Determination of phenolic compounds and antioxidant potential of aqueous extracts of Curcuma longa L., Piper nigrum L. and Cuminum cyminum: an experimental and a quantum-mechanical study / Determinação de compostos fenólicos e potencial antioxidante dos extratos aquosos de Curcuma longa L., Piper nigrum L. e Cuminum cyminum: um estudo experimental e quanto-mecânico

Objectives: Evaluation of phenolic compounds and antioxidant activities of aqueous extracts of C. longa, P. nigrum and C. cyminum. In addition to proposing a quantum-mechanical model to evaluate the antioxidant activity. Methods: The aqueous extracts were prepared using roots of the Curcuma longa L., seeds of the Piper nigrum L. and seeds of Cuminum cyminum. The extracts were subjected to tests to detect and quantify phenolic compounds and to assess their antioxidant capacity by different methods. Furthermore, to investigate the electronic nature of the antioxidant activity of the main compounds present in these extracts, frontier molecular orbitals (FMOs) were obtained by the DFT/B3LYP/6-31G(d,p) level of theory. Results: After statistical analysis of the results, a greater number of phenolic compounds and better antioxidant activity was identified in the aqueous extracts of cumin (C. cyminum) in all three assays performed, when compared to the other extracts tested. The theoretical model based on the Pietro method is in agreement with the experimental results. Conclusion: This study has an innovative proposal with the trivial antioxidant activity combined with theoretical quantum-mechanical calculations that can serve to reduce costs and time and to predict the antioxidant activity of subsequent studies.

Objetivos: avaliar os compostos fenólicos e atividades antioxidantes dos extratos aquosos de C. longa, P. nigrum e C. cyminum bem como propor um modelo quanto-mecânico para avaliar a atividade antioxidante. Métodos: os extratos aquosos foram preparados por meio da utilização de raízes de Curcuma longa L., sementes de Piper nigrum L. e sementes de Cuminum cyminum. Os extratos foram submetidos a ensaios para detectar e quantificar compostos fenólicos e atividade antioxidante por diferentes métodos. Além disso, com objetivo de investigar a natureza eletrônica da atividade antioxidante dos principais compostos presentes nesses extratos, orbitais moleculares de fronteira (OMFs) foram obtidos pelo nível de teoria DFT/B3LYP/6-31G(d,p). Resultados: após as análises estatísticas dos resultados, a maior quantidade de compostos fenólicos com maior atividade antioxidante foi identificada no extrato aquoso do cominho (C. cyminum) em todos os ensaios realizados, quando comparados com os outros extratos testados. O modelo teórico baseado no método de Pietro está concordante com os resultados experimentais. Conclusão: este estudo possui uma proposta inovadora com a atividade antioxidante trivial combinada com cálculos quanto-mecânicos que podem servir para reduzir custos e tempo para predizer a atividade antioxidante de estudos futuros.

Bioelectrical impedance vector analysis and phase angle in boys with Duchenne muscular dystrophy.

BACKGROUND: Duchenne muscular dystrophy (DMD) is a disease characterized by progressive loss of functional muscle mass followed by changes in body composition. OBJECTIVE: This study aimed to describe and evaluate bioimpedance parameters in boys with DMD. DESIGN: This cross-sectional, descriptive study investigated children and adolescents diagnosed with DMD. Age, weight, height, resistance, and reactance data were collected. Phase angle and bioelectrical impedance vector analysis were calculated based on resistance and reactance values. RESULTS: We analyzed 43 boys aged between 2.7 and 19.8 years. Low-phase angle values were observed during the investigation of bioimpedance parameters. Bioelectrical impedance vector analysis showed that approximately 87% of the subjects presented vectors outside the tolerance ellipses, and only one patient presented vectors located within the 50% tolerance ellipse, indicating normally hydrated and a good body cell mass. Compared with the reference population, boys with DMD had lower levels of body cell mass. CONCLUSION: Based on the evidence, compared with the reference population, patients with DMD had lower levels of body cell mass. This evidence points to bioimpedance parameters as useful tools for the nutritional evaluation and clinical management of patients with DMD.

