Elsevier

Journal of Ethnopharmacology

Volume 236, 23 May 2019, Pages 129-135
Journal of Ethnopharmacology

Chemical constituents of Patrinia heterophylla Bunge and selective cytotoxicity against six human tumor cells

https://doi.org/10.1016/j.jep.2019.03.005Get rights and content

Abstract

Ethnopharmacological relevance

Patrinia heterophylla Bunge, known as “Mu-Tou-Hui” in China, is distributed in most provinces and regions of China. As a traditional medicinal plant, which was first found in <Ben- Cao-Gang- Mu>. In many traditional herbal books, there are records of “Mu-Tou-Hui” of treatment for uterine bleeding, cancer, swelling pain, leukemia, etc. However, there are few studies on the chemical constituents of Patrinia heterophylla Bunge.

Aim of the study

To investigate the chemical constituents of P. heterophylla and the basis of their antitumor activity.

Materials and methods

15 compounds were isolated from roots and rhizomes of P. heterophylla by repeating various column chromatography techniques, whose structures were determined by organic spectrum analysis methods and compared with published data. The cytotoxicities were evaluated by MTT assay on six cancer cell lines: human melanoma cell (A375), human hepatocellular carcinoma cell (SMMC-7721), human gastric cancer cell (SGC-7901), human cervical cancer cell (HeLa), human colon cancer cell (HCT-116), and human breast cancer cell (MDA-MB-231). The apoptosis-inducing activities of compounds 1, 5, 12 and 15 in A375 tumor cell determined by flow cytometry.

Result

Five phenylpropanoids, ethyl caffeate (1), coniferaldehyde (5), trans-p-coumaryl aldehyde (6), caffeic acid methyl ester (12), and 3,4-dihydroxycinnamic acid (15), four acetophenones, 1-(2,4-dihydroxyphenyl) ethanone (2), 2′,5′-Dihydroxyacetophenone (3), cynanchone A (8), and cynandione A (10), two phenols, vanillin (4) and catechol (9), two iridoids, sarracenin (7) and patriscabrol (11), one alkane, tetracosane (14), and one coumarin, scopoletin (13), were isolated from the EtOH extracts. Of them, compounds 110, 12 and 1415 were isolated for the first time from the roots of P. heterophylla. Compounds 1 and 15 were reported for the first time with in vitro inhibitory activity against tumor cells. MTT assay showed that compounds 1, 59, 1213 and 15 had selective cytotoxic activities (IC50 27.20–163.03 μM) against tumor cells. Apoptosis detected by flow cytometry revealed that compounds 1, 5, 12 and 15 can induce apoptosis for A375 at low concentrations when the concentrations of compounds 1, 5, 12 and 15 are the value of 14, 40, 34, 108 μM, the percentages of apoptotic cells were about 50%.

Conclusions

Compounds 110, 12 and 1415 were isolated for the first time from the P. heterophylla. This result enriches the previous studies on the chemical constituents of P. heterophylla. Compounds 1 and 15 were reported for the first time to have cytotoxic activities against tumor cells. Compounds 1, 5, 6, 7, 12, 15 showed cytotoxic activities against tumor cells. This result reveals that the active ingredient of P. heterophylla are composed of phenylpropanoids, iridoids and coumarins. This study provides some theoretical basis for the anti-tumor research of P. heterophylla.

Introduction

Patrinia belongs to the Valerianaceae family which includes 20 species and is mainly produced in eastern and central Asia and northern North America. There are 10 species, 3 subspecies and 2 varieties in China, which have abundant resources and produced all over the country (He et al., 2017). This genus has a long history of medicinal use in China with anti-tumor, anti-inflammatory, antibacterial, antiviral, anti-oxidant, sedative and hypnotic, and hepatoprotective activities (He et al., 2017).

The source of “Mu-Tou-Hui" recorded in the <National Grass Compilation> is Patrinia heterophylla Bunge and Patrinia scabra Bunge, take medicine with its roots or whole grass (Chinese Flora Editorial Board of the Chinese Academy of Sciences, 1975). “Mou-Tou-Hui” has a reputation for returning to life in ancient China, in many local floras, there are records about the “Mu-Tou-Hui”, which is used by folks to treat cervical cancer, malaria, typhoid, limb pain, etc. Malaria is known as the “Da-Bai-Zi” in the folk, and the “Mu-Tou-Hui” has a miraculous effect on the treatment of malaria, so “Mou-Tou-Hui” was also called “Bai-Zi-Cao” (Chen and Du, 2004). “Mou-Tou-Hui” has a significant therapeutic effect on uterine bleeding, people have been using it to treat cervical cancer, so it was called “Xue-Yun-Cao” in the folk (Chen and Du, 2004). At present, the pharmacological research of the P. heterophylla mainly focuses on anti-tumor effects, improving immunity, sedation, etc. (Ma et al., 1987; Tang, 1998; Yang et al., 2011) and is clinically used for the treatment of leukemia, uterine cancer, cervical cancer, acute appendicitis, irregular menstruation, etc.

