ERIC Number: EJ759209
Record Type: Journal
Publication Date: 2007
HPLC Determination of Caffeine and Paraxanthine in Urine: An Assay for Cytochrome P450 1A2 Activity
Furge, Laura Lowe; Fletke, Kyle J.
Biochemistry and Molecular Biology Education, v35 n2 p138-144 Mar-Apr 2007
Cytochrome P450 enzymes are a family of heme-containing proteins located throughout the body with roles in metabolism of endogenous and exogenous compounds. Among exogenous compounds, clinically relevant pharmaceutical agents are nearly all metabolized by P450 enzymes. However, the activity of the different cytochrome P450 enzymes varies among individuals and, therefore, so does drug efficacy as well as susceptibility to side effects and toxicity. Thus, assessing P450 activity is of great interest in drug development and clinical pharmacology. This study investigates the phenotyping of a single P450 activity by analyzing urine samples using isocratic reverse-phase HPLC. Specifically, the activity of human P450 1A2, which converts caffeine into paraxanthine, can be investigated by measuring the change in caffeine and paraxanthine concentrations in urine over time following a single dose of caffeine. There is an observable relationship between caffeine intake and paraxanthine formation that varies among individuals. This laboratory exercise provides a means for simple assessment of P450 1A2 metabolic activity using an HPLC method without additional extraction or purification steps and introduces students to the complexities of individualized medicine as well as the basic biochemical techniques of sample preparation and quantitative HPLC. Furthermore, students may design and test their own hypothesis using these methods.
Descriptors: Pharmacology, Drug Education, Laboratory Experiments, Biochemistry, Hypothesis Testing, Individual Differences, Undergraduate Students
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Publication Type: Guides - Classroom - Teacher; Journal Articles; Reports - Research
Education Level: Higher Education; Postsecondary Education
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