Positive GABA(A) receptor modulators from Acorus calamus and structural analysis of (+)-dioxosarcoguaiacol by 1D and 2D NMR and molecular modeling.

In a two-microelectrode voltage clamp with Xenopus laevis oocytes, a petroleum ether extract of Acorus calamus rhizomes enhanced the GABA-induced chloride current through GABA(A) receptors of the α1ß2γ(2S) subtype by 277% ± 9.7% (100 µg/mL). ß-Asarone (1), (+)-dioxosarcoguaiacol (2), (+)-shyobunone (3), and (+)-preisocalamenediol (4) were subsequently identified as main active principles through HPLC-based activity profiling and targeted isolation. The compounds induced maximum potentiation of the chloride current ranging from 588% ± 126% (EC50: 65.3 ± 21.6 µM) (2) to 1200% ± 163% (EC(50): 171.5 ± 34.6 µM) (1), whereas (-)-isoshyobunone (5) and (-)-acorenone (6) exhibited weak GABA(A) modulating properties (5: 164% ± 42.9%; EC50: 109.4 ± 46.6 µM and 6: 241% ± 23.1%; EC50: 34.0 ± 6.7 µM). The relative configuration of 2 was established as 4R*8S*10R* by NOESY experiments and conformational analysis.

HPLC-based activity profiling of Angelica pubescens roots for new positive GABAA receptor modulators in Xenopus oocytes.

A petroleum ether extract of the traditional Chinese herbal drug Duhuo (roots of Angelica pubescens Maxim. f. biserrata Shan et Yuan), showed significant activity in a functional two-microelectrode voltage clamp assay with Xenopus oocytes which expressed recombinant γ-aminobutyric acid type A (GABA(A)) receptors of the subtype α(1)ß(2)γ(2S). HPLC-based activity profiling of the active extract revealed six compounds responsible for the GABA(A) receptor modulating activity. They were identified by microprobe NMR and high resolution mass spectrometry as columbianetin acetate (1), imperatorin (3), cnidilin (4), osthol (5), and columbianedin (6). In concentration-dependent experiments, osthol and cnidilin showed the highest potentiation of the GABA induced chloride current (273.6%±39.4% and 204.5%±33.2%, respectively at 300 µM). Bisabolangelone (2) only showed minor activity at the GABA(A) receptor. The example demonstrates that HPLC-based activity profiling is a simple and efficient method to rapidly identify GABA(A) receptor modulators in a bioactive plant extract.