Antrodia Cinnamomea ameliorates neointimal formation by inhibiting infl ammatory cell infi ltration through downregulation of adhesion molecule expression in vitro and in vivo

The increased vascular infl ammation is a key event in the development of atherosclerotic lesions.Antrodia cinnamomea has been shown to promote anticancerogenic activity through decreasing infl ammation.However,the potential role of A.cinnamomea in cardiovascular diseases remains unexplored.Herein,using carotid arterial ligation models,we found that ethanol extract from A.cinnamomea(EEAC)signifi cantly inhibited neointimal hyperplasia in a dose-dependent manner,accompanied with the reduced expression of activated p65 and infl ammatory cytokines.We also show that EEAC ameliorated TNF-α-induced phosphorylation of p65 and pro-infl ammatory cytokine expression in both vascular smooth muscle cells(VSMCs)and macrophages in vitro.Mechanistically,EEAC suppressed expression levels of intercellular adhesion molecule-1(ICAM-1)and vascular cell adhesion molecule(VCAM-1)in VSMCs,which attenuates the ability of monocytes/macrophages adhesion to VSMCs.Furthermore,the expression level of these adhesion molecules and infi ltration of monocytes/macrophages were also decreased in neointimal VSMCs of arteries pretreated with EEAC.Altogether,our results reveal a novel function of A.cinnamomea in suppressing vascular infl ammation upon ligation injury during neointimal formation,likely through inhibition of infl ammatory cell infi ltration via downregulating the adhesion molecules in VSMCs.Thus,A.cinnamomea may offer a pharmacological therapy to slow down disease progression in patients with vascular injury.

Intestinal Absorption of Ergostane and Lanostane Triterpenoids from Antrodia cinnamomea Using Caco-2 Cell Monolayer Model

Antrodia cinnamomea is a precious medicinal mushroom.It exhibits promising therapeutic effects on cancer,intoxication,hypertension,hepatitis,and inflammation.Its major bioactive constituents are ergostane and lanostane triterpenoids.In this study,we used intestinal Caco-2 cell monolayer model to reveal the intestinal absorption property of 14 representative triterpenoids from A.cinnamomea.The bidirectional transport through the monolayer at different time points was monitored by a fully validated LC/MS/MS method.In the case of pure compounds,ergostanes 5(25R-antcin H),6(25Santcin H)and 10(25R-antcin B)could readily pass through the Caco-2 cell layer,whereas lanostanes 13(dehydroeburicoic acid)and 14(eburicoic acid)could hardly pass through.When the cells were treated with A.cinnamomea extract,antcins A,B,C,H and K(1–6 and 9–11)were absorbed via passive transcellular diffusion,and showed high PAB and PBA values(>2.5×10^(-5) cm/s).Meanwhile,the lanostanes dehydrosulphurenic acid(8),15a-acetyldehydrosulphurenic acid(12),13 and 14 exhibited poor permeability.Transport features of these compounds were consistent with their pharmacokinetic behaviors in rats.This study could also be helpful in predicting the intestinal absorption of A.cinnamomea in human.

The protoplast two-way fusions and fusant characteristics of Antrodia cinnamomea and Cordyceps militaris

This study generated two fused protoplasts of Antrodia cinnamomea and Cordyceps militaris in two ways.The protoplasts of A.cinnamomea were inactivated by heat to inactivate biochemical processes and enzymatic activities in the cytoplasm,and the protoplasts of C.militaris were inactivated by UV radiation to invalidate their genome function,then they were fused under optimal conditions to get a fusion rate as(7.42±0.8)×10^(-6) fusants/mL;the new fusants were abbreviated as Ac-Cm.On the other hand,when A.cinnamomea and C.militaris were treated with heat and UV oppositely using similar experiments,the fusion rate was(9.70±0.68)×10^(-5) fusants/mL,and the new fusants were abbreviated as Cm-Ac.We selected each of two best-growing fused colonies Ac-Cm-1,Ac-Cm-2,Cm-Ac-1,and Cm-Ac-2,together with parental A.cinnamomea and C.militaris,and studied their morphology,growth antagonism tests,and genetic relationships by 18 S rRNA sequencing.In comparison with the initial cultures of 4 fusants,the yields of adenosine,biomass,cordycepic acid,cordycepin,total polysaccharide,and total triterpenoids were increased up 1.305-50.1563 times in the optimal medium conditions.For gene stability tests,those of the four fusants and their outputs were stabilized within 10 generations.

