I 0733r I 0734 acids. Methanol containing 0.001 M tetramethylammonium hydroxide (J. T. Baker, Phillipsburg, NJ) served as the modifier. A kit of PTH amino acids was obtained from Sigma Chemical Co. (St. Louis, MO).
3 Results and Discussion
Twenty-four PTH-amino acids have been rapidly and efficiently separated on a packed cyanopropyl column by gradient elution of supercritical C02 and tetramethylammonium hydroxide (TMA0H)-modified methanol. A typical optimized elution profileof the mixture is shown in Figure 1. Complete or partial resolution of 22 derivatives is observed with only valine (V) co-eluting with norleucine (0) and lysine (K) co-eluting with asparagine (N). A wide variety of stationary phases and modifiers were investigated with supercritical CO2 in attempting to achieve the separation of 24 relevant PTH-amino acids in less than 15 minutes. The Zorbax cyanopropyl phase and methanol containing 0.001 M TMAOH modifier gave the best results. Critical to achieving a rapid and efficient separation was the control of modifier flow into the C02. Modified COP varied from 0.1% MeOHlTMAOH by volume at the beginning of the separation to 33% MeOHlTMAOH by volume at the end of the run.
No modifier is required for elution of neutral PTH-AA's. The addition of TMAOH, however, plays a major role in the elution of acidic and basic PTH-AAs. Both peaktailing is minimized and the elution order of several peaks is altered upon incorporation of the ionpairing reagent into the mobile phase. Good run-to-run reproducibility could be obtained with the Zorbax cyanopropyl phase by equilibrating the column with COP that contains 0.1% by volume modifier. The role of TMAOH in the separation of these derivatives and the complete resolution ofall 24speciesare major goals of on-going work in our laboratories.
An Alternative Gas Sample introduction
System for Gas 10734 Chromatography P. A. Hilling (decreased) Scientific Glass Engineering,