天然小分子誘生機(jī)體抗病毒細(xì)胞因子研究

胡小鵬，金 哲，賀震旦

深圳大學(xué)醫(yī)學(xué)部醫(yī)學(xué)院藥學(xué)系 深圳大學(xué)生物治療研究所 深圳天然小分子創(chuàng)新藥物工程實(shí)驗(yàn)室，深圳 518060

中醫(yī)藥具有五千多年的歷史，國內(nèi)外科學(xué)家對中藥、天然藥、民間藥和民族藥物進(jìn)行了大量的研究，具有抗病毒藥用植物約100 余種［1］，如金銀花(Lonicera japonica)、黃芩(Scutellaria baicalensis)、黃芪(Astragalus membranaceus)、五味子(Schisandra chinensis(Turcz.))、夏枯草(Prunella vulgaris)、連翹(Forsythia suspensa)等(見表1)，據(jù)迄今為止的研究表明天然小分子(Natural micromolecule)為這些抗病毒藥用植物活性的物質(zhì)基礎(chǔ)。

抗病毒細(xì)胞因子(Antiviral cytokines)曾在Nasu K［2］和Fakruddin JM［3］等人發(fā)表的學(xué)術(shù)論文中提及過，但未曾對其作出具體的定義。為系統(tǒng)地開展細(xì)胞因子抗病毒作用及其作用機(jī)制研究的研究，有必要對抗病毒細(xì)胞因子進(jìn)行明確地定義:抗病毒細(xì)胞因子是由機(jī)體細(xì)胞分泌的，具有聯(lián)動機(jī)體固有免疫和特異性免疫應(yīng)答，進(jìn)而抑制或捕殺體內(nèi)病毒的小分子蛋白?？共《炯?xì)胞因子可分兩大類:一類是可直接作用體細(xì)胞產(chǎn)生抗病毒蛋白，即1)干擾素(Interferon):IFN-α、IFN-β、IFN-γ 和IFN-λ(IL-28、IL-29)［4-7］;2)腫 瘤 壞 死 因 子(Tumor Necrosis Factor)［8］:TNF-α 和TNF-β;干擾素和腫瘤壞死因子可直接激活細(xì)胞內(nèi)JAK(Janus activated kinase)-STAT(signal transducer and activator of transcription)信號通路［9-13］，隨后引起下游信號級聯(lián)反應(yīng)，包括MTOR信號通路、PI3K 信號通路、MAPK 信號通路均被激活，最終啟動細(xì)胞核中一些關(guān)鍵的蛋白基因進(jìn)行轉(zhuǎn)錄，翻譯出抗病毒蛋白進(jìn)而對細(xì)菌、病毒以及癌細(xì)胞等產(chǎn)生防御作用［14］(見圖1)。另一類是能激活巨噬細(xì)胞、自然殺傷細(xì)胞(NK 細(xì)胞)和毒性T 淋巴細(xì)胞(CTL)等細(xì)胞，進(jìn)而捕殺或吞噬體內(nèi)病毒的細(xì)胞因子(見圖1)，即白介素(Interleukin):IL-1β、IL-2、IL-6、IL-8、IL-12、IL-15、IL-17、IL-18、IL-21、IL-27等［15-20］。

基于細(xì)胞因子對捕殺或抑制病毒方面的作用，本文綜述了國內(nèi)外迄今為止報道的可誘生抗病毒細(xì)胞因子的天然小分子(見圖2，圖3)，其中以多酚類、苷類居多，還有部分物質(zhì)基礎(chǔ)未明確的提取物，有待進(jìn)一步研究。

1 多酚類化合物(Polyphenolic compounds)

白藜蘆醇(1，Resveratrol，10 mol/L)能上調(diào)人外周血單核細(xì)胞IL-1β、IL-6、TNF-α，調(diào)節(jié)免疫細(xì)胞功能［21］。從姜黃(Curcuma longa)根莖分離的天然多酚姜黃素(2，Curcumin)，通過激活人細(xì)胞內(nèi)MAPK以及MTOR 等信號通路，調(diào)節(jié)下游STATs 蛋白磷酸化水平，控制TNF-α、IL-1 和IL-6 的表達(dá)分泌［22］，姜黃素還可以顯著的調(diào)控TRAIL(TNF 相關(guān)凋亡誘導(dǎo)配基)，有研究顯示姜黃素(10 μmol/L 和25 μmol/L)和TRAIL(20 ng/mL)聯(lián)合使用能夠促進(jìn)胞內(nèi)半胱天冬酶的前體Procaspase-3，8，9 的分裂，并使得線粒體釋放細(xì)胞色素C，導(dǎo)致細(xì)胞毒性的增強(qiáng)［23］。茶多酚復(fù)方制劑(50～150 mg/kg)可增強(qiáng)小鼠淋巴細(xì)胞增殖能力，提高血清溶菌酶活性，上調(diào)血清中IFN-γ 分泌［24］。番茄(Lycopersicum esculentum)的多酚提取物能夠調(diào)節(jié)牛中性粒細(xì)胞TNF-α 基因的表達(dá)，加熱處理的提取物可顯著上調(diào)IFN-γ 的分泌［25］，具有增強(qiáng)動物和人類固有免疫的潛在活性。由咖啡酸(3，Caffeic acid)、阿魏酸(4，F(xiàn)erulic acid)和對香豆酸(5，p-Coumaric acid)酶促聚合的多酚(20～100 μg/mL)，通過和T 細(xì)胞表面CD4+蛋白結(jié)合，激活下游信號通路后上調(diào)小鼠脾臟細(xì)胞中的IFN-γ 和GM-CSF(Granulocyte-macrophage Colonystimulating Factor)分泌量［26］，增強(qiáng)機(jī)體固有免疫。