Stigmurin and TsAP-2 from Tityus stigmurus scorpion venom: Assessment of structure and therapeutic potential in experimental sepsis.

Microbial resistance to conventional antibiotics is a public health problem worldwide, motivating the search for new therapeutic alternatives in varied natural sources. Cationic peptides without disulfide bridges from scorpions have been targeted in this context, mainly due to their multifunctional action and the limited ability of microorganisms to develop resistance against them. The present study was focused on Stigmurin and TsAP-2, cationic peptides found in the transcriptome of the venom gland from the scorpion Tityus stigmurus. The aims were: to assess the secondary structure of TsAP-2 and the structural stability of both peptides by circular dichroism; to evaluate their antiproliferative effect, and antimicrobial activities in vitro, ex vivo and in vivo; and to investigate their therapeutic potential in a murine model of polymicrobial sepsis. Stigmurin and TsAP-2 secondary structures responded similarly to environment polarity changes, and were stable to temperature and pH variation. Both peptides showed antiproliferative effect on tumor cells. TsAP-2 showed lower cytotoxicity to normal cells, and had a mitogenic activity on murine macrophages. Stigmurin demonstrated bactericidal and bacteriostatic activity, depending on the microorganism, whereas TsAP-2 had bactericidal action upon different bacterial strains analyzed. Both peptides were able to reduce leukocyte migration, TNF-α levels and microorganism load in the peritoneal cavity after induction of experimental sepsis, decreasing inflammation in the lung and cecum of septic animals. TsAP-2 also reduced the release of nitric oxide in the peritoneal cavity. Taken together, these data suggest that Stigmurin and TsAP-2 are structurally stable molecules and are efficient in the control of the infectious focus in polymicrobial sepsis, with potential use as a prototype for the rational design of novel therapeutic agents.

Characterization of TistH, a multifunctional peptide from the scorpion Tityus stigmurus: Structure, cytotoxicity and antimicrobial activity.

The presence of bioactive peptides in animal venoms has been targeted in scientific research for assessing biological activities, as well as mechanisms of action. A recent study by our group observed hypotensive action of TistH (Tityus stigmurus Hypotensin), a peptide deduced from the transcriptome of T. stigmurus venom gland. The present study aims to analyze TistH structure properties and to evaluate its toxicity on normal and tumor cells, its in vitro antimicrobial activity, as well as its inflammatory effect. Circular dichroism analyses of TistH showed a general predominance of α-helix conformation in TFE (20-70%) and structural stability to pH variations. TistH was not cytotoxic to normal cell lines (3T3, RAW and HEK), and also not to cancer cell lines (HeLa, B16, 786-0, SiHa and HepG2). The peptide did not present inflammatory activity up to 6 h after administered subcutaneously to Swiss mice. It was observed that concentrations of 4-1024 µg/mL of TistH produced no inhibition against the bacteria Staphylococcus aureus, Staphylococcus epidermidis e Pseudomonas aeruginosa. The results of antifungal assays showed a moderate activity of TistH against Candida albicans strain LM-108 and the filamentous fungus Trichophyton rubrum LM-640, with growth inhibition at a concentration of 1024 µg/mL. In contrast, the peptide presented a greater activity (MIC 128 µg/mL) against C. albicans LM-106, Candida tropicalis ATCC 13308 and Aspergillus flavus strains LM-247 and LM-26, fungi that cause oral and vaginal infections, candidiasis and respiratory allergies, respectively. The present data contribute to a better understanding of TistH and its possible use as a bioactive compound. This multifunctional peptide is capable of acting as anti-hypertensive, as well as to inhibit the growth of fungal strains, having low toxicity, which suggests its safety for using as a pharmacological agent.

Supplementation with a new trypsin inhibitor from peanut is associated with reduced fasting glucose, weight control, and increased plasma CCK secretion in an animal model.