Convergent synthesis and immunological study of oligosaccharide derivatives related to galactomannan from Antrodia cinnamomea

The galactomannan from Antrodia cinnamomea(AC)is characterized as one of the important bioactive components that exhibits potential immunostimulatory propriety.The biological function of its corresponding oligosaccharide fragments has not been revealed yet.In this study,we reported the first chemical synthesis of the series of oligosaccharide fragments related to AC galactomannan via the convergent glycosylation strategy.The preliminary immunological evaluation of these synthesized AC oligosaccharides disclosed that the backbone tetrasaccharide 1d showed the best immunomodulatory ability on enhancing proliferation,phagocytosis and cytokines secretion of Raw264.7 macrophages in vitro,indicating its immense potential as an immunostimulant candidate.

Antrodia cinnamomea exerts an anti-hepatoma effect by targeting PI3K/AKT-mediated cell cycle progression in vitro and in vivo

Antrodia cinnamomea is extensively used as a traditional medicine to prevention and treatment of liver cancer.However,its comprehensive chemical fingerprint is uncertain,and the mechanisms,especially the potential therapeutic target for anti-hepatocellular carcinoma(HCC)are still unclear.Using UPLC-Q-TOF/MS,139 chemical components were identified in A.cinnamomea dropping pills(ACDPs).Based on these chemical components,network pharmacology demonstrated that the targets of active components were significantly enriched in the pathways in cancer,which were closely related with cell proliferation regulation.Next,HCC data was downloaded from Gene Expression Omnibus database(GEO).The Cancer Genome Atlas(TCGA)and Dis Ge NET were analyzed by bioinformatics,and 79 biomarkers were obtained.Furtherly,nine targets of ACDP active components were revealed,and they were significantly enriched in PI3 K/AKT and cell cycle signaling pathways.The affinity between these targets and their corresponding active ingredients was predicted by molecular docking.Finally,in vivo and in vitro experiments showed that ACDPs could reduce the activity of PI3 K/AKT signaling pathway and downregulate the expression of cell cycle-related proteins,contributing to the decreased growth of liver cancer.Altogether,PI3 K/AKT-cell cycle appears as the significant central node in anti-liver cancer of A.Cinnamomea.

Anti-Obesity and Anti-Hyperlipidemia Effects of Antrodia cinnamomea Mycelial Extract in High-Fructose Diet-Fed Mice

This study investigated hypolipidemic,weight-reducing,and hepatoprotective effects of Antrodia cinnamomea mycelial extract obtained from solid-state culture(ACME)in an HFr D-induced obese/hyperlipidemic mouse model.Following 4-week ACME treatment,body weight,epididymal fat index,and some serum biochemical indices were measured.Expression levels of some related genes involved in cholesterol and lipid metabolism were analyzed by q RT-PCR.Moreover,histological studies of hepatic tissues were also conducted.After ACME treatment,body weight and epididymal fat index were significantly lower than that in model control group.ACME and simvastatin significantly reduced serum total cholesterol(T-CHO)and low-density lipoprotein cholesterol(LDL-C)levels,and increased high-density lipoprotein cholesterol(HDL-C)level.Subsequent experiments showed that:(i)ACME regulated transcriptional expression of 3-hydroxy-3-methylglutaryl-Co A reductase(HMGR),low-density lipoprotein receptor(LDLR),adipose triglyceride lipase(ATGL),and fatty acid synthase(FAS),with consequent reduction of blood lipid levels and body weight;(ii)ACME enhanced total antioxidant capacity(T-AOC)and superoxide dismutase(SOD)activity in hepatic tissues;(iii)ACME reduced malondialdehyde(MDA)level and ameliorated lipid oxidative damage in liver.Our findings indicate that ACME is a strong candidate for development as a novel anti-hyperlipidemia and anti-obesity health product.

