Meiosis,SC-formation,and karyotype structure in diploidPaeonia tenuifolia and tetraploidP. officinalis

The meiosis of the diploidPaeonia tenuifola and the allotetraploidP. officinalis was studied after conventional methanol/acetic acid-fixation and synaptonemal complex (SC) spreading. Meiosis inP. tenuifolia (2n = 10) is normal with five bivalents in metaphase I, and the SCs in pachytene show regular features. InP. officinalis (2n = 4x = 20) univalents, bivalents and multivalents are found in metaphase I. The SCs reveal several abnormalities: a high number of unpaired lateral elements, partner exchanges between three and four lateral elements, loops and lateral element thickenings. These characteristics are compared with the situations found in other polyploid and hybrid species. It is noteworthy that the abnormalities in meiosis ofP. officinalis are not reflected in its somatic karyotype. Its features were analysed after silver staining and fluorescent staining with chromomycin and compared with those ofP. tenuifolia. Synaptonemal Complex Spreading in Plants2; for part1 see Pl. Syst. Evol.154, 129–136 (1986).     

Bioactive metabolites from Alternaria brassicicola ML-P08, an endophytic fungus residing in Malus halliana

A total of 48 strains were isolated from the normal tissues of Malus halliana and the EtOAc extracts of their cultures were subjected to primary antimicrobial screening against four test bacteria and three fungi. As a result, 22 strains exhibited antimicrobial activity against at least one test microbe. Among them, Alternaria brassicicola ML-P08 showing strong activity (MICs: 0.31–2.50 mg/ml) was selected for further investigation on its secondary metabolites. Bioassay-guided fractionation of the EtOAc extract of its liquid culture afforded seven compounds, which were identified as alternariol (1), alternariol 9-methyl ether (2), altechromone A (3), herbarin A (4), cerevisterol (5), 3β,5α-dihydroxy-(22E,24R)-ergosta-7,22-dien-6-one (6) and 3β-hydroxy-(22E,24R)-ergosta-5,8,22-trien-7-one (7), respectively, by spectral means (MS, IR, ¹H- and ¹³C-NMR). In vitro antimicrobial assay showed that compound 3 was substantially active against Bacillus subtilis, Escherichia coli, Pseudomonas fluorescens and Candida albicans with the MICs of 3.9, 3.9, 1.8, and 3.9 μg/ml, respectively. Compound 4 also showed pronounced antifungal activity against Trichophyton rubrum and C. albicans with MICs of both 15.6 μg/ml. In addition, compound 1 exhibited strong xanthine oxidase inhibitory activity with the IC₅₀ of 15.5 μM, comparable to that of positive control, allopurinol (IC₅₀: 10.7 μM).     

Design, synthesis and in vitro evaluation of novel chroman-4-one, chroman, and 2H-chromene derivatives as human rhinovirus capsid-binding inhibitors

As part of an effort to generate broad-spectrum inhibitors of rhinovirus replication, novel series of (E)-3-[(E)-3-phenylallylidene]chroman-4-ones 1a–e, (E)-3-(3-phenylprop-2-yn-1-ylidene)chroman-4-ones 2a and 2b, (Z)-3-[(E)-3-phenylallylidene]chromans 3a–e, and (E)-3-(3-phenylprop-1-en-1-yl)-2H-chromenes 4a–d were designed and synthesized. All the compounds were tested in vitro for their efficacy against infection by human rhinovirus (HRV) 1B and 14, two representative serotypes for rhinovirus group B and A, respectively. Most of the analogues were found to be potent and selective inhibitors of both HRVs, although HRV 1B was generally more susceptible than HRV 14. Mechanism of action studies of (E)-6-chloro-3-(3-phenylprop-1-en-1-yl)-2H-chromene 4b, the most potent compound on HRV 1B infection, suggested that 4b behaves as a capsid-binder probably acting at the uncoating level.     

