Amino acid analysis - technique and history
As E. Brand discovered in 1946, if we used specific procedures to determine each amino acid, the analysis of a protein would be extremely tedious and exceptionally time consuming.
Today's techniques, which are mainly chromatographic, owe much to A.J.D Martin and R.L.M Synge's research, which were developed to today's advanced state by S. Moore and W. H. Stein together with D.H. Spackman back in 1958.
These techniques form an integral part of almost all biochemical structure investigations. Amino acids give evidence as to the purity of proteins.
The mean of the molar ratios of all accurately measurable amino acids in the acid hydrolysate was used to calculate the concentration of the protein or peptide.
Accuracy of determination is of most importance in studying newly isolated proteins. When calculation is the correlation of its molecular weight with integral numbers of amino acid residues which are present in small molar properties.
Investigations not involving large peptides or proteins, sophisticated techniques may not always be required.
For example:
When studying bee venom, particularly phospolipase obtained from the venom, it was found that for small peptides, qualitative analyses by the method of dansylation* gave satifactory results.
The residue of amino acids after NH2 analysis was completely dansylated and the resultant mixture of the dansyl amino acids separated by thin layer chromatography.
Composition of the mixture was assessed by visual comparison of the intensity of flourescence of the separated spots.
* Dansyl is a contraction for dimethylaminorapthalenesulfonyl.
The example above is an extract from the Introduction of 'amino acid determination' by S. Blackburn.
|
