Preparation and Characterization of a Rifampicin Coamorphous Material with Tromethamine Coformer: An Experimental-Theoretical Study.

Rifampicin (RIF) is an antibiotic used to treat tuberculosis and leprosy. Even though RIF is a market-available drug, it has a low aqueous solubility, hindering its bioavailability. Among the strategies for bioavailability improvement of poorly soluble drugs, coamorphous systems have been revealed as an alternative in the increase of the aqueous solubility of drug systems and at the same time also increasing the amorphous state stability and dissolution rate when compared with the neat drug. In this work, a new coamorphous form from RIF and tromethamine (TRIS) was synthesized by slow evaporation. Structural, electronic, and thermodynamic properties and solvation effects, as well as drug-coformer intermolecular interactions, were studied through density functional theory (DFT) calculations. Powder X-ray diffraction (PXRD) data allowed us to verify the formation of a new coamorphous. In addition, the DFT study indicates a possible intermolecular interaction by hydrogen bonds between the available amino and carbonyl groups of RIF and the hydroxyl and amino groups of TRIS. The theoretical spectra obtained are in good agreement with the experimental data, suggesting the main interactions occurring in the formation of the coamorphous system. PXRD was used to study the physical stability of the coamorphous system under accelerated ICH conditions (40 °C and 75% RH), indicating that the material remained in an amorphous state up to 180 days. The thermogravimetry result of this material showed a good thermal stability up to 153 °C, and differential scanning calorimetry showed that the glass temperature (Tg) was at 70.0 °C. Solubility studies demonstrated an increase in the solubility of RIF by 5.5-fold when compared with its crystalline counterpart. Therefore, this new material presents critical parameters that can be considered in the development of new coamorphous formulations.

Optimization and validation of LLE-LTP to determine florpyrauxifen-benzyl herbicide in water samples by HPLC-DAD.

Florpyrauxifen-benzyl is a systemic herbicide which acts on weeds of common occurrence in rice cultivation areas using flooded or rainfed systems. The Brazilian Health Regulatory Agency (ANVISA) authorized the commercialization of this pesticide, but did not establish guidelines with an extraction and quantification method of residues of this compound as a form of environmental monitoring. Therefore, the present study aimed to optimize and validate the liquid-liquid extraction with low temperature purification (LLE-LTP) to determine florpyrauxifen-benzyl content in water samples by high performance liquid chromatography with diode array detection (HPLC-DAD). The recovery ranged from 95.84 to 105.4% with a relative standard deviation less than 1.5. LLE-LTP was selective, precise, accurate, linear in the range from 4.00 to 150 µg L-1, and the limit of quantification was 4.00 µg L-1. The stability study of the compound in water revealed its degradation in 25 days and DT50 in approximately 5 days. LLE-LTP coupled to HPLC-DAD presented itself as a simple, easy and efficient method of extracting and analyzing it in water samples.

l-Glutamine supplementation enhances glutathione peroxidase and paraoxonase-1 activities in HDL of exercising older individuals.

BACKGROUND: Oxidative stress is an important factor in the formation of atherosclerotic plaques. High-density lipoprotein (HDL) harbors paraoxonase-1 (PON-1) and glutathione peroxidase (GPx), key enzymes in the protection against the harmful effects of oxidative stress. Although exercise training can increase both HDL-c content and its antioxidant action, and glutamine (Gln) intake also promotes GPx-based defenses, the association between exercise training and Gln in the regulation of PON-1 activity was not explored. Therefore, the objective of this study was to investigate the effects of Gln supplementation on the redox balance and on the total HDL antioxidant capacity by evaluation of the activity of PON-1 and GPx enzymes in physically exercised elderly individuals compared to non-exercised ones. METHODS: Fifty-one practitioners of a combined exercise training program (CET, age: 71.9 ± 5.7 years) and 32 non-practitioners (NP, age: 73 ± 6.3 years) participated in the study. CET and NP groups were separated into 2 subgroups according to the supplementation: Gln, 0.3 g/kg/day + 10 g maltodextrin (CET-Gln, n = 26; and NP-Gln, n = 16) or placebo, 10 g maltodextrin (CET-PL, n = 25; and NP-PL, n = 16). Blood samples were drawn at baseline and after 30 days after commencement of the supplementation for biochemical and enzyme activity analyses. RESULTS: Increased HDL-c, total peroxidase (PRx), and GPx activities were found in both CET-Gln and NP-Gln after the supplementation period, compared to baseline, in opposition to CET-PL and NP-PL groups. PON-1 activity increased only in CET-Gln. In both CET-Gln and NP-Gln groups, there was a reduction of the total peroxides/PRx, iron/PRx, and total peroxides/GPX ratios after supplementation. In CET-Gln, thiobarbituric acid-reactive substances (TBARS)/PRx and TBARS/GPx ratios were also lower after supplementation. CET-Gln and CET-PL subgroups had lower glycemia than NP-Gln and NP-PL, either at baseline or after the supplementation periods. The other parameters were unchanged after supplementation [total cholesterol, LDL-c, triglycerides, non-HDL cholesterol, total peroxides, TBARS, iron serum, Trolox-equivalent antioxidant capacity (TEAC), and uric acid]. CONCLUSIONS: Gln supplementation can increase glutathione peroxidase activity regardless the individuals were physically active or sedentary, but the PON-1 activity only increased in physically active individuals. These results show the potential of Gln supplementation in the maintenance of the vascular redox balance, with potential implications for atherogenesis protection.

