Trichomonas vaginalis is a flagellated protozoan that affects the human urogenital tract causing 276.4 million new infections a year. The parasite elicits a vaginal mucosal infiltration of immune cells, especially neutrophils which are considered to be primarily responsible for cytological change observed at the infection site as well as the major contributor in the inflammatory response against the parasite. Extracellular nucleotides and their nucleosides are signaling compounds involved in several biological processes, including inflammation and immune responses. Once in the extracellular space, the nucleotides and nucleosides can directly activate the purinergic receptors. Herein, we investigated the involvement of purinergic signaling on the production of reactive oxygen species (ROS) and cytokines by T. vaginalis-stimulated neutrophils. Parasites were able to induce an increase in ROS and IL-8 levels while they did not promote IL-6 secretion or neutrophil elastase activity. Adenine and guanine nucleotides or nucleosides were not able to modulate ROS and cytokine production; however, when T. vaginalis-stimulated neutrophils were incubated with adenosine and adenosine deaminase inhibitor, the levels of ROS and IL-8 were significantly reduced. These immunosuppressive effects were probably a response to the higher bioavailability of adenosine found in the supernatant as result of inhibition of enzyme activity. The involvement of P1 receptors was investigated by immunofluorescence and A1 receptor was the most abundant. Our data show that the influence of purinergic signaling, specifically those effects associated with adenosine accumulation, on the modulation of production of proinflammatory mediators by T. vaginalis-stimulated neutrophils contribute to the understanding of immunological aspects of trichomoniasis.

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A simple, stability-indicating and compatible method by liquid chromatography with ultraviolet (LC-UV) and mass spectrometry (LC-MS) detections was developed for the evaluation of delapril (DEL) and manidipine (MAN) degradation products formed under alkaline and photolytic forced conditions, respectively. The chromatographic separation of all compounds was obtained within 7 min and carried out on a C 18 column with mobile phase composed by ammonium acetate and acetonitrile for both techniques. Thereby, the main degradation products detected were studied by LC coupled to electrospray ionization mass spectrometry (LC-ESI-MS). Based on retention times and molecular weight confirmation, a comprehensive degradation pathway for both drugs and the identity of its major products formed could be suggested, without complicated isolation or purification processes. The degradation kinetics of the drugs was also evaluated and could be best described as first-order process for DEL (R 2 = 0.9991) and zero-order process for MAN (R 2 = 0.9867). Furthermore, no evidence of cytotoxicity in human mononuclear cells was observed for DEL and MAN degraded samples. The proposed Downloaded by [New York University] at 09:09 18 June 20152 LC-UV method was successfully validated according ICH guidelines and thus helping to improve the quality control of pharmaceuticals.

The present study describes the development and validation of a dissolution method for delapril (DEL) and manidipine (MAN) combination tablets, using a simulated absorption profile based on in vivo data for MAN. The suitable in vitro dissolution profile for this formulation was obtained using 900 mL of citrate buffer pH 3.2 at 37 °C±0.5 °C as dissolution medium and USP apparatus 2 (paddle) at 75 rpm. All samples were analyzed by a liquid chromatography (LC) method. Under these conditions, a significant linear relationship between the absorbed (calculated by deconvolution approach) and dissolved fractions of MAN was obtained (R=0.997) and an in vivo-in vitro (IVIV) correlation for this particular formulation containing MAN can be established. Validation parameters for dissolution methodology such as the specificity, linearity, accuracy and precision were also evaluated according to the international guidelines, giving results within the acceptable range. Therefore, the proposed dissolution conditions can be applied for the simultaneous release analysis of DEL and MAN from the solid dosage form, contributing to the improvement of the quality control of pharmaceutics and minimizing the number of bioavailability studies.