Current Pharmaceutical Analysis - Volume 8, Issue 2, 2012

The biological and toxicological effects caused by nitric oxide and nitrogen dioxide, more commonly known as reactive nitrogen species (RNS), have increasingly stirred scientific curiosity about the molecular mechanisms of RNS activity in human body. These radicals possess certain characteristics that complicate their detection, namely their short lifetime and lack of specificity as they react with a variety of endogenous compounds present in biological samples. The development of reliable detection and quantification methods that are sensitive and specific is thus important. Today, nitric oxide can be detected and determined quantitatively by various analytical methods, such as colorimetry, spectrophotometry, electrochemistry, fluorescence, chemiluminescence, electron paramagnetic resonance, gas and liquid chromatography, electrophoresis and mass spectrometry. On the other hand, there are only a few methods capable of measuring nitrogen dioxide. The aim of the present article is to give an overview of the commonly used analytical methods for measuring ·NO and ·NO2 in biological systems. The underlying mechanisms and principles of these methods are discussed.

Simvastatin, is a lipid regulating drug, which is a competitive inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-CoA reductase), obtained by synthesis from lovastatin. It is indicated in the treatment of hyperlipidemias, including hypercholesterolaemias, combined mixed hyperlipidemia hyperlipoproteinemias, hypertriglyceridemia, and primary dysbetalipoproteinemia. This review describes a brief overview of the chemistry including impurities and different synthetic schemes, pharmacokinetic, pharmacodynamic and adverse effect of simvastatin as well as the recent trends in drug–drug interaction studies of simvastatin with various drug categories such as antibacterials, anticoagulants, antidepressants, antifungals, antivirals, calcium-channel blockers, immunosuppressants, lipid regulating drugs and grapefruit juice. This review also focuses distinctly on comprehensive update of various analytical methods for the quantification of simvastatin and/or its metabolites, impurities, degradants and other drugs.

Principal component analysis (PCA) has been performed for some acridinone derivatives with antitumor activity to model relationships between structural descriptors and lipophilicity and their antileukemia activity. Principal component analysis obtained with the use of parameters of lipophilicity led to extract two main factors with eigenvalue higher than 1. The first principal component (factor 1) accounted for, by 84.3% of the variance in data and the second principal component (factor 2) explained 10.0% of data variance indicating that total data variance at the level 94.5% can be explained by the first two principal components. On the other hand, principal component analysis obtained with the use of parameters of lipophilicity together with some other molecular descriptors led to extract five main factors with eigenvalue higher than 1. In this case, the first principal component (factor 1) accounted for, by 62.64% of the variance in data, the second principal component (factor 2) explained 19.04% and the third principal component (factor 3) explained 8.49% of data variance indicating that total data variance at the level 90.17% can be explained by the first three principal components. Moreover, one of the most significant influences on the value of factors possessed parameters describing lipophilicity. However, some other descriptors considered in the study (some of constitutional, topological, molecular and geometrical one) could play an important role in the reflection of acridinone properties, too. More importantly, distribution of individual drugs on the plane determined by two and three principal components produced patterns in good agreement with their chemical structures as well as with their antitumor activity.

The formation of trans-esterification byproducts of methyl- and propylparaben with glycerin was observed during formulation of an oral solution drug product. To obtain a stable product that is physically, chemically and microbial stable during storage at room temperature conditions, the pH of the formulation and the concentration level of the parabens were optimized meticulously to obtain a robust manufacturing process. An HPLC method was developed and fully validated for the determination of methyl- and propylparaben in the formulation. The method was able to detect the levels of the transesterifcation byproducts as well. Typical validation parameters such as accuracy, precision, and linearity, etc. were assessed, along with demonstration of the robustness, stability of solution, evaluation of forced degradation studies and setting of system suitability tests and criteria.

