Capillary electrophoresis method for determination of D-serine and its application for monitoring of serine racemase activity
Serine racemase (SR) is an enzyme responsible for the biosynthesis of D‐serine, the coagonist of the N‐methyl‐D‐aspartate receptor, in the brain. Therefore, it has been suggested as a possible therapeutic target for the treatment of various neurodegenerative diseases. To develop a potent inhibitor o...
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Veröffentlicht in: | Electrophoresis 2006-07, Vol.27 (13), p.2558-2566 |
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Sprache: | eng |
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Zusammenfassung: | Serine racemase (SR) is an enzyme responsible for the biosynthesis of D‐serine, the coagonist of the N‐methyl‐D‐aspartate receptor, in the brain. Therefore, it has been suggested as a possible therapeutic target for the treatment of various neurodegenerative diseases. To develop a potent inhibitor of SR, a simple, sensitive, fast, and robust assay is needed. In this paper, a new CE method for the determination of D‐serine is described. Serine enantiomers are resolved in the form of o‐phthaldialdehyde (OPA)/2‐mercaptoethanol (2‐ME) derivatives in an alkaline BGE composed of 50 mM sodium tetraborate, pH 9.7, and containing 40 mM 2‐hydroxypropyl‐γ‐CD as a chiral selector. The problem of time‐limited stability of OPA/2‐ME derivatives has been overcome by employing in‐capillary derivatization of the sample, i.e., the derivatization reaction was carried out directly in the separation capillary in the first phase of the CE run. UV‐absorption detection at 230 nm allowed concentration detection limit of 3 μM. Baseline resolution of D‐ and L‐serine derivatives was achieved in less than 10 min. This fact, together with the simple sample pretreatment, allowed application of the method to medium‐throughput monitoring of SR activity, such as the screening of potential SR inhibitors. A good agreement was achieved between the developed CE method and the previously established HPLC method for determination of the inhibition constant, Ki, of a new SR inhibitor, L‐erythro‐3‐hydroxyaspartate. |
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ISSN: | 0173-0835 1522-2683 |
DOI: | 10.1002/elps.200500946 |