Assessment of catechol-O-methyltransferase activity and its inhibition in erythrocytes of animals and humans

A universal competitive fluorescence polarization activity assay for S-adenosylmethionine utilizing methyltransferases

“…7). The average IC 50 value and standard deviation derived from three separate experiments was 12 ± 9 nM. Thus, the IC 50 value determined with this COMT FP assay was consistent with the range of IC 50 values reported in the literature.…”

“…A majority of the assays require separation of labeled reaction products from labeled substrate. Currently, HPLC is widely employed for separation in conjunction with detection of fluorescent [43][44][45][46][47][48][49][50][51][52] or radiolabeled [53,54] product, UV detection [55][56][57][58], electrochemical detection [59][60][61], colorimetric detection [62], or mass spectrometry [63]. Assays that require physical separation of product from substrate are not well suited for high-throughput purposes due to the number of manipulations and amount of time required for the separation process.…”

“…FP was determined 1 h after terminating the reaction. The average IC 50 value from three independent experiments was 12 nM.…”

“…FP was measured 1 h after the addition of anti-AdoHcy antibody/tracer complex to the reaction. For IC 50 determination, serial dilutions of the Ro 41-0960 COMT inhibitor were added to the reaction mixture at concentrations ranging from 1.5 nM to 10 μM with a final dimethyl sulfoxide (DMSO) concentration of 1%.…”

Pharmacodynamic Response of Entacapone in Rats after Administration of Entacapone Formulations and Prodrugs with Varying Bioavailabilities

“…Secondly, there are species differences in the metabolism and elimination of entacapone between rats and man (Wikberg et al 1993). Finally, the COMT activity in rat erythrocytes is markedly higher than that found in human erythrocytes (Schultz et al 1989; Zürcher et al 1996).…”