An ultra high performance liquid chromatography-electrospray ionization-tandem mass spectrometric method (UHPLC/ESI-Q-TOF-MS) for the analysis of terbutaline (TB) in Wistar rat plasma has been developed and validated. The chromatographic separation was achieved on a Waters ACQUITY UPLCTM BEH C18 (100.0 mmx2.1 mm; 1.7 μm) column using isocratic mobile phase, consisting of 2 mM ammonium acetate and acetonitrile (90: 10; v/v), at a flow rate of 0.25 mL min-1. The transitions occurred at m/z 226.19→152.12 for TB, and m/z 260.34→183.11 for the internal standard. The recovery of the analytes from Wistar rat plasma was optimized using liquid- liquid extraction technique (LLE) in ethyl acetate. The total run time was 3.0 min and the elution of TB occurred at 1.85±0.05 min. The linear dynamic range was established over the concentration range 1–1000 ng mL-1 (r2; 0.9938±0.0005) for TB. The intra-assay and inter-assay accuracy in terms of % CV was in between 1.8–3.5. The lower limit of quantitation (LLOQ) for TB was 1.0 ng mL-1. Analytes were stable under various conditions (in autosampler, during freeze–thaw, at room temperature, and under deepfreeze conditions). The developed method was successfully applied for pharmacokinetic profiling in rodents.

A HPLC method for the simultaneous determination of three widely used antihistaminic drugs, diphenhydramine, oxatomide and promethazine, in surface water samples and a toxicological assay using the bioluminescent bacterium Vibrio fischeri to assess a comprehensive contamination were developed. The analytical procedure was extended to the by-products from promethazine photodegradation. The method involved pre-concentration and clean-up by solid phase extraction and HPLC analysis with diode array detection. Validation of the method was performed on synthetic mixtures and surface water samples spiked with the drugs, showing mean recovery ranging from 93 and 107%, with RSD below 5.5%. Limits of quantitation in surface waters, calculated on 1.01 of sample, were in the range 1.9 – 2.5 μg/l. The toxicological tests demonstrated toxicity due to promethazine from a concentration of 3.94 μg/ml while the other antihistamines showed no significant toxicity up to 50.0 μg/ml. However, the toxicity of drug mixtures was found greater than the sum of the values from samples containing single drugs. The presence of the studied drugs was monitored in waters sampled from two rivers in Calabria (Italy), collected along a period of seven months. Diphenhydramine and oxatomide were not detected in any samples. On the contrary, promethazine was found in two samples taken in July at a concentration of 2.0 and 2.3 μg/l, significantly below the values causing toxicity.

An HPLC-UV method was developed for simultaneously quantifying 13 major components in Semen Oroxyli, a commonly used traditional Chinese medicinal herb, including scutellarin, quercetin-3-O-α-L-arabinopyranoside, oroxin B, chrysin-7-O-β-D-gentiobioside, baicalin, oroxin A, scutellarein, chrysin-7-O-β-D-glucuronide, quercetin, norwogonine, baicalein, chrysin and oroxylin A. The optimal conditions of separation and detection were achieved on an Agilent Zorbax SB-C18 column (250 mm x 4.6 mm, 5 μm) with a gradient elution of acetonitrile and 0.3% (v/v) acetic acid at a flow rate of 1.0 ml/min, with a detection wavelength of 277 nm. A complete separation was obtained within 50 min for the 13 target compounds. All calibration curves showed good linearity (r2 > 0.9997) within the testing range. The assay was reproducible with overall intra-and inter-day variation of less than 3%. The mean spiked recovery of all analytes was 100 ± 10%, with RSD less than 5%. The validated method was successfully applied to quantitatively analyse 13 flavonoids for quality evaluation of commercial Semen Oroxyli samples from different locations.

The aim of the present study was to evaluate the potential hepatoprotective effect of α-tocopherol and/or selenium on some plasmatic biochemical indices of liver profile and hepatic damage induced in adult male rats exposed to subchronic dose of malathion (MTN) equivalent to 1/50 LD50. Oral administration of MTN for 45 days significantly induced severe hepatic injury as revealed by increased activity of plasmatic biochemical indices, including alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH) and gamma-glutamyl transferase (GGT) enzymes. Oral administration of α-tocopherol (α-T) and selenium (Se) concomitant with MTN resulted in a significant ameliorative effect by lowering the elevated plasma levels of the previous enzymes. Light microscopic investigation revealed that MTN exposure was associated with necrosis of hepatocytes and marked degenerative changes of liver tissues. Coadministration of (α-T) and Se concomitant with MTN to rats improved the histopathological severity score from severe to normal. However the individual treatment of (α-T) or Se correlated with a relative protection as reflected in the change of the histopathological severity score from severe to mild and moderate, respectively. Thus, it appears that the treatment with (α-T) and/or Se improves MTN hepatotoxicity but is not completely protective.