Constitutive nitric oxide synthase-like enzyme in two species involved in cutaneous and mucocutaneous leishmaniasis.

Leishmania is an obligate intracellular parasite that primarily inhabits macrophages. The destruction of the parasite in the host cell is a fundamental mechanism for infection control. In addition, inhibition of the leishmanicidal activity of macrophages seems to be related to the ability of some species to inhibit the production of nitric oxide (NO) by depleting arginine. Some species of Leishmania have the ability to produce NO from a constitutive nitric oxide synthase-like enzyme (cNOS-like). However, the localization of cNOS-like in Leishmania has not been described before. As such, this study was designed to locate cNOS-like enzyme and NO production in promastigotes of Leishmania (Leishmania) amazonensis and Leishmania (Viannia) braziliensis. NO production was initially quantified by flow cytometry, which indicated a significant difference in NO production between L. (L.) amazonensis (GMFC = 92.17 +/- 4.6) and L. (V.) braziliensis (GMFC = 18.89 +/- 2.29) (P < 0.05). Analysis of cNOS expression by immunoblotting showed more expression in L. (L.) amazonensis versus L. (V.) braziliensis. Subsequently, cNOS-like immunolabeling was observed in promastigotes in regions near vesicles, the flagellar pocket and mitochondria, and small clusters of particles appeared to be fusing with vesicles suggestive of glycosomes, peroxisome-like-organelles that compartmentalize the glycolytic pathway in trypanosomatid parasites. In addition, confocal microscopy analysis demonstrated colocalization of cNOS-like and GAPDH, a specific marker for glycosomes. Thus, L. (L.) amazonensis produces greater amounts of NO than L. (V.) braziliensis, and both species present the cNOS-like enzyme inside glycosomes.

Recent computational methods for white blood cell nuclei segmentation: A comparative study.

BACKGROUND AND OBJECTIVE: Leukaemia is a disease found worldwide; it is a type of cancer that originates in the bone marrow and is characterised by an abnormal proliferation of white blood cells (leukocytes). In order to correctly identify this abnormality, haematologists examine blood smears from patients. A diagnosis obtained by this method may be influenced by factors such as the experience and level of fatigue of the haematologist, resulting in non-standard reports and even errors. In the literature, several methods have been proposed that involve algorithms to diagnose this disease. However, no reviews or surveys have been conducted. This paper therefore presents an empirical investigation of computational methods focusing on the segmentation of leukocytes. METHODS: In our study, 15 segmentation methods were evaluated using five public image databases: ALL-IDB2, BloodSeg, Leukocytes, JTSC Database and CellaVision. Following the standard methodology for literature evaluation, we conducted a pixel-level segmentation evaluation by comparing the segmented image with its corresponding ground truth. In order to identify the strengths and weaknesses of these methods, we performed an evaluation using six evaluation metrics: accuracy, specificity, precision, recall, kappa, Dice, and true positive rate. RESULTS: The segmentation algorithms performed significantly differently for different image databases, and for each database, a different algorithm achieved the best results. Moreover, the two best methods achieved average accuracy values higher than 97%, with an excellent kappa index. Also, the average Dice index indicated that the similarity between the segmented leukocyte and its ground truth was higher than 0.85 for these two methods This result confirms the high level of similarity between these images but does not guarantee that a method has segmented all leukocyte nuclei. We also found that the method that performed best segmented only 58.44% of all leukocytes. CONCLUSIONS: Of the techniques used to segment leukocytes, we note that clustering algorithms, the Otsu threshold, simple arithmetic operations and region growing are the approaches most widely used for this purpose. However, these computational methods have not yet overcome all the challenges posed by this problem.

