Flow injection analysis using carbon film resistor electrodes for amperometric determination of ambroxol

Flow injection analysis (FIA) using a carbon film sensor for amperometric detection was explored for ambroxol analysis in pharmaceutical formulations. The specially designed flow cell designed in the lab generated sharp and reproducible current peaks, with a wide linear dynamic range from 5 x 10(-7) to 3.5 x 10(-4) mol L-1, in 0.1 mol L-1 sulfuric acid electrolyte, as well as high sensitivity, 0.110 A mol(-1) L cm(-2) at the optimized flow rate. A detection limit of 7.6 x 10(-8) mol L-1 and a sampling frequency of 50 determinations per hour were achieved, employing injected volumes of 100 mu L and a flow rate of 2.0 mL min(-1). The repeatability, expressed as R.S.D. for successive and alternated injections of 6.0 x 10(-6) and 6.0 x 10(-5) mol L-1 ambroxol solutions, was 3.0 and 1.5%, respectively, without any noticeable memory effect between injections. The proposed method was applied to the analysis of ambroxol in pharmaceutical samples and the results obtained were compared with UV spectrophotometric and acid-base titrimetric methods. Good agreement between the results utilizing the three methods and the labeled values was achieved, corroborating the good performance of the proposed electrochemical methodology for ambroxol analysis. (C) 2008 Elsevier B.V. All rights reserved.

Determination of ambroxol in syrups using diffuse reflectance spectroscopy

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

Fabrication of Electrochemical Sensors for the Determination of Pharmaceuticals

Voltammetric sensors are an important class of electrochemical sensors in which the analytical information is obtained from the measurement of current obtained as a result of electrochemical oxidation/reduction.This current is proportional to the concentration of the analyte.Chemically modified electrodes(CMEs) have great significance as important analytical tools for the electrochemical determination of pharmaceuticals.The modification of electrode results in efficient determination of electro-active biomolecules at very lower potential without its major interferences.The operation mechanism of CMEs depends on the properties of the modifier materials that are used to promote selectivity towards the target analytes.Modified electrodes can be prepared by deposition of various compounds such as organic compounds ,conducting polymers,metal oxides,etc. on the various electrode surfaces.The thesis presents the development ,electrochemical characterization and analytical application studies of eight voltammetric sensors developed for six drugs viz.,Ambroxol,Sulfamethoxazole,PAM Chloride, Lamivudine,Metronidazole and Nimesulide.The modification techniques adopted as part of the present work include Multiwalled Carbon Nanotube(MWCNT) based modification.Electropolymerisation and Gold Nanoparticle (AuNP) based modifications.

Fabrication of Electrochemical Sensors for Pharmaceutical Analysis

Potentiometric chemical sensors,an important class of electro-chemical sensors are widely used in pharmaceutical analysis because of its inherent advantages.The present study was aimed at fabrication of potentiometric sensors for the drugs mebendazole,pefloxacin,ambroxol,sildenafil citrate,dextro-methorphan and tetracycline.A total of 18 sensors have been developed for the determination of theses drugs.The major step in the fabrication of the sensor was the preparation of the ion association.Two types of sensors viz:PVC membrane sensor and carbon paste electode (CPE) were fabricated.The response characteristics of the different sensors fabricated were studied.Various response parameters studied include response time,selectivity and the effect of pH.The developed sensors were also employed for the determination of the drugs in pharmaceutical formulations and also for the recovery of the drug from urine samples.The selectivity studies reveal that the developed sensors are highly selective to the drug even in prescence of foreign ions.

