Pharmacognostic specification and rotenone content in derris elliptica stems, macroscopic, microscopic and molecular identification of selected derris species in Thailand

Abstract

The genus Derris belonging to Fabaceae family are distributed along the equatorial zone in Africa and Asia. In Thailand, 16 species of Derris species can be found throughout the country. Derris elliptica (Hanglai-Dang), is well-known among fishermen and gardeners as fish poison and insecticide, especially the stem and root which contain isoflavonoid compound named rotenone. This study established the pharmacognostic specification of Derris elliptica dried stem crude drug and developed the quantitative analysis of rotenone content in D. elliptica stem using TLC-densitometry and image analysis. The physico-chemical parameters of D. elliptica stem included total ash, acid-insoluble ash, loss on drying and water contents that should not be more than 7, 1, 5 and 8 g/ 100g crude drug, respectively. The ethanol and water extractive values should not be less than 4 and 11 g/ 100 g crude drug, respectively. Microscopic characteristics (transverse section and powder) of D.elliptica stem were illustrated by hand drawing. Rotenone contents obtained from both quantitative TLC were 0.2870 ± 0.1242 and 0.2844 ±0.1209 g/100 g crude drug, respectively which was not statistically significantly different (P > 0.05). In vitro biological activities of the stem ethanolic extract and rotenone revealed the antioxidant, α-glucosidase inhibitory properties and brine shrimp toxicity. MTT assay revealed that the ethanolic extract and rotenone were marginal cytotoxic (LC50 > 20 µg/ml) against the cancer cells (MAD-231, HepG2, HT-29, CaCo-2). Microscopic leaf characteristics including leaf constant numbers of 8 Derris ( D. amonea, D. malaccensis, D. elliptica, D. reticulate, D. trifoliate, D. indica, D. solorioides ) were shown that stomatal number, stomata index, palisade ratio, epidermal cell area, trichome number could be used as the identification key for selected 8 species. DNA fingerprinting using ISSR 11, 12, 14, 17 and 35 as primers produced a total of distinct and reproducible 256 bands. The genetic relationship among selected Derris species presented the similarity index between 0.11 to 0.43. The information from this research can be applied for identification of D. elliptica stems in Thailand as well as providing the information of pharmacogenetic specification, including rotenone content in and in vitro biological properties of D. elliptica stem.