Quantitative analysis of plaunotol in the leaves and tissue cultures of Croton sublyratus Kurz

Abstract

A simple, rapid and accurate method of TLC-densitometry was developed to quantitate plaunotol, an antipeptic ulcer substance, in the leaves and tissue cultures of Croton sublyratus Kurz (Euphorbiaceae). The developed method could detect plaunotol at the amount of as low as 50 ng and its standard curve showed linearity up to 1.0 μg of the compound. In the evaluation of plaunotol content in the leaves of Thai C. sublyratus , 68 samples of the leaves were collected from various sources, including Rayong, Prachuap Khiri Khan, Nakhon Pathom, Bangkok and Chachoengsao. The analysis revealed that C. sublyratus leaves contained plaunotol in the range of 0.139-0.789% (w/w) dry weight with the majority (39.71%) falling between 0.3-0.4%. This is the first report on the quantitative analysis of leaf plaunotol in C. sublyratus. Since the plants from the same geographical location also showed high variation of leaf plaunotol content, it was suggested that the genetic background of each individual plant is the major factor contributing to the appearent plaunotol content. Screening for high plaunotol-containing plant was, therefore, a crucial step for quality improvement of this plant. In addition, the developed TLC-densitometry was also used for the detection of plant materials which potentially contain plaunotol such as the leaves of other Croton species and callus and cell cultures of C. sublyratus. However, no detectable plaunotol was found in these materials. Starting with high plaunotol-containing C. sublyratus plant, we could regenerate the plant via callus culture. Multiple shoot formation was observed when the callus was cultured on MS medium containing 10 g/l sucrose, 1.0 mg/l BA, 0.05 mg/l GA₃ and 0.2% (wlv) gellan gum. The efficiency of the adventitious shoot formation was 87.5% with the number 13 shoots per callus.