Synthesis, characterization, and evaluation of a novel 99mTc(CO)3 pyrazolyl conjugate of a peptide nucleic acid sequence

The 16-mer peptide nucleic acid sequence H-A GAT CAT GCC CGG CAT-Lys-NH₂ (1), which is complementary to the translation start region of the N-myc oncogene messenger RNA, was synthesized and conjugated to a pyrazolyl diamine bifunctional chelator (pz). The novel conjugate pz-A GAT CAT GCC CGG CAT-Lys-NH₂ (2) was labeled with technetium tricarbonyl, yielding quantitatively the complex fac-[^99mTc(CO)₃(κ³-pz-A GAT CAT GCC CGG CAT-Lys-NH₂)]²⁺ (4). Complex 4 was obtained with high radiochemical purity and high specific activity, revealing high stability in human serum and in cell culture medium. The identity of 4 was confirmed by comparing its reversed-phase high performance liquid chromatography profile with that of the rhenium analog fac-[Re(CO)₃(κ³-pz-A GAT CAT GCC CGG CAT-Lys-NH₂)]²⁺ (3), prepared by conjugation of fac-[Re(CO)₃(3,5-Me₂pz(CH₂)₂N((CH₂)₃COOH)(CH₂)₂NH₂)]⁺ to 1, using solid-phase techniques. UV melting experiments of 1 and 3 with the complementary DNA sequence led to the formation of stable duplexes, indicating that the conjugation of 1 to the pyrazolyl chelator and to the metal fragment fac-[M(CO)₃]⁺ did not affect the recognition of the complementary sequence as well as the duplex stability. For a first screening, SH-SY5Y human neuroblastoma cells, which express N-myc, were treated with 4. The results show that 4 internalizes (7% of the activity goes into the cells, after 4 h at 37 °C), presenting also a relatively high cellular retention (only 40% of internalized activity is released from the cells after 5 h). An erratum to this article can be found at     

The synthesis of 2-nitroaryl-1,2,3,4-tetrahydroisoquinolines, nitro-substituted 5,6-dihydrobenzimidazo[2,1-a]isoquinoline N-oxides and related heterocycles as potential bioreducible substrates for the enzymes NAD(P)H: quinone oxidoreductase 1 and E. coli nitroreductase

A series of 2-nitroaryl-1,2,3,4-tetrahydroisoquinolines 10 and nitro-substituted 5,6-dihydrobenzimidazo[2,1-a]isoquinoline N-oxides 11 have been synthesised and evaluated as potential bioreducible substrates for the enzymes NAD(P)H: quinone oxidoreductase 1 (NQO1) and Escherichia coli nitroreductase (NR). Also prepared and evaluated were 2-(3,5-dinitropyridin-2-yl)-1,2,3,4-tetrahydroisoquinoline 12 and 5,6-dihydro-10-nitropyrido[3″,2″:4′,5′]imidazo[2′,1′-a]isoquinoline 12-oxide 13. Both compounds 10b and 13 were reduced faster by human NQO1 than by CB-1954 [5-(aziridin-1-yl)-2,4-dinitrobenzamide].     

Pictolejeunea reginac, a new species of Lejeuneaceae (Hepaticae) from Venezuela

Pictolejeunea reginae, a new species from Venezuela is described and illustrated. It is the type of the new subgenusNeopitolejeunea Ilkiu-Borges, which is characterized by entire underleaves. A key to the two subgenera and four known species ofPictolejeunea is provided.

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Ilkiu-Borges, A. L. (Museu Paraense Emílio Goeldi, C. P. 399, Belém, Pará, CEP 66040-170. Brasil: email: ilkiuborges@hotmail.com).Pictolejeunea reginae, a new species of Lejeuneaceae (Hepaticae) from Venezuela. Brittonia 54: 318–321. 2002.—Pictolejeunea reginae, uma nova espécie encontrada na Venezuela, é descrita e ilustrada. A espécie é tratada como tipo do novo subgêneroNeopictolejeunea Ilkiu-Borges, caracterizada pelos anfigastros inteiros. é apresentada uma chave para os dois subgêneros e as quatro espécies dePictolejeunea.

