Study on the chemical components of lignans and their liver cell protective effects in Chinese wolfberry
Leonuri Fructus is derived from the dried and ripe fruit of Leonurus japonicus Houtt, a plant in the family Lamiaceae. Its medicinal properties include promoting blood circulation, regulating menstruation, clearing the liver, and improving vision. It was first recorded in the “Shennong Bencao Jing” and is known for its pungent and mild taste, as well as its ability to improve vision, nourish essence, and remove moisture. It has a long history of medicinal use. Yimucao and Wuweizi are two traditional Chinese medicines from the same plant with different medicinal parts. Their traditional effects also have similarities and differences. The “Compendium of Materia Medica” distinguishes the functions and indications of the two Chinese medicines: “Yimucao promotes blood circulation and nourishes blood… It is truly the holy medicine of the blood family. Wuweizi is excellent for treating women’s irregular meridians, fetal birth, and all kinds of blood and qi diseases. Its roots, stems, flowers, and leaves are all used as medicine and can be used together. If it treats hand and foot stagnation, yin and blood separation, wind and heat, clearing and purging liver heat, improving vision and essence, and regulating women’s meridians, then Wuweizi alone is good. If it treats swelling, toxins, sores, and ulcers, it can eliminate inflammation. Water promotes blood circulation, and if a woman suffers from various illnesses during pregnancy, it is advisable to use it in combination for good. From this, it can be seen that both motherwort and wolfberry have the function of promoting blood circulation and removing blood stasis, but the former is good at promoting blood circulation and removing blood stasis, while the latter is good at clearing the liver and improving vision. However, the breadth and depth of current research on Polygonatum sibiricum are far less than those on Leonurus heterophyllus. Chemical studies have found that more than 300 chemical components have been isolated and identified in Leonurus heterophyllus, and there are many literature explaining its pharmacological effects and mechanisms in vasodilation, antiplatelet aggregation, and regulation of uterine smooth muscle. However, there are less than 70 compounds reported in Polygonatum sibiricum, with the main component types being triterpenoids, cyclic peptides, flavonoids, sterols, phenylpropanoids, lignans, and alkaloids. Modern pharmacological studies have shown that Polygonatum sibiricum has effects such as uterine contraction, liver cell protection, inhibition of tyrosinase and acetylcholinesterase activity, anti-cancer, anti-inflammatory, neuroprotective, etc. However, most pharmacological studies only focus on the pharmacological effects of the extract, and there is relatively little research on the pharmacological effects and mechanisms of individual components. In the preliminary research, the research team found that tetrahydrofuran lignans in the seeds of Chinese wolfberry have significant liver cell protective activity, and explored the possible mechanism of action through fluorescence staining, Western blot and other methods. Therefore, in order to further investigate the active ingredients with liver protective effects in Fructus Polygonatum sibiricum, this study used various chromatographic and spectroscopic techniques to isolate and identify 8 lignans from the n-butanol fraction of Fructus Polygonatum sibiricum. Then, MTT, Hoechst 33342 fluorescence staining, and Western blot techniques were used to characterize the liver cell injury protective effects and mechanisms of the compounds.

 

In this study, eight lignans were isolated and identified from the n-butanol fraction of Polygonatum sibiricum, and compounds 1-8 were all isolated from Polygonatum sibiricum for the first time. At present, triterpenoids are the main compounds isolated from the seeds of Polygonatum sibiricum, and the research team has isolated 23 lignans from Polygonatum sibiricum so far. Literature research has found that the monomeric compounds isolated from Leonurus and Leonurus are mainly terpenoids, but there are few reports of lignans in Leonurus. Therefore, it is speculated that lignans are differential substances between Leonurus and Leonurus.

Acetaminophen, also known as paracetamol, is an aniline based antipyretic and analgesic drug. Overuse can easily cause liver damage or even liver failure, mainly by causing mitochondrial dysfunction and cell apoptosis in liver cells, resulting in severe liver toxicity. Therefore, APAP is often used as a modeling drug for drug-induced liver injury. This experiment used APAP to establish an in vitro liver injury model and screened the isolated compounds for liver cell protective activity. The results showed that compounds 1 and 3 significantly inhibited APAP induced liver cell injury, with EC50 values of 10.23 ± 2.45 μ mol/L and 5.20 ± 1.28 μ mol/L, respectively. Meanwhile, it was observed through fluorescence staining experiments that compound 3 can inhibit apoptosis in HL-7702 cells.

The intrinsic pathway of cell apoptosis is regulated by the B-cell lymphoma (Bcl-2) family proteins, which form a protein-protein interaction network and regulate cell apoptosis by affecting the permeability of the mitochondrial outer membrane. Bcl-2 and Bax are a pair of key proteins that regulate cell apoptosis. Bcl-2 is an anti apoptotic protein that enhances the resistance of liver cells to most DNA damage factors, while Bax is a pro apoptotic protein that induces increased mitochondrial membrane permeability and releases cytochrome c, leading to cell apoptosis. The ratio of Bcl-2 to Bax protein expression reflects the survival ability of cells after being stimulated by apoptosis. To further explore the mechanism of compound 3 in inhibiting cell apoptosis, Western blot technology was used to detect its expression levels of Bcl-2 and Bax proteins. The results showed that compound 3 could upregulate Bcl-2 protein expression, downregulate Bax protein expression, and increase the Bcl-2/Bax ratio, suggesting that compound 3 may exert anti apoptotic effects by regulating the expression levels of Bcl-2 and Bax proteins, thereby exerting a protective effect on liver cells.

This study explored the pharmacological substance basis and mechanism of action of liver cell protective activity in Radix Polygonatum sibiricum, providing a basis for elucidating the traditional efficacy of Radix Polygonatum sibiricum and facilitating the rational development and utilization of medicinal resources in the future. So far, the research team has isolated six lignans with liver cell protective activity from Fructus Ranunculi, including frutoligide A fruleoligide B、lariciresinol-4-O-β-D-glucoside、lariciresinol-4′-O-β-D-glucoside、 (+) – Isoquercetin -9 ‘- O – β – D-glucoside, artemisinin. The “Compendium of Materia Medica” suggests that the Chinese wolfberry is good at clearing the liver and improving vision, but there is currently limited research on its material basis and mechanism of action for clearing the liver and improving vision. Through the research of the research team, it was found that the lignans in the Chinese wolfberry have good liver cell protection effects, which may be the active substance basis for the Chinese wolfberry’s clearing the liver and improving vision.