Tryptophane separation from cystine

Figure 4 HPLC separation of 17 PTC AAs (Pico-Tag column 150 x 3.9mm, eluent A 0.14moll NaAc, pH 6.4, containing 0.5ml TEAI B 60% acetonitrile in water flow rate 1 ml minpeaks 1 = aspartic, 2=glutamic acids, 3 = serine, 4=glycine, 5=his-istidine, 6 = arginine, 7=threonine, 8 = alanine, 9 = proline, 10 = ammonia, l1=tyrosine, 12=valine, 13=methionine, 14=cystine, 15 = isoleucine, 16 = r>leucine, 17 = phenylalanine, 18=tryptophan. (Reproduced with permission from Bidlingmayer BA ef a/. (1984) Journal of Chromatography 336 Elsevier.)...

Since biological value is dependent primarily upon essential amino acid constitution, it would seem logical to assess the nutritive value of a protein by determining its essential amino acid constitution and then comparing this with the known amino acid requirements of a particular class of animal. Application of modern chromatographic techniques coupled with automated procedures allows relatively quick and convenient resolution of mixtures of amino acids. However, the acid hydrolysis used to produce such mixtures from protein destroys practically all the tryptophan and a considerable proportion of the cystine and methionine. Tryptophan has to be released by a separate alkaline hydrolysis, and cystine and methionine have to be oxidised to cysteic acid and methionine sulphone to ensure their quantitative recovery. Losses of amino acids and the production of artefacts, which are greater with foods of high carbohydrate content, are reduced if the hydrolysis is carried out in vacuo. Evaluations of proteins in terms of each individual amino acid would be laborious and inconvenient, and several attempts have been made to state the results of amino acid analyses in a more useful and convenient form. 

Mondino and Bongiovanni 262) observed that cystine, tryptophan and to a lesser extent threonine and serine were degraded when a mixture of all the common amino acids found in proteins was hydrolyzed in 6N HCl by an open-flask technique and analyzed on an automatic amino acid analyzer. More recently, Gruen 157) carefully studied the separate effect of each of the other commonly occuring amino acids on the recovery of tryptophan, using the standard procedure for amino acid analysis, whereby tryptophan is eluted from the short column along with the basic amino acids as a well resolved peak appearing before lysine 135, 369). 

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