Determination of glutathione disulfide levels in biological samples using thiol-disulfide exchanging agent, dithiothreitol

A reverse-phase HPLC method incorporating dithiothreitol (DTT) reduction for quantitative determination of oxidized glutathione (GSSG) in biological samples is described here. This method is based on our previous enzymatic reduction technique that uses N-1-(pyrenyl) maleimide (NPM) as a derivatizing agent. In our earlier method, glutathione disulfide (GSSG) was measured by first reducing it to GSH with glutathione reductase (GR) in the presence of NADPH. However, this is a very costly and time-consuming technique. The method described here employs a common and inexpensive thiol-disulfide exchanging agent, DTT, for reduction of GSSG to GSH, followed by derivatization with NPM. The calibration curves are linear over a concentration range of 25-1250 nm (r(2) > 0.995). The coefficients of variations for intra-run precision and inter-run precision range from 0.49 to 5.10% with an accuracy range of 1.78-6.15%. The percentage of relative recovery ranges from 97.3 to 103.2%. This new method provides a simple, efficient, and cost-effective way of determining glutathione disulfide levels with a 2.5 nm limit of detection per 5 microL injection volume.     

Microwave‐assisted synthesis of 1,3′‐diaza‐flavanone/flavone and their alkyl derivatives with antimicrobial activity

A simple environmentally friendly solid‐phase microwave‐assisted method was used to synthesis of the 1,3′‐diazaflavanone ( 2 ) and 1,3′‐diazaflavone ( 3 ) from the cyclization of 2′‐amino (E)‐3″‐azachalcone ( 1 ). Ten new N‐alkyl (C_5–12,14,15)‐substituted 1,3′‐diazaflavanonium bromides ( 2a–j ) were prepared from compound 2 with corresponding alkyl halides in acetonitrile under reflux. In addition, nine new N,N′‐dialkyl (C_5–12,14)‐substituted 1,3′‐diazaflavonium bromides ( 3a–i ) were also synthesized from compound 3 with corresponding alkyl halides using basic silica in acetonitrile. The antimicrobial activities of compounds 1–3 , 2a–j , and 3a–i were tested against Gram‐positive (G+) (Bacillus subtilis, Staphylococcus epidermidis, Staphylococcus aureus, and Enterococcus faecalis) and Gram‐negative (G?) (Escherichia coli, Klebsiella pneumonia, Pseudomonas aeruginosa, Proteus vulgaris, Salmonella typhimirium, Yersinia pseudotuberculosis, and Enterobacter cloaceae) microorganisms. They showed good antimicrobial activity against the Gram‐positive bacteria tested with the minimal inhibitory concentration values less than 7.8 μg/mL in most cases. The optimum length of the alkyl chain for better and broader activity is situated in the range of 9–12 carbon atoms in the series of compounds 2a–j and five to six carbon atoms in the series of compounds 3a–i . The nonalkylated compounds 1–3 were not effective, as were the ones alkylated with five or six C alkyl groups ( 2a and 2b ) and 8–13 C alkyl groups for N,N′‐dialkyl compounds ( 3c–3i ). The antimicrobial activity increased as the length of the alkyl substitution increased from 8 to 12 carbons in compounds 2a–j . However, antimicrobial activity decreased as the length of the alkyl substitution increased from 7 to 13 carbons in compounds 3c–i . J. Heterocyclic Chem., (2012)     

Determination of magnesium,calcium, sodium,and potassium in blood plasma samples by capillary zone electrophoresis

A capillary zone electrophoretic assay has been developed and validated for analysis of magnesium, calcium, sodium, and potassium in blood plasma samples. Optimum results were obtained with 20 mmol L⁻¹ imidazole (pH 2.8) and 0.5 mmol L⁻¹ oxalic acid containing 5% methanol, capillary temperature 25°C, applied voltage 30 kV, hydrodynamic injection time 3 s, and a poly(vinyl alcohol)-coated capillary (i.d. 50 μm, total length 64.5 cm and effective length 56 cm). Indirect detection was performed at 214 nm. Cadmium was used as internal standard. The migration times of magnesium, calcium, sodium, and potassium were 4.25, 3.79, 3.96, and 2.79 min, respectively. The method was applied to the determination of magnesium, calcium, sodium, and potassium in blood plasma samples. The results were compared with those from atomic absorption spectrophotometry and no statistically significant difference was found (P>0.05). This study was supported by the Turkish Republic, Prime Ministry State Planning Organization (Project Number: 98K121730)