Frans_M_Everaerts_Isotachophoresis_378342.pdf

336 AMINO ACIDS, PEPTIDES AND PROTEINS
13.3.2. Experimental
The operational system used was that specified in Table 13.3. L(+)-Alanine was used as
the terminating electrolyte, adjusted to pH 9.8 by addition of barium hydroxide. The
leading ion, 5-bromo-2,4-dihydroxybenzoic acid (0.004 M), was adjusted to pH 9.05 by
addition of L-lysine. The current was stabilized at 100 MA, and the time of analysis was
approximately 8 min. About 0.01 mole of glutathione (Merck, Darmstadt, G.F.R.),
glycylglycine hydrochloride, glycylglycylglycylglycine and D-leucyl-L-tyrosine (Nutritional
Biochemicals, Cleveland, Ohio, U.S.A.) was injected. The isotachopherogram of the
analysis is shown in Fig.13.16. One should note the chloride*, which is more mobile than
the 5-bromo-2,4-dihydroxybenzoic acid, which has passed the first separation boundary.
Because the chloride is coming from the cathode compartment, it is not a pH disturbance,
which may originate from the semi-permeable membrane, especially as the zone is
reasonably well defined.
It can clearly be seen in the linear traces of both the conductivity detector and the
W absorption detector that the concentration of the leading electrolyte is not changed
after the passage of the mobile chloride ion.
REFERENCES
1 A. Niederwasser and H. Curtius, Z. Klin. Chem. Klin. Biochem., 5 (1969) 4G4.
2 D.H. Spackman, W.H. Stein and S. Moore, Anal. Chem, 30 (1958) 90.
3 F.M. Everaerts and A.J.M. van der Put, J. Chromatogr., 52 (1970) 415.
4 A. Kopwillem, J. Chromatogr., 82 (1973) 407.
5 A.J. de Kok, Graduation Rep., University of Technology, Eindhoven, 1975.
6 F.E.P. Mikkers, Graduation Rep., University of Technology, Eindhoven, 1974.
*The chloride is derived from the sample component glycylglycine hydrochloride.