RESUMEN
During the development of a specific dipeptidyl peptidase 4 (DPP4) inhibitor to treat type 2 diabetes, a fluorogenic kinetic analysis for DPP4 enzymatic activity using Gly-Pro-Aminomethylcoumarin (AMC) as a substrate was optimized and validated for recombinant DPP4 and human plasma samples. The sensitivity, calibration curve, detection range, accuracy, precision, recovery efficiency, Km constant, short/long-term stability, and stability after freezing-thawing cycles were analyzed. DPP4 enzymatic activity (mU/min) was measured as the initial velocity (Vo) of the enzymatic reaction over time. The sensitivity of the Vo value was 14,488 mU/min for recombinant DPP4 and 17,995 mU/min for human plasma samples. The dynamic ranges of the calibration curve were linear and reliable between 1.11 × 104-1.86 × 106 mU/min of the mean Vo value and in the DPP4 concentration range of 23.4-3,000 ng/mL. The assay's accuracy and precision met acceptance criteria for all samples. Plasma DPP4 was stable under various storage temperatures, even after three freeze-thaw cycles. Our optimized, validated bioanalytic method for measuring DPP4 activity in plasma samples was successfully employed to evaluate the effect of evogliptin (DA-1229) tartrate, which irreversibly and dose-dependently inhibits DPP4 enzymatic activity, without the dilution effect of human plasma samples and irrespective of the co-treated metformin.
Asunto(s)
Dipeptidil Peptidasa 4/sangre , Pruebas de Enzimas/métodos , Espectrometría de Fluorescencia/métodos , Calibración , Cumarinas/metabolismo , Dipeptidil Peptidasa 4/análisis , Inhibidores de la Dipeptidil-Peptidasa IV/administración & dosificación , Inhibidores de la Dipeptidil-Peptidasa IV/metabolismo , Inhibidores de la Dipeptidil-Peptidasa IV/farmacología , Humanos , Cinética , Límite de Detección , Piperazinas/administración & dosificación , Piperazinas/metabolismo , Piperazinas/farmacología , Estabilidad ProteicaRESUMEN
In the title compound, [Cu(NO(3))(2)(C(15)H(19)N(3)O)], the Cu(II) ion is coordinated by the N atoms of the tetra-dentate 3-[bis-(2-pyridyl-meth-yl)amino]-propanol ligand and two O atoms from two monodentate nitrate anions, resulting in a distorted square-pyramidal environment. An inter-molecular O-Hâ¯O hydrogen-bonding inter-action between the free hy-droxy group of the ligand and a nitrate O atom of an adjacent complex unit, gives a chain structure which extends across the (101) planes.