     

Pollen morphology ofSonchus and related genera,and a general discussion

The pollen morphology of the taxa belonging to the generaAetheorhiza Cass.,Launaea Cass.,Reichardia Roth andSonchus L. in the Iberian Peninsula has been studied with light and electron microscopy. The pollen is 3(-4)-zonocolporate and echinolophate (without polar lacunae, but in general with prelacunae), with equatorial ridges and 15–20 lacunae: 3–4 poral, 6–8 abporal and 6–8 paraporal. Small to medium size, P × E = 19–36 × 23–42 µm; sometimes two different sizes have been found. Exine 3–9 µm thick and ornamentation microreticulate and echinate. The results clearly show the relationships between genera. Moreno-Socías, E., Mejías, J. A., Díez, M. J., 1994: Morfología polínica deLactuceae (Asteraceae) en la Península Ibérica, I.Lactuca y géneros relacionados. — Acta Bot. Malacitana.19: 103–113.     

A revision of Casimirella,including Humirianthera (Icacinaceae)

Casimirella Hassler (1913) is accepted and Humirianthera Huber (1914) considered a synonym. Casimirella diversifolia and C. lanata from Brazil are described as new species. Casimirella ampla (Miers) based on Leretia ampla Miers, C. crispula (Howard) based on Humirianthera crispula Howard, and C. rupestris (Ducke) based on Humirianthera rupestris Ducke are new combinations.     

Agouticarpa, a new neotropical genus of Tribe Gardenieae (Rubiaceae)

Agouticarpa, a new genus of the informalAlibertia group (Rubiaceae), is described and illustrated. It comprises six species, and occurs from Costa Rica to Bolivia. Three species are here described as new:Agouticarpa grandistipula, A. hirsuta, andA. velutina. Additionally, three new combinations are made:A. curvifolia andA. williamsii (previously inGenipa) andA. isernii (previously inAlibertia). Agouticarpa is characterized by being dioecious, having elliptic to abovate, membranaceous stipules, male flowers in a branched dichasial or thyrse-like inflorescence, a poorly developed cup-shaped calyx, pollen grains with 3–7 apertures, and large globose fruits.     

The cell wall of Chlamydomonas reinhardtii gametes: Composition,structure and autolysin-mediated shedding and dissolution

The cell walls of Chlamydomonas gametes are multilayered structures supported on frameworks of polypeptides extending from the plasma membrane. The wall-polypeptide catalogue reported by Monk et al. (1983, Planta 158, 517–533) and extended by U.W. Goodenough et al. (1986, J. Cell Biol. 103, 405–417) was re-evaluated by comparative analysis of mechanically isolated cell walls purified from several strains. The extracellular locus of wall polypeptides was verified by in vivo iodogen-catalysed iodination and by autolysin-mediated elimination of the bulk of these polypeptides from the cell surface. Three (w15, w16, w17) and possibly four (w14) polypeptides were located to the most exterior aspect of the wall because of their susceptibility to Enzymobeadcatalysed iodination and their retention by a cell-wall-less mutant. The composition of shed walls stabilised with ethylenediaminetetraacetic acid during natural mating and kinetic analysis of the dissolution of walls purified from a bald-2 mutant demonstrated the rapid and specific destruction of polypeptide w3. Differential solubilisation of wall polypeptides occurred after loss of w3. Wall dissolution, characterised by the generation of fishbone structures from the W2 layer, gave as many as four additional polypeptides. Charged detergents and sodium perchlorate extracted a comparable range of polypeptides at room temperature from mechanically isolated walls, i.e. components of the W4–W6 layers, hot sodium dodecyl sulphate solubilised framework polypeptides, while reducing agent was required to solubilise the W2 layer. A model of wall structure is presented.Abbreviations DTE dithioerythritol - EDTA ethylenediaminetetraacetic acid - M_r relative molecular mass - SDS-PAGE sodium dodecyl sulphate-polyacrylamide gel electrophoresis - Tris 2-amino-2-(hydroxymethyl)-1,3-propanediol     

Agave rzedowskiana, a new species in subgenus Littaea (Agavaceae) from western Mexico

Agave rzedowskiana is described and illustrated from the states of Sinaloa and Jalisco, Mexico. This species is a member of subgenusLittaea (Tagliabue) Baker “groupStriatae” of Baker. It is compared withA. petrophila García-Mend. & E. Martínez andA. dasylirioides Jacobi & Bouché.

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Resumen  Se describe e ilustraAgave rzedowskiana de los estados de Sinaloa y Jalisco, México. La especie pertenece al subgéneroLittaea (Tagliabue) Baker “Grupo Striatae” de Baker. Se le compara conA. petrophila García-Mend. & E. Martínez yA. dasylirioides Jacobi & Bouché.