從Cocoa 提取的黃酮類化合物，能夠減少小鼠腹膜巨噬細(xì)胞活性氧和NO 的生成，上調(diào)IFN-γ 和TNF-α 的作用［27］。中藥黃芪(Astragalus membranaceus)和黃芩(Scutellaria baicalensis)是傳統(tǒng)的抗病毒藥材，主含槲皮素(6，Quercetin)、黃芩素(7，Baicalein)等天然小分子。Nair［28］等人報道，槲皮素(5 μmol/L)可調(diào)節(jié)外周血單核細(xì)胞IFN-γ 和IL-4 的分泌，從而促使Th0 細(xì)胞向Th1 細(xì)胞分化;隨后進(jìn)行了攻毒試驗(yàn)，發(fā)現(xiàn)誘生的IFN-γ 具有很強(qiáng)的抗VSV(vesicular stomatitis virus)活性。Johari 等［29］報道槲皮素和黃芩素抑制JEV(Japanese Encephalitis Virus)的IC50分別為212.1、14.28 μg/mL，提示黃芩素比槲皮素具有更好的抗JEV 作用。有研究表明，黃芩素可直接抑制病毒血凝素和神經(jīng)氨酸酶［30］。

桂皮醛(13，Cinnamaldehyde，1 mg/kg)［31，32］可上調(diào)小鼠血清中IL-2、IL-10 的分泌量，其誘生機(jī)制可能與Toll 樣受體的激活有關(guān)。從百屈菜中提取得到 血 根 堿(14，6-acetonyl-5，6-dihydrosanguinarine)［33］，體外實(shí)驗(yàn)證實(shí)其可誘導(dǎo)巨噬細(xì)胞和樹突狀細(xì)胞分泌TFN-α、IL-6、IL-8 等抗病毒細(xì)胞因子，對其誘生機(jī)制的研究表明，血根堿可激活MAPK 通路中ERK，NF-kB 循環(huán)活化機(jī)制也參與其中，共同調(diào)控了下游抗病毒細(xì)胞因子的表達(dá)。朱曉新等人［34］從紫萁貫眾分離得到四種多酚化合物:4＇-hydroxybenzalacetone(HBAc)，3＇，4＇-dihydroxybenzalacetone(DHBAc)、4-hydroxybenzaldehyde(HBAI)、3，4-dihydroxybenzaldehyde(DHBAI)，其中HBAI(15，10-100 ug/mL)、DHBAI(16，10-100 ug/mL)可激活胞內(nèi)NF-kB 通路，可顯著上調(diào)小鼠腹膜細(xì)胞分泌TNF、IL-1β、IFN-γ 等，誘生機(jī)制可能和iNOS(一氧化氮合成酶)的激活和NO 的生成有關(guān)。

2 苷類化合物(Glycosides)

研究表明苷類化合物具有抗腫瘤抗病毒活性［35］;桔梗(Platycodon grandiflorum)中分離得到的單體桔梗苷D(8，Platycodin D，0.0016～0.2 μg/mL)［36］，能夠上調(diào)IL-2、IFN-γ 的mRNA 表達(dá)水平，調(diào)控Th1/Th2 細(xì)胞的轉(zhuǎn)錄因子T-bet 和CATA-3 表達(dá)，同時，桔梗苷D 可以提高OVA 致敏小鼠抗體水平，具有免疫調(diào)節(jié)作用。王先遠(yuǎn)等［37］研究苦瓜總皂苷對衰老小鼠免疫功能的影響，發(fā)現(xiàn)苦瓜總皂苷不僅可上調(diào)血清中IL-2 水平，而且促進(jìn)脾臟分泌IL-2及腹腔巨噬細(xì)胞分泌TNF-α，其機(jī)制可能與DNA 多聚酶II 活性增強(qiáng)有關(guān);苦瓜總皂苷促進(jìn)IL-2 分泌，改變T 細(xì)胞亞群的組成，使機(jī)體免疫狀態(tài)趨向年輕化。徐先祥等［38］人報道甘草皂苷(9，0.5 mg/mL)可抑制98%以上的HIV 增殖，0.125 mg/mL 時，可抑制98%以上的HIV 的增殖，并可抑制50%斑塊的形成，其作用機(jī)制是誘生干擾素和增強(qiáng)NK 細(xì)胞的活性抵抗HIV。柚皮苷(17，naringin，20，40 and 80 mg/kg)［39］可上調(diào)Th2 型細(xì)胞分泌IL-4、IL-6、IL-10等。

表1 抗病毒藥用植物Table 1 Antivial Medical Plants

注:* 已報道的可誘生抗病毒細(xì)胞因子的天然小分子Note:* These natural micromolecules have been reported to induce antiviral cytokines.

