In-capillary nanoreactor based on transverse diffusion of
laminar flow profiles for cytochrome P450 2C9 isoform reaction
with diclofenac

D 2010

In-capillary nanoreactor based on transverse diffusion of laminar flow profiles for cytochrome P450 2C9 isoform reaction with diclofenac

ŘEMÍNEK, Roman, Aleš MÁDR and Zdeněk GLATZ

Basic information

Original name

In-capillary nanoreactor based on transverse diffusion of laminar flow profiles for cytochrome P450 2C9 isoform reaction with diclofenac

Authors

ŘEMÍNEK, Roman, Aleš MÁDR and Zdeněk GLATZ

Edition

Baltimore, MD, USA, Book of Abstracts, 17th International Symposium on Electro- and Liquid Phase-Separation Techniques ITP 2010, p. 62-62, 1 pp. 2010

Publisher

Prof. Ziad El Rassi, Oklahoma State University, Stillwater, OK, SA

Other information

Language

English

Type of outcome

Stať ve sborníku

Field of Study

10600 1.6 Biological sciences

Country of publisher

United States of America

Confidentiality degree

není předmětem státního či obchodního tajemství

Organization unit

Faculty of Science

Keywords (in Czech)

CYP 2C9, diklofenak, on-line reakce, kapilární elekroforéza

Keywords in English

CYP 2C9, diclofenac, on-line reaction, capillary electrophoresis

Abstract

ORIG CZ

V originále

A fused silica capillary as an important part of capillary electrophoresis system represents a favourable instrument not only for highly effective separations but also for creating nanoscale reaction chambers. Today commonly used methodology of electrophoretically mediated microanalysis does not allow simple changing of an incubation mixture composition, nevertheless, foreclosing in-capillary reactions from screening studies. To solve this limitation, Krylov et al. introduced the generic methodology for mixing the reactants inside the capillary based on transverse diffusion of laminar flow profiles (TDLFP) [1]. Conceptually, solutions of incubation mixture components are injected by pressure as a series of consecutive plugs which have parabolic profiles due to the laminar nature of flow inside the capillary. Resulting longitudinal interfaces between the plugs then enable the plugs mixing by transverse diffusion within minutes. In this study a method for cytochrome P450 2C9 (CYP2C9) reaction with diclofenac as a probe substrate was developed. 30 min incubation time and sandwich layout, where a plug of CYP2C9 is surrounded by two plugs of diclofenac and NADPH mixture, were selected as appropriate incubation conditions for final method within the optimizing. Although bare silica capillary was used for performing the analyses of samples consisting of enzyme preparations, strong rinsing procedure with 1 M H3PO4 and 1 M NaOH applied between following runs provides good method’s repeatability with RSD values of 1.52 % and 4.83 % (n = 6) for migration times and peak areas of 4’-hydroxydiclofenac, respectively. Since methodology of TDLFP allows simple implementation of changes in incubation mixture composition, introduced method represents a promising tool for in-capillary inhibition studies of CYP2C9.

In Czech

Byla zavedena metoda pro on-line reakci cytochromu P450 2C9 se specifickým substrátem diklofenakem.

Links

GAP206/10/0057, research and development project

Name: Miniaturizovaný systém pro "on-line" studie metabolismu léčiv založený na kapilární elektroforéze

Investor: Czech Science Foundation, Miniaturized on-line drug metabolism system based on capillary electrophoresis

LC06023, research and development project

Name: Integrované bioanalytické technologie pro mikroanalýzy a diagnostiku s využitím LIF a hmotnostní spektrometrie

Investor: Ministry of Education, Youth and Sports of the CR

MSM0021622413, plan (intention)

Name: Proteiny v metabolismu a při interakci organismů s prostředím

Investor: Ministry of Education, Youth and Sports of the CR, Proteins in metabolism and interaction of organisms with the environment

Citovat

ŘEMÍNEK, Roman, Aleš MÁDR and Zdeněk GLATZ. In-capillary nanoreactor based on transverse diffusion of laminar flow profiles for cytochrome P450 2C9 isoform reaction with diclofenac. In Book of Abstracts, 17th International Symposium on Electro- and Liquid Phase-Separation Techniques ITP 2010. Baltimore, MD, USA: Prof. Ziad El Rassi, Oklahoma State University, Stillwater, OK, SA, 2010, p. 62-62.

