Isolation of Chemical Components from Cushioned Cypress and Their Inhibition of Tumor Cell Proliferation Activity
Selaginella pulvinata (Hook.&Grev.) Maxim. is a perennial herbaceous plant of the Selaginella genus in the family Selaginellaceae. It is also known as Evergreen, Longevity Grass, Nine Dead Resurrection Grass, etc. There are over 50 species of Selaginella plants in China, widely distributed in Guangxi, Liaoning, Yunnan, Fujian, Zhejiang, Shandong, Jilin, Hunan, Shaanxi, Hebei and other places. The book “Seeking Truth from Materia Medica” records that its treatment is divided into two categories: mature and raw, slightly cold when raw, and strong enough to break through the blood and meridians, thus treating symptoms such as stagnation and lymph nodes; If roasted, it will be warm and effective in stopping bleeding, thus treating symptoms such as intestinal redness and prolapse. Modern pharmacological research has shown that cushion shaped Selaginella has the effects of clearing heat and detoxifying, anti-inflammatory, anti-tumor, anti-aging, immune regulation, and treating cardiovascular diseases. The main components of Platycodon grandiflorus include flavonoids, flavones, alkynes, alkaloids, organic acids, etc. Among them, flavonoids and alkynes are the main active ingredients of Platycodon grandiflorus, and selaginelin compounds are a new type of alkynes skeleton compounds found only in plants of the Platycodon grandiflorus genus. Therefore, it is of great significance to purify the flavonoids and alkyne phenols in the cushioned cypress and conduct in-depth pharmacological studies on their mechanisms of action.

Due to the similar structures of selaginelin and flavonoids, it is difficult to separate them using conventional separation methods. Therefore, using mass spectrometry guided automatic purification and separation technology, the molecular weights and fragments of selaginellines and flavonoids were detected by mass spectrometry, and the corresponding compounds were prepared online in liquid phase. At the same time, vacuum and atmospheric pressure column chromatography were used to selectively separate flavonoids and alkynephenols from the ethyl acetate extraction part of the 95% ethanol extract of Selaginella cuspidata. Four flavonoids and seven selaginellines were obtained, and their structures were determined by chemical and spectral data. The in vitro proliferation inhibitory activity of these compounds on human non-small cell lung cancer cell (NSCLC) H322 was evaluated.

Using mass spectrometry guided PHPL-MS separation technology, combined with vacuum and atmospheric pressure column chromatography, selective preparation and separation of flavonoids and alkyne phenols in cushioned cypress were carried out, resulting in four flavonoids (1-4) and seven alkyne phenols (5-11). This method can quickly and efficiently obtain high-purity flavonoids and alkyne phenols. The evaluation results of inhibiting tumor cell proliferation activity showed that these natural compounds have good proliferation inhibitory activity on human non-small cell lung cancer cell line H322, with IC50 values of 16.70 μ mol/L for compound 3 and 9.54 μ mol/L for compound 10. Both flavonoids and alkynes have shown strong activity in anti-tumor and COX-2 inhibition, while alkynes have shown in vitro synergistic effects on the antifungal activity of fluconazole and ketoconazole. Therefore, the compounds prepared in this article have value for further research and structural modification. Especially, the alkyne phenolic structures are currently only found in plants of the Selaginella genus. The established rapid preparation method also provides an effective means for the rapid and large-scale separation and purification of flavonoids and alkyne phenolic compounds in cushioned Selaginella.