Characterization of Marker Compounds in Ziziphi Fructus Using TLC

Abstract

Ziziphi Fructus (the fruit of Ziziphus jujuba Miller var. inermis Rehder; Japanese name “Taiso”), listed in the 18th edition of the Japanese Pharmacopoeia (JP18), is widely used in Kampo (Japanese traditional) medicine. However, a specific identification test for Ziziphi Fructus was not prescribed in JP18. Hence, in this study, we establish a TLC-based identification test for quality assurance of the crude drug Ziziphi Fructus. The HPLC experiments identified 3-O-cis-p-coumaroylalphitolic acid and 3-O-trans-p-coumaroylaliphitolic acid as suitable indicators. After developing the samples to a distance of 10 cm using ethyl acetate (EtOAc)/n-hexane/acetic acid (AcOH)/H₂O (60 : 40 : 2 : 1) as the TLC-developing solvent and UV light at 254 nm, two spots with an Rf value of approximately 0.4–0.6 were detected. Additionally, based on the TLC-based experiments, alphitolic and maslinic acids emerged as potential indicator components. Treatment of samples with EtOAc/n-hexane/AcOH (60 : 40 : 1) serving as the developing solvent, followed by exposure to dilute sulfuric acid solution, heat, and UV light at 365 nm, revealed two clear spots with Rf values of approximately 0.2–0.3. These findings potentially serve as a basis for the development of an identification test to identify Ziziphi Fructus.