Lasibidoupins A and B, two new compounds from the stems of Lasianthus bidoupensis V.S. Dang & Naiki

Abstract From the Lasianthus bidoupensis stems, two new compounds, including one new 9,10-anthraquinone, lasibidoupin A (1), and one new 6,7-benzocoumarin, lasibidoupin B (2), together with one known compound, 11-O-methyldamnacanthol (3) were isolated using chromatographic method. Their structures were determined by extensive HRMS, and NMR assignments. Compound 3 was reported for the first time from this species. New compounds (1 & 2) were tested for the cytotoxicity against three human cancer cell lines (MCF-7, HeLa, and NCI-H460) by SRB assay. As results, 1 & 2 exhibited significant cytotoxic activity against all cancer cell lines (IC50 ranged from 0.058 ± 0.003 to 0.177 ± 0.014 μM). Graphical Abstract


Introduction
The Lasianthus genus (Rubiaceae) comprising more than 180 species mostly presents in tropical Asia (Zhu et al. 2012). Fourty species and one subspecies were found in Vietnam (Dang et al. 2019).
Lasianthus bidoupensis V.S. Dang & Naiki, a new species was discovered in Vietnam (Dang et al. 2019). There is no report on the phytochemical components and biological properties of this species. Continuing our excited studies on cytotoxic constituents of new species explored in Vietnam (Nguyen et al. 2016;Ngo et al. 2017Ngo et al. & 2020Le et al. 2020;Mai et al. 2021), this paper detailed the isolation, structural determination, and cytotoxic effect of two new secondary metabolites from the stems of L. bidoupensis collected in Lam Dong province, Vietnam.

General experimental procedures
The HR-ESI-MS data were recorded on a Bruker spectrometer (Bruker Daltonik GmbH, Germany). The IR data were measured on a Bruker spectrometer (Tensor 27 FT-IR, Germany). The NMR spectra were performed on a Bruker spectrometer (Avance III, 500 MHz for 1 H NMR and 125 MHz for 13 C NMR). TLC was carried out on silica gel 60 F 254 or silica gel 60 RP-18 F 254 S (Merck) and spots were visualized by spraying with a solution of 5% vanillin in ethanol, followed by heating at 100 C. Analytical TLC was eluted using the increased polarity of solvent mixtures n-hexane-chloroform (from 80-20 to 50-50, v/v) and chloroform-methanol (from 99-1 to 90-10, v/v). Column chromatography was performed with silica gel 60 (0.040-0.063 mm, Merck).

Plant material
The stems of Lasianthus bidoupensis were collected in Bidoup Nui Ba National Park, Lam Dong Province, Viet Nam (December 2020), and certificated by Dr. Van Son Dang, Institute of Tropical Biology, VAST. The voucher specimen of species (No.SGU-001LB) was deposited in laboratory of Faculty of Environmental Science, Saigon University.

Srb assay
The cytotoxic effect of two new compounds (1-2) against three human cancer cells (HeLa, NCI-H460, and MCF-7) was exiamed using the SRB method according to Do et al. 2020;Huynh et al. 2021;Pham et al. 2021; and camptothecin was used as the positive control.
HeLa, NCI-H460, and MCF-7 were purchased from the American Type Culture Collection (Manassas, Rockville). Cells were cultured at 37 C and 5% CO 2 in Eagle's Minimal Essential Medium (EMEM) supplemented with 10% (v/v) FBS (Sigma), 2 mM L glutamine (Sigma), 20 mM HEPES (Sigma), 0.025 lg/mL amphotericin B (Sigma), 100 IU/ mL penicillin G (Sigma), and 100 lg/mL streptomycin (Sigma). Optical density values were determined with a 96-well micro-titer plate reader (Synergy HT, Biotek Instruments) at the wavelengths of 492 nm and 620 nm. Cells were seeded at a density of 10,000 cells/well. The treated cells were fixed with cold 50% (w/v) trichloroacetic acid (Merck) solution for 1-3 h, washed, and stained with 0.2% (w/v) SRB (Sigma) for 20 min. After five washes with 1% acetic acid (Merck), protein-bound dye was solubilized in 10 mM Tris base solution (Promega). The samples were dissolved in DMSO (Merck). Camptothecin was purchased from Calbiochem. The percentage of growth inhibition (Inh %) was calculated according to the formula: I % ¼ (1 À [ODt/ ODc] Â 100) %, in which ODt and ODc are the optical density value of the test sample and the control sample, respectively.

Disclosure statement
No potential conflict of interest was reported by the authors.

Funding
This research is funded by Sai Gon University (SGU) under grant number T -D2021-14.