ABSTRACT
The amino-epoxyquinols 6a and 6b were synthesized as soluble derivatives of an NF-κB inhibitor DHMEQ (1). In spite of the opposite configuration from 1, 6b rather than 6a affected the deactivation of NF-κB, based on NO secretion and MALDI-TOF MS analysis. It was indicated that 6b inhibited the activation by different manner from that of 1.
Subject(s)
Benzamides/chemistry , Cyclohexanones/chemistry , Epoxy Compounds/chemistry , Hydroquinones/chemistry , NF-kappa B/antagonists & inhibitors , Animals , Cell Line, Tumor , Cyclohexanones/chemical synthesis , Cyclohexanones/pharmacology , Epoxy Compounds/chemical synthesis , Epoxy Compounds/pharmacology , Mice , NF-kappa B/metabolism , Nitric Oxide/metabolism , Spectrometry, Mass, Matrix-Assisted Laser Desorption-IonizationABSTRACT
We have developed a simple and reliable method for determining plasma concentration of dehydroxymethylepoxyquinomicin (DHMEQ), a new low molecular weight NF-kappaB inhibitor, using high performance liquid chromatography with mass spectrometry (LC-MS). An experiment of mass spectrometry with electrospray ionization in the negative ionization mode was performed to detect ion transitions at m/z 260.05 [M-H](-) for DHMEQ and 240.29 for mefenamic acid as an internal standard. The samples were purified using liquid-liquid extraction with ethyl acetate. The method yielded a standard curve which was linear for the concentration range of 0.1-125 ng/mL when 0.05 mL plasma was used. The correlation coefficients of all standard curves were greater than or equal to 0.999. The limit of detection was 50 pg/mL (signal/noise >3). Daily fluctuation of plasma standard curve was small. The intra- and inter-assay precision ranged from 2.84 to 4.76% (n=6) and 2.91 to 7.03% (n=6), respectively. The LC-MS technique described provides a simple and reliable liquid chromatographic method for the determination of DHMEQ level and for use in studies involving pharmacokinetics.
Subject(s)
Benzamides/blood , Chromatography, High Pressure Liquid/methods , Cyclohexanones/blood , Mass Spectrometry/methods , Animals , Calibration , Humans , Male , Mice , Mice, Inbred C57BL , Reproducibility of Results , Sensitivity and SpecificityABSTRACT
Several synthetic N-substituted N-nitrosohydroxylamines were found to inhibit mushroom tyrosinase in a pH-dependent manner regardless of the N-substituent. The inhibitory activity, or pI(50) ( - log [IC(50), M]) value, linearly decreased as the pH of the media increased. The inhibitory activities of tested N-substituted N-nitrosohydroxylamines at pH 6.8 and 5.8 were found to be almost 10 times and 100 times greater than at pH 7.8, respectively. The types of inhibition were different at pH 6.8 and 5.8. These results suggest that the inhibitory effect of N-substituted N-nitrosohydroxylamines is caused by the non-ionized form of the inhibitor. Furthermore, the mechanism of inhibition depends on the interaction between the inhibitor and the active site of tyrosinase at different pH values.
Subject(s)
Agaricales/enzymology , Hydroxylamines/pharmacology , Monophenol Monooxygenase/antagonists & inhibitors , Nitroso Compounds/pharmacology , Benzoic Acid/pharmacology , Hydrogen-Ion Concentration , Hydroxamic Acids/pharmacology , Kinetics , Pyrones/pharmacology , Tropolone/pharmacologyABSTRACT
Several novel N-substituted N-nitrosohydroxylamines were synthesized. They all inhibited mushroom tyrosinase, but the type of inhibition was different depending on the substituent. Some N-(mono- or dihydroxybenzyl)-N-nitrosohydroxylamines exhibited uncompetitive inhibition with respect to L-dopa. Among them, compound 6 was also a competitive inhibitor with respect to oxygen. This observation suggests that another interaction by the meta- or para-hydroxyl group might stabilize the binding of the inhibitor to the enzyme through the oxygen binding site.
