Role of the triad N46, S106 and T107 and the surface charges in the determination of the acidic pH optimum of digestive lysozymes from Musca domestica

Structures of digestive lysozymes 1 and 2 from housefly (MdL1 and MdL2) show that S106-T107 delimit a polar pocket around E32 (catalytic acid/base) and N46 contributes to the positioning of 050 (catalytic nucleophile), whereas those residues are replaced by V109-A110 and D48 in the non-digestive lysozyme from hen egg-white (HEWL). Further analyses revealed that MdL1 and MdL2 surfaces are less positively charged than HEWL surface. To verify the relevance of these differences to the acidic pH optimum of digestive lysozymes it was determined that pKas of the catalytic residues of the triple mutant MdL2 (N46D-S106V-T107A) are similar to HEWL pKas and higher than those for MdL2. In agreement, triple mutant MdL2 and HEWL exhibits the same pH optimum upon methylumbelliferylchitotrioside. In addition to that, the introduction of six basic residues on MdL1 surface increased by 1 unit the pH optimum for the activity upon bacterial walls. Thus, the acidic pH optimum for MdL2 and MdL1 activities upon methylumbelliferylchitotrioside is determined by the presence of N46, S106 and T107 in the environment of their catalytic residues, which favors pKas reduction. Conversely, acidic pH optimum upon bacterial walls is determined by a low concentration of positive charges on the MdL2 and MdL1 surfaces. (C) 2010 Elsevier Inc. All rights reserved.

Purification, characterization and sequencing of the major beta-1,3-glucanase from the midgut of Tenebrio molitor larvae

The major beta-1,3-glucanase from Tenebrio molitor (TLam) was purified to homogeneity (yield, 6%; enrichment, 113 fold; specific activity, 4.4 U/mg). TLam has a molecular weight of 50 kDa and a pH optimum of 6. It is an encloglucanase that hydrolyzes beta-1,3-glucans as laminarin and yeast beta-1,3-1,6-glucan, but is inactive toward other polysaccharides (as unbranched beta-1,3-glucans or mixed beta-1,3-1,4-glucan from cereals) or disaccharides. The enzyme is not inhibited by high substrate concentrations and has low processivity (0.6). TLam has two ionizable groups involved in catalysis, and His, Tyr and Arg residues plus a divalent ion at the active site. A Cys residue important for TLam activity is exposed after laminarin binding. The cDNA coding for this enzyme was cloned and sequenced. It belongs to glycoside hydrolase family 16, and is related to other insect glucanases and glucan-binding proteins. Sequence analysis and homology modeling allowed the identification of some residues (E174, E179, H204, Y304, R127 and R181) at the active site of the enzyme, which may be important for TLam activity. TLam efficiently lyses fungal cells, suggesting a role in making available walls and cell contents to digestion and in protecting the midgut from pathogen infections. (C) 2009 Elsevier Ltd. All rights reserved.

Anthrone and oxanthrone C,O-diglycosides from Picramnia teapensis

Two C,O-diglycosylated compounds, the anthrone picramnioside F, and the oxanthrone mayoside C, were isolated from the stem bark of Picramnia teapensis, along with the previously reported anthraquinones, 1-O-beta -D- and 8-O-beta -D-glucopyranosyl emodin. The compounds were separated by recycling-HPLC, and their structures were determined on the basis of spectroscopic analysis. CD measurements were used to establish the absolute configuration of the anthrone and oxanthrone. The antifungal activity of 1-O-beta -D- and 8-O--D-glucopyranosyl emodin against Leucoagaricus gongilophorus was shown to be similar to that of the lignan sesamin. (C) 2000 Elsevier B.V. Ltd. All rights reserved.

Chemical Composition and Antioxidant and Antimycobacterial Activities of Bromelia balansae (Bromeliaceae)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Intraspecific variability of flavonoid glycosides and styrylpyrones from leaves of Cryptocarya mandioccana Meisner (Lauraceae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Metabolite fingerprint of "capim dourado" (Syngonanthus nitens), a basis of Brazilian handcrafts

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Antioxidant flavonol glycosides from Nectandra grandiflora (Lauraceae)

O fracionamento cromatográfico do extrato etanólico das folhas de Nectandra grandiflora resultou no isolamento de dois flavonóides glicosilados que apresentaram atividade antioxidante inibindo a oxidação do beta -caroteno em CCDC. As substâncias isoladas tiveram suas estruturas elucidadas através de técnicas espectrométricas de RMN uni- e bidimensional e foram identificadas como 3-O-beta -ramnosy kaempferol and 3-O-beta -ramnosylquercetina.

Xanthones and flavonoids from Leiothrix curvifolia and Leiothrix flavescens

8-Carboxymethyl-1,6-dihydroxy-3,5-dimethoxyxanthone 8-carboxymethyl-1,5,6-trihydroxy-3-methoxyxanthone and 8-carboxy-methyl-1,3,5,6-tetrahydroxyxanthone were isolated from the capitula of Leiothrix curvifolia and Leiothrix flavescens and characterized by spectroscopic methods, mainly 1D and 2D NMR experiments, as well as by electrospray mass spectrometry. Eight known flavonoids were also isolated and they were identified by 1D and 2D NMR experiments and comparison with literature data. (C) 2001 Elsevier B.V. Ltd. All rights reserved.

