Gradient separation of PTH-amino acids employing supercritical CO2 and modifiers.
Twenty-four PTH-amino acids are rapidly and efficiently separated on a packed cyanopropyl Zorbax column by gradient elution of supercritical CO2 and tetramethylammonium hydroxide-modified methanol. Complete or partial resolution of 22 derivatives is observed with only valine coeluting with norleucine and lysine coeluting with asparagine. A wide variety of stationary phases and modifiers are investigated with supercritical CO2 in attempting to achieve the separation in less than 15 min. Critical to achieving a rapid and efficient separation is the control of modifier flow into the CO2. No modifier is required for elution of neutral PTH-amino acids. The addition of base plays a major role in the elution of acidic and basic PTH-amino acids. Peak tailing is minimized and the elution order of several peaks is altered upon incorporation of reagent into the mobile phase.
Berger
T A
TA
Hewlett-Packard, Inc., Avondale Division, Pennsylvania 19311-0900.
Deye
J F
JF
Ashraf-Khorassani
M
M
Taylor
L T
LT
D000596:Amino Acids / Q000032:analysis*; D002845:Chromatography / Q000379:methods; D007202:Indicators and Reagents; D010669:Phenylthiohydantoin
D000596:Amino Acids; D007202:Indicators and Reagents; D010669:Phenylthiohydantoin