921 resultados para denaturing high-performance liquid chromatography (DHPLC) sequencing


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This study conducted chemical analyzes of the fruits of Licania tomentosa Benth belonging to the Chrysobalanaceae family, order and superorder Rosiflorae Rosales. Samples were collected in February 2011 in the city of Natal - RN. The fruits were separated into peel, pulp and seed, and the samples were analyzed on moisture and ash by thermogravimetric methods, reducing and total sugars, lipids by Soxhlet extraction, minerals by ICP / OES and other analyzes by physical measurements of the fruit. The fruits for the analysis of determination of flavonoids and phenolic compounds were dried at 40 ° C and ground. For the analyzes to determine the content of phenolics and flavonols was used spectroscopy in the visible region, and also performed the characterization of ethanol extracts by high performance liquid chromatography and test for antioxidant activity hydroethanolic extracts. The results obtained are about 83.9% of the fruits have a length between 4.0 to 6.0 cm with respect to the diameter of the fruit about 97.4% of the fruits have a diameter between 2.0 - 4, 0 cm, mean weight of 36.14 g (7.86), the bark showed 60.5 (1.5)% relative humidity, the flesh 72.3 (0.9) and 44.4 seeds (0.4 )%, the bark showed 0.22 (0.00)% ash, the pulp 1.41 (0.01)% and the seed 1.18 (0.01)%, the bark showed 3.73 (0, 09) mg lipids / 100 g sample of the pulp 0.40 (0.04) and the seed 0.05 (0.00). With respect to reducing sugars and total gave the results, 16.23 (0.23) g/100g and 20.70 (0.24) g/100g. The fruit is presented with a good source of nutrients especially carbohydrates and lipids. The analysis for determining the antioxidant activity tests were performed reducing power and antioxidant capacity, the antioxidant capacity of hydroethanolic extracts of peel and pulp showed similar results, with higher concentrations however hydroethanolic bark extract. Regarding the content of phenolics and flavonoids hidroetanílico bark extract showed higher concentration values. The characterization of the extracts by high performance liquid chromatography identified the following compounds by retention time indicated by the race of the standards: galic acid, kaempferol and kaempferol derivatives. Considering the results of the fruit Licania tomentosa Benth may be an alternative food by analysis of their nutritional content and has substances with antioxidant action

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Introduction .The renal prostaglandins (PGs), vasodilators, preserve kidney function during increased activity of the renin-angiotensin system or renal sympathetic nerves (renal PG-dependent state [RPGD]). Ketoprofen (Ket) inhibits cyclooxygenase and, therefore, the synthesis of PGs. The aim of this study was to determine, in the rat, the action of Ket in the renal histology and function in a RPGD state (stress of anesthesia and hemorrhage). Material and Methods . Twenty male Wistar rats, anesthetized with sodium pentobarbital, were randomly divided into two groups: G1-control ( n = 10) and G2-Ket ( n = 10) submitted to arterial hemorrhage of 30% of volemia (estimated as 6% of body weight) three times (10% each 10 min), 65 min after anesthesia. G2 animals received Ket, 1.5 mg. kg -1 , venously, 5 min after anesthesia and 60 min before the first hemorrhage moment (first moment of the study [M1]). Medium arterial pressure (MAP), rectal temperature (T), and heart rate were monitored. G1 and G2 received para-aminohippurate sodium (PAH) and iothalamate sodium (IOT) solutions during the entire experimental time in order to determine clearance of PAH (effective renal plasma flow [ERPF]) and clearance of IOT (glomerular filtration rate [GFR]) without urine collection (determination of blood concentrations of PAH and IOT through the high-performance liquid chromatography), filtration fraction (FF), and renal vascular resistance (RVR). The animals were sacrificed in M3, 30 min after the third hemorrhage (M2) moment, and the kidneys and blood collected during the hemorrhage periods were utilized for histological study and determinations of hematocrit (Ht), serum creatinine (S Cr ), ERPF, GFR, FF, and RVR, respectively. Results . There were significant reductions of MAP, T, and Ht and a significant increase of S Cr . During the experiment, ERPF and GFR did not change, but ERPF was always higher in G1 than in G2. Ket did not alter FF, which increased in G1 over the duration of experiment. The Ket group had significantly higher RVR than the control group. The histology verified that both G1 and G2 were similar for tubular dilation and necrosis, but they were significantly different for tubular degeneration: G1 > G2. Conclusion . The changes observed in kidney histology probably were determined by hemorrhage and hypotension. Ket inhibited the synthesis of PGs and diminished tubular degeneration.

