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  • Amino acid analysis: Total and Free amino acid analysis

AltaBioscience offers two main types of analysis, ‘total’ and ‘free’ amino acid analysis for a diverse range of samples.
Types of analysis : for amino acids bound in proteins or amino acids existing as free entities.

 

Total amino acid analysis determines the amino acids which are bound in proteins as well as those amino acids that may be present as individual unbound units.

Free amino acid analysis determines the amount of each unbound individual amino acid i.e. not bound in a protein.

Some application may require both analyses to determine the difference in bound and unbound amino acids.

Total Amino Acid Analysis

Where the amino acid composition of a protein or peptide is of interest, or if the total protein concentration of a sample is required, then this is the method of choice. The proteins and/or peptides within the sample are broken down by acid hydrolysis into their individual amino acids, the extract is therefore a sum of amino acids which were free in solution and those which were previously incorporated into proteins before hydrolysis. The amino acids within the hydrolysate are separated using a sodium citrate buffer system prior to detection.

This method measures 18 of the 20 standard amino acids.

Figure 1. Example chromatogram of an amino acid standard separated on the protein hydrolysate (sodium) system

Tryptophan and cystine are usually lost during acid hydrolysis, however, estimates can be given for these amino acids when the sequence is known. For specific quantification of tryptophan or cystine, separate additional testing methods are available.

Free Amino Acid Analysis

For the analysis of samples such as physiological fluids including serum or cell growth media, the amino acids incorporated into proteins or peptides are not of interest, and it is the concentration of the individual unbound amino acids which is required. Therefore the protein and interfering compounds are removed, leaving the amino acids free in solution. The free amino acids are then separated using a lithium citrate buffer system prior to detection. This high resolution method quantifies up to 40 components, including many non-standard amino acids.

Figure 2.   Example chromatogram of an amino acid standard separated on the physiological (lithium) system

For more information, please contact us

E:  info@altabioscience.com T:  +44 (0)1527 584495

    Author:  Sarah Hawker, Senior Analyst

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