圖1 天然小分子誘生細(xì)胞因子抗病毒作用機(jī)制Fig.1 The mechanisms of antiviral activity of antiviral cytokines induced by natrual micromolecule

圖2 誘生抗病毒細(xì)胞因子的天然小分子1～12 的化學(xué)結(jié)構(gòu)式Fig.2 The chemical structure of natural micromolecule 1-12 with the function of inducing antiviral cytokines

圖3 誘生抗病毒細(xì)胞因子的天然小分子13～19 的化學(xué)結(jié)構(gòu)式Fig.3 The chemical structure of natural micromolecule 13-19 with the function of inducing antiviral cytokines

另外，本課題組開展天然小分子誘生機(jī)體抗病毒細(xì)胞因子的研究中，發(fā)現(xiàn)天麻素(10，Gastrodin，0～1000 μmol/L)、迎春花素(11，Jasminin，1-100 μmol/L)能夠顯著上調(diào)RAW246.7 細(xì)胞分泌TNFα;從苦丁茶紫莖女貞(Ligustrum purpurascens)葉提取得到苯丙素總苷(crude phenylethanoid glycosides)，屬多酚類化合物，活性測試表明，苯丙素總苷(0.22～1.32 g/kg)具有免疫增強(qiáng)作用［49］;進(jìn)一步研究還發(fā)現(xiàn)，從苯丙素總苷分離得到的單體阿克苷(12，acteoside，0.8～200 μg/mL)，能夠顯著上調(diào)Balb/c 小鼠體內(nèi)IFN-γ 的表達(dá)水平，動物攻毒實(shí)驗(yàn)證實(shí)可顯著抑制小鼠體內(nèi)病毒載量。

3 其他

大蒜辣素(18，Allitridin，25 mg/kg/day)［40］可調(diào)節(jié)CMV 感染小鼠體內(nèi)Foxp3 蛋白表達(dá)，上調(diào)機(jī)體IFN-γ 和IL-4 分泌，恢復(fù)CMV 攻擊造成的免疫系統(tǒng)紊亂。青蒿素(19，Artemisinin)［41］可顯著刺激巨噬細(xì)胞分泌IL-12，其誘生機(jī)制和JNK 磷酸化的抑制有關(guān)。

滑菇(Pholiota nameko)提取物(20～60 mg/kg)［42］能夠上調(diào)白介素、腫瘤壞死因子、干擾素等多種抗病毒細(xì)胞因子的分泌。

此外，許多中藥提取物具有免疫增強(qiáng)作用［43］，如黃芪提取物(100～200 μg/mL)、板藍(lán)根提取物(100～200 μg/mL)能夠刺激雞外周血淋巴細(xì)胞IFN-γ 的mRNA 的表達(dá)量，并對NDV(Newcastle disease virus)感染有抑制作用。人參葉提取物［44］可顯著上調(diào)感染H5N1 小鼠和雪貂肺部IFN-γ 分泌，與空白對照組相比，動物的存活率可提高至40%～45%。五味子［45］和太子參［46］提取物，均可顯著誘導(dǎo)正常細(xì)胞生成TNF-α。商路提取物(50 mg/kg)［47］處理小鼠，每周喂食一次，可顯著刺激小鼠淋巴細(xì)胞增殖，產(chǎn)生IL-2，如果改變喂食頻率，一周三次，IL-2 的分泌量則開始下降;此外，當(dāng)劑量調(diào)整為10 mg/kg，每周三次喂食，則可以刺激巨噬細(xì)胞分泌IL-1，TNF，推測商路提取物可增強(qiáng)小鼠中巨噬細(xì)胞的活性，殺傷病變細(xì)胞。蓼大青葉提取物(1～100 mg/kg)［48］可顯著增加小鼠脾臟IFN-γ 的分泌。

4 結(jié)語

目前，用于臨床的抗病毒藥物主要有化學(xué)藥物和疫苗兩類，而通過誘生抗病毒細(xì)胞因子途徑的藥物有望成為第三類新型抗病毒藥物。藥用植物中的天然小分子結(jié)構(gòu)復(fù)雜，資源豐富，成藥性強(qiáng)，然而目前有關(guān)天然小分子誘生抗病毒細(xì)胞因子的研究缺乏系統(tǒng)性，尚待進(jìn)行深入研究。對抗病毒藥用植物進(jìn)行提取分離及純化，進(jìn)行體外細(xì)胞模型篩選，并開展動物免疫學(xué)和動物病毒學(xué)研究，進(jìn)而發(fā)現(xiàn)活性天然小分子，通過系統(tǒng)的基礎(chǔ)研究，有望研發(fā)出誘生機(jī)體抗病毒細(xì)胞因子的抗感染性疾病的臨床藥物，我們預(yù)計(jì)此類相關(guān)的研究將成為抗病毒藥物研發(fā)的新途徑。

1 Chinese Pharmacopoeia Commission(國家藥典委員會).Pharmacopoeia of the People’s Republic of China(中華人民共和國藥典).Beijing:China Medical Science Press，2010.Vol Ⅰ，1-362.

