In Silico and Chromatographic Methods for Analysis of Biotransformation of Prospective Neuroprotective Pyrrole-Based Hydrazone in Isolated Rat Hepatocytes.

In the current study, chromatographic and in silico techniques were applied to investigate the biotransformation of ethyl 5-(4-bromophenyl)-1-(2-(2-(2-hydroxybenzylidene) hydrazinyl)-2-oxoethyl)-2-methyl-1H-pyrrole-3-carboxylate (11b) in hepatocytic media. The initial chromatographic procedure was based on the employment of the conventional octadecyl stationary phase method for estimation of the chemical stability. Subsequently, a novel and rapid chromatographic approach based on a phenyl-hexyl column was developed, aiming to separate the possible metabolites. Both methods were performed on a Dionex 3000 ThermoScientific (ACM 2, Sofia, Bulgaria) device equipped with a diode array detector set up at 272 and 279 nm for analytes detection. An acetonitrile: phosphate buffer of pH 3.5: methanol (17:30:53 v/v/v) was eluted isocratically as a mobile phase with a 1 mL/min flow rate. A preliminary purification from the biological media was achieved by protein precipitation with methanol. A validation procedure was carried out, where the method was found to correspond to all ICH (Q2) and M10 set criteria. Additionally, an in silico-based approach with the online server BioTransformer 3.0 was applied in an attempt to predict the possible metabolites of the title compound 11b. It was hypothesized that four CYP450 isoforms (1A2, 2C9, 3A4, and 2C8) were involved in the phase I metabolism, resulting in the formation of 12 metabolites. Moreover, docking studies were conducted to evaluate the formation of stable complexes between 11b and the aforementioned isoforms. The obtained data indicated three metabolites as the most probable products, two of which (M9_11b and M10_11b) were synthesized by a classical approach for verification. Finally, liquid chromatography with a mass detector was implemented for comprehensive and summarized analysis, and the obtained results revealed that the metabolism of the 11b proceeds possibly with the formation of glucuronide and glycine conjugate of M11_11b.

Correlation of Disease Activity and Quality of Life of Patients with Psoriasis after Narrow-band Ultraviolet B Therapy.

INTRODUCTION: Treatment with ultraviolet light is a well-established and effective treatment option for mild to moderate psoriasis. The aims of the study were to measure the psoriasis area and severity index (PASI) reduction after narrow-band ultraviolet B (NB UVB) therapy, to evaluate the quality of life before and after treatment using the dermatology life quality index (DLQI), and to compare the clinical effectiveness with quality of life improvement. MATERIAL AND METHODS: Twenty two patients (13 male and 9 female patients), aged between 21 to 70 years (mean age 40±14.65 years) were enrolled in the study. NB UVB treatment was performed with 10 to 25 (mean 18.5; SD 3.39) procedures with cumulative doses of 5 to 19.4 J/cm2. The baseline median PASI score was 20.027 which decreased after therapy to 11.11. More than PASI 50% reduction was achieved in 40.91% of the patients after at least 6 weeks of treatment and the results are highly statistically significant. Quality of life (QoL) assessed using DLQI was found moderately affected by disease pretreatment. NB UVB therapy significantly increased DLQI score in spectrum of 'symptoms and feelings' and 'treatment'. DISCUSSION: The PASI score reduction that we observed after NB-UVB therapy is consistent with the results reported by other authors. Baseline DLQI scores were indicative of moderate QoL impairments associated with disease. At the same time, the reduction of the DLQI index corresponding to improved QoL correlated with the objective clinical symptom assessment. CONCLUSION: Our data suggest that DLQI and PASI indexes are important complementary methods for comprehensive health assessment of patients with psoriasis.

Air pollution by carcinogenic PAHs and plasma levels of p53 and p21(WAF1) proteins.

We analyzed the effect of exposure to carcinogenic polycyclic aromatic hydrocarbons (c-PAHs) in ambient air on the plasma levels of p53 and p21(WAF1) proteins among city policemen, bus drivers and controls in three European cities: Prague (Czech Republic), Kosice (Slovakia) and Sofia (Bulgaria). p53 and p21(WAF1) proteins are key regulators of the cell cycle and are accepted as universal markers of genotoxic stress and DNA damage. In total 204 exposed subjects (100 smokers, 104 nonsmokers) and 152 controls (54 smokers, 98 nonsmokers) were analyzed. Personal exposure to c-PAHs was evaluated using personal samplers during the working shift. The levels of p53 and p21(WAF1) proteins were assessed by ELISA assay. There were no differences between the levels of either protein between exposed and controls, or smokers and nonsmokers, in any city. However, we observed significant differences in p53 plasma levels in all subjects regardless of the exposure status between the individual cities (median values: 5, 31, 234pg/ml, p<0.001, for Prague, Kosice and Sofia, respectively). The levels correspond to the differences in exposure levels to c-PAHs and benzo[a]pyrene (B[a]P) in the individual cities. A multiple linear regression analysis confirmed that c-PAHs exposure is a variable significantly affecting levels of both proteins in all locations. When all subjects were divided into the group exposed to below-median levels of c-PAHs and the group exposed to above-median levels of c-PAHs we found significantly higher p53, as well as p21(WAF1) levels in the above-median exposure group (p53, 167pg/ml versus 25pg/ml, p<0.001; p21(WAF1), 2690pg/ml versus 2600pg/ml, p<0.05). Among all subjects p53 plasma levels were positively correlated with p21(WAF1) levels, exposure to B[a]P, c-PAHs and levels of total DNA adducts; for p21(WAF1) levels we observed the positive correlation with cotinine, c-PAHs exposure, total and B[a]P-like DNA adduct levels. In conclusion our results suggest that p53 and p21(WAF1) proteins plasma levels may be useful biomarkers of c-PAHs environmental exposure.