Document Detail


Amino acid analysis by reverse-phase high-performance liquid chromatography: precolumn derivatization with phenylisothiocyanate.
MedLine Citation:
PMID:  6711815     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Methods for the quantitative derivatization of amino acids with phenylisothiocyanate and for the separation and quantitation of the resulting phenylthiocarbamyl derivatives by reverse-phase high-performance liquid chromatography are described. Phenylthiocarbamylation of amino acids proceeds smoothly in 5 to 10 min at room temperature. Coupling solvents, reagent, and some byproducts are removed by rotary evaporation under high vacuum, and the phenylthiocarbamyl derivatives are dissolved in 0.05 M ammonium acetate, pH 6.8, for injection onto the octyl or octadecylsilyl reverse-phase column. Columns are equilibrated with the same solvent and the effluent stream is monitored continuously at 254 nm for detection of the amino acid derivatives. Elution of all of the phenylthiocarbamyl amino acids is achieved in about 30 min utilizing gradients of increasing concentrations of ammonium acetate and acetonitrile or methanol. This approach to amino acid analysis offers select advantages, both with respect to methods which employ reverse-phase separation of prederivatized samples and to the classical ion-exchange procedure. All amino acids, including proline, are converted quantitatively to phenylthiocarbamyl compounds and these are stable enough to eliminate any need for in-line derivatization. Furthermore, results comparable in sensitivity and precision to those obtained by state-of-the-art ion-exchange analyzers may be generated with equipment that need not be dedicated to a single application.
Authors:
R L Heinrikson; S C Meredith
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S.    
Journal Detail:
Title:  Analytical biochemistry     Volume:  136     ISSN:  0003-2697     ISO Abbreviation:  Anal. Biochem.     Publication Date:  1984 Jan 
Date Detail:
Created Date:  1984-05-11     Completed Date:  1984-05-11     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  0370535     Medline TA:  Anal Biochem     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  65-74     Citation Subset:  IM    
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MeSH Terms
Descriptor/Qualifier:
Amino Acids / analysis*
Animals
Cattle
Chromatography, High Pressure Liquid / methods
Isothiocyanates
Muramidase
Peptides
Ribonuclease, Pancreatic
Thiocyanates*
Chemical
Reg. No./Substance:
0/Amino Acids; 0/Isothiocyanates; 0/Peptides; 0/Thiocyanates; 103-72-0/phenylisothiocyanate; EC 3.1.27.5/Ribonuclease, Pancreatic; EC 3.2.1.17/Muramidase

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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