2 Nasu K，et al.Human oviductal epithelial cells express Tolllike receptor 3 and respond to double-stranded RNA:Fallopian tube-specific mucosal immunity against viral infection.Hum reprod，2007，22:356-361.

3 Fakruddin JM，et al.Noninfectious papilloma virus-like particles inhibit HIV-1 replication:implications for immune control of HIV-1 infection by IL-27.Blood，2007，109:1841-1849.

4 Poecka H，Ruland J.2012.From virus to inflammation:Mechanisms of RIG-I-induced IL-1β production.Eur J Cell Biol，91:59-64.

5 Brand S，et al.IL-28A and IL-29 mediate antiproliferative and antiviral signals in intestinal epithelial cells and murine CMV infection increases colonic IL-28A expression.Am J Physiol，2005，289:960-968.

6 Rossol-Voth R，et al.In vivo protective effect of tumour necrosis factor alpha against experimental infection with herpes simplex virus type 1.J Gen Virol，1991，72:143-147.

7 Sharma B，et al.Protein Kinase R as Mediator of the effects of interferon(IFN)γ and tumor necrosis factor(TNF)α on normal and dysplastic hematopoiesis.J Biol Chem，2011，286:27506-27514.

8 Xu SH(許沈華)，Zhang ZX(張宗顯).The advance research of the effect of tumor necrosis factor on anti-tumor.Foreign Medical Sciences(國外醫(yī)學(xué))，1989，(3):134-138.

9 Claudio A.Bonjardim，et al.Interferons:Signaling，antiviral and viral evasion.Immunology Letters，2009，122:1-11.

10 Hsu YM，et al.The adaptor protein CARD9 is required for innate immune responses to intracellular pathogens.Nat Immunol，2007，8:198-205.

11 Colonna M.TLR pathways and IFN-regulatory factors:to each its own.Eur J Immunol，2007，37:306-309.

12 Baccala R，et al.TLR-dependent and TLR-independent pathways of type I interferon induction in systemic autoimmunity.Nat Med，2007，13:543-551.

13 Schindler C，PlumLee C.Inteferons pen the JAK-STAT pathway.Semin Cell Dev Biol，2008，19:311-318.

14 Bartee E，et al.The addition of tumor necrosis factor plus beta interferon induces a novel synergistic antiviral state against poxviruses in primary human fibroblasts.J Virol，2009，83:498-511.

15 Vijay AK，et al.Inflammasomes and anti-viral immunity.J Clin Immunol，2010，30:632-637.

16 Sutlu T，et al.Inhibition of intracellular antiviral defense mechanisms augments lentiviral transduction of human natural killer cells:Implications for gene therapy.Hum Gene Ther，2012，23:1090-1100.

17 Liao Y，et al.Regulation of the p38 mitogen-activated protein kinase and dual-specificity phosphatase 1 feedback loop modulates the induction of interleukin 6 and 8 in cells infected with coronavirus infectious bronchitis virus.Virology，2011，420:106-116.

18 de Groot R，et al.Viral dsRNA-activated human dendritic cells produce IL-27，which selectively promotes cytotoxicity in naive CD8+T cells.J Leukoc Biol，2012，92:605-610.

19 Verbist KC，Lonowski KD.Functions of IL-15 in anti-viral immunity:Multiplicity and variety.Cytokine，2012，9:467-478.

20 Ryzhakov G，et al.IL-17 Boosts Proinflammatory outcome of antiviral response in human cells.J Immunol，2011，187:5357-5362.

21 Bergman M，et al.Resveratrol affects the cross talk between immune and colon cancer cells.Biomed ＆ Pharmacother，2013，67:43-47.

22 Zhou HY，et al.Targets of curcumin.Curr Drug，2011，12:332-347.

23 Deeb D，et al.Curcumin sensitizes prostate cancer cells to tumor necrosis factor-related apoptosis-inducing ligand/Apo2L by inhibiting nuclear factor-kB through suppression of IkBαphosphorylation.Mol Cancer Ther，2004，3:803-812.

24 Wang FR(王芙蓉)，et al.Effect of drug combinations from tea polyphenol on immune system of mice.China Animal Husbandry ＆ Veterinary Medicine(中國畜牧獸醫(yī))，2006，33(9):11-12.

25 Gyenai K，et al.Influence of Heirloom tomato polyphenol extracts on expression of inflammation genes In Bovine.American Journal of Animal and Veterinary Sciences，2012，7:126-135.

26 Yamanaka D，et al.The effect of enzymatically polymerised polyphenols on CD4 binding and cytokine production in murine splenocytes.Plos One，2013，7(4):e36025.

27 Ramos RS，et al.Effect of a cocoa flavonoid-enriched diet on experimental autoimmune arthritis.Br J Nutr，2012，107:523-532.

28 Nair MP，et al.The flavonoid，quercetin，differentially regulates Th-1(IFNgamma)and Th-2(IL4)cytokine gene expression by normal peripheral blood mononuclear cells.Biochimica et Biophysica Acta，2002，1593:29-36.

29 Johari J，et al.Antiviral activity of baicalein and quercetin against the japanese encephalitis virus.International journal of molecular sciences，2012，13:16785-16795.