Buffalo Cheese Whey Proteins, Identification of a 24 kDa Protein and Characterization of Their Hydrolysates: In Vitro Gastrointestinal Digestion.

Milk whey proteins are well known for their high biological value and versatile functional properties, characteristics that allow its wide use in the food and pharmaceutical industries. In this work, a 24 kDa protein from buffalo cheese whey was analyzed by mass spectrometry and presented homology with Bos taurus beta-lactoglobulin. In addition, the proteins present in buffalo cheese whey were hydrolyzed with pepsin and with different combinations of trypsin, chymotrypsin and carboxypeptidase-A. When the TNBS method was used the obtained hydrolysates presented DH of 55 and 62% for H1 and H2, respectively. Otherwise for the OPA method the DH was 27 and 43% for H1 and H2, respectively. The total antioxidant activities of the H1 and H2 samples with and without previous enzymatic hydrolysis, determined by DPPH using diphenyl-p-picrylhydrazyl radical, was 4.9 and 12 mM of Trolox equivalents (TE) for H2 and H2Dint, respectively. The increased concentrations for H1 and H2 samples were approximately 99% and 75%, respectively. The in vitro gastrointestinal digestion efficiency for the samples that were first hydrolyzed was higher compared with samples not submitted to previous hydrolysis. After in vitro gastrointestinal digestion, several amino acids were released in higher concentrations, and most of which were essential amino acids. These results suggest that buffalo cheese whey is a better source of bioavailable amino acids than bovine cheese whey.

Optimization of ß-glucosidase, ß-xylosidase and xylanase production by Colletotrichum graminicola under solid-state fermentation and application in raw sugarcane trash saccharification.

Efficient, low-cost enzymatic hydrolysis of lignocellulosic residues is essential for cost-effective production of bioethanol. The production of ß-glucosidase, ß-xylosidase and xylanase by Colletotrichum graminicola was optimized using Response Surface Methodology (RSM). Maximal production occurred in wheat bran. Sugarcane trash, peanut hulls and corncob enhanced ß-glucosidase, ß-xylosidase and xylanase production, respectively. Maximal levels after optimization reached 159.3 ± 12.7 U g-1, 128.1 ± 6.4 U g-1 and 378.1 ± 23.3 U g-1, respectively, but the enzymes were produced simultaneously at good levels under culture conditions optimized for each one of them. Optima of pH and temperature were 5.0 and 65 °C for the three enzymes, which maintained full activity for 72 h at 50 °C and for 120 min at 60 °C (ß-glucosidase) or 65 °C (ß-xylosidase and xylanase). Mixed with Trichoderma reesei cellulases, C. graminicola crude extract hydrolyzed raw sugarcane trash with glucose yield of 33.1% after 48 h, demonstrating good potential to compose efficient cocktails for lignocellulosic materials hydrolysis.

Population structure of the malaria vector Anopheles darlingi in Rondônia, Brazilian Amazon, based on mitochondrial DNA

Anopheles darlingi is the most important Brazilian malaria vector, with a widespread distribution in the Amazon forest. Effective strategies for vector control could be better developed through knowledge of its genetic structure and gene flow among populations, to assess the vector diversity and competence in transmitting Plasmodium. The aim of this study was to assess the genetic diversity of An. darlingi collected at four locations in Porto Velho, by sequencing a fragment of the ND4 mitochondrial gene. From 218 individual mosquitoes, we obtained 20 different haplotypes with a diversity index of 0.756, equivalent to that found in other neotropical anophelines. The analysis did not demonstrate significant population structure. However, haplotype diversity within some populations seems to be over-represented, suggesting the presence of sub-populations, but the presence of highly represented haplotypes complicates this analysis. There was no clear correlation among genetic and geographical distance and there were differences in relation to seasonality, which is important for malarial epidemiology.

