Isolation and in vitro hydrolysis of lentil protein fractions by trypsin

The albumin and globulin fractions from lentil seeds were isolated and characterised by gel filtration. The latter was shown to be homogeneous and the former heterogeneous on PAGE. The aminoacid analysis revealed high values of amidic amino acids for both fractions with great differences in the sulphur-containing amino acids. Native albumin, globulin and salt-soluble proteins were markedly resistant to trypsin hydrolysis compared to casein. The SDS-PAGE of native salt-soluble proteins indicated that the globulin fragments (20 to 30 kD) were slowly digested in the presence of albumin. The heating increased the hydrolysis of the proteins in the order: salt-soluble, albumin and globulin. The facilitated hydrolysis of the heated salt-soluble fraction seemed to be due to protein-protein interactions induced by heat.

Isolation and characterization of microsatellite loci in the woolly mouse opossum, Micoureus paraguayanus (Marsupialia : Didelphimorphia)

Five microsatellite loci were isolated and characterized within the woolly mouse opossum (Micoureus paraguayanus), a Neotropical marsupial, using an enrichment cloning procedure. Between four and seven alleles were detected per locus, with expected heterozygosity ranging from 0.358 to 0.560. These microsatellites should provide useful markers in a variety of genetic analyses to examine parentage, inbreeding, population structure and population dynamics in fragmented forest habitats.

Isolation and structure elucidation of two new flavonoid glycosides from the infusion of Maytenus aquifolium leaves. Evaluation of the antiulcer activity of the infusion

Droplet countercurrent chromatography and high-performance liquid chromatography fractionation of the aqueous infusion from Maytenus aquifolium Martius leaves afforded two flavonoid tetrasaccharides: quercetin 3-O-alpha-L-rhamnopyranosyl(1-->6)-O-[beta-D-glucopyranosyl(1-->3)-O-alpha-L-rhamnopranosyl( 1-->2)-O-beta-D-galactopyranoside and kaempferol 3-O-alpha-L-rhamnopyranosyl(1-->6)-O-[beta-D-glucopyranosyl( 1-->3)-O-alpha-L-rhamnopyranosy(1-->2-2)-O-beta-D-galactopyranoside. All structures were elucidated by spectroscopic methods. Pharmacological essays of the infusion showed antiulcer activity in rats.

PRODUCTIVITY IN MASS CROSSES OF DROSOPHILA-STURTEVANTI - A COMPARATIVE-STUDY OF LABORATORY STOCKS AND RECENTLY COLLECTED FLIES

Drosophila sturtevanti from several geographic origins were analyzed for their capacity to intercross and to yield progeny. Mass intercrosses involving laboratory stocks and recently collected strains were fertile, which suggests that the genetic differentiation among these geographically isolated populations did not affect their reproductive patterns sufficiently to lead to reproductive isolation. Analysis of the number of progeny (productivity) in intracrosses and intercrosses was informative as to the amount of variation this feature exhibits in the laboratory stocks and in the recently collected strains. Also laboratory stocks and recently collected flies shared a positive correlation in that the greater the control productivity of a strain the greater the number of its intercrosses which exhibited reduced productivity.

ISOLATION AND IN-VITRO HYDROLYSIS OF GLOBULIN G1 FROM LENTILS (LENS-CULINARIS, MEDIK)

The major globulin fraction from lentil seeds was investigated with respect td in vitro hydrolysis by trypsin and chymotrypsin. Globulin was isolated by a NaCl-ascorbate extraction procedure and purified by DEAE-cellulose chromatography and gelfiltration chromatography on Sepharose CL-6B. The purity and identification of the protein were performed by PAGE. The native globulin, with a molecular weight of 375 kD, was resolved by SDS-PAGE into twelve polypeptides with molecular weights ranging from 61 to 14.5 kD. Native and heated globulin GI was hydrolyzed with trypsin and chymotrypsin. SDS-PAGE indicated that native globulin was more resistant to digestion than heated protein. Amino acid analysis of the major globulin revealed that glutamic acid was present in the largest concentration, followed by aspartic acid, arginine and leucine. As is also the case for other legumin-like globulins, lentil GI was deficient in sulfur-containing amino acids.

Isolation of Campylobacter jejuni from viscera and bile secretion of broiler chickens with diarrhea

Since chickens are important reservoirs of Campylobacter jejuni and their meat is the most frequent route of transmission for human campylobacteriosis, the purpouse of the present study was to investigate the presence of Campylobacter in viscera of chickens with diarrhea, evaluating the frequency of isolation of this microorganism from organs considered to be preferential for its isolation. A total of 107 visceral samples from chickens with diarrhea from different farms in the Ribeirao Preto region, SP, were examined for the presence of Campylobacter jejuni. The material consisted of 73 livers and 34 spleens, plus 29 bile secretion samples. The frequency of Campylobacter jejuni isolation was 54.79% for the liver samples, 35,29% for the spleens and 6,89% for the bile secretion samples. The data suggest that, under the conditions of the present study, the liver may be the organ of choice for the isolation of Campylobacter in the presence of diarrhea and liver involvement in chickens.

