The stability of Z-ligustilide and its relevance for the biological evaluation of Angelica botanicals

Abstract

Originating from traditional Chinese medicine, Angelica species are commonly used herbals [1]. Phthalide monomers such as cis-3-butylidene-4,5-dihydro-phthalide (Z-ligustilide, 1) are often associated with the active principle in botanical preparations [2, 3]. Because pure 1 is known to undergo rapid chemical degradation [4, 5], the purpose of this study was to elucidate whether the observed bioactivity may be attributed to 1 by itself, or associated with its degradation products. Therefore, a three-prong approach was taken, which involved: 1) the isolation of pure 1, 2) careful stability monitoring, and 3) the evaluation of biological activity using a quinone reductase (QR) assay. The use of an optimized liquid-liquid chromatography protocol employing 2-step FCPC/HSCCC permitted the isolation of high-purity 1 (99.4 by GC, 99.6% by qHNMR), from the roots of Angelica sinensis (Oliv.) Diels. (Apiaceae). When the stability of 1 was monitored by GC-MS and [q]NMR to evaluate the impact of different storage conditions, 1 was found stable when stored in organic solutions at temperatures -30o C or below. However, once dried, 1 undergoes significant degradation within 24 hours, even when kept in the dark at -30° C. Interestingly, 1 of high-purity, when subjected to biological testing, showed no measurable activity in the QR assay, while chemically degraded samples exhibited significant activity (CD value: 6.5–11.7µM±0.4–1.2) in the same assay. Our results demonstrate that the storage conditions can have a profound impact on the quality of phthalide-containing herbals as well as on the biological activities attributed to the phthalide class of compound.