °C and reached peaks between 30 °C and 32 °C, followed by a considerable drop when increasing the incubation temperature up to 40 °C. The experiments reveal that primary processes of energy formation and consumption are much less affected by temperature influences than cell shape formation and division rate

The structure of the xanthophyll pigments lutein and zeaxanthin differs in the position of one double bond and refers to one of the ionon rings. Specific antibodies to zeaxanthin were used to analyse the localisation and orientation of these two xanthophyll pigments in lipid membranes formed with egg yolk lecithin. Bimolecular and monomolecular layers were used. Antibody-antigen interaction was demonstrated and analysed by the bathochromic shift of the absorption spectra of both pigments and by the increase of light-scattering of the pig- mented liposome suspension. It appeared that the extent of the spectral effects accompanying the interaction of the antiserum to zeaxanthin, injected to the liposome suspension which was pigmented with either zeaxanthin or lutein, was different in spite of their similar molecular structures. The results are interpreted in terms of a localisation and distribution of lutein, in the hydrophobic phase of liposomes within two essentially different pigment pools, one oriented horizontally and the other vertically with respect to the membrane plane. This inter- pretation is supported by the analysis of isotherms of the compression of monomolecular layers of lutein and zeaxanthin formed at the air-water interface and of mixed xanthophyll- lipid monolayers as well as by analysis of the penetration of antibody proteins dissolved in the subphase into the mixed xanthophyll-lipid films.

Relationships of the width of annual rings of 75-85-year old Pinns sylvestris L. and Picea abies (L.) Karst, with average monthly temperature, amount of precipitation and a complex climatic indicator developed on their basis was studied against the background of pollution load in the zone influenced by a cement plant and in a control area. Multiple regression analysis (equations with two and three independent variables) showed a significant correlation between precipitation and temperatures with increment, especially for pine: however, the prediction capability of the models is modest, describing usually 35-40% of the variation in radial increment. The calculations suggest that precipitation amounts are more important and temperature parameters less important and can be replaced by one another in the models. A direct correlation with the pollution load can be observed: at probability (P) near zero the coefficients for precipitation were the highest (0.45-0.51) in the area strongly affected by the cement plant and the lowest (0.31 -0.35) in the weakly affected and control areas. In case of spruce shortage of air humidity during summer months was important for increment, especially so in the heavily polluted area.

The binuclear Co(II) and Mn(II) complexes with H 5 (HXTA). where H 5 (HXTA) repre­sents N,N′-(2-hydroxy-5-methyl-1,3-xylylene)bis(N-carboxymethylglycine), induced a strong ethylene evolution from 1-aminocyclopropane-l-carboxylic acid (ACC) in the presence of hydrogen peroxide, whereas activities of the corresponding Fe(III), Ni(II), and V(III) com­plexes were found negligible. Based on spectroscopic results and mass-spectral data it is proposed that a peroxide adduct of binuclear Co(II) (and Mn(II)) complex with η 1 -coordina­tion mode interacts with ACC, which is chelated to a binuclear cobalt complex leading to facile oxidative degradation of ACC and to evolution of ethylene.

Although Mg 2+ is an important cofactor for the specific degradation of RNA by ribozymes, it is not considered as a typical chemical nuclease. In this study we show that in combination with common buffers such as tris(hydroxymethyl)aminomethane and sodium borate. Mg 2+ is a powerful catalyst for the degradation of RNA. pH and temperature are found to be the principal factors for the efficient degradation of RNA. Whereas in Tris-HCl/Mg 2+ the effi­cient cleavage starts at pH values higher than 7.5 and temperatures higher than 37 °C, in sodium borate RNA degradation begins at pH 7.0 and at 37 °C. RNA hydrolysis promoted under the combined catalytic activity of buffer/Mg 2+ results in partially degraded RNA and negligible amounts of acid-soluble material. Reaction is insensitive to the concentration of monovalent cations but is completely prevented by chelating agents (EDTA and citrate) at concentrations exceeding that of Mg 2+ . Borate-magnesium reaction is inhibited also by some polyvalent alcohols (glycerol) and sugars.

Ethanolic extract of propolis exerts a strong anti-bacterial activity, in addition to antifungal. antiviral and antiprotozoal properties. In previous studies from these laboratories we have demonstrated that the intensity of the bactericidal activity of EEP is correlated with the virulence of the mycobacteria tested, and that EEP has a synergistic effect with antibiotics on growth of staphylococcus aureus. In the present study we investigated whether the same synergism and correlation exists between EEP and some anti-tuberculosis drugs on tubercu­losis mycobacteria with different degrees of virulence. Six standard strains and 11 wild strains of mycobacteria were exposed for 30 days to EEP, with or without streptomycin, rifamycin, isoniazid or ethambutol. Out of the 17 strains, 8 were resistant to at least two standard antibiotics, and were considered “multi-resistant strains”. The rest were either susceptible or resistant to only one of the antimycobacterial drugs. Antagonism was recorded only in one case, when Staphylococcus aureus were treated with a mixture of EEP and ethambutol, sug­gesting that a chemical bond could have been formed between this anti-tuberculosis anti­ biotic and one of the active components of the ethanol extract of propolis.