@inproceedings{898948,
   author = {Řemínek, Roman and Mádr, Aleš and Glatz, Zdeněk},
   address = {Baltimore, MD, USA},
   booktitle = {Book of Abstracts, 17th International Symposium on Electro- and Liquid Phase-Separation Techniques ITP 2010},
   keywords = {CYP 2C9, diclofenac, on-line reaction, capillary electrophoresis},
   language = {eng},
   location = {Baltimore, MD, USA},
   pages = {62-62},
   publisher = {Prof. Ziad El Rassi, Oklahoma State University, Stillwater, OK, SA},
   title = {In-capillary nanoreactor based on transverse diffusion of laminar flow profiles for cytochrome P450 2C9 isoform reaction with diclofenac},
   year = {2010}
}

TY  - JOUR
ID  - 898948
AU  - Řemínek, Roman - Mádr, Aleš - Glatz, Zdeněk
PY  - 2010
TI  - In-capillary nanoreactor based on transverse diffusion of laminar flow profiles for cytochrome P450 2C9 isoform reaction with diclofenac
PB  - Prof. Ziad El Rassi, Oklahoma State University, Stillwater, OK, SA
CY  - Baltimore, MD, USA
KW  - CYP 2C9, diclofenac, on-line reaction, capillary electrophoresis
N2  - A fused silica capillary as an important part of capillary electrophoresis system represents a favourable instrument not only for highly effective separations but also for creating nanoscale reaction chambers. Today commonly used methodology of electrophoretically mediated microanalysis does not allow simple changing of an incubation mixture composition, nevertheless, foreclosing in-capillary reactions from screening studies. To solve this limitation, Krylov et al. introduced the generic methodology for mixing the reactants inside the capillary based on transverse diffusion of laminar flow profiles (TDLFP) [1]. Conceptually, solutions of incubation mixture components are injected by pressure as a series of consecutive plugs which have parabolic profiles due to the laminar nature of flow inside the capillary. Resulting longitudinal interfaces between the plugs then enable the plugs mixing by transverse diffusion within minutes. In this study a method for cytochrome P450 2C9 (CYP2C9) reaction with diclofenac as a probe substrate was developed. 30 min incubation time and sandwich layout, where a plug of CYP2C9 is surrounded by two plugs of diclofenac and NADPH mixture, were selected as appropriate incubation conditions for final method within the optimizing. Although bare silica capillary was used for performing the analyses of samples consisting of enzyme preparations, strong rinsing procedure with 1 M H3PO4 and 1 M NaOH applied between following runs provides good method’s repeatability with RSD values of 1.52 % and 4.83 % (n = 6) for migration times and peak areas of 4’-hydroxydiclofenac, respectively. Since methodology of TDLFP allows simple implementation of changes in incubation mixture composition, introduced method represents a promising tool for in-capillary inhibition studies of CYP2C9.
ER  -

ŘEMÍNEK, Roman, Aleš MÁDR and Zdeněk GLATZ. In-capillary nanoreactor based on transverse diffusion of laminar flow profiles for cytochrome P450 2C9 isoform reaction with diclofenac. In \textit{Book of Abstracts, 17th International Symposium on Electro- and Liquid Phase-Separation Techniques ITP 2010}. Baltimore, MD, USA: Prof. Ziad El Rassi, Oklahoma State University, Stillwater, OK, SA, 2010, p.~62-62.

Detailed Information on Publication Record