Population structure of the malaria vector Anopheles darlingi in Rondônia, Brazilian Amazon, based on mitochondrial DNA.

Anopheles darlingi is the most important Brazilian malaria vector, with a widespread distribution in the Amazon forest. Effective strategies for vector control could be better developed through knowledge of its genetic structure and gene flow among populations, to assess the vector diversity and competence in transmitting Plasmodium. The aim of this study was to assess the genetic diversity of An. darlingi collected at four locations in Porto Velho, by sequencing a fragment of the ND4 mitochondrial gene. From 218 individual mosquitoes, we obtained 20 different haplotypes with a diversity index of 0.756, equivalent to that found in other neotropical anophelines. The analysis did not demonstrate significant population structure. However, haplotype diversity within some populations seems to be over-represented, suggesting the presence of sub-populations, but the presence of highly represented haplotypes complicates this analysis. There was no clear correlation among genetic and geographical distance and there were differences in relation to seasonality, which is important for malarial epidemiology.

Recent observations on the sand fly (Diptera: Psychodidae) fauna of the State of Rondônia, Western Amazônia, Brazil: the importance of Psychdopygus davisi as a vector of zoonotic cutaneous leishmaniasis.

Sand flies were collected in the central region of the state of Rondônia (W 64 degrees 30' to 63 degrees 00' and S 10 degrees 00'to 11 degrees 00') using Shannon and CDC light traps from October 1997 to August 2000. A total of 85,850 specimens representing 78 named species were captured. Of these 14 were new records for Rondônia. The proportion of males/females was 1/1.131. Trypanosomatids, that are presently being identified, were detected in 11 species. Leishmania (Viannia) naiffi was recorded from Psychodopygus davisi and P. hirsutus. In the present study the dominant species was P. davisi (39.6%) followed by Lutzomyia whitmani (13.1%), P. carrerai (11.6%), and P. hirsutus (10.2%). The importance of P. davisi as a vector of zoonotic cutaneous leishmaniasis is discussed.

Recent observations on the sand fly (Diptera: Psychodidae) fauna of the State of Rondônia, Western Amazônia, Brazil: the importance of Psychdopygus davisi as a vector of zoonotic cutaneous leishmaniasis

Sand flies were collected in the central region of the state of Rondônia (W 64 30' to 63 00' and S 10 00'to 11 00') using Shannon and CDC light traps from October 1997 to August 2000. A total of 85,850 specimens representing 78 named species were captured. Of these 14 were new records for Rondônia. The proportion of males/females was 1/1.131. Trypanosomatids, that are presently being identified, were detected in 11 species. Leishmania (Viannia) naiffi was recorded from Psychodopygus davisi and P. hirsutus. In the present study the dominant species was P. davisi (39.6 percent) followed by Lutzomyia whitmani (13.1 percent), P. carrerai (11.6 percent), and P. hirsutus (10.2 percent). The importance of P. davisi as a vector of zoonotic cutaneous leishmaniasis is discussed

Grau de conhecimento sobre diabetes mellitus: um estudo populacional / Knowledge about diabetes Mellitus: a populational study

O conhecimento que o indivíduo com diabetes mellitus tenha sobre a própria doença é importante tanto para seu manejo como para a adoçäo de medidas preventivas. Com o objetivo de avaliar conhecimentos básicos, foi realizado um estudo de base populacional, com 117 indivíduos com diabetes - 75 com diagnóstico prévio e 42 em que o diabetes foi detectado durante o estudo. Através da aplicaçäo de um questionário padronizado observou-se que o grau de conhecimento sobre diabetes nos dois grupos foi semelhante (56,2% de acertos no grupo com diagnóstico prévio e 47,4% no grupo detectado durante o estudo; p-0,38), havendo diferenças significativas apenas nas questöes sobre faixa glicêmica aceitável para controle, atitude frente a hipoglicemia e cuidados para prevençäo da gangrena. No entanto, o grau de conhecimento dos dois grupos é insatisfatório, sugerindo que os serviçÝs de saúde enfatizam pouco os aspectos educacionais no atendimento ao diabetes