The (μ-oxo)(μ-carbonato)diiron(III) complex with H 2 (tfda) (H 2 (tfda) = 2-aminomethyl-tetrahydrofuran-N,N-diacetic acid) exhibited high activity for hydroxylation of 2′-deoxygua-nosine in the presence of hydrogen peroxide, giving 8-hydroxydeoxyguanosine, but its hy­ droxylation activity towards other nucleosides such as 2′-deoxyadenosine, adenosine or thym­ idine was found negligible. In the case of the Fe(III)-(ed a) complex (H 2 (eda) = 2-methoxyethylamine-N,N-diacetic acid), hydroxylation occurred mainly at the sugar site, con­verting 2′-deoxyguanosine to guanosine. Based on the spectroscopic and structural properties of these iron(III) compounds, it seems most likely that an intrinsic active species for hydrox­ylation should be an electrophilic peroxide adduct of the (μ-oxo)diiron(III) core with η 1 coordination mode, while the contribution of OH· sides is ruled out.

Using indicators for the production of reactive oxygen species (ROS) such as the a) OH- radical type (α-keto-γ-methiolbutyric acid. KMB) or b) hypochlorous acid (1-amino-cyclo- propvl-1-carboxylic acid, ACC) neutrophil activities can be both quantified and differentiated in whole blood via ethene production, Ethene is trapped in the head space of blood samples incubated in the presence of zymosan and the respective indicators, KMB or ACC. This procedure allows the detection of effects of aminoadamantanes (AAD) such as amantadine or memantine, compounds frequently used for the treatment of Morbus Parkinson and Morbus Alzheimer. In this report we describe the detection of OH-type oxidants produced by isolated activated neutrophils and whole blood. Immunomodulatory activities of AAD are deduced from the following observations: AAD-stimulated ethene formation from (KMB) as an indicator for production of OH′-type reactive oxygen species by zymosan-stimulated neutrophils (“respiratory burst”) is detectable with isolated neutrophils. In whole blood, how- ever, this reaction is only measurable in the presence of Fe-EDTA-complex. Stimulating effects of AAD are observed within a concentration range between 10 -8 and 10 -4 m with a maximum at 1 μΜ. Ethene release from (ACC) as indicator for the myeloperoxidase reaction after degranulation is not stimulated by AAD but inhibited at concentrations higher than 100 μΜ. The presented results suggest that submicromolar concentrations of AAD only stimulate the respiratory burst and apparently not degranulation of zymosan-prestimulated poly- morphonuclear neutrophils (PMN).

In this communication we present a comparative investigation of the dielectric properties of native E. coli 70S and ribosomal cores obtained by LiCl treatment. Previous data obtained in our laboratory showed that ribosomes exhibit two different dielectric dispersions. We show that elimination of some select proteins modifies only the first one and therefore the overall dielectric properties of the ribosome result altered. Ribosomal RNA and proteins remaining in the core particle are mainly responsible for the second dielectric dispersion. Our experimental approach allows an estimation of the size of RNA traits exposed to solvent both in native ribosomes and in core particles where a higher portion of rRNA interacts with the external environment. Furthermore our results are consistent with the idea that proteins remaining after high salt treatment are necessary and sufficient for the maintenance of the basic structural properties of the ribosome.

We established a relationship between the toxic effects of cadmium on the expression of the endoplasmic reticulum (E R) chaperone GRP94 (glucose regulated protein 94) and cell survival in cultured rat-thyrocytes of FRTL5 cells. There are no data reporting that the en­hanced expression of GRP94 by Cd stimulation is detectable in thyrocytes. Western blot analysis revealed higher levels of GRP94 expression in those cells post-treated with low concentrations of Cd, following a step-down treatment method, than in Cd pre-treated cells or cells not treated with any Cd, due to changes in cellular sensitivity after pre-treatment with Cd and the possible induction of GRP94 expression after removal of a low concentration of Cd. Elevated GRP94 expression in thyrocytes post-treated with Cd confers a survival advantage by rendering them resistant to cytotoxic stress, and the existence in the thyrocytes of a Cd-specific pathway regulates the expression of stress proteins by Cd.

Differential Display (DD) PCR, Thyroid Stimulating Hormone (TSH), Rat Thyrocytes (FR TL5 cells) Differential display (DD) PCR (Liang and Pardee, 1992) is a recently described technique to identify genes whose expression has changed during a biological process. We used this method to detect genes thyroid stimulating hormone-dependently regulated in a rat thyroid cell line, because thyroid stimulating hormone (TSH) is the most important hormone for cell proliferation and differentiation including prehormonal proteins secretion in thyrocytes (Kim and Arvan. 1991: Kim and Arvan, 1993). Following DD-PCR experimentation, thyroid stimu­ lating hormone -dependently regulated gene fragments of 15 species were obtained. The genes were used as molecular probes in Northern blot analysis and then sequenced. Two of the clones (#123 and #205) were up-regulated and two more (#107 and #111) were down-regulated thyroid stimulating hormone-dependently in the thyroid cells, as demonstrated by Northern blot analysis. Following partial sequencing, each of the clones #107, #111 and #205 were shown to be homologues of the apoptosis-related gene, aldolase A, and a-2 collagen (IV), respectively, while clone #123 showed no homology with known genes. These findings suggest that the four genes mentioned above may have an a important physiological function in the thyrocytes, which is thyroid stimulating hormone-dependently up-/down-regulated.