Implementing stepwise solvent elution in sequential injection chromatography for fluorimetric determination of intracellular free amino acids in the microalgae Tetraselmis gracilis

The concept of sequential injection chromatography (SIC) was exploited to automate the fluorimetric determination of amino acids after pre-column derivatization with ophthaldialdehyde (OPA) in presence of 2-mercaptoethanol (2MCE) using a reverse phase monolithic C(18) stationary phase. The method is low-priced and based on five steps of isocratic elutions. The first step employs the mixture methanol: tetrahydrofuran: 10 mmol L(-1) phosphate buffer (pH 7.2) at the volumetric ratio of 8:1:91; the other steps use methanol: 10 mmol L-1 phosphate buffer (pH 7.2) at volumetric ratios of 20:80, 35:65, SO:SO and 65:35. At a flow rate of 10 mu L s(-1) a 25 mm long-column was able to separate aspartic acid (Asp), glutamic acid (Glu), asparagine (Asn), serine (Ser), glutamine (Gln), glycine (Gly), threonine (Thr), citruline (Ctr), arginine (Arg), alanine (Ala), tyrosine (Tyr), phenylalanine (Phe), ornithine (Orn) and lysine (Lys) with resolution >1.2 as well as methionine (Met) and valine (Val) with resolution of 0.6. Under these conditions isoleucine (Ile) and leucine (Leu) co-eluted. The entire cycle of amino acids derivatization, chromatographic separation and column conditioning at the end of separation lasted 25 min. At a flow rate of 40 mu L s(-1) such time was reduced to 10 min at the cost of resolution worsening for the pairs Ctr/Arg and Orn/Lys. The detection limits varied from 0.092 mu mol L(-1) for Tyr to 0.51 mu mol L(-1) for Orn. The method was successfully applied to the determination of intracellular free amino acids in the green alga Tetraselmis gracilis during a period of seven days of cultivation. Samples spiked with known amounts of amino acids resulted in recoveries between 94 and 112%. (C) 2008 Elsevier B.V. All rights reserved.

CD8+ T cells and NK cells from blister fluid of an EEM/SJS overlap highly express Fas ligand and Granulysin

INTRODUCTION Erythema exsudativum multiforme majus (EEMM) and Stevens-Johnson Syndrome (SJS) are severe cutaneous reaction patterns caused by infections or drug hypersensitivity. The mechanism by which widespread keratinocyte death is mediated by the immune system in EEMM/SJS are still to be elucidated. Here, we characterized the blister cells isolated from a patient with EEMM/SJS overlap and investigated its cause. METHODS Clinical classification of the cutaneous eruption was done according to the consensus definition of severe blistering skin reactions and histological analysis. Common infectious causes of EEMM were investigated using standard clinical techniques. T cell reactivity for potentially causative drugs was assessed by lymphocyte transformation tests (LTT). Lymphocytes isolated from blister fluid were analyzed for their expression of activation markers and cytotoxic molecules using flow cytometry. RESULTS The healthy 58 year-old woman suffered from mild respiratory tract infection and therefore started treatment with the secretolytic drug Ambroxol. One week later, she presented with large palmar and plantar blisters, painful mucosal erosions, and flat atypical target lesions and maculae on the trunc, thus showing the clinical picture of an EEMM/SJS overlap (Fig. 1). This diagnosis was supported by histology, where also eosinophils were found to infiltrate the upper dermis, thus pointing towards a cutaneous adverse drug reaction (cADR). Analysis of blister cells showed that they mainly consisted of CD8+ and CD4+ T cells and a smaller population of NK cells. Both the CD8+ T cells and the NK cells were highly activated and expressed Fas ligand and the cytotoxic molecule granulysin (Fig. 2). In addition, in comparison to NK cells from PBMC, NK cells in blister fluids strongly upregulated the expression of the skin-homing chemokine receptor CCR4 (Fig 4). Surprisingly, the LTT performed on PBMCs in the acute phase was positive for Ambroxol (SI=2.9) whereas a LTT from a healthy but exposed individual did not show unspecific proliferation. Laboratory tests for common infectious causes of EEMM were negative (HSV-1/-2, M. pneumoniae, Parvovirus B19). However, 6 weeks later, specific proliferation to Ambroxol could no longer be observed in the LTT (Fig 4.).