牛樟树水提物中促进樟芝无性产孢的效应物鉴定及其添加量优化

为明确牛樟树水提物(aqueous extract of Cinnamomum kanehirae Hay,CWE)中促进樟芝(Antrodia cinnamomea)深层发酵无性产孢的效应物。首先,采用醇沉法及不同有机溶剂分级萃取法对CWE进行分离,并以产孢量为检测指标,考察不同组分对樟芝无性产孢的影响。结果表明,30μg/mL的CWE醇沉上清氯仿萃取物LFE和50μg/mL的醇沉上清乙酸乙酯萃取物YZE对樟芝无性产孢有明显的促进效果,且LFE的促进效果明显优于YZE,表明效应物主要存在于LFE和YZE中。随后,采用液相色谱-质谱联用仪对CWE不同萃取组分进行成分分析,并结合不同组分对樟芝产孢的影响效果,确定主要效应物为赤藓糖醇。最后,考察赤藓糖醇(纯度98%)对樟芝深层发酵性能的影响,结果表明,赤藓糖醇对樟芝无性产孢有显著促进效果,且在最佳添加量(1.0μg/mL)下,产孢量较对照组提高55.17%;同时,1.0μg/mL赤藓糖醇显著促进樟芝深层发酵菌丝体生长及胞内多糖的合成,生物量和胞内多糖产量较对照组分别提高18.65%和260.13%,但对三萜的合成无显著影响。

三种单萜对皿式培养樟芝三萜合成的影响

三萜是药食用真菌樟芝野生子实体中主要的活性成分之一,但是人工皿式培养的樟芝菌丝体中三萜含量较低。为了提高樟芝皿式培养三萜的产量,该研究在培养基中添加了3种单萜作为刺激因子。通过添加浓度的优化,得到了对生物量具有显著促进作用的添加方式是0.08 g/L的α-松油醇;对三萜产率具有显著促进作用的最佳添加方式为0.16 g/L的松油烯、0.08 g/L的α-松油醇以及0.12 g/L的4-萜烯醇,三萜产率分别较对照组提高了55.68%、50.39%和28.56%;且在添加0.16 g/L的松油烯时,有两种三萜(去氢硫色多孔菌酸和去氢齿孔菌酸)在樟芝菌丝体中的含量显著升高,分别较对照组提高了5.56倍和0.96倍,在添加0.08 g/L的α-松油醇时,去氢硫色多孔菌酸的含量显著升高,较对照组提高了3.20倍。该研究发现了3种可以促进樟芝皿式培养三萜生成的单萜,为樟芝皿式培养提供了新的思路。

牛樟芝的质量标准研究

目的:建立牛樟芝药材的质量标准。方法:建立牛樟芝药材性状及显微鉴别特征、水分、总灰分、黄曲霉毒素、基因序列鉴定及浸出物含量以制定相应项目的合理限度。结果:性状及显微鉴别方法专属性强,15批样品的水分、总灰分和浸出物平均值分别为13.0%、3.1%、40.4%。结论:制定的鉴别项、检查项、含量项等方法可应用于牛樟芝药材的质量控制。

