Document Detail


Analysis of femtomole concentrations of alpha-ketoisocaproic acid in brain tissue by precolumn fluorescence derivatization with 4,5-dimethoxy-1,2-diaminobenzene.
MedLine Citation:
PMID:  8238877     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
An extremely sensitive fluorimetric HPLC method used to determine in vivo cerebral alpha-ketoisocaproic acid (KIC) concentration with less than 2 g of tissue (e.g., a single rat brain, approximately 1.8 g) is reported. Removal of unwanted lipids and amino acids and isolation of alpha-ketoacids in an optimal derivatization buffer are achieved by C18 solid-phase extraction of the acid-soluble fraction of brain tissue. Quantitation of KIC to the femtomole level is accomplished by reversed-phase HPLC using 4,5-dimethoxy-1,2-diaminobenzene precolumn fluorescence derivatization and on-line fluorescence detection. These techniques are applicable to femtomole quantitation of other alpha-ketoacids in various tissue and blood matrices. Combination of this fluorimetric method with simple nonchromatographic procedures to measure in vivo cerebral [1-14C]alpha-ketoisocaproic acid radioactivities in tissue provides estimates of specific activities.
Authors:
R E Keen; C H Nissenson; J R Barrio
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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.; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Analytical biochemistry     Volume:  213     ISSN:  0003-2697     ISO Abbreviation:  Anal. Biochem.     Publication Date:  1993 Aug 
Date Detail:
Created Date:  1993-12-03     Completed Date:  1993-12-03     Revised Date:  2007-11-14    
Medline Journal Info:
Nlm Unique ID:  0370535     Medline TA:  Anal Biochem     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  23-8     Citation Subset:  IM    
Affiliation:
Department of Pharmacology, UCLA School of Medicine.
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MeSH Terms
Descriptor/Qualifier:
Animals
Brain Chemistry*
Carbon Radioisotopes / diagnostic use
Chromatography, High Pressure Liquid / methods
Fluorescent Dyes
Fluorometry
Keto Acids / analysis*,  blood,  isolation & purification
Leucine / metabolism
Male
Microchemistry
Phenylenediamines / chemistry*
Rats
Rats, Sprague-Dawley
Reproducibility of Results
Sensitivity and Specificity
Swine
Grant Support
ID/Acronym/Agency:
NIMH R01-MH-37916/MH/NIMH NIH HHS; P01-NS-15654/NS/NINDS NIH HHS
Chemical
Reg. No./Substance:
0/Carbon Radioisotopes; 0/Fluorescent Dyes; 0/Keto Acids; 0/Phenylenediamines; 27841-33-4/1,2-diamino-4,5-dimethoxybenzene; 61-90-5/Leucine; 816-66-0/alpha-ketoisocaproic acid

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


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