     

Recognition of glycoconjugates byHelicobacter pylori: an apparently high-affinity binding of human polyglycosylceramides,a second sialic acid-based specificity

Helicobacter pylori has been reported to agglutinate erythrocytes and to bind to various other cells in a sialic acid-dependent way. The binding was inhibited by sialyllactose or fetuin and other sialylated glycoproteins. The specificity apparently requires bacterial growth on agar, since we found that it was lost after growth in the nutrient mixture Ham's F12. Instead, the bacteria bound with high affinity and in a sialic acid-dependent way to polyglycosylceramides of human erythrocytes, a still incompletely characterized group of complex glycolipids.Bacteria grown in F12 medium were metabolically labelled with³⁵S-methionine and analysed for binding to glycolipids on thin-layer chromatograms and to glycoproteins on blots after electrophoresis, with human erythrocyte glycoconjugates in focus. There was no binding to simpler gangliosides including GM3 or sialylparagloboside, or to a mixture of brain gangliosides. In contrast, polyglycosylceramides of human erythrocyte membranes bound at a pmol level. The activity was eliminated by mild acid treatment, mild periodate oxidation or sialidase hydrolysis. Erythrocyte proteins as well as a range of reference glycoproteins did not bind, except band 3, which was weakly active. However, this activity was resistant to periodate oxidation.These results indicate a second and novel sialic acid-recognizing specificity which is expressed independently of the previously described specificity. Abbreviations: PGCs, polyglycosylceramides; TLC, thin-layer chromatography; C, chloroform; M, methanol; EI/MS, electron impact ionization mass spectrometry, SDS PAGE, sodium dodecylsulfate polyacrylamide gel electrophoresis; PBS, phosphate-buffered saline; PVDF, polyvinylidene difluoride; BSA, bovine serum albumin. The carbohydrate and glycosphingolipid nomenclatures are according to recommendations of IUPAC-IUB Commission on Biochemical Nomenclature (Lipids (1977)12: 455–68;J Biol Chem (1982)257: 3347–51 andJ Biol Chem (1987)262: 13–18).This paper is dedicated to Professor S.-i. Hakomori and is paper no. 1 from our research onHelicobacter pylori.     

New triterpenoid saponins from Patrinia scabiosaefolia

Phytochemical investigation of the methanol extract from the whole plants of Patrinia scabiosaefolia Fisch. resulted in the isolation of four new triterpenoid saponins (1–4) along with six known compounds (5–10). On the basis of spectroscopic and chemical methods, the structures of the new compounds were established as 3-O-β-d-xylopyranosyl-(1→3)-α-l-rhamnopyranosyl-(1→2)-β-d-xylopyranosyl-12β,30-dihydroxy-olean-28,13β-olide (1), 3-O-α-l-rhamnopyranosyl-(1→2)-β-d-xylopyranosyl-12β,30-dihydroxy-olean-28,13β-olide (2), 3-O-β-d-xylopyranosyl-(1→2)-β-d-glucopyranosyl-12β, 30-dihydroxy-olean-28,13β-olide (3), and 3-O-β-d-glucopyranosyl-(1→4)-β-d-xylopyranosyl-(1→3)-α-l-rhamnopyranosyl-(1→2)-β-d-xylopyranosyl-oleanolic acid 28-O-β-d-glucopyranoside (4), respectively. Compounds 1–3 possess a novel 12β,30-dihydroxy-olean-28,13β-lactone aglycone and a 12β-hydroxy substituent that is rarely found in this kind of triterpenoid saponin.     