30 Dou J，et al.Effects of baicalein on Sendai virus in vivo are linked to serum baicalin and its inhibition of hemagglutininneuraminidase.Arch Virol，2011，156:793-801.

31 Lin SS，et al.In vivo cytokine modulatory effects of cinnamaldehyde，the major constituent of leaf essential oil from Cinnamomum osmophloeum Kaneh.Phytother Res，2011，25:1511-1518.

32 Ding YY(丁媛媛)，et al.Influence of cinnamaldehyde on coxsackie virus B3 induced viral myocarditis in mice.Nat Prod Res Dev(天然產(chǎn)物研究與開發(fā))2010，22:769-776.

33 Kim DH，et al.6-Acetonyl-5，6-dihydrosanguinarine(ADS)from Chelidonium majus L.triggers proinflammatory cytokine production via ROS-JNK/ERK-NF-κB signaling pathway.Food Chem Toxicol，2013，58:273-279.

34 Zhu XX，et al.Divergent immunomodulatory effects of extracts and phenolic compounds from the Osmunda japonica Thunb.Chin J Integr Med.2013，19:761-770.

35 Lee J，et al.Saponin inhibits hepatitis C virus propagation by upregulating suppressor of cytokine signaling 2.Plos One，2012，7(6):e39366.

36 Xie Y，et al.Contribution of the glycidic moieties to the haemolytic and adjuvant activity of platycodigenin-type saponins from the root of Platycodon grandi?orum.Vaccine，2008，26:3452-3460.

37 Wang XY(王先遠(yuǎn))，et al.Effects of Momordca charantia L saponins on immune system of senile mice.Acta Nutrimenta Sinica(營養(yǎng)學(xué)報)，2001，23:263-265.

38 Xu XX(徐先祥)，Dai Y(戴岳).The overview of the effect of saponins from traditional Chinese medicine on anti-virus.Pharmacology and Clinics of Chinese Materia Medica(中藥藥理與臨床)，2008，24:110-111.

39 Ahmad SF1，Zoheir KM2，Abdel-Hamied HE3，Amelioration of autoimmune arthritis by naringin through modulation of T regulatory cells and Th1/Th2 cytokines.Cell Immunol，2014 287:112-120.

40 Li YN，et al.Allium sativum-derived allitridin inhibits Treg amplification in cytomegalovirus infection.J Med Virol.2013，85:493-500.

41 Cho YC，et al.Enhanced IL-12p40 production in LPS-stimulated macrophages by inhibiting JNK activation by artemisinin.Arch Pharm Res，2012，35:1961-1968.

42 Li HP，et al.Effects of the polysaccharide from Pholiota nameko on human cytokine network in serum.Int J Biol Macromol，2012，50:164-170.

43 Qiu Y(邱妍).Studies on immune-enhancing and antivirial action and mechanism of four Chinese herbal polysaccharides.Nanjing:Nanjing Agricultural University(南京農(nóng)業(yè)大學(xué))，PhD.2007.

44 Park EH，et al.Red Ginseng-containing diet helps to protect mice and ferrets from the lethal infection by highly pathogenic H5N1 influenza virus.J Ginseng Res.2014，38:40-46.

45 Zhao T，et al.Schisandra polysaccharide evokes mmunomodulatory activityh through TLR 4-mediated activation of macrophages.Int J Biol Macromol.2014，10，65C:33-40.

46 Wong CK，et al.Tumor necrosis factor eliciting fractions separated from Pseudostellaria heterophylla.Int J Immunopharmacol，1994，16:271-277.

47 Wang HB，Zheng QY.Effects of Phytolacca acinosa polysaccharides I with different schedules on its antitumor efficiency in tumor bearing mice and production of IL-1，IL-2，IL-6，TNF，CSF activity in normal mice.Immunotoxicol，1997，19:197-213.

48 Jeong HJ，et al.Inhibition of IL-32 and TSLP production through the attenuation of caspase-1 activation in an animal model of allergic rhinitis by Naju Jjok(Polygonum tinctorium).Int J Mol Med，2014，33:142-150.

49 Song X，et al.Immunomodulatory effects of crude phenylethanoid glycosides from Ligustrum purpurascens.J Ethnopharmacol，2012，144:584-591.

50 Kim YJ，et al.Functional analysis of HMGR encoding genes in triterpene saponin-producing Panax ginseng Meyer.Plant Physiol，2014，25.［Epub ahead of print］

51 Jian YQ，et al.Two new euglobals from the leaves of Eucalyptus robusta.J Asian Nat Prod Res.2012，14:831-837.

52 Zou P，Koh HL.Determination of indican，isatin，indirubin and indigotin in Isatis indigotica by liquid chromatography/electrospray ionization tandem mass spectrometry.Rapid Commun Mass Spectrom，2007，21:1239-1246.

53 Gu YC(顧玉誠)，Tu YY(屠呦呦).Chemical constituents of Cirsium Japonicum D.C.China J Chin Mat Med(中國中藥雜志)，1992，17:489-490.

54 Kim SR，et al.Trichosanthes kirilowii Ethanol Extract and Cucurbitacin D Inhibit Cell Growth and Induce Apoptosis through Inhibition of STAT3 Activity in Breast Cancer Cells.Evid Based Complement Alternat Med，2013:975350.

