Simultaneous Determination of Catalpol, Aucubin, and Geniposidic Acid in Different Developmental Stages of Rehmannia glutinosa Leaves by High Performance Liquid Chromatography

Table 1

Optimization of HPLC-UV conditions for three analytes.

Chromatographic factor	Compared parameter	Optimized conditions	Main advantages
Column	Phenomenex Hydro_RP (4.6 mm × 250 mm, 4 μm), Kinetex-C18; (4.6 mm × 100 mm, 2.6 μm)	Kinetex-C18	Improved column efficiency, speed, separation, and sensitivity
	203~210 nm for CA and AU, 235~240 nm for GPA	210 nm for CA and AU; 240 nm for GPA	No difference in 203~210 nm; less miscellaneous peaks and stabler baseline at λ 240 nm
Mobile phase	Methanol-H2O, ACN-H2O	ACN-H2O	Better peak shape and separation, stabler baseline
	Adding or not adding of formic acid in H2O	(0.1% formic acid)	Better peak shape and separation for GPA; no obvious influence on CA and AU
	Linear gradient or isocratic	Isocratic	Smooth baseline, better separation, less miscellaneous peaks
ACN : H2O (0.1% FA)	1 : 99, 2 : 98, 3 : 97, 5 : 95, 7 : 93, 10 : 90	5 : 95	Shorter elution time; however, CA overlapped with solvent peak when ACN was >5%; longer elution time for AU and GPA when ACN was less than 5%