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Objective: To compare the efficiency of an Aeroneb Pro vibrating plate and an Atomisor MegaHertz ultrasonic nebulizer for providing ceftazidime distal lung deposition.Design: In vitro experiments. One gram of cetazidime was nebulized in respiratory circuits and mass median aerodynamic diameter of particles generated by ultrasonic and vibrating plate nebulizers was compared using a laser velocimeter. In vivo experiments. Lung tissue concentrations and extrapulmonary depositions were measured in ten anesthetized ventilated piglets with healthy lungs that received 1 g of ceftazidime by nebulization with either an ultrasonic (n = 5), or a vibrating plate (n = 5) nebulizer.Setting: A two-bed Experimental Intensive Care Unit of a University School of Medicine.Intervention: Following sacrifice, 5 subpleural specimens were sampled in dependent and nondependent lung regions for measuring ceftazidime lung tissue concentrations by high-performance liquid chromatography.Measurements and results: Mass median aerodynamic diameters generated by both nebulizers were similar with more than 95% of the particles between 0.5 and 5 mu m. Lung tissue concentrations were 553 +/- 123 [95% confidence interval: 514-638] mu g g(-1) using ultrasonic nebulizer, and 452 +/- 172 [95% confidence interval: 376-528] mu g g(-1) using vibrating plate nebulizers (NS). Extrapulmonary depositions were, respectively, of 38 +/- 5% (ultrasonic) and 34 +/- 4% (vibrating plate) (NS).Conclusions: Vibrating plate nebulizer is comparable to ultrasonic nebulizers for ceftazidime nebulization. It may represent a new attractive technology for inhaled antibiotic therapy.

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Alpha thalassemia, the most common monogenic disorder in the world, is characterized by deletions of one (+-thalassemia) or both alpha genes (0-thalassemia) located on human chromosome 16 (16p13.3). The most common case of +-thalassemia is a deletion of 3.7 kb of DNA (-3.7 deletion). It is most prevalent in African and Middle East regions. In the few studies carried out in Brazilian population -3.7 deletion was the most common deletion, mainly in African descendants. This study was conducted to determine the prevalence of +- thalassemia (deletion 3.7kb) in adult population from Rio Grande do Norte. We obtained blood samples from 713 unrelated individuals of both genders, aged between 18 and 59 years old. All individuals were born in Rio Grande do Norte. The hematological indices were obtained in an automatic cell counter (Micros 60, ABX Diagnostics). The hemoglobin measurement (A2 and Fetal hemoglobin) and the profile confirmation were carried out by high performance liquid chromatography (HPLC) methodology. Genomic DNA was obtained from peripheral blood leukocytes using Illustra Blood GenomicPrep Mini Spin kit and -3.7 deletion was investigated by PCR. Among the 713 individuals studied, 80 (11,2%) presented +- thalassemia: 79 (11,1%) were heterozygous and 1 (0,1%) homozygous for the -3.7 deletion. Considering the ethnic group, negroes showed the greatest prevalence of +-thalassemia (12,5%), followed by mulattoes (12,3%) and caucasian (9,6%). Statistical comparison of hematological parameters between normal individuals and heterozygous to +-thalassemia showed significant differences in RBC (p<0,001), MCV (p<0,001), MCH (p<0,001), Hb A2 (p=0,007) as well as female hemoglobin concentration (p=0,003). This is one of the first studies to research +-thalassemia in general population of Rio Grande do Norte state and these results attest the importance of investigation of this condition to define the etiology of microcytosis and hypochromia.

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The pharmacokinetics of enrofloxacin (ENRO), a fluoroquinolone antimicrobial agent, was studied in male broiler chickens (Cobb) after single oral administration of 10 mg of ENRO/kg b.w. A high-performance liquid chromatography-photodiode array detector (DAD) (HPLC-DAD) method was developed and validated and used for quantitation of ENRO and its major metabolite ciprofloxacin in plasma. The HPLC analyses were carried out using a cationic-octadecyl mixed column and 0.05 mol/L phosphate buffer (pH 2.5)/acetonitrile as mobile phase. The sample preparation of plasma consisted of the precipitation of proteins followed by solid phase extraction on cationic-octadecyl mixed cartridges. The method was validated considering linear range, linearity, selectivity, sensitivity, limit of detection (LOD), limit of quantitation (LOQ), intra- and inter-day precisions and accuracy. The LOD and LOQ for both fluoroquinolones were 60 and 200 ng/mL for plasma. The plasma concentration vs. time graph was characteristic of a two-compartment open model. The maximal plasma concentration of 1.5 +/- 0.2 mg/mL was achieved at 9 +/- 2 h. The elimination half-life and the mean residence time of ENRO were 1.5 +/- 0.2 and 15.64 h, respectively. The area under the concentration-time curve was calculated as 35 +/- 4 mg(.)h/mL.

