Structurally diverse lignans from Solanum lyratum: chemical evidence for their acetylcholinease inhibitory activity

Abstract A chemical investigation of Solanum lyratum Thumb. (Solanace) afforded two new lignans (1b and 3) and eleven known lignan analogues (1a, 2a/2b and 4–11). Compounds 1a/1b and 2a/2b were separated as two pairs of enantiomers by chiral high-performance liquid chromatography (HPLC). Their structures were elucidated by detailed spectroscopic and comparative literature data analysis. The absolute configurations of compounds 1a/1b and 2a/2b were determined by comparing the experimental ECD data with the calculated values. All compounds were evaluated for their acetylcholinesterase (AChE) inhibitory activity. Notably, compared to the positive control, compounds 4 and 9 showed obvious AChE inhibition with their IC50 values of 1.30 ± 0.25 and 0.89 ± 0.04 μM, respectively. In addition, the possible interaction between acetylcholinesterase and the active compounds was also investigated by molecular docking. Graphical Abstract


Introduction
Solanum lyratum Thumb. (Solanace) is a common herbal plant that is natively distributed in India, Korea and China . The whole plant of S. lyratum., was well-known as BaiYing in traditional Chinese medicine and Back-Mo-Deung in traditional Korean medicine (Xu et al. 2018), and it has been used for the treatment of fever, jaundice hepatitis, cholelithiasis, cancers, nephritis edema, and arthritis diseases (Yao et al. 2013). Up to now, a variety of active components have been reported in S. lyratun, including saponins, flavonoids, lignans, sesquiterpenoids, steroidal alkaloids, coumarins, and polysaccharides (Yue et al. 2012).
Levels of the neurotransmitter acetylcholine are proved to be one of the key factors closely associated with the etiology of Alzheimer's disease (AD) (Mella et al. 2022). Modern pharmacological studies showed one of the most important strategies for treating AD is to enhance the acetylcholine levels in the brain using acetylcholinesterase (AChE) inhibitors (Yang et al. 2012).
In order to search for more bioactive constituents, a further phytochemical investigation on the whole plant of S. lyratum has been undertaken. It led to the isolation of two new lignans (1b and 3) and 11 known structurally diverse lignans (1a, 2a/2b and 4-11). Herein, the isolation, chiral resolution and structure elucidation together with their acetylcholinesterase inhibitory activity are described. And molecular docking was employed to study the interactions between AChE and active compounds. In the HMBC spectrum of 1 ( Figure S2), the spectral correlations for H-2, H-6/C-7, H-7/C-9, C-4 0 , H-8/C-4 0 , H-9/C-5 0 indicated compound 1 was a phenylpropanoid. According to the HMBC cross peaks of H-2 0 , H-6 0 and C-7 0 , the aldehyde group was connected to C-1 0 . Additionally, the HMBC correlations from d H 3.73 to C-3/5 (d C 56.1) and from d H 3.86 to C-5 0 (d C 55.8), suggested the position of the three methoxy groups. The spectra data above was identical to a known compound tomentosanan A (Liu et al. 2014) and the trans configuration of 1 was established by the NOESY correlations from H-7 to H-9a ( Figure S3) (Liu et al. 2019).

Results and discussion
The absence of cotton effect in the ECD spectrum and optical rotation indicated compound 1 was a racemic mixture ). Subsequently, the enantiomers 1a and 1 b with the opposite specific rotation and mirror-imaged ECD Cotton effects were successfully separated by using the Daicel Chiralpak AD-H chiral column ( Figure  S4). On the basis of the comparison between the experimental and calculated ECD spectra, the absolute configurations of 1a and 1 b were assigned and named as (7S,8R)-tomentosanan A (1a) (Liu et al. 2014) and (7 R,8S)-tomentosanan A (1b), respectively.
In the NOESY experiment of 3, a significant correlation of H-7 0 /H-9a, H-8 0 /H-9b and H-7a/H-9a implied the 7,8-trans and 8,8 0 -cis orientation (Zhao et al. 2020) (Fig. S3). However, the structure of 3 contains more than two chiral carbons and each fragment is separated from others, only the part relative configuration can be determined. Thus, compound 3 was confirmed and named solasesquilin B.
The spectrophotometric method of Ellman was used to evaluate AChE inhibitory activity of all compounds with donepezil hydrochloride used as a reference compound (Gao et al. 2021). The results (IC 50 ) were summarized in Table S2. The compounds 4 and 9 dose-dependently inhibited AChE inhibitory with an IC 50 value of 1.30 ± 0.25 and 0.89 ± 0.04 lM (positive control donepezil, exhibiting an IC 50 value of 2.40 ± 0.93 lM). Furthermore, compounds 3, 5 and 11 exhibited equivalent inhibitory actives with donepezil.
Molecular docking was performed to investigate possible mechanisms of interactions between compounds 4, 9 and AChE ( Figure S19). The docking results have shown that the compounds could easily fit into the active catalytic pocket of AChE. Taken together, the results of molecular modeling supported the possible interactions between compounds 4 and 9 and the enzyme AChE. More importantly, The fact that compound 9 have stronger AChE inhibitory action than compound 4 was supported scientifically.

Conclusion
In summary, thirteen lignans (1a/1b, 2a/2b, and 3-11), including two new lignans (1b and 3), were isolated from the whole plant of S. lyratum. Compounds 1a/1b and 2a/ 2b were obtained on a chiral chromatographic column. Their gross structures and configurations were unequivocally assigned as different types of lignins by detailed spectroscopic analyses, NMR and ECD spectral calculations. Notably, the inhibition of AChE activity was evaluated, compounds 4 and 9 exhibited more potent inhibitory activity than positive control donepezil. In addition, molecular docking results revealed that compounds 4 and 9 were involved in a large number of favorable interactions with the active site residues of the acetylcholinesterase protein, which can stabilize the ligands in the active site and increase their affinities. Based on these results, compounds 4 and 9 may be developed as an effective and favorable potential ACE inhibitory agent.