2种樟树叶片基质添加对牛樟芝三萜含量及抗氧化活性影响

比较分析在大米培养基中添加2种樟树不同浓度叶片匀浆对牛樟芝三萜含量及抗氧化活性的影响,筛选出最适的培养条件,为牛樟芝人工培养及樟属植物资源开发利用提供参考。以大米培养基为对照,分别添加0、0.5、1、2、3、4 g共6个浓度的牛樟和香樟叶片匀浆基质,测量了6种不同处理下牛樟芝的生长特性及三萜含量,并比较其对DPPH自由基清除能力和总还原能力,以评价牛樟芝的抗氧化活性。结果表明:不同浓度的牛樟和香樟叶片匀浆基质添加均能显著提高牛樟芝的生长和生物活性,各处理分别在9~27 d完成高峰生长。经过生长特性因子综合评价得出,在大米培养基中添加3 g香樟基质显著促进了牛樟芝的生长,较对照提高了4倍,为促进牛樟芝生长的最优处理;添加1 g牛樟基质显著提高了牛樟芝的三萜含量、DPPH清除能力及总还原能力,较对照分别提高了5.2、15.8和23倍,为提高牛樟芝三萜含量及抗氧化活性的最优处理。

牛樟芝培养物对产蛋高峰后期蛋鸡生产性能、蛋品质及血清抗氧化指标的影响

试验旨在探究牛樟芝培养物对产蛋高峰后期蛋鸡生产性能、蛋品质及血清抗氧化指标的影响。选取体况和产蛋性能相近的处于产蛋高峰后期(400日龄)海兰白蛋鸡288只,随机分为4组,分别为对照组及牛樟芝低、中、高剂量组。对照组蛋鸡饲喂基础饲粮,牛樟芝低、中、高剂量组蛋鸡分别在基础饲粮中添加50、100 mg/kg和200 mg/kg牛樟芝培养物。预试期7 d,正试期28 d。结果显示:与对照组相比,牛樟芝培养物组产蛋率、日均采食量和日均产蛋重均略有提高,50 mg/kg和100 mg/kg牛樟芝培养物组料蛋比略有降低,但差异不显著(P>0.05);牛樟芝培养物对蛋品质无显著影响(P>0.05);随牛樟芝培养物添加量的增加暗红色肝脏占比逐渐增大,200 mg/kg牛樟芝培养物组暗红色肝脏占比达到83.33%;随牛樟芝培养物添加量的增加,蛋鸡肝脏组织切片病变情况得以减轻或消失;200 mg/kg牛樟芝培养物组超氧化物歧化酶(SOD)含量显著高于50 mg/kg组和对照组(P<0.05),丙二醛(MDA)含量和谷胱甘肽过氧化物酶(GSH-Px)活性各组间差异不显著(P>0.05);饲粮中添加50 mg/kg和100 mg/kg牛樟芝培养物,72只蛋鸡饲喂28 d分别可以额外获得34.4元和58.4元利润。综上所述,牛樟芝培养物可改善产蛋高峰后期蛋鸡生产性能,减少蛋鸡脂肪肝发生率,改善血清抗氧化指标,并增加养殖户的养殖收益,其中以牛樟芝100 mg/kg组经济效益最好。

牛樟芝总三萜提取工艺优化及其抗氧化活性研究

牛樟芝是我国台湾特有的珍稀药食两用真菌,含有丰富活性成分,具有抗癌、抗肿瘤和保肝等多种药理作用,三萜类化合物被认为是牛樟芝中的重要药用物质,不同提取工艺对牛樟芝总三萜得率影响不同.为探究牛樟芝总三萜最佳提取工艺及其抗氧化活性.选取不同提取溶剂、温度、时间、液料比,采用单因素试验方法和正交试验方法探究不同因素对牛樟芝总三萜得率的影响,并通过高效液相色谱(HPLC)分析不同溶剂提取工艺之间的成分差异,比较分析其抗氧化活性.结果表明,牛樟芝总三萜提取最佳工艺为:二氯甲烷为提取溶剂、液料比1∶320 g·mL^(-1)、45℃提取5 h.在此条件下,牛樟芝总三萜得率为10.16±0.077 6%,通过HPLC分析,所得粗提物成分丰富且具有较好的抗氧化活性,其中DPPH清除能力的IC50值达0.137 6,总还原能力达58.03%.通过对牛樟芝提取工艺优化显著提高了牛樟芝总三萜得率,为牛樟芝活性成分提取、化合物分离及开发利用提供理论参考.