Today, cancer has become the primary enemy of the world's serious threat to human life and health. The incidence is still rising year by year. Traditional Chinese medicine (TCM) treatment of cancer has the characteristics of low toxicity and high cure rate, attracting more and more people to invest in the research of Chinese herbal anti-cancer medicine, hoping to find anti-cancer active substances from Chinese herbal medicine. However, as a traditional anti-cancer Chinese medicine, there have been many reports on the anti-tumor research in vitro and in vivo about the crude extracts of P. heterophylla, little is known about which compounds are responsible for the anti-tumor effects. The extracts of P. heterophylla can inhibit the growth of U14 cervical cancer solid tumor mice (Lu et al., 2010). P. heterophylla has the function of inducing apoptosis of nude mouse transplantation tumor of human colon cancer HT-29 (Chen et al., 2007). The total glycoside of P. heterophylla had a strong inhibitory effect on ehrlich ascites, which could control the amount of ascites and significantly reduce the density of sarcoma cells per unit volume, and it also showed a good inhibitory effect on S180 sarcoma, and the tumor inhibition rate was up to 41.12% (Chen, 1999). Studies have shown that the active ingredients of P. heterophylla can resist human cervical cancer Hela, HL-60, K562, PC3, SGC-7901, Jurkat cell growth in vitro (Lu et al., 2009; Yang et al., 2011, 2013; Liu et al., 2017).

There is a systematic study on the chemical constituents of P. scabra. It also isolated dozens of compounds such as iridoids, triterpenoid saponins, flavonoids, coumarins, lignans, alkaloids, etc. (Li et al., 2003; Kouno et al., 1994; Yang et al., 2010; Feng et al., 2014; Liu et al., 2006; Gu et al., 2002). But the research report on the chemical constituents of P. heterophylla is still very lacking. So far, there are only 7 triterpenoids, 4 iridoid esters, 3 lignans, 2 steroids, 3 flavonoids, 5 triterpenoid saponin and 1 coumarin are separated from this plant in addition to volatile oil components (Yang et al., 2013; Lei et al., 1995, 1997; Ding et al., 2009; Chen et al., 2013; Liu et al., 2017; Xu, 2007). P. heterophylla has a good anti-tumor pharmacological effect, the separation and identification of further chemical constituents of this plant and the pharmacological studies of the monomeric compounds are very necessary.

A systematic chemical study was carried out on the P. heterophylla, and 15 monomer compounds were isolated. Their chemical structures were determined by NMR spectroscopy, Mass spectrometry and compared with literature data. The cytotoxic activities were tested against six human cancer cell lines including human melanoma cell (A375), human hepatocellular carcinoma (SMMC-7721), human gastric cancer (SGC-7901), human cervical cancer (Hela), human colon cancer (HCT-116), and human breast cancer (MDA-MB-231). The apoptosis-inducing activities of compounds 1, 5, 12 and 15 in A375 tumor cell determined by Annexin V-FITC/PI double staining assay.

Section snippets

Plant material

The roots of P. heterophylla Bunge were purchased in September 2015 in Bozhou, Auhui, China. The species was identified by Prof. Kai-Jin Wang from the School of Life Sciences, Anhui University, where a voucher specimen (No. 20150901) was deposited.

Instruments and reagents

Column chromatography silica gel material: 200–300 mesh, Qingdao Ocean Chemical Plant. HSGF 254 and GF 254 thin layer chromatography silica gel plate: Yantai Jiangyou Silicone Development Co. Ltd. MCI-gel CHP-20P (75–150 μm): Mitsubishi Chemical

Results and discussion

Fifteen compounds were isolated from P. heterophylla and identified by spectroscopic means (1H NMR, 13C NMR, 13C DEPT-135, MS) and compared with published data, they were ethyl caffeate (1) (Dai et al., 2006), 1-(2,4-dihydroxyphenyl) ethanone (2) (Zhao et al., 2007; El-Razek, 2006), 2′,5′-Dihydroxyacetophenone (3) (Miao et al., 2014), vanillin (4) (Du et al., 2010), coniferaldehyde (5) (Ragasa et al., 2013; Sun et al., 2017), trans-p-coumaryl aldehyde (6) (Stange Jr et al., 1999), sarracenin (7

Conclusion

In conclusion, this study reported 15 compounds isolated from the EtOH extract of P. heterophylla and evaluated their cytotoxicities on six cancer cell lines (A375, SMMC-7721, SGC-7901, HeLa, HCT-116 and MDA-MB-231). To the best of our knowledge, this is the first report that compounds 110, 12 and 15 were isolated from Patrinia, compounds 1 and 15 were reported for the first time with in vitro inhibitory activities on -tumor cells. Our experimental results not only found more chemical

Author contributions

LS carried out the plant extraction, the isolation of compounds, data analysis and manuscript preparation. YY provided supervision for the biological studies. LS and YZ assisted with bioassay. NL projected design, interpretation of data and manuscript editing. All authors read and approved the final version of the manuscript.

Conflicts of interest

The authors declare no conflict of interest.

Acknowledgements

The work was supported by the National Science Foundation of China (31670353), Major Project of Science and Technology of Anhui Province, China (16030801112), and National Science Foundation of Anhui Province, China (1708085MH206).

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