Studying Mass Balance and the Stability of ( Z)-Ligustilide from Angelica sinensis Helps to Bridge a Botanical Instability-Bioactivity Chasm

Abstract

Numerous reports assigning ( Z)-ligustilide ( 1) the role of a major bioactive principle in Apiaceae botanicals are called into question by the recurrent demonstrations of 1 being an unstable, rapidly degrading compound, ultimately leading to dynamic Residual Complexity. While Angelica sinensis is recognized for its therapeutic value in (peri-)menopausal symptom management, its purported active principle, ligustilide, represents a typical example for the instability-bioactivity chasm of botanicals. To help bridge the gap, this study used both the essential oil and purified 1 obtained from A. sinensis to investigate the factors that influence the chemical transformation of 1, the products formed, and the rationale for monitoring 1 in natural product preparations. Countercurrent separation was used to purify 1 from a supercritical fluid extract of A. sinensis, achieving 93.4% purity in a single step. Subsequent purification by preparative HPLC afforded 1 with a 98.0% purity. Providing a mass balance setting, we monitored chemical changes occurring to highly purified 1 under various conditions and at different time points, in sealed NMR tubes by quantitative ¹H NMR (qHNMR). The non-destructive nature of NMR enabled a comprehensive assessment of degradation products. Moreover, in being a mole-based determination, the total intensity (integral) of all NMR signals intrinsically represent the theoretical mass balance within the sample solution. The results demonstrated that 1 is most stable while within the original plant material. Exposure to light had a profound impact on the chemical transformation of 1 leading to the formation of ligustilide dimers and trimers, as verified by both NMR and LC-HRMS studies. Moreover, the results shown for 1, augmented by other recent outcomes, have serious implications for the meaningful biological evaluation of NPs which exhibit instability/reactivity, while having a plethora of “promising” bioactivities reported in the literature and being frequently associated with unsubstantiated health claims.

Graphical Abstract