Cocktail探针法评价牛樟芝醇提物对大鼠肝CYP450酶活性的作用

目的:研究牛樟芝醇提物对大鼠肝微粒体CYP450亚酶活性的影响。方法:建立大鼠肝微粒体体外代谢的HPLC法,采用Cocktail探针法评价牛樟芝醇提物对大鼠肝微粒体CYP450亚酶的作用。(1)分别精密称取美托洛尔、奥美拉唑、非那西汀、氯唑沙宗、甲苯磺丁脲、硝苯吡啶各适量,配制Cocktail混合探针底物溶液。(2)设活性组、无活性组和抑制剂组,活性组:大鼠肝微粒体活性未进行灭活处理;无活性组:大鼠肝微粒体被灭活,其他同活性组;抑制剂组(分别为奎宁丁、噻氯匹定、α-萘黄酮、二乙氨基二硫代甲酸钠、磺胺苯吡唑、酮康唑组):在活性组的基础上加抑制剂各2.5μl。检测各组CYP2D6、CYP2C19、CYP1A2、CYP2E1、CYP2C9、CYP3A4/5亚酶的相对活性。结果:与无活性组比较,活性组中CYP2D6、CYP2C19、CYP1A2、CYP2E1、CYP2C9、CYP3A4/5相对酶活性升高,差异均有统计学意义(P<0.05)。与活性组比较,牛樟芝高、中、低剂量组中CYP1A2和CYP3A4/5相对酶活性降低,差异有统计学意义(P<0.05);CYP2C9相对酶活性降低,但差异无统计学意义(P>0.05);高、中剂量组中CYP2D6相对酶活性降低,高剂量组中CYP2E1、低剂量组中CYP2C19相对酶活性降低,高剂量组中CYP2C19相对酶活性升高,差异均有统计学意义(P<0.05)。结论:牛樟芝醇提物对大鼠肝微粒体CYP450亚酶中CYP1A2、CYP3A4/5、CYP2C9、CYP2D6、CYP2E1的代谢活性在一定浓度范围内有抑制作用,而对CYP2C19亚酶的代谢活性在一定浓度范围内有促进作用。

不同碳氮源对牛樟芝菌丝体生长的影响

为研究不同碳氮源对牛樟芝菌丝体生长的影响,采用不同碳源（甘露醇、麦芽糖、山梨醇、蔗糖、乳糖、葡萄糖、肌醇、果糖、可溶性淀粉）、不同氮源（麦芽浸粉、酵母提取物、番茄浸粉、马铃薯浸粉、大豆蛋白胨、胰蛋白胨、牛肉浸粉、酪蛋白胨、麦芽浸粉肉汤）及不同浓度果糖（0．2%~3．2%）对牛樟芝菌丝体进行培养,观测其菌落大小、菌落长势、生长速率及菌丝生长指数。结果表明,以麦芽浸粉与酵母提取物为氮源,9种糖类中果糖作为碳源时,牛樟芝菌丝生长速度最快,培养30d时的菌丝生长速率为1．47mm/d,且菌丝呈红色,与野生牛樟芝子实体颜色接近。以麦芽糖和果糖为碳源,9种提取物中牛肉浸粉作为氮源时,牛樟芝生长最佳,培养30d时的菌丝生长速率为1．51mm/d,但其菌丝呈黄白色。果糖浓度在4%以内,生长的各个指标均随着果糖浓度的升高而升高。本研究为今后研究牛樟芝菌种扩繁、大规模发酵及椴木接种提供了实验依据。