New Substrates for Enteropeptidase: I. Biologically Active Hepta-, Octa-, and Nonapeptides

Enteropeptidase (enterokinase, EC 3.4.21.9) hydrolyzes peptide bonds formed by carboxyl groups of Lys or Arg residue if less than four negatively charged amino acid residues are in positions P ₂–P ₅ of its substrate. We determined the kinetic parameters of three substrates of this type: human angiotensin II (AT) (DR VYIHPF) and the Hb(2–8) (LTAEEK A) and Hb(1–9) (MLTAEEK AA) peptides of the cattle hemoglobin -chain. The K _m values for all the substrates (10^–3 M) were one order of magnitude higher than those of the typical synthetic substrates of enteropeptidase or chimeric proteins with the –DDDDK– full-size linker (K _m 10^–4 M). The k _cat values for AT and Hb(2–8) were also close and low (30 min^–1). The general hydrolysis efficiency of such substrates is no more than 1% of the corresponding value for the typical peptide and protein substrates of the enteropeptidase. However, the elongation of Hb(2–8) peptide by one amino acid residue from both its N- and C-termini results in a dramatic increase in the catalytic efficiency of the hydrolysis: the k _cat value for Hb(1–9) is 1510 min^–1, which means that it is hydrolyzed only three times less effective than the chimeric protein with the full-size linker.     

Metabolic changes in tomato fruits and seeds after viral, bacterial and fungal infections

The metabolic changes in tomato fruits and seeds separately infected with cucumber mosaic virus, Pseudomonas syringae pv. tomato or Botrytis cinerea were investigated cytochemically. The changes of peroxidase (E.C. 1.11.1.7) and β-glucosidase (E.C. 3.2.1.21) were investigated biochemically as well. Tomato fruits were involved in the study because of their high economic value. Tomato seeds were investigated since they have been used most extensively as a model system for studying the physiology and biochemistry of seed development. The diseases caused by the pathogens under study are of special importance for yield reduction in tomato. The three pathogens provoked local changes in the activities of enzymes under study that affect the infected pericarp tissues and neighboring seeds. It was established non-specific and specific responses. The non-specific responses of invaded tissues were expressed as a local enhancement of peroxidase activity in both pericarp tissues and seeds as well as a decrease in activities of: i. enzymes taking part in aerobic and anaerobic respiration, ii. hydrolases esterase and acid phosphatase involved in the basic metabolism as well as an enhancement of their activities in neighboring tissues. Furthermore, it was observed an enhancement of α-galactosidase activity in infected area was observed. The specific responses depending on the type of the pathogen consisted in an enhancement of glucose-6-phosphate dehydrogenase activity by virus infection and an increase of β-glucosidase activity by fungal invasion.     

Pinus georginae (Pinaceae), a new species from western Jalisco, Mexico

Pinus georginae is described and illustrated as a new endemic species from the valleys in the southern basin of Río Ameca, Jalisco, western Mexico. It is closely related to Pinus praetermissa; however, after studying the cones and anatomy of the leaves in detail, characteristics were revealed that identify P. georginae as a distinct species.

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Resumen  Se describe e ilustra Pinus georginae como una especie nueva, endémica de algunos valles intermontanos al sur del Río Ameca, Jalisco, en el Occidente de México. Los árboles se parecen a Pinus praetermissa; sin embargo, al estudiar con detalle los conos y la anatomía de las acículas lo ubican a Pinus georginae como una entidad biológica con categoría específica independiente.

     

Antimicrobial Butyrolactone I Derivatives from the Ecuadorian Soil Fungus Aspergillus terreus Thorn. var terreus

Summary The fungus Aspergillus terreus Thorn var. terreus isolated from an Ecuador soil sample was cultured in liquid and solid media and yielded three main metabolites identified as terreic acid (1), butyrolactone I (2) and lovastatin (3). The natural products as well as three synthetic butyrolactone I derivatives were assessed for antimicrobial activity against Gram-positive and Gram-negative bacteria and fungi as well as for seed germination and seedling growth. Furthermore, the compounds were assessed as inhibitors towards the enzymes acetylcholinesterase, β-glucosidase, and β-glucuronidase. Terreic acid, butyrolactone I, butyrolactone 4′,4′′-diacetate (2.1), and 3′-(3-methylbutyl)-butyrolactone II (2.2) were active towards the phytopathogenic bacteria Erwinia carotovora with IC₅₀ of 5 and 4–18 μg/ml, respectively. Under the same experimental conditions, the IC₅₀ of streptomycin was 1.9 μg/ml. 3′-(3-Methylbutyl)-butyrolactone II was moderately active against Pseudomonas syringae and Botrytis cinerea with IC₅₀ of 21μg/ml and MIC of 15.6 μg/ml, respectively. Butyrolactone I also inhibited germination of the dicot Lactuca sativa with an IC₅₀ of 5 × 10⁻⁵ M. The IC₅₀ of reference herbicide acetochlor was 1 × 10⁻⁵ M. The effect of 2.2 and 2.3, known as butyrolactone III on Panicum millaceum germination and growth was stronger than that of 2 and 2.1. Reduction of the double bond in the isoprenyl side chain of butyrolactone I increased the antibacterial effect against E. carotovora as well as acetylation. To our best knowledge, this is the first report on the antibacterial effect of butyrolactone derivatives towards Erwinia carotovora and the phytopathogenic fungus Botrytis cinerea. The butyrolactone I derivative 2.2 presented a moderate inhibitory effect against the enzyme acetylcholinesterase with an IC₅₀ of 47 μg/ml. Under the same experimental conditions, the reference inhibitor galanthamine had an IC₅₀ of 3 μg/ml.     