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Equine antivenom is considered the only treatment for animal-generated envenomations, but it is costly. The study aimed to produce Apis mellifera (Africanized honeybee) and Crotalus durissus terrificus (C.d.t.) antivenoms using nanostructured silica (SBA-15) as adjuvant and cobalt-60 (60Co)-detoxified venoms utilizing young sheep. Natural and 60Co-irradiated venoms were employed in four different hyperimmunization protocols. Thus, 8 groups of 60- to 90-d-old sheep were hyperimmunized, enzyme-linked immunosorbent assay (ELISA) serum titers collected every 14 d were assessed clinically daily, and individual weight were measured, until d 84. Incomplete Freund's (IFA) and nanostructured silica (SBA15) adjuvants were compared. The lethal dose (LD50) for both venoms was determined following intraperitoneal (ip) administration to mice. High-performance liquid chromatography on reversed phase (HPLC-RP) was used also to measure the 60Co irradiation effects on Apis venom. At the end of the study, sheep were killed in a slaughterhouse. Kidneys were histologically analyzed. LD50 was 5.97 mg/kg Apis and 0.07 mg/kg C.d.t. for native compared to 13.44 mg/kg Apis and 0.35 mg/kg C.d.t. for irradiated venoms. HPLC revealed significant differences in chromatographic profiles between native and irradiated Apis venoms. Native venom plus IFA compared with SBA-15 showed significantly higher antibody titers for both venoms. Apis-irradiated venom plus IFA or SBA-15 displayed similar antibody titers but were significantly lower when compared with native venom plus IFA. Weight gain did not differ significantly among all groups. 60Co irradiation decreased toxicity and maintained venom immunogenic capacity, while IFA produced higher antibody titers. SBA-15 was able to act as an adjuvant without producing adverse effects. Hyperimmunization did not affect sheep weight gain, which would considerably reduce the cost of antiserum production, as these sheep were still approved for human consumption even after being subjected to hyperimmunization.

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(1) Venom pools from Bothrops neuwiedi (Bn) and from two subspecies, namely Bothrops neuwiedi pauloensis (Bnp) and Bothrops neuwiedi urutu (Bnu), collected in the States of São Paulo (SP) and Minas Gerais (MG), Brazil, were electrophoretically examined. Basic toxins with different isoelectric points were identified in the venom collected in São Paulo (BnSP). These toxins were absent in the corresponding pools from Minas Gerais (BnMG, BnpMG and BnuMG). (2) BnSP, but not BnMG, BnpMG or BnuMG, showed two myotoxins (pI congruent to 8.6 and 8.8, respectively) which were isolated by ion-exchange chromatography on CM-Sepharose. (3) From BnMG, three myotoxic isoforms (pI congruent to 8.2 and M-r = 13600) were isolated by chromatography on CM-Sepharose followed by reversed-phase high-performance liquid chromatography. (4) the chemical and biological characterization of these toxins showed a high similarity with the Lys-49 myotoxins from other bothropic venoms. (5) Doses up to 5 LD50 (i.p.) of p-bromophenacyl bromide alkylated BnSP-7 caused a total loss of lethality in 18-22-g mice, thus indicating that the LD50 was increased by greater than 5-fold. At this dose myotoxicity was also not detectable, but the edematogenic activity on the rat paw apparently did not change. (C) 1998 Elsevier B.V. All rights reserved.

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O presente estudo consiste em uma caracterização preliminar da atividade proteolítica de frações de proteínas purificadas a partir de lisados de trofozoítos de cepa isolada e axenizada no Brasil. Frações obtidas por cromatografia líquida (FPLC) foram analisadas quanto ao perfil eletroforético em géis de poliacrilamida (SDS-PAGE) e a atividade proteolítica foi avaliada em géis contendo gelatina como substrato. A caracterização das enzimas foi realizada a partir da análise do efeito de inibidores sintéticos de cisteína-proteases (E-64, IAA), serina-proteases (PMSF), serina e cisteína-proteases (TPCK, TLCK, elastatinal), metalo-proteases (EDTA) e aspartil proteases (pepstatina) sobre a degradação do substrato. Entre 30 frações eluídas, bandas de proteínas foram observadas em oito delas, entretanto, atividade proteolítica foi detectada apenas nas frações 23, 24, 25 e 26. O perfil eletroforético das proteínas revelou poucas bandas distribuídas na faixa de 45 a 18 kDa. Os zimogramas revelaram zonas de proteólise na faixa de aproximadamente 62 a 35 kDa, entretanto destacaram-se as bandas de hidrólise de 62, 55, 53, 50, 46 e 40 kDa. Nos ensaios de inibição, a proteólise foi marcantemente inibida por E-64, TPCK, TLCK e elastatinal. Redução discreta da proteólise foi observada com IAA e PMSF, enquanto que EDTA e pepstatina não promoveram alteração dos perfis de hidrólise. Estas observações são relevantes, especialmente se considerarmos que para elucidar o envolvimento das proteases na relação parasita-hospedeiro, a purificação dessas moléculas é um requisito importante.

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