55 Lin RJ，et al.Cytotoxic activity of Ipomoea cairica.Nat Prod Res，2008，22:747-753.

56 Chyau CC，et al.Schisandra chinensis Peptidoglycan-Assisted Transmembrane Transport of Lignans Uniquely Altered the Pharmacokinetic and Pharmacodynamic Mechanisms in Human HepG2 Cell Model.PLoS One，2014，9:e85165.

57 Wang Z，et al.Protective effects of fractions from Pseudostellaria heterophylla against cobalt chloride-induced hypoxic injury in H9c2 cell.J Ethnopharmacol，2013，147:540-545.

58 Escobar FM，et al.Antiviral effect and mode of action of methanolic extract of Verbascum thapsus L.on pseudorabies virus(strain RC/79).Nat Prod Res，2012，26:1621-1625.

59 Huh JE，et al.Combined prescription(OAH19T)of Aralia cordata Thunb and Cimicifuga heracleifolia Komar and its major compounds inhibit matrix proteinases and vascular endothelial growth factor through the regulation of mitogen-activated protein kinase pathway.J Ethnopharmacol，2011，135:414-421.

60 Yim SH，et al.Cytotoxic caffeic acid derivatives from the rhizomes of Cimicifuga heracleifolia.Arch Pharm Res，2012，35:1559-1565.

61 Lai HY，et al.Blechnum orientale Linn-a fern with potential as antioxidant，anticancer and antibacterial agent.BMC Complement Altern Med，2010，10:15.

62 Cui CW(崔傳文)，et al.Preliminary study on chemical constituents seperated from Cayratia japonica(中國中藥雜志)，2012，37:2906-2909.

63 Jia SS，et al.Studies on flavonoid constituents isolated from the leaves of Glycyrrhiza uralensis Fisch.Acta Pharmaceutica Sinica，1990，25:758-762.

64 Wang N(王楠)，et al.Chemical constituents of Pyrrosia petiolosa.Journal of Shenyang Pharmaceutical University(沈陽藥科大學(xué)學(xué)報)，2003，20:425-438.

65 Xuan SQ，et al.Polymorphic microsatellite loci for the genetic analysis of Lycoris radiata(Amaryllidaceae)and cross-amplification in other congeneric species.Genet Mol Res，2011，10:3141-3145.

66 Piacente S，et al.Constituents of Ardisia japonica and their in vitro anti-HIV activity.J Nat Prod，1996，59:565-569.

67 Tang X，et al.Screening anti-HIV Chinese materia medica with HIV and equine infectious anemic virus reverse transcriptase.J Tradit Chin Med，1994，14:10-13.

68 Zhang J，et al.Paeoniflorin abrogates DSS-induced colitis via a TLR4-dependent pathway.Am J Physiol Gastrointest Liver Physiol，2014，306:27-36.

69 Ye XY，et al.Dolichin，a new chitinase-like antifungal protein isolated from field beans(Dolichos lablab).Biochem Biophys Res Commun，2000，269:155-159.

70 Zhang XM(張雪梅)，et al.Simultaneous determination of five constituents in Scrophularia ningpoensis by HPLC.China Journal of Chinese Materia Medica(中國中藥雜志)，2011，36:709-711.

71 Nagao T，et al.Studies on the constituents of Luffa acutangula Roxb.I.Structures of acutosides A-G，oleanane-type triterpene saponins isolated from the herb.Chem Pharm Bull(Tokyo)，1991，39:599-606.

72 Cheng HB，et al.Chemical constituents of ethyl acetate extract from Polygonum perfoliatum.J Chin Med Mater，2012，35:1088-1090.

73 Zhou XW(周興旺)，et al.Chemical constituents in the roots of Lycium chinense Mill.China Journal of Chinese Materia Medica(中國中藥雜志)，1996，21:675-676.

74 Szymańska G，et al.Field cultivation and in vitro cultures，root-forming callus cultures and adventitious root cultures，of Panax quinquefolium as a source of ginsenosides.Z Naturforsch C，2013，68:482-488.

75 Yang XZ，et al.Alkaloids from roots of Stemona sessilifolia and their antitussive activities.Planta Med，2009，75:174-177.

76 Hai LQ(海力茜)，et al.Studies on chemical constituents from root of Hedysarum polybotrys.China Journal of Chinese Materia Medica(中國中藥雜志)，2004，29:432-434.

77 Qi HY，et al.Chemical constituents of Polygonum cillinerve.The 3～(rd)International Conference on Natural Products:Natural Products-a Must for Human Survival，Nanjing，China，2004，10:190.

78 Kan S，et al.Chemical constituents from the roots of Xanthium sibiricum.Nat Prod Res，2011，25:1243-1249.

79 Wang K，et al.Phenolic constituents from Brainea insignis.Chem Pharm Bull(Tokyo)，2010，58:868-871.

80 Fang X，et al.Microwave-assisted extraction followed by RPHPLC for the simultaneous extraction and determination of forsythiaside A，rutin，and phillyrin in the fruits of Forsythia suspensa.J Sep Sci，2013，36:2672-2679.

