Document Detail


Determination of tryptophan as the reduced derivative by acid hydrolysis and chromatography.
MedLine Citation:
PMID:  6528998     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
A new procedure for the analyses of tryptophan and the total amino acid composition of proteins was based on the observations that pyridine borane reduces tryptophan in trifluoroacetic acid, while other amino acids remain intact [M. Kurata, Y. Kikugawa, T. Kuwae, I. Koyama, and T. Takagi (1980) Chem. Pharm. Bull. 28, 2274-2275; W.S.D. Wong, D.T. Osuga, and R.E. Feeney (1984) Anal. Biochem. 139, 58-67]. Concentrated HCl was used instead of trifluoroacetic acid for analytical purposes. The products were stable to hydrolysis in 6 N HCl, and the reduction did not interfere with hydrolysis and subsequent analyses. Quantitative recovery was achieved with most proteins when they were subjected to acid reduction in ice-cooled concentrated HCl with two incremental additions of pyridine borane. The reaction was terminated after 10 min by dilution with an equal volume of H2O, vacuum sealing, and hydrolyzing at 110 degrees C for 22 h. The yields of the expected values for cytochrome c, catalase, bovine serum albumin, subtilisin BPN', trypsin, chymotrypsin, beta-lactoglobulin, lysozyme, and pepsin were obtained. Ovotransferrin and ovalbumin, however, yielded values for tryptophan lower than literature values. With two different ion-exchange methods, the recoveries of all other amino acids were comparable to those obtained by acid hydrolysis with 6 N HCl. Since the same hydrolysate can be analyzed for both tryptophan and all the other amino acids, the procedure is a more convenient method than those requiring separate determinations. Initial results indicate that the method may be applied to high-performance liquid chromatographic procedures with adaptations of the protocols if necessary.
Authors:
W S Wong; D T Osuga; T S Burcham; R E Feeney
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Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Analytical biochemistry     Volume:  143     ISSN:  0003-2697     ISO Abbreviation:  Anal. Biochem.     Publication Date:  1984 Nov 
Date Detail:
Created Date:  1985-03-27     Completed Date:  1985-03-27     Revised Date:  2008-11-21    
Medline Journal Info:
Nlm Unique ID:  0370535     Medline TA:  Anal Biochem     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  62-70     Citation Subset:  IM    
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MeSH Terms
Descriptor/Qualifier:
Amino Acids / analysis*
Chemical Phenomena
Chemistry
Chromatography, High Pressure Liquid
Chromatography, Thin Layer
Hydrolysis
Oxidation-Reduction
Proteins / analysis*
Sulfhydryl Compounds
Tryptophan / analogs & derivatives,  analysis*
Grant Support
ID/Acronym/Agency:
GM 23817/GM/NIGMS NIH HHS; HL 18619/HL/NHLBI NIH HHS
Chemical
Reg. No./Substance:
0/Amino Acids; 0/Proteins; 0/Sulfhydryl Compounds; 73-22-3/Tryptophan; 7536-97-2/2,3-dihydrotryptophan

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


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