Forty-three stool samples and rectal swabs were collected from diarrhoeic and 20 non-diarrhoiec patients under 5 years of age at various general hospitals in Lagos. A total of 110 faecal samples from animals (101 from chickens and 9 from pigs) were obtained from Mitchel farm, Agege and Oshodi -Isolo Local Government farms at Ejigbo. Campylobacter species were isolated from 6 (13.9%) of 43 children with gastroenteritis and none from 20 asymptom­atic subjects. Forty-nine (48.5%) isolates from the hundred and one faecal specimen were isolated from chicken, while 3 (33.33%) out of 9 were from pigs. Campylobacter jejuni was the most prevalent accounting for 79% of the total isolates and C. coli accounted for 25% . All the human strains were gentamicin sensitive, while streptomicin and cloxacillin were resistant. Erythromycin had a high activity (83.3%) on human strains and only 59.6% activity on animal strains. About ten percent (9.6%) of the strains produced beta-lactamase.

Bombycidae, Saturniidae, mtDNA RFLP, Cytochrome Oxidase I Gene The phylogenetic relationships between Bombyx mori and Bombyx mandarina species of Bombycidae, and Antheraea yamamai and Antheraea pernyi species of Saturniidae were investigated based on mtDNA RFLP and cytochrome oxidase I gene. The sizes of the mtDNA of all the species were estimated at approximately 16 kbp ± 500 bp by total length of all the restricted fragments and no variation in size was recognized. Of the fourteen different restriction endonucleases used, BamHl, Hindlll, Pstl, EcoRl and Xbal showed RFLP. Among these, only Hindlll showed RFLP between B. mori and B. mandarina. A comparative analysis of sequences was also conducted with the mitochondrial cytochrome oxidase I genes of each species. The results indicated that B. mori shared a 97% , 85% and 87% sequence identity with B. mandarina, A. yamamai and A. pernyi, respectively. B. mandarina shared a 87% and 88% sequence identity with A. yamamai and A. penyi, respectively. A. yamamai shared 92% sequence identity with A. pernyi. The results of the phylogenetic analysis exhib­ited monophyly and confidence limits of more than 99% in all trees for both Bombycidae and Saturniidae.

The female sex pheromone of Bonagota (=Phthteochroa) cranaodes (Meyrick) is a blend of (E,Z)-3,5-dodecadienyl ajcetate (E3,Z5-12:Ac) and (Z)-9-hexadecenyl acetate (Z9-16:Ac) according to analysis of pheromone - gland extracts and field trapping in apple orchards. This is the first time that E3,Z5-12:Ac has been identified as a lepidopteran sex pheromone. Traps baited with 100 μg E3,Z5-12:Ac were attractive over 15 weeks in the field and were as effective as traps baited with virgin females. Addition of Z9-16:Ac to E3,Z5-12:Ac at ratio of 1:10 had a significantly increase of male moths. The addition of the Z,E and Z,Z isomers to rubber septa baited with E3,Z5-12:Ac did not modify B. cranaodes male attraction, but 10% of EE enhanced trap catch.

Leaf and stem exudate of Ozothamnus scutellifolius, previously reported to contain 21 flavonoid aglycones, was now investigated for its terpenoid composition. Seven structurally related diterpenoids were identified. One of them, 16a,19-diacetoxy-enf-kaurane, is a novel natural product.

Antimicrobial activity in some lichens from south Spain has been studied. Some lichenical substances are also identified. The structures of all compounds were elucidated by physical, spectral and chemical methods. A very high activity against Gram-positive bacteria has been observed in lichens containing usnic acid.

Eight known flavonoids were isolated from an ethanol extract of hop cones (Humulus lupulus L., Cannabaceae) together with two new hop chalcones. The new compounds were identified as a,β-dihydroxanthohumol and iso-dehydrocycloxanthohumol hydrate

Sulfate-reducing bacteria (SRB) were isolated from soils around corroded pipelines and tanks. High num­bers of the organisms occurred in areas closest to the corroded tanks and pipelines. Morphological types cor­ responding to rod, spirilloid, vibriod and coccoid were encountered. All the organisms utilized lactate as carbon and energy source. None could grow at temperatures higher than 40 °C. All the isolates grew at 1% (w/v) NaCl while none could grow at 8% (w/v) NaCl. All the isolates grew at pH 7 .0 -7 .5 . Growth was not recorded at pH below 5.5 and above 8.0. These factors may be useful in manipulating tropical soil environments to reduce activities of SRB in corrosion of pipelines and tanks.