牛樟芝药用功能成分作用及应用进展

牛樟芝(Antrodia cinnamomea)是珍贵的药用大型真菌,野生子实体专性寄生于台湾特有牛樟树心材内壁。文章综述了牛樟芝的分类、形态、生境生态等方面的研究进展,并阐述了其在药用化学成分、药理作用、毒性安全评价及药物应用等方面的研究应用前景。

牛樟芝ITS序列分析及液体发酵菌丝体总三萜含量测定

为筛选富萜牛樟芝(Antrodia cinnamomea)菌株,对收集的3株牛樟芝菌株(AC620,AC1404,AC001)进行ITS序列分析,通过香草醛-冰醋酸法对牛樟芝液体发酵菌丝体总三萜含量进行分析。ITS序列分析结果表明菌株AC1404和AC001的ITS序列完全相同,长度均为593 bp,AC 620菌株的长度为581 bp。供试菌株AC001、AC1404和AC620菌丝体中的三萜含量分别为34.30、18.47和16.68 mg·g-1(DW),菌株AC001与AC1404和AC620间菌丝三萜含量差异极显著(P<0.01)。

牛樟芝发酵液提取物抗菌活性研究

牛樟芝作为一种珍稀食用和药用菌,具有极大的开发潜力。该研究以麦芽浸粉肉汤液体培养基(BD,美国BD公司)对牛樟芝菌丝体进行摇床培养60 d后,收获发酵液并用乙酸乙酯对其进行萃取,浓缩至干获得提取物;同时,采用抑菌圈法评价培养物对13种致病细菌抗菌活性(蜡样芽孢杆菌、缓慢芽孢杆菌、无乳链球菌、短小芽孢杆菌、福氏志贺氏菌、枯草芽孢杆菌、金黄色葡萄球菌、藤黄微球菌、副溶血性弧菌、溶血性葡萄球菌、铜尿假单胞菌、乙型副伤寒沙门氏菌、大肠埃希菌),并检测相应致病细菌的最低抑制浓度(MIC)。结果表明:牛樟芝麦芽浸粉肉汤发酵液提取物对供试的13种致病菌均有抑菌活性;在供试的13种致病菌中,提取物对缓慢芽孢杆菌、短小芽孢杆菌、枯草芽孢杆菌、副溶血性弧菌、藤黄微球菌5种致病菌的最低抑制浓度值均小于80μg·m L^(-1),其中对藤黄微球菌的最低抑制浓度最低为66.5μg·m^(-1);随着培养时间的增加,提取物的抗菌活性也增加。这说明牛樟芝菌丝体在液体培养条件下,能够产生广谱高效抑菌活性的次生代谢产物。该研究结果为牛樟芝进一步的有效利用开发奠定了理论基础。

牛樟芝对易卒中的自发性高血压大鼠脑蛋白质组学的影响

目的研究牛樟芝能否延长易卒中的自发性高血压大鼠的生存时间,并运用蛋白质组学方法对其作用机制进行探讨。方法先选取80只雄性易卒中的自发性高血压大鼠,随机分为给药组和对照组(每组40只),给药组每天用牛樟芝(150mg/kg)灌胃治疗,记录卒中大鼠的自然死亡时间。另外再选取6只易卒中的自发性高血压大鼠,随机分为给药组和对照组(每组3只),给药组给予牛樟芝灌胃治疗(150mg/kg),连续90d后取大鼠脑组织进行蛋白质组学研究,用WKY大鼠作为正常对照,对用药前后产生的差异蛋白点进行质谱鉴定,并用蛋白质印迹法进一步验证组学的鉴定结果。结果牛樟芝延长了易卒中的自发性高血压大鼠的生存时间(P<0.05),蛋白质组学鉴定结果显示牛樟芝上调了脑组织抗氧化酶谷胱甘肽巯基转移酶(GST)和超氧化物歧化酶(SOD)的表达(P<0.05),这一结果