Determination of anticonvulsants in human plasma using SPME in a heated interface coupled online to liquid chromatography (SPME-LC)

A simple and sensitive method using solid phase microextraction (SPME) and liquid chromatography (LC) with heated online desorption (SPME-LC) was developed and validated to analyze anticonvulsants (AEDs) in human plasma samples. A heated lab-made interface chamber was used in the desorption procedure, which allowed the transference of the whole extracted sample. The SPME conditions were optimized by applying an experimental design. Important factors are discussed such as fiber coating types, pH, extraction time and desorption conditions. The drugs were analyzed by LC, using a C18 column (150 mm 4.6 mm 5 mm); and 50 mmol L1 , pH ¼ 5.50 ammonium acetate buffer : acetonitrile : methanol (55 : 22 : 23 v/v) as the mobile phase with a flow rate of 0.8 mL min1 . The suggested method presented precision (intra-assay and inter-assay), linearity and limit of quantification (LOQ) all adequate for the therapeutic drug monitoring (TDM) of AEDs in plasma.

Analysis of fluoxetine and norfluoxetine in human plasma by HPLC-UV using a high purity C18 silica-based SPE sorbent

This paper reports on the development and validation of a simple and sensitive method that uses solid phase extraction (SPE) and liquid chromatography with ultraviolet detection to analyze fluoxetine (FLX) and norfluoxetine (NFLX) in human plasma samples. A lab-made C18 SPE phase was synthesized by using a sol–gel process employing a low-cost silica precursor. This sorbent was fully characterized by nuclear magnetic resonance (NMR), Fourier-transform infrared spectroscopy (FT-IR), and scanning electron microscopy (SEM) to check the particles' shape, size and C18 functionalization. The lab-made C18 silica was used in the sample preparation step of human plasma by the SPE-HPLC-UV method. The method was validated in the 15 to 500 ng mL 1 range for both FLX and NFLX using a matrix matched curve. Detection limits of 4.3 and 4.2 ng mL 1 were obtained for FLX and NFLX, respectively. The repeatability and intermediary precision achieved varied from 7.6 to 15.0% and the accuracy ranged from 14.9 to 9.1%. The synthesized C18 sorbent was compared to commercial C18 sorbents. The average recoveries were similar (85–105%), however the lab-made C18 silica showed fewer interfering peaks in the chromatogram. After development and validation, the method using the lab-made C18 SPE was applied to plasma samples of patients under FLX treatment (n ¼ 6). The concentrations of FLX and NFLX found in the samples varied from 46.8–215.5 and 48.0–189.9 ng mL 1 , respectively.

New LLLT protocol to speed up the bone healing process-histometric and immunohistochemical analysis in rat calvarial bone defect

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Curcumin pharmacokinetic and pharmacodynamic evidences in streptozotocin-diabetic rats support the antidiabetic activity to be via metabolite(s)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Myosin heavy chain expression and atrophy in rat skeletal muscle during transition from cardiac hypertrophy to heart failure

The purpose of this investigation was to determine whether changes in myosin heavy chain (MHC) expression and atrophy in rat skeletal muscle are observed during transition from cardiac hypertrophy to chronic heart failure (CHF) induced by aortic stenosis (AS). AS and control animals were studied 12 and 18 weeks after surgery and when overt CHF had developed in AS animals, 28 weeks after the surgery. The following parameters were studied in the soleus muscle: muscle atrophy index (soleus weight/body weight), muscle fibre diameter and frequency and MHC expression. AS animals presented decreases in both MHC1 and type I fibres and increases in both MHC2a and type IIa fibres during late cardiac hypertrophy and CHF. Type IIa fibre atrophy occurred during CHF. In conclusion, our data demonstrate that skeletal muscle phenotype changes occur in both late cardiac hypertrophy and heart failure; this suggests that attention should be given to the fact that skeletal muscle phenotype changes occur prior to overt heart failure symptoms.

Norepinephrine responses in rat renal and femoral veins are reinforced by vasoconstrictor prostanoids

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Periapical lesions decrease insulin signaling in rat skeletal muscle

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

TNF-alpha and IL-6 immunohistochemistry in rat renal tissue experimentaly infected with Leptospira interrogans serovar Canicola

Leptospirosis is a public health problem worldwide and its etiology remains unclear. Its pathogenesis involves a complex interaction between host and infecting microorganism. The inflammatory reaction that controls the infection process also underscores many pathophysiological events occurring in leptospirosis. We investigated the presence of tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) in renal tissues by immunohistochemical and histopathological examination in animals experimentally inoculated with Leptospira serovar Canicola. All the tests were carried out 2, 7, 14, 21 or 28 days after inoculation. Although TNF-alpha and IL-6 had been detected in tissues throughout the observation period, these cytokines appeared more intensely during the initial phase of infection. Therefore, both TNF-alpha and IL-6 were associated with the immunopathogenesis of leptospirosis. This profile suggests a high immunocellular response throughout the early infection stages followed by subsequent humoral response.

Development and validation of an LC-MS/MS method for quantitative analysis of mirtazapine in human plasma

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The anti-ulcerogenic effects of Curatella americana L.

