Bioactive compounds from the seeds of Amomum tsaoko Crevost et Lemaire, a Chinese spice as inhibitors of sphingosine kinases, SPHK1/2

Amomum tsaoko Crevost et Lemaire (Zingiberaceae), a traditional Chinese spice also known as “Caoguo” or “tsao-ko,” has been considered to have many health benefits. As part of our continuous efforts to screen natural resources exhibiting potential bioactivity, we examined the seeds of A. tsaoko and found that its EtOH extract inhibited sphingosine kinases 1 and 2 (SPHK1/2). Bioactivity-based analysis and chemical investigation of the EtOH extract led to the isolation and identification of four aliphatic alcohols (1–4), five fatty acids (5–9), 12 phenolics (10–21), and four terpenoids (22–25), including four new compounds, an acetylated aliphatic alcohol (2), a fatty acid (5), and two phenolics (10–11). In addition, compound 1 was isolated for the first time from natural sources in this study. The structures of all compounds were elucidated based on spectroscopic analysis, including 1D and/or 2D NMR and HR-ESIMS as well as LC/MS analysis. A recently developed method using competing enantioselective acylation (CEA) coupled with LC/MS analysis was applied for the assignment of absolute configuration of compound 5. The absolute configurations of compounds 10 and 11 were determined using ECD calculations. All of the compounds (1–25) isolated from the active fraction were evaluated for their SPHK1/2 inhibitory effects at the concentration of 10 μM. Aliphatic alcohols 2–4, fatty acids 7 and 9, and phenolic compounds 13–15 and 21 showed inhibition against the activity of SPHK1 up to 20% and aliphatic alcohols 2 and 4, fatty acid 8, and phenolic compounds 10, 11, 18, and 22 showed inhibition against the activity of SPHK2 up to 40% compared with the control. Compound 2 showed the highest potency to inhibit SPHK1 enzymatic activity, by 59.75%, and compound 22 showed the highest potency in inhibiting the activity of SPHK2, by 22.75%, in comparison with the control, where both exhibited higher inhibition compared to those of positive controls. Docking modeling studies were conducted to suggest the binding mode of 2 and 22 in the substrate-binding pocket of SPHK1 and SPHK2, respectively.