Automated Method, Based on Micro-Sequential Injection, for the Study of Enzyme Kinetics and Inhibition
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- CHEN Yan
- Department of Chemistry, University of Washington
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- CARROLL Andrea D.
- Department of Chemistry, University of Washington
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- SCAMPAVIA Louis
- Department of Chemistry, University of Washington
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- RUZICKA Jaromir
- Department of Chemistry, University of Washington
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Abstract
A micro-reactor system with continual spectrophotometric detection has been operated in Sequential Injection lab-on-valve (SI-LOV) mode and applied to enzyme kinetics and inhibition studies, using acetylcholinesterase (AChE) and angiotensin-converting enzyme (ACE) as model systems. With the advantages of automation, real-time kinetic measurement, and thorough mixing, the SI-LOV micro-reactor system allows for the monitoring of initial reaction rates and determination of reactant concentrations in the reaction mixture, both of which are essential for the determination of kinetic constants for enzymes and inhibitors. Enzyme, substrate, and inhibitor are precisely metered by the syringe pump and delivered to a stirred micro-reactor, followed by a reference scan that establishes the baseline for the following reaction rate measurement. Michaelis constants (Km) for AChE and ACE were determined to be 0.16 mM and 0.30 mM, respectively, which are consistent with literature values. The type of inhibition (competitive, uncompetitive, or mixed), the dissociation constants for the inhibitors, and the inhibitor dose-response curves were also determined.
Journal
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- Analytical Sciences
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Analytical Sciences 22 (1), 9-14, 2006
The Japan Society for Analytical Chemistry
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Details 詳細情報について
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- CRID
- 1390001204255781888
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- NII Article ID
- 130004441136
- 10017165749
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- NII Book ID
- AA10500785
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- COI
- 1:CAS:528:DC%2BD28XntVyjtw%3D%3D
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- ISSN
- 13482246
- 09106340
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- NDL BIB ID
- 7763281
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- Text Lang
- en
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- Data Source
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- JaLC
- NDL
- Crossref
- CiNii Articles
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- Abstract License Flag
- Disallowed