A study of habitat fragmentation in Southeastern Brazil using remote sensing and geographic information systems (GIS)

The purpose of this work was to study fragmentation of forest formations (mesophytic forest, riparian woodland and savannah vegetation (cerrado)) in a 15,774-ha study area located in the Municipal District of Botucatu in Southeastern Brazil (São Paulo State). A land use and land cover map was made from a color composition of a Landsat-5 thematic mapper (TM) image. The edge effect caused by habitat fragmentation was assessed by overlaying, on a geographic information system (GIS), the land use and land cover data with the spectral ratio. The degree of habitat fragmentation was analyzed by deriving: 1. mean patch area and perimeter; 2. patch number and density; 3. perimeter-area ratio, fractal dimension (D), and shape diversity index (SI); and 4. distance between patches and dispersion index (R). In addition, the following relationships were modeled: 1. distribution of natural vegetation patch sizes; 2. perimeter-area relationship and the number and area of natural vegetation patches; 3. edge effect caused by habitat fragmentation, the values of R indicated that savannah patches (R = 0.86) were aggregated while patches of natural vegetation as a whole (R = 1.02) were randomly dispersed in the landscape. There was a high frequency of small patches in the landscape whereas large patches were rare. In the perimeter-area relationship, there was no sign of scale distinction in the patch shapes, In the patch number-landscape area relationship, D, though apparently scale-dependent, tends to be constant as area increases. This phenomenon was correlated with the tendency to reach a constant density as the working scale was increased, on the edge effect analysis, the edge-center distance was properly estimated by a model in which the edge-center distance was considered a function of the to;al patch area and the SI. (C) 1997 Elsevier B.V. B.V.

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).

A rapid procedure for the isolation of the Lys-49 myotoxin II from Bothrops moojeni (caissaca) venom: Biochemical characterization, crystallization, myotoxic and edematogenic activity

A, M. Soares, V, M, Rodrigues, M. I. Homsi-Brandeburgo, M. H. Toyama, F, R, Lombardi, K. Arni and J. R, Giglio. A rapid procedure for the isolation of the Lys-49 myotoxin II from Bothrops moojeni (caissaca) venom: Biochemical characterization, crystallization, myotoxic and edematogenic activity. Toxicon 36, 503-514, 1998.-Bothrops moojeni snake venom was fractionated on a CM-Sepharose column which was previously equilibrated with 0.05 M ammonium bicarbonate buffer at pH 8.0 and subsequently eluted with an ammonium bicarbonate concentration gradient from 0.05 to 0.5 M at constant pH (8.0) and temperature (25 degrees C). The fraction which eluted last (M-VI) showed, after direct lyophilization, a single band by polyacrylamide gel electrophoresis (PAGE) and SDS-PAGE, indicating an approximate M,. of 14 000 and 77 000, in the presence and absence of dithiothreitol, respectively. Its amino acid composition revealed a high level of hydrophobic and basic amino acids as well as 13 half-cystine residues. Its isoelectric point and extinction coefficient (E-1.0cm(1.0mg/ml) at 278 nm and pH 7.0) were 8.2 and 1.170, respectively. M-VI was devoid of phospholipase A(2) (PLA(2)) activity on egg yolk, as well as of hemorrhagic, anticoagulant and coagulant activities, but could induce drastic necrosis on skeletal muscle fibres as well as rapid and transient edema on the rat paw. Its N-terminal sequence: SLFELGKMILQETGKNPAKSYGVYGCNCGVGGRGKPKDATDRCCYVHKCCYK.... revealed high homology with other Lys 49 PLA(2)-like myotoxins from other bothropic venoms. Orthorhombic crystals of M-VI? which diffracted to a maximal resolution of 1.6 Angstrom. were obtained and indicated the presence of a dimer in the asymmetrical unit. (C) 1998 Elsevier B.V. Ltd. All rights reserved.

A SIMPLE TECHNIQUE FOR ISOLATION OF DNA SUITABLE FOR PCR AMPLIFICATION FROM CYTOGENETIC PREPARATIONS

In order to rescue molecular information from chromosome preparations, we describe a rapid procedure to obtain DNA from cytogenetic preparations in microscope slides, stored for one to live years at room temperature. This technique was modified from previously described procedures and the DNA obtained was shown to be suitable for PCR amplification.

Isolation and sequence determination of peptides in the venom of the spider wasp (Cyphononyx dorsalis) guided by matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry

Micro-scale (sub-pmol) isolation and sequence determination of three peptides from the venom of the solitary spider wasp Cyphononyx dorsalis is described. We isolated two novel peptides Cd-125 and Cd-146 and a known peptide Thr(6)-bradykinin from only two venom sacs of solitary spider wasp Cyphononyx dorsalis without bioassay-guided fractionation. but instead guided by MALDI-TOF MS. The MALDI-TOF MS analysis of each fraction showed the purity and molecular weight of the components, which led to the isolation of the peptides virtually without loss of sample amount. The sequences of the novel peptides Cd-125 (Asp-Thr-Ala-Arg-Leu-Lys-Trp-His) and Cd-146 (Ser-Glu-Thr-Gly-Asn-Thr-Val-Thr-Val-Lys-Gly-Phe-Ser-Pro-Leu-Arg) were determined by Edman degradation together with mass spectrometry. and finally corroborated by solid-phase synthesis. The known peptide Thr(6)-bradykinin (Arg-Pro-Pro-Gly-Phe-Thr-Pro-Phe-Arg) was identified by comparison with the synthetic authentic specimen. This is the first example for any kinins to be found in Pompilidae wasp venoms. The procedure reported here can be applicable to studies on many other components of solitary wasp venoms with limited sample availability. (C) 2001 Elsevier B.V. Ltd. All rights reserved.