81 Yuan Y，et al.Functional diversity of genes for the biosynthesis of paeoniflorin and its derivatives in Paeonia.Int J Mol Sci，2013，14:18502-18519.

82 Schühly W，et al.Chemical characterization of Magnolia biondii(Flos Magnoliae，Xin Yi).Nat Prod Commun，2009，4:231-234.

83 Li HY，et al.Antioxidant activities of extracts and fractions from Lysimachia foenum-graecum Hance.Bioresour Technol，2009，100:970-974.

84 Chen QZ(陳秋竹)，et al.Studies on chemical constituents of the extract of Lonicera japonica.Journal of Chinese Medicinal Materials(中國中藥雜志)，2010，33:920-922.

85 Chen CX(陳常興).Studies on the chemical constituents from the fruit of Aristolochia debilis J Chin Med Mater(中國中藥雜志)，2010，33:1260-1261.

86 Agatonovic-Kustrin S，Loescher CM.Qualitative and quantitative high performance thin layer chromatography analysis of Calendula officinalis using high resolution plate imaging and artificial neural network data modelling.Anal Chim Acta，2013，798:103-108.

87 Scragg AH，Allan EJ.Production of the triterpenoid quassin in callus and cell suspension cultures of Picrasma quassioides Bennett.Plant Cell Rep，1986，5:356-359.

88 Patel R，et al.Analgesic and antipyretic activities of Momordica charantia Linn.fruits.J Adv Pharm Technol Res，2010;1:415-418.

89 Shao H，et al.Matrine effectively inhibits the proliferation of breast cancer cells through a mechanism related to the NFκB signaling pathway.Oncol Lett，2013，6:517-520.

90 Chang SJ，et al.Antiviral Activity of Isatis indigotica Extract and Its Derived Indirubin against Japanese Encephalitis Virus.Evid Based Complement Alternat Med，2012，925830.

91 Jin S，et al.Biotransformation of polydatin to resveratrol in Polygonum cuspidatum roots by highly immobilized edible Aspergillus niger and Yeast.Bioresour Technol，2013，136:766-770.

92 Li N(李娜)，et al.Studies on anti-inflammation chemical constitutes of Patrinia villosa.Journal of Chinese Medicinal Materials(中國中藥雜志)，2008，31:51-53.

93 Wang Y，et al.The genus Anemarrhena Bunge:A review on ethnopharmacology，phytochemistry and pharmacology.J Ethnopharmacol，2014，S0378-8741(14)00114-7.

94 Jiang HX，et al.Benzofuran derivatives from Eupatorium fortunei.Nat Prod Res，2008;22:937-941.

95 Zhang HB，et al.Flavonoids from Inula britannica reduces oxidative stress through inhibiting expression and phosphorylation of p47(phox)in VSMCs.Pharm Biol，2011，49:815-820.

96 Xiong Y，et al.Quality control of Lonicera japonica stored for different months by electronic nose.J Pharm Biomed Anal，2014，91:68-72.

97 Zhao RZ，et al.Determination of isofraxidin and astilbin by HPLC in rat plasma and its application after orally administration the extract of Sarcandra glabra.Pak J Pharm Sci，2013，26:1-6.

98 Yang ZN，et al.Quality evaluation of Houttuynia cordata Thunb.by high performance liquid chromatography with photodiode-array detection(HPLC-DAD).Pak J Pharm Sci，2014，:223-231.

99 Zomorodian K，et al.Chemical composition and antimicrobial activities of essential oil of nepeta cataria L.Against common causes of oral infections.J Dent(Tehran)，2013，10:329-337.

100 Lee LS，et al.Metabolomic analysis of the effect of shade treatment on the nutritional and sensory qualities of green tea.J Agric Food Chem，2013，61:332-338.

101 Nugroho A，et al.vivo sedative and gastroprotective activities of Salvia plebeia extract and its composition of polyphenols.Arch Pharm Res，2012，35:1403-1411

102 Hou HC(侯惠嬋)，Liang SZ(梁少珍).Determination of indirubin and indigo in Baphicacanthus cusia(Nees)Bremek by HPLC.J Chin Med Mater(中國中藥雜志)，2006，29:681-682.

103 Ogata S，et al.Apoptosis induced by the flavonoid from lemon fruit(Citrus limon BURM.f.)and its metabolites in HL-60 cells.Biosci Biotechnol Biochem，2000，64:1075-1078.

104 Zhao L，et al.Phytochemical and Biological Activities of an Anticancer Plant Medicine:Brucea javanica.Anticancer Agents Med Chem，2014，14:440-458.

105 Xie C，et al.Flavonol glycosides from Lysimachia capillipes.J Asian Nat Prod Res，2002，4:17-23.

106 Yu H，et al.Protective effects of the total Saponins from Dioscorea nipponica Makino against carbon tetrachloride-induced liver injury in mice through suppression of apoptosis and inflammation.Int Immunopharmacol，2014，19:233-244.

107 Sulaiman CT，Balachandran I.Total phenolics and total flavonoids in selected Indian medicinal plants.Indian J Pharm Sci，2012，74:258-260.

108 Park NI，et al.Anthocyanin accumulation and expression of anthocyanin biosynthetic genes in radish(Raphanus sativus).J Agric Food Chem，2011，59:6034-6039.

