Various steviol glycosides which are used as sweetener were analyzed using Asahipak NH2P-40 2D, a polymer-based amino column, with LC/MS/MS detection. Steviolbioside and Rebaudioside B which have carboxyl group can be eluted under alkaline conditions. Furthermore, alkaline conditions promote the deprotonation of hydroxyl groups at the time of ionization. Therefore, alkaline conditions are suitable for high sensitive detection of substances with hydroxyl groups such as saccharides under the negative mode.
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Sample : 5 μL
50 ng/mL each
(in H₂O/CH₃CN=1/1)
Isosteviol
Stevioside
Rebaudioside C
Rebaudioside F
Rebaudioside A
Steviolbioside
Rebaudioside D
Rebaudioside B

Column       : Shodex Asahipak NH2P-40 2D (2.0 mm I.D. x 150 mm)
Eluent       : (A); 0.1 % NH3 aq./(B); CH3CN
               Isocratic ; (B %) 78 %
Flow rate    : 0.2 mL/min
Detector     : ESI-MS (SIM Negative)
Column temp. : 30 °C