Document Detail


Microwave-assisted high-throughput acid hydrolysis in silicon carbide microtiter platforms--a rapid and low volume sample preparation technique for total amino acid analysis in proteins and peptides.
MedLine Citation:
PMID:  21056423     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
An efficient microwave-assisted high-throughput protein hydrolysis protocol was developed utilizing strongly microwave absorbing silicon carbide-based microtiter platforms. The plates are equipped with 20 bore holes having the proper dimensions for holding standard screw-capped HPLC/GC vials. Due to the possibility of heating up to four heating platforms simultaneously (80 vials), parallel microwave-assisted acid hydrolyses can be performed under carefully controlled conditions significantly reducing the overall time required for protein hydrolysis and the subsequent evaporation step required for larger volumes of acid. An extensive optimization of the hydrolysis conditions has demonstrated that 5min irradiation at 160°C with 6N HCl leads to comparable results in terms of total and individual amino acid recovery as the traditional method requiring 24h heating at 110°C. Complete hydrolysis of several proteins and synthetic peptides was performed using 25μg of sample material and 100μL of 6N HCl in a dedicated low-volume HPLC/GC vial. Since the hydrolysis and subsequent analysis can be performed from the same vial, errors caused by sample transfer can be minimized. Control experiments have demonstrated that the observed rate enhancements are the result of a purely thermal/kinetic effect as a consequence of the considerable higher reaction temperatures.
Authors:
Markus Damm; Michael Holzer; Günther Radspieler; Gunther Marsche; C Oliver Kappe
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2010-10-21
Journal Detail:
Title:  Journal of chromatography. A     Volume:  1217     ISSN:  1873-3778     ISO Abbreviation:  J Chromatogr A     Publication Date:  2010 Dec 
Date Detail:
Created Date:  2010-11-24     Completed Date:  2011-03-11     Revised Date:  2012-10-01    
Medline Journal Info:
Nlm Unique ID:  9318488     Medline TA:  J Chromatogr A     Country:  Netherlands    
Other Details:
Languages:  eng     Pagination:  7826-32     Citation Subset:  IM    
Copyright Information:
Copyright © 2010 Elsevier B.V. All rights reserved.
Affiliation:
Christian Doppler Laboratory for Microwave Chemistry (CDLMC) and Institute of Chemistry, Karl-Franzens-University Graz, Heinrichstraße 28, A-8010 Graz, Austria.
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MeSH Terms
Descriptor/Qualifier:
Amino Acids / analysis*
Animals
Carbon Compounds, Inorganic / chemistry*
Cattle
Chromatography, Ion Exchange / methods*
High-Throughput Screening Assays / instrumentation*,  methods*
Hot Temperature
Hydrochloric Acid
Hydrolysis
Microwaves*
Miniaturization
Peptides / chemistry
Protein Processing, Post-Translational
Proteins / chemistry*
Serum Albumin, Bovine / chemistry
Silicon Compounds / chemistry*
Grant Support
ID/Acronym/Agency:
P 21004-B02//Austrian Science Fund FWF
Chemical
Reg. No./Substance:
0/Amino Acids; 0/Carbon Compounds, Inorganic; 0/Peptides; 0/Proteins; 0/Serum Albumin, Bovine; 0/Silicon Compounds; 409-21-2/silicon carbide; 7647-01-0/Hydrochloric Acid

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


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