109 Kocevski D，et al.Antifungal effect of Allium tuberosum，Cinnamomum cassia，and Pogostemon cablin essential oils and their components against population of Aspergillus species.J Food Sci，2013，78:731-737.

110 Feng L，et al.Antioxidant activities of total phenols of Prunella vulgaris L.in vitro and in tumor-bearing mice.Molecules，2010，15:9145-9156.

111 Lu CN，et al.Saikosaponin a and its epimer saikosaponin d exhibit anti-inflammatory activity by suppressing activation of NF-κB signaling pathway.Int Immunopharmacol，2012，14:121-6.

112 Jin A，et al.Four new compounds from the bulbs of Lycoris aurea with neuroprotective effects against CoCl2 and H 2O 2-induced SH-SY5Y cell injuries.Arch Pharm Res，2014 Mar;37:315-323.

113 Jun HJ1，et al.Iristectorigenin B isolated from Belamcanda chinensis is a liver X receptor modulator that increases ABCA1 and ABCG1 expression in macrophage RAW 264.7 cells.Biotechnol Lett，2012，34:2213-2221.

114 Xue Z(薛智)，et al.Studies on the minor alkaloids of Cephalotaxus hainanensis Li(author＇s transl).Acta Pharmaceutica Sinica(藥學(xué)學(xué)報)，1981，16:752-756.

115 Kristanti RA，et al.Accelerated biodegradation of nitrophenols in the rhizosphere of Spirodela polyrrhiza.J Environ Sci(China)，2012，24:800-807.

116 Lee WJ，Choi SW.Quantitative Changes of Polyphenolic Compounds in Mulberry(Morus alba L.)Leaves in Relation to Varieties，Harvest Period，and Heat Processing.Prev Nutr Food Sci，2012，17:280-285.

117 Li L，et al.Antibacterial activity of Lansiumamide B to tobacco bacterial wilt(Ralstonia solanacearum).Microbiol Res，2014，S0944-5013(13)00196-1.

118 Wangchuk P，et al.Antimalarial alkaloids from a Bhutanese traditional medicinal plant Corydalis dubia.J Ethnopharmacol，2012，143:310-313.

119 Zhang Z，et al.Baicalein Reduces the Invasion of Glioma Cells via Reducing the Activity of p38 Signaling Pathway.PLoS One，2014，9:e90318.

120 Liu R，et al.Expression profile of a PAL gene from Astragalus membranaceus var.Mongholicus and its crucial role in flux into flavonoid biosynthesis.Plant Cell Rep，2006，25:705-710.

121 Chen L(陳亮)，et al.Non-alkaloid chemical constituents from Coptis chinensis.China Journal of Chinese Materia Medica(中國中藥雜志)，2012，37:1241-1244.

122 Li XC，et al.Steroid saponins from Polygonatum kingianum.Phytochemistry，1992，31:3559-3563.

123 Mishra MR，et al.Antidiabetic and Antioxidant Activity of Scoparia dulcis Linn.Indian J Pharm Sci，2013，75:610-614.

124 Zang X，et al.A-type proanthocyanidins from the stems of Ephedra sinica(Ephedraceae)and their antimicrobial activities.Molecules，2013，18:5172-5189.

125 Li YB(李遇伯)，et al.Chemical constituents from herb of Epimedium brevicornum.China Journal of Chinese Materia Medica(中國中藥雜志)，2005，30:586-588.

126 Lim DW，et al.Anti-inflammatory effects of total isoflavones from Pueraria lobata on cerebral ischemia in rats.Molecules，2013，18:10404-10412.

127 Duelund L，et al.Influence of the active compounds of Perilla frutescens leaves on lipid membranes.J Nat Prod，2012，75:160-166.

128 Numata A，et al.Plant constituents biologically active to insects.VI.Antifeedants for larvae of the yellow butterfly，Eurema hecabe mandarina，in Osmunda japonica.(2).Chem Pharm Bull(Tokyo)，1990，38:2862-2865.

129 Fu GM(浮光苗)，et al.Studies on the chemical constituents from the aerial parts of Breynia fruticosa.China Journal of Chinese Materia Medica(中國中藥雜志)，2004，29:1052-1054.

130 Oh HA，et al.Evaluation of the effect of kaempferol in a murine allergic rhinitis model.Eur J Pharmacol，2013，718(1-3):48-56.

131 Rubio-Pina J，et al.Pharmaceutical applications of the benzylisoquinoline alkaloids from Argemone mexicana L.Curr Top Med Chem，2013，13:2200-2207.

132 He ZD，et al.Antioxidant activity of phenylethanoid glycosides from Brandisia hancei.J Ethnopharmacol，2000，71:483-486.

133 Dong W，et al.A novel near-infrared spectroscopy and chemometrics method for rapid analysis of several chemical components and antioxidant activity of mint(Mentha haplocalyx Briq.)samples.Appl Spectrosc，2014，68:245-254.

134 Wang D(王大海)，et al.Nonvolatile chemical constituentsfrom Pogostemon cablin.China Journal of Chinese Materia Medica(中國中藥雜志)，2010，35:2704-2707.