Ingestion of peanuts may have a beneficial effect on weight control, possibly due to the satietogenic action of trypsin inhibitors. The aim of this study was to isolate a new trypsin inhibitor in a typical Brazilian peanut sweet (paçoca) and evaluate its effect in biochemical parameters, weight gain and food intake in male Wistar rats. The trypsin inhibitor in peanut paçoca (AHTI) was isolated. Experimental diets were prepared with AIN-93G supplemented with AHTI. Animals had their weight and food intake monitored. Animals were anesthetized, euthanized, and their bloods collected by cardiac puncture for dosage of cholecystokinin (CCK) and other biochemical parameters. Supplementation with AHTI significantly decreased fasting glucose, body weight gain, and food intake. These effects may be attributed to increased satiety, once supplemented animals showed no evidence of impaired nutritional status and also because AHTI increased CCK production. Thus, our results indicate that AHTI, besides reducing fasting glucose, can reduce weight gain via food intake reduction.

Homology modeling, vasorelaxant and bradykinin-potentiating activities of a novel hypotensin found in the scorpion venom from Tityus stigmurus.

In a recent work by our group involving a transcriptomics approach applied to the venom glands from Tityus stigmurus we identified a new family of peptides called Hypotensins (TSTI0006C) (Almeida et al., 2012). The cluster TSTI0006C was analyzed in the main 25 amino acid residues and named T. stigmurus Hypotensin (TistH), showing a molecular mass of 2.7 kDa, an absence of cysteines and the presence of two C-terminal proline residues, which are a bradykinin-potentiating peptide (BPP) signature. Here, we describe the homology modeling of the three-dimensional structure of TistH. In addition, we evaluated the cardiovascular effects elicited by TistH in normotensive rats. Firstly, TistH showed no cytotoxic effect on horse erythrocyte. Furthermore, in normotensive rats TistH was able to potentiate the hypotensive action of bradykinin (BK) and induced a vasorelaxant effect in mesenteric artery rings by endothelium-dependent release of nitric oxide (NO) and demonstrated independent inhibition of angiotensin converting enzyme (ACE). Our data can contribute to a better understanding of the structural and functional characteristics of TistH and suggest its potential use in cardiovascular diseases.

Characterization and pharmacological properties of a novel multifunctional Kunitz inhibitor from Erythrina velutina seeds.

Inhibitors of peptidases isolated from leguminous seeds have been studied for their pharmacological properties. The present study focused on purification, biochemical characterization and anti-inflammatory and anticoagulant evaluation of a novel Kunitz trypsin inhibitor from Erythrina velutina seeds (EvTI). Trypsin inhibitors were purified by ammonium sulfate (30-60%), fractionation followed by Trypsin-Sepharose affinity chromatography and reversed-phase high performance liquid chromatography. The purified inhibitor showed molecular mass of 19,210.48 Da. Furthermore, a second isoform with 19,228.16 Da was also observed. The inhibitor that showed highest trypsin specificity and enhanced recovery yield was named EvTI (P2) and was selected for further analysis. The EvTI peptide fragments, generated by trypsin and pepsin digestion, were further analyzed by MALDI-ToF-ToF mass spectrometry, allowing a partial primary structure elucidation. EvTI exhibited inhibitory activity against trypsin with IC50 of 2.2×10(-8) mol.L(-1) and constant inhibition (Ki) of 1.0×10(-8) mol.L(-1), by a non-competitive mechanism. In addition to inhibit the activity of trypsin, EvTI also inhibited factor Xa and neutrophil elastase, but do not inhibit thrombin, chymotrypsin or peptidase 3. EvTI was investigated for its anti-inflammatory and anti-coagulant properties. Firstly, EvTI showed no cytotoxic effect on human peripheral blood cells. Nevertheless, the inhibitor was able to prolong the clotting time in a dose-dependent manner by using in vitro and in vivo models. Due to anti-inflammatory and anticoagulant EvTI properties, two sepsis models were here challenged. EvTI inhibited leukocyte migration and specifically acted by inhibiting TNF-α release and stimulating IFN-α and IL-12 synthesis. The data presented clearly contribute to a better understanding of the use of Kunitz inhibitors in sepsis as a bioactive agent capable of interfering in blood coagulation and inflammation.