Studies on KAKI CALYX: A Method for Quantitative Analysis of Betulinic Acid, Ursolic Acid and Oleanolic Acid Using HPLC and Application of the Method to Market and Hand-picked Samples

Abstract

KAKI CALYX is defined in the Japanese standards for non-Pharmacopoeial crude drugs 2018 (non-JPS 2018) as the persistent calyx of mature fruit of Diospyros kaki Thunberg　(Ebenaceae), and is used for Kampo formulae such as Shiteito and Choukoushiteito that are prescribed for continuous hiccups. In order to help better understand and facilitate quality control of KAKI CALYX, HPLC methods for quantitative analyses of three triterpenoids, i.e. betulinic, ursolic and oleanolic acids in KAKI CALYX were elaborated. The calyces from various types of cultivated persimmon trees were collected and analyzed to clarify the difference affected by harvesting periods and sweet/astringent types of fruits. Market samples were also investigated and compared. As a result, betulinic acid content was larger in sweet persimmons than in astringent persimmons (p<0.05). Ursolic and oleanolic acids contents of cultivated persimmons were significantly higher than those of market samples (p<0.01).