Cytotoxic and antioxidant triterpene saponins from Butyrospermum parkii (Sapotaceae)

Three new triterpenoid saponins, elucidated as 3-O-β-d-glucopyranosyloleanolic acid 28-O-β-d-xylopyranosyl-(1→4)-α-l-rhamnopyranosyl-(1→2)-β-d-xylopyranoside (parkioside A, 1), 3-O-[β-d-apifuranosyl-(1→3)-β-d-glucopyranosyl]oleanolic acid 28-O-[β-d-apifuranosyl-(1→3)-β-d-xylopyranosyl-(1→4)-[α-l-rhamnopyranosyl-(1→3)]-α-l-rhamnopyranosyl-(1→2)β-d-xylopyranoside (parkioside B, 2) and 3-O-β-d-glucuronopyranosyl-16α-hydroxyprotobassic acid 28-O-α-l-rhamnopyranosyl-(1→3)-β-d-xylopyranosyl-(1→4)-α-l-rhamnopyranosyl-(1→2)-β-d-xylopyranoside (parkioside C, 3), were isolated from the n-BuOH extract of the root bark of Butyrospermum parkii, along with the known 3-O-β-d-glucopyranosyloleanolic acid (androseptoside A). The structures of the isolated compounds were established on the basis of chemical and spectroscopic methods, mainly 1D and 2D NMR data and mass spectrometry. The new compounds were tested for both radical scavenging and cytotoxic activities. Compound 2 showed cytotoxic activity against A375 and T98G cell lines, with IC₅₀ values of 2.74 and 2.93 μM, respectively. Furthermore, it showed an antioxidant activity comparable to that of Trolox or butylated hydroxytoluene (BHT), used as controls, against 2,2-diphenyl-1-picryl hydrazyl (DPPH), 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), oxygen and nitric oxide radicals.     

Synthesis of p-nitrophenyl sulfated disaccharides with beta-D-(6-sulfo)-GlcNAc units using beta-N-acetylhexosaminidase from Aspergillus oryzae in a transglycosylation reaction

When α-d-GlcNAc-OC₆H₄NO₂ -p and β-d-(6-sulfo)-GlcNAc-OC₆H₄NO₂-p (2) were used as substrates, β-N-acetylhexosaminidase from Aspergillus oryzae transferred the β-d-(6-sulfo)-GlcNAc(unit from 2 to α-d-GlcNAc-OC₆H₄NO₂ -p to afford β-d-(6-sulfo)-GlcNAc-(1→4)-α-d-GlcNAc-OC₆H₄NO₂-p (3) in a yield of 94% based on the amount of donor, 2, added. β-d-(6-sulfo)-GlcNAc-(1→4)-α-d-Glc-OC₆H₄NO₂-p (4) was obtained with α-d-Glc-OC₆H₄NO₂ -p as acceptor in a similar manner. With a reaction mixture of 2 and β-d-GlcNAc-OC₆H₄NO₂-p (1) in a molar ratio of 6:1, the enzyme mediated the transfer of β-d-GlcNAc from 1 to 2, affording disaccharide β-d-GlcNAc-(1→4)-β-(6-sulfo)-d-GlcNAc-OC₆H₄NO₂-p (5) in a yield of 13% based on the amount of 1 added.