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Complete Analysis of a Biologically Active Tetrapeptide: A Project Utilizing Thin-Layer Chromatography and Tandem Quadrupole Mass Spectrometry
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Abstract
The biologically active tetrapeptide d-Ala-Gly-l-Phe-d-Leu ([des-Tyr1-d-Ala2-d-Leu5]enkephalin) was analyzed for its amino acid content and stereochemistry by normal and reversed-phase thin-layer chromatography (TLC), and its sequence was determined by tandem quadrupole mass spectrometry. The project involved sequential N-dansylation of a portion of the tetrapeptide, hydrolysis, isolation, and identification of the N-terminal amino acid as dansyl-alanine by comparison with standards using normal-phase TLC. A second portion of the tetrapeptide was hydrolyzed and the resulting four free amino acids were converted to their corresponding dansyl derivatives and purified by preparative normal-phase TLC. The three dansyl amino acids not identified previously were identified by TLC. The stereochemistry of each was determined by comparison with dansyl-dl-amino acid standards using reversed-phase TLC in the presence of ß-cyclodextrin, a chiral mobile phase additive. Finally, the correct amino acid sequence was determined by tandem quadrupole mass spectrometry. This project gives students valuable experience in microscale synthesis, both normal and reversed-phase TLC, stereochemical analysis, and mass spectrometry.
Keywords (Audience):
Upper-Division UndergraduateKeywords (Domain):
Laboratory InstructionKeywords (Feature):
The Microscale LaboratoryKeywords (Pedagogy):
Hands-On Learning / ManipulativesKeywords (Subject):
Proteins / PeptidesCiting Articles
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This article has been cited by 1 ACS Journal articles (1 most recent appear below).
Using the Mini-Session Course Format To Train Students in the Practical Aspects of Modern Mass Spectrometry
Dale A. Rosado, Jr., Tina S. Masterson, and Douglas S. MastersonJournal of Chemical Education2011 88 (2), 178-183Using the Mini-Session Course Format To Train Students in the Practical Aspects of Modern Mass Spectrometry
Dale A. Rosado, Jr., Tina S. Masterson, and Douglas S. MastersonJournal of Chemical Education2011 88 (2), 178-183Mass spectrometry (MS) has been gaining in popularity in recent years owing in large part to the development of soft-ionization techniques such as matrix-assisted laser desorption ionization (MALDI) and electrospray ionization (ESI). These soft-ionization ...
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- Received: August 03, 2009
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