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Plasma concentrations and placental immunostaining of interleukin-10 and tumor necrosis factor-alpha as predictors of alterations in the embryo-fetal organism and the placental development of diabetic rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Influence of acute pancreatitis on the in vitro responsiveness of rat mesenteric and pulmonary arteries

Background: Acute pancreatitis is an inflammatory disease characterized by local tissue injury and systemic inflammatory response leading to massive nitric oxide (NO) production and haemodynamic disturbances. Therefore, the aim of this work was to evaluate the vascular reactivity of pulmonary and mesenteric artery rings from rats submitted to experimental pancreatitis.Male Wistar rats were divided into three groups: saline (SAL); tauracholate (TAU) and phospholipase A(2) (PLA(2)). Pancreatitis was induced by administration of TAU or PLA(2) from Naja mocambique mocambique into the common bile duct of rats, and after 4 h of duct injection the animals were sacrificed. Concentration-response curves to acetylcholine (ACh), sodium nitroprusside (SNP) and phenylephrine (PHE) in isolated mesenteric and pulmonary arteries were obtained. Potency (pEC(50)) and maximal responses (E(MAX)) were determined. Blood samples were collected for biochemical analysis.Results: In mesenteric rings, the potency for ACh was significantly decreased from animals treated with TAU (about 4.2-fold) or PLA(2) (about 6.9-fold) compared to saline group without changes in the maximal responses. Neither pEC(50) nor E(MAX) values for Ach were altered in pulmonary rings in any group. Similarly, the pEC(50) and the E(MAX) values for SNP were not changed in both preparations in any group. The potency for PHE was significantly decreased in rat mesenteric and pulmonary rings from TAU group compared to SAL group (about 2.2- and 2.69-fold, for mesenteric and pulmonary rings, respectively). No changes were seen in the E(MAX) for PHE. The nitrite/nitrate (NO(x)(-)) levels were markedly increased in animals submitted to acute pancreatitis as compared to SAL group, approximately 76 and 68% in TAU and PLA(2) protocol, respectively.Conclusion: Acute pancreatitis provoked deleterious effects in endothelium-dependent relaxing response for ACh in mesenteric rings that were strongly associated with high plasma NO(x)(-) levels as consequence of intense inflammatory responses. Furthermore, the subsensitivity of contractile response to PHE in both mesenteric and pulmonary rings might be due to the complications of this pathological condition in the early stage of pancreatitis.

Tissue inhibitor of metalloproteinase-2 (TIMP-2) location in the ventral, lateral, dorsal and anterior lobes of rat prostate by immunohistochemistry

Matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) play a major role in extracellular matrix component degradation in several normal and abnormal tissue situations; they are also found in human seminal plasma. MMPs have been found in rat prostate secretions and are nearly lobe specific in expression pattern. The aim of this study was to evaluate whether TIMP-2, like other semen components, is expressed differently from different rat prostatic lobes. Immunohistochemical staining was performed in both young and adult rat ventral (VP), lateral (LP), dorsal (DP), and anterior (AP) prostatic lobes and confirmed by western blotting. TIMP-2 expression was found in the epithelial cells in the following sequence: LP > AP > DP > VP, in both young and adult rats. In this study, 100% of adult LP presented histological signs of prostatitis, where TIMP-2 immunostaining was positive in normal epithelium even with intraluminal neutrophils, but was reduced or absent in the epithelium with intraepithelial leukocytes or with periductal stroma disorganization associated with mononuclear cell infiltration. However, TIMP-2 expression in LP was not induced by prostatitis, since younger rat LPs were also strongly TIMP-2 positive. The distal and intermediate VP regions were TIMP-2 negative, but the proximal regions were strongly stained. Western blotting results confirmed the high TIMP-2 expression in the LP lobe. Thus, TIMP-2 is expressed differently between the prostatic lobes and is another nearly lobe-specific protein, which plays a role in the regulation of MMP activity in seminal plasma and glandular homeostasis. TIMP-2 is also another regional ductal variation of VP. Further studies should address whether TIMP-2 expression is related to the highest incidence of rat LP prostatitis and adenocarcinoma. © 2006 International Federation for Cell Biology.

Healing of fresh frozen bone allograft with or without platelet-rich plasma: A histologic and histometric study in rats

Purpose: This study histomorphometrically analyzed the effect of autogenous platelet-rich plasma (PRP) on healing of fresh frozen bone allograft (FFBA) in bony defects in rat calvaria. Materials and Methods: A 5mm-diameter defect was created in the calvarium of 30 rats. Animals were divided into three groups: C (defect was filled by blood clot only), FFBA (defect was filled with 0.01mL of FFBA), and FFBA/PRP (defect was filled with 0.01mL of FFBA combined with 100μL of PRP). All animals were euthanized at 30 days postoperatively. Histomorphometry and histology analyses were performed. Data were statistically analyzed (analysis of variance, Tukey, p<.05). Results: FFBA had a statistically smaller new bone area than groups FFBA/PRP and C. No statistically significant differences were observed between groups FFBA and FFBA/PRP with regard to remaining bone graft particle area. Conclusion: It can be concluded that (1) PRP improved the incorporation of FFBA, increasing the amount of new bone formed; (2) PRP has not influenced the resorption of nonviable particles of the FFBA; and (3) presence of remaining FFBA particles might have accounted for the smaller amount of new bone observed in group FFBA when compared with control group. © 2011 Wiley Periodicals, Inc.

Hepatic mRNA expression and plasma levels of insulin-like growth factor-I (IGF-I) in broiler chickens selected for different growth rates

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)