Studies on the constituents of a Brazilian folk infusion. Isolation and structure elucidation of new triterpene saponins from Ilex amara leaves

The isolation of three new triterpene saponins 3beta-O-beta-D-glucopyranosyl-(1-->3)-alpha-L-2-O-acetylara-binopyranosylolean-12-en-28-oic acid 28-O-beta-D-glucopyranosyl ester (2), 3beta-O-beta-D-glucopyranosyl-(1-->2)-alpha-L-O-arabinopyranosylurs-12-en-28-oic acid (3), and 3beta-O-beta-D-glucopyranosyl-(1-->2)-beta-D-O-galactopyranosylurs-12-en-28-oic acid (4) together with five known saponins and one flavonoid glycoside from the aqueous infusion of flex amara (Vellozo) Loes. leaves is reported. All structures were elucidated by spectroscopic methods, including the concerted application of one-dimensional (H-1, TOCSY, C-13, and C-13 DEPT NMR) and two-dimensional NMR techniques (DQF-COSY, HSQC, and HMBC).

Antifungal iridoids from the stems of Tocoyena formosa

A new antifungal iridoid compound, together with three active known iridoids, galioside, galioside aglucone, and apodanthoside, as well as 3-O-beta-D-glucopyranosyl-28-O-beta-D-glucopyranosyl quinovic acid and quercetin-3-O-methyl-3-O-beta-D-rhamnopyranoside, were isolated from the stems of Tocoyena formosa. This new compound was characterized by spectral data as 11-O-trans-feruloylteucrein.

Pharmacological and toxicological studies of Drimys angustifolia Miers. (Winteraceae)

Drimys angustifolia Miers. (Winteraceae) is a Brazilian medicinal plant used as analgesic, antiulcer and anti-inflammatory without studies to assure its efficacy and safety Leaf and stem bark extracts were evaluated to determine the antiulcer, analgesic, antiinflammatory and antioxidant activities. Preliminary toxic effects and qualitative phytochemical profile were also performed. The antiulcer activity was detected in both extracts. Administration of the leaf extract at 250 mg/kg inhibited total lesion area by 76.50% (p < 0.01 in ethanol/HCl method), while carbenoxolone at 250 mg/kg reduced lesions by 69.48%. Stem bark extract (250 mg/kg) inhibited lesion by 81.42%, while carbenoxolone by 74.10%. Similar effects were observed in the ethanol-induced ulcer method, but no activity was observed in piroxican model. The effects involve nitric oxide in gastric protection, since the L-NAME treatment reversed the protection given by the extracts. Antioxidant effects suggest an involvement against oxidative stress. In the pain (writhing, tail-flick and hot-plate tests) and inflammation (carrageenan-induced paw edema) models, the extracts did not present any effect. The phytochemical studies demonstrated that both extracts contain flavonoids, saponins, glycosilated triterpenoids, fixed acids, cyanogenic glycosides, quinones, tannins, xanthone and steroidal aglycones. Toxicological studies showed that the extracts are safe at the effective antiulcer doses. (c) 2006 Elsevier B.V.. All rights reserved.

Planifolin, a new naphthopyranone dimer and flavonoids from Paepalanthus planifolius

A new naphthopyranone dimer (I) named planifolin was isolated from a methylene chloride extract of the capitula of Paepalanthus planifolius. The structure of 1 has been determined by chemical and spectroscopic means. In addition, a known dihydronaphthopyranone glycoside and seven known flavonoids were isolated from an ethanolic extract of the leaves of P. planifolius.

Naphthopyranone glycosides from Paepalanthus bromelioides

Two new naphtho[2,3-C]pyran-1-one glycosides, paepalantine-9-O-β-D- glucopyranoside and paepalantine-9-O-β-D-allopyranosyl(1 → 6)glucopyranoside, were isolated from an ethanolic extract of capitula from Paepalanthus bromelioides and identified from their spectrometric data.

Constituintes de Sorocea bomplandii Baillon

The chemical investigation of the leaves of Sorocea bomplandii Baillon (Moraceae) led to the identification of pentacyclic triterpenes, fatty acid ester and isoprenoids. Chromatographyc comparison between the infusion S. bomplandii with that of Maytenus aquifolium - a Celastraceae with proved antiulcer activity - has shown that they have different compositions: the first one is based on sugars and the last one is based on flavonoid glycosides.

A new application of humic substances: Activation of supports for invertase immobilization

Invertase was immobilized on aminopropyl silica (APTS-SiO2) activated with humic substances (APTS-SiO2-HS) and on aminopropyl silica activated with glutaraldehyde (APTS-SiO2-GA). The resulting activity of both systems was compared. Humic substances (HS) used for the activation of the silica were extracted from soil of Cananéia, São Paulo State, Brazil, according to the procedure recommended by the International Humic Substances Society. Activity was determined by measuring the rate of formation of reduced sugars using the reaction with dinitrosalicylic acid (DNS). The amount of HS bound on the APTS-SiO2 was equal to 50 mg. The maximum amount of invertase immobilized on APTS-SiO2-HS was 15200 U/g while in the system APTS-SiO2-GA it was 13400 U/g. The experimental enzymatic activity was 3700 and 3300 U/g, for the systems APTS-SiO2-HS and APTS-SiO2-GA, respectively. Considering the increased amount and activity of immobilized enzyme compared with the glutaraldehyde method, it was concluded that this technique opens a new perspective in the preparation of supports for enzyme immobilization employing humic substances. © Springer-Verlag 2000.