| Helianthus annuus L. (sunflower) was widely planted in China. The byproduct resources from sunflower such as heads, leaves and stalks are rich, but generally are used as feed, fuel or waste, lead to serious environmental pollution, resource waste and with low industrial added value. In Chinese traditional medicine field, sunflower has been recognized with high medicinal value, and the whole plant can be used as medicine for cancer prevention, hypotensive effectivity, immunity improvement, et al. But the current validation studies of medicinal components in sunflower and the associated functional activity are scarce. Therefore, the research and development of medicinal components resources from sunflower, explore their effective medicinal functions, are important for the medicinal modernization applications of sunflower.In this paper, accumulation and distribution of medicinal components in sunflower resources (except seed) has been studied, anti-tumor activity medicinal component screening and activity mechanism research were carried out, expected providing the basis for sunflower medicinal resources and anti-tumor activity components exploitation. The main results are as follows:(1) The distribution and accumulation of total polyphenols, total flavonoids, polysaccharides and chlorogenic acid, which are also medicinal components, in various growth stages of sunflower plants were studied, results showed that polysaccharide substances should be harvested from mature head, the flowering head and budding leaves are the best sunflower medicinal resources.(2) Homogenates extraction method was used for preparation of root extract, stem pith extract and head extract from sunflower which were reported previously with anti-tumor activity. The yield of total head extract was up to 13.33%, which was the highest one. Extracts of sunflower head were fractionated according to polarity, and obtained dichloromethane extract, ethyl acetate extract, n-butanol extract and water extract-four polarity extracts, simultaneously the highest yield was water extract, followed by dichloromethane extract.(3) The in vitro and in vivo anti-tumor activity of sunflower root extract, stem pith extract and head extract, and also the dichloromethane extract, ethyl acetate extract, n-butanol extract and water extract of sunflower head extract were investigated by MTT assay and S180 tumor-bearing models. The results showed that head extract has the best in vitro and in vivo anti-tumor activity in the extract of various parts, and dichloromethane extract has more superior in vitro and in vivo antitumor activity than sunflower head extract, it is indicated that the dichloromethane extract is the best anti-tumor active medicinal composition in sunflower head extract.(4) The anti-tumor mechanism of sunflower head dichloromethane extract (HHDE) has been studied by the human gastric cancer cell SGC-7901 cell proliferation assays, fluorescence microscopy, flow cytometry and Western-blot method. The results show that HHDE has a significant inhibition of tumor cell proliferation, and its dose-dependent anti-tumor activity significantly greater than the time-dependent. While under the condition of HHDE concentration ≥ 0.25mg/mL after 24 hours, the tumor cell growth inhibit rate was ≥58%. Accompanied by HHDE concentration increase a dose-dependent morphological changes of cell apoptosis can be observed by inverted microscope and fluorescence microscope, the tumor cells showed significant apoptosis signals in Annexin-V-FITC/PI assay and cell cycle test results, Meanwhile mitochondrial membrane potential was decreased, ROS, Ca2+ levels were significantly increased. The results of Western-blot showed that the expression of mitochondrial cytochrome C was decline, cytoplasmic cytochrome C-expressing cells was increased, indicating that cytochrome C release from mitochondria into the cytosol, and Caspase-3, Caspase-9 precursor protein expression decreased to prove Caspase-3, Caspase-9 has been activated. In addition, Bcl-2 protein expression decreased, Bax protein expression increased, Bax/Bcl-2 ratio was rised. Based on the above results, the anti-tumor mechanism of HHDE may be achieved mainly due to induction of apoptosis through mitochondrial pathway.(5) Chemical compositions of HHDE have been studied by LC-MS/MS method. The results show that HHDE may contain 14 kinds of compounds, including 8 sesquiterpene compounds and 6 diterpenoids compositions, in which grandifloric acid has been reported with significant anti-tumor activity, and the remaining compounds were unreported.(6) The 35 kinds of compounds has been identified by GC-MS method from HHDE, accounting for 93.26% HHDE total volatile substance, and in which the 8 highest kinds of compounds accounted for 63.26% HHDE total volatile substances. Comprehensive literature survey results, and verified in vitro anti-tumor activity of pimpinellin, pimaric acid and β-sitosterol by MTT assay, the results show these three compounds have significant anti-tumor activity in vitro, were effective anti-tumor compounds in sunflower, even more pimpinellin and pimaric acid were greater than HHDE.In summary, sunflower head is one of the mainly medicinal component distributed organs in sunflower plant. HHDE which comes from the sunflower head extract has significant anti-tumor activity and with a clear activity mechanism. Thus effectively exploit anti-tumor medicinal component of sunflower head, promote the modernization of traditional Chinese medicine applications of medicinal resources in sunflower, with significant economic and social benefits. |
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