A.M. Soares

Direct capture of lactoferrin from cheese whey on supermacroporous column of polyacrylamide cryogel with copper ions

Lactoferrin is a protein that is present in cheese whey (a waste product from the dairy industry) and has several biological activities. However, its production from whey must have a high yield and low cost for industrial applications. As such, this study reports the use of polyacrylamide cryogel, loaded with Cu(2+) (through the bond with iminodiacetic acid (IDA)), as an adsorbent for the chromatographic process to capture lactoferrin whey. Ultrafiltered cheese whey was passed through the cryogel-IDA-Cu(2+) system. The eluates were subjected to analysis of total protein, SDS-PAGE and size exclusion chromatography. The results showed an axial dispersion coefficients, at different superficial velocities of liquid, in a range of 10(-6)-10(-5)m(2)/s. The cryogel demonstrated good hydraulic permeability (4.7086×10(-13)m(2)) and a porosity of approximately 78.2%. The IDA-Cu(2+) cryogel system was also able to capture lactoferrin in high purity.

Microcalorimetric study of the adsorption of lactoferrin in supermacroporous continuous cryogel with immobilized Cu2+ ions

The adsorption affinity of lactoferrin from whey in monolithic supermacroporous cryogel was analyzed using equilibrium data adsorptive isothermal titration microcalorimetry to measure thermodynamic information governing the process. Isotherm data was obtained at temperatures of 20, 30 and 40°C, pH 6, 7 and 8, and ionic strength of 200, 600 and 1000mmolL(-1) NaCl. The Langmuir model was fitted to equilibrium data. The binding was tighter at higher temperatures. The adsorption of protein was observed as spontaneous in all cases analyzed. The microcalorimetric study indicated that, in most cases examined, the adsorption of the protein in the matrix was entropy and enthalpy favored and entropy driven. Results provide data to enable the improvement of technical processes for the affinity separation of proteins.

Vascular effects and electrolyte homeostasis of the natriuretic peptide isolated from Crotalus oreganus abyssus (North American Grand Canyon rattlesnake) venom

Crotalus oreganus abyssus is a rattlesnake that is usually found in the Grand Canyon, United States of America. Knowledge regarding the composition of C. o. abyssus venom is scarce. New natriuretic peptides (NPs) have been isolated and characterized from the venoms of members of the Crotalinae family. The NP family comprises three members, ANP (atrial natriuretic peptide), BNP (b-type natriuretic peptide) and CNP (c-type natriuretic peptide), and has an important role in blood pressure regulation and electrolyte homeostasis. The aim of the present study was to characterize a novel natriuretic-like peptide (Coa_NP2), isolated from C. o. abyssus venom. The Coa_NP2 presents an average molecular mass of 3419.88Da (theoretical average molecular mass 3418.94Da, monoisotopic molecular mass 3416.66Da and theoretical PI 7.78) and its amino acid sequence presents the loop region that is characteristic of natriuretic peptides. The peptide has 32 amino acids and its complete sequence is SYGISSGCFGLKLDRIGTMSGLGCWRLLQDSP. Coa_NP2 is a natriuretic peptide of the ANP/BNP-like family, since the carboxyterminal region of CNP has its own NP domain. We demonstrate, herein, that Coa_NP2 produces a dose-dependent decrease in mean arterial pressure in rats, followed by significant increases in concentrations of markers of nitric oxide formation measured in the plasma and vasorelaxation in a thoracic aortic ring bath. The structural and biological aspects confirm Coa_NP2 as a new natriuretic peptide, isolated from snake venom.

Exploring and understanding the functional role, and biochemical and structural characteristics of an acidic phospholipase A2, AplTx-I, purified from Agkistrodon piscivorus leucostoma snake venom

ABSTRACT Phospholipases A2 (PLA2s) constitute a class of extensively studied toxins, isolated from snake venoms. Basic PLA2 isoforms mediate various toxicological effects, while the acidic isoforms generally have higher enzymatic activities, but do not promote evident toxic effects. The functions of these acidic isoforms in snake venoms are still not completely understood and more studies are needed to characterize the biological functions and diversification of acidic toxins in order to justify their abundant presence in these secretions. Recently, Lomonte and collaborators demonstrated, in a proteomic and toxicological study, high concentrations of PLA2s in the venom of Agkistrodon piscivorus leucostoma. We have, herein, purified and characterized an acidic PLA2 from this snake venom, denominated AplTx‐I, in order to better understand its biochemical and structural characteristics, as well as its biological effects. AplTx‐I was purified using two chromatographic steps, in association with enzymatic and biological assays. The acidic toxin was found to be one of the most abundant proteins in the venom of A. p. leucostoma; the protein was monomeric with a molecular mass of 13,885.8 Da, as identified by mass spectrometry ESI‐TOF and electrophoresis. The toxin has similar primary and tridimensional structures to those of other acidic PLA2s, a theoretical and experimental isoelectric point of ≈5.12, and a calcium‐dependent enzyme activity of 25.8985 nM/min/mg, with maximum values at 37 °C and pH 8.0. Despite its high enzymatic activity on synthetic substrate, AplTx‐I did not induce high or significant myotoxic, coagulant, anticoagulant, edema, neuromuscular toxicity in mouse phrenic nerve‐diaphragm preparations or antibacterial activities. Interestingly, AplTx‐I triggered a high and selective neuromuscular toxicity in chick biventer cervicis preparations. These findings are relevant to provide a deeper understanding of the pharmacology, role and diversification of acidic phospholipase A2 isoforms in snake venoms. HIGHLIGHTSAplTx‐I is an acidic and monomeric phospholipase A2, isolated from Agkistrodon piscivorus leucostoma venom.AplTx‐I has high catalytic activity on synthetic and non‐micellar substrate.The amino acid sequence of AplTx‐I was identified by mass spectrometry.In vitro and in vivo, AplTx‐I presented low‐level, or lack of, toxicity to mammals and bacteria.AplTx‐I promoted an irreversible and selective neuromuscular blockade in biventer cervicis preparations.

Results of low power DRM+ field trial in the FM band II in Brasília

This work presents the results of DRM+ field tests in the VHF band II (FM band) using a low power FM station, which is known in Brazil as Community Radio. The purpose is the assessment of on-channel simulcast transmission, which will exist during the transition period to digital transmission (the so called “analog switch-off”). The results from trials performed in Brasília showed that the achievement of the same analog coverage area with digital system is conditioned to a ratio of 10 dB between analog and digital power levels, which can impairment in the current channelization.