5] DISCUSSION ON PROFESSOR SORM'S PAPER 91 



With compartments of 10 ml. capacity each, up to 1 g. of material may 

 be fractionated. 



There are two further techniques which I would like to mention: one for 

 the quantitative determination of amino acids, and the second for carrying 

 out dinitrophenylation analysis. 



The most precise method for the quantitative analysis of amino acid 

 mixtures today is, of course, the technique of Moore and Stein; however, 

 for the routine quantitative determination of the amino acid composition 

 of peptide fractions we have worked out four years ago a simpler pro- 

 cedure.^ This required, first, an adaptation of the paper-chromatographic 

 technique which would permit the separation of as many of the amino acids 

 as possible in a single direction, so that a number of samples might be 

 analysed in parallel (Fig. 3). 



This problem was essentially solved by repeated development of the 

 chromatogram, with drying in between each solvent run. After detection, 

 the chromatogram is photographed on cinefilm and the blackening of the 

 film is measured using a precision spectrographic microphotometer. A blank 

 reading for any part of the paper is readily obtained from the photographic 

 record. Since the photographs are taken by reflected light the inhomogeneity 

 of the paper fibres is of no consequence. The same technique can be used 

 for example for the direct quantitative determination of glutamic, aspartic, 

 or cysteic acid, histidine, etc., after paper-electrophoretic separation, or, in 

 general, for the estimation of any compound giving a coloured spot. In 

 another laboratory of our Institute this technique is for instance being used 

 for the determination of elementary sulphur. The areas under the curves 

 obtained — which resemble those obtained by the Moore and Stein method 

 — are converted graphically into equivalents of each particular compound. 



Finally, a few words about the dinitrophenylation technique: With lower 

 peptides on a micro scale, i.e. with amounts of about 5-30 fj-g, we use various 

 modifications of the following procedure :2 



The peptide is dinitrophenylated in the microflask and then selectively 

 retained on a microcolumn of Zerolite H, consisting of about 20 yug. of the 



