An improved design to capture magnetic microparticles for
in-line CE in a liquid based capillary coolant system

a 2017

An improved design to capture magnetic microparticles for in-line CE in a liquid based capillary coolant system

RAMANA, Pranov; Jan SCHEJBAL; Erwin ADAMS; Jan PREISLER; Zdeněk GLATZ et al.

Základní údaje

Originální název

An improved design to capture magnetic microparticles for in-line CE in a liquid based capillary coolant system

Název česky

Vylepšený způsob zachycení magnetických mikročástic pro in-line kapilární elektroforézu chlazenou kapalinou

Autoři

RAMANA, Pranov; Jan SCHEJBAL; Erwin ADAMS; Jan PREISLER; Zdeněk GLATZ a Ann VAN SCHEPDAEL

Vydání

45th International symposium on high performance liquid phase separations and related techniques - HPLC 2017, 2017

Další údaje

Typ výsledku

Konferenční abstrakt

Utajení

není předmětem státního či obchodního tajemství

Označené pro přenos do RIV

Klíčová slova česky

kapilární lektroforéza, magnetické mikročástice, IMER, hFMO3

Klíčová slova anglicky

capillary electrophoresis, magnetic microparticles, IMER, hFMO3

Příznaky

Mezinárodní význam

Změněno: 26. 1. 2018 08:32, prof. RNDr. Zdeněk Glatz, CSc.

Anotace

V originále

Capillary electrophoresis as an analytical technique has emerged in recent years in the field of enzyme kinetics and inhibition. This is mainly attributed to the lower amounts of enzyme and substrate that can be used for reaction. One of the main advantages of CE is its ability to use capillary for both reaction and separation also known as in-line capillary electrophoresis. Since the introduction of this technique, a number of enzymatic reactions were performed using in-line capillary electrophoresis. In-line capillary electrophoresis is known for its lower sample consumption and organic solvents are not required to stop the reaction. Immobilizing drug-metabolizing enzymes on insoluble supports on either magnetic microparticles or capillary to develop CE based IMERs (Immobilized microenzyme reactors) have been attractive for their ability to repeated use with relatively small sample volumes. In the following work, we demonstrate the effectiveness of a new 3D printed magnet holder to hold magnets against the capillary that enables capture of magnetic microparticles in a commercially available liquid based coolant system. We discuss the design as well as the method to capture magnetic microparticles capture inside the capillary. This setup was tested at higher temperature and pH suitable for performing enzymatic reactions without any leakage of magnetic microparticles. To demonstrate its applicability in CE-IMER development, human Flavin monooxygenase 3 (hFMO3) was immobilized on amino functionalized magnetic microparticles using glutaraldehyde. In general, CYP enzymes are the major contributors of oxidative xenobiotic metabolism. However, flavin monoxygenases (FMOs) also contribute in these kinds of reactions which is less well explored than CYP counterparts. The significance of FMO should not be overlooked because FMO and CYP have overlapping substrate specificities. However, they often yield distinct metabolites with potentially significant toxicological/pharmacological consequences. hFMO3 is known to have limited stability when compared to CYP counterparts making it a suitable enzyme to develop an IMER that allows its reusability. Clozapine was used as a model substrate. This setup could be used further to employ CE-IMERs on other drug metabolic enzymes in a commercially available liquid based capillary coolant system.

Návaznosti

GA16-06106S, projekt VaV

Název: Vysoce efektivní systém založený na kapilární elektroforéze pro screening inhibitorů beta-sekretázy jako terapeutického cíle pro Alzheimerovu chorobu (Akronym: Alzheimer)

Investor: Grantová agentura ČR, Vysoce efektivní systém založený na kapilární elektroforéze pro screening inhibitorů beta-sekretázy jako terapeutického cíle pro Alzheimerovu chorobu

MUNI/A/1278/2016, interní kód MU

Název: Podpora biochemického výzkumu v roce 2017

Investor: Masarykova univerzita, Podpora biochemického výzkumu v roce 2017, DO R. 2020_Kategorie A - Specifický výzkum - Studentské výzkumné projekty

Citovat

RAMANA, Pranov; Jan SCHEJBAL; Erwin ADAMS; Jan PREISLER; Zdeněk GLATZ a Ann VAN SCHEPDAEL. An improved design to capture magnetic microparticles for in-line CE in a liquid based capillary coolant system. In 45th International symposium on high performance liquid phase separations and related techniques - HPLC 2017. 2017.

@proceedings{1398358,
   author = {Ramana, Pranov and Schejbal, Jan and Adams, Erwin and Preisler, Jan and Glatz, Zdeněk and Van Schepdael, Ann},
   booktitle = {45th International symposium on high performance liquid phase separations and related techniques - HPLC 2017},
   keywords = {capillary electrophoresis, magnetic microparticles, IMER, hFMO3},
   title = {An improved design to capture magnetic microparticles for in-line CE in a liquid based capillary coolant system},
   year = {2017}
}

TY  - CONF
ID  - 1398358
AU  - Ramana, Pranov - Schejbal, Jan - Adams, Erwin - Preisler, Jan - Glatz, Zdeněk - Van Schepdael, Ann
PY  - 2017
TI  - An improved design to capture magnetic microparticles for in-line CE in a liquid based capillary coolant system
KW  - capillary electrophoresis, magnetic microparticles, IMER, hFMO3
N2  - Capillary electrophoresis as an analytical technique has emerged in recent years in the field of enzyme kinetics and inhibition. This is mainly attributed to the lower amounts of enzyme and substrate that can be used for reaction. One of the main advantages of CE is its ability to use capillary for both reaction and separation also known as in-line capillary electrophoresis. Since the introduction of this technique, a number of enzymatic reactions were performed using in-line capillary electrophoresis. In-line capillary electrophoresis is known for its lower sample consumption and organic solvents are not required to stop the reaction. Immobilizing drug-metabolizing enzymes on insoluble supports on either magnetic microparticles or capillary to develop CE based IMERs (Immobilized microenzyme reactors) have been attractive for their ability to repeated use with relatively small sample volumes. In the following work, we demonstrate the effectiveness of a new 3D printed magnet holder to hold magnets against the capillary that enables capture of magnetic microparticles in a commercially available liquid based coolant system. We discuss the design as well as the method to capture magnetic microparticles capture inside the capillary. This setup was tested at higher temperature and pH suitable for performing enzymatic reactions without any leakage of magnetic microparticles. To demonstrate its applicability in CE-IMER development, human Flavin monooxygenase 3 (hFMO3) was immobilized on amino functionalized magnetic microparticles using glutaraldehyde. In general, CYP enzymes are the major contributors of oxidative xenobiotic metabolism. However, flavin monoxygenases (FMOs) also contribute in these kinds of reactions which is less well explored than CYP counterparts. The significance of FMO should not be overlooked because FMO and CYP have overlapping substrate specificities. However, they often yield distinct metabolites with potentially significant toxicological/pharmacological consequences. hFMO3 is known to have limited stability when compared to CYP counterparts making it a suitable enzyme to develop an IMER that allows its reusability. Clozapine was used as a model substrate. This setup could be used further to employ CE-IMERs on other drug metabolic enzymes in a commercially available liquid based capillary coolant system.
ER  -

RAMANA, Pranov; Jan SCHEJBAL; Erwin ADAMS; Jan PREISLER; Zdeněk GLATZ a Ann VAN SCHEPDAEL. An improved design to capture magnetic microparticles for in-line CE in a liquid based capillary coolant system. In \textit{45th International symposium on high performance liquid phase separations and related techniques - HPLC 2017}. 2017.

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