Development of Identification and Purity Tests for Bamboo Leaf by Using TLC and the Identification of the Indicator Components

Abstract

Bamboo Leaf, the leaf of Phyllostachys nigra Munro var. henonis Stapf ex Rendle, P. bambusoides Siebold et Zuccarini, Bambusa textilis McClure or B. emeiensis L. C. Chia et H. L. Fung, is used as a crude drug in the Kampo medicine Chikuyo-sekko-to. Lophatherum Herb, which is derived from the leaves of Lophatherum gracile Brongniart, is a crude drug similar to bamboo leaf. There has been a concern over the misuse of these crude drugs. Recently, because Chikuyo-sekko-to has been approved by the Ministry of Health, Labour and Welfare as an OTC Kampo formula, we investigated the standardization of bamboo leaf and decided to list it in the non-JP crude drug standards 2015. In the process of the standardization, we designed identification and purity tests for the crude drug by using TLC. In addition, each indicator spot of the tests was isolated from bamboo leaf and Lophatherum Herb using repeated column chromatography and HPLC. The chemical structures of these spots were elucidated as p-coumaric acid (1) and trans-aconitic acid (2) based on m.p. and spectroscopic data including 1D- and 2D-NMR and MS.

The established TLC conditions were chromatographic support, silica gel; developing solvent, EtOAc/hexane/acetic acid (20/20/1) for the identification test, EtOAc/H₂O/formic acid (10/1/1) for the purity test; developing length, 7 cm; visualization, 4-methoxybenzaldehyde-sulphuric acid reagent for the identification test, UV (254 nm) for the purity test; R_f values, 0.4 (p-coumaric acid; 1) for the identification test, 0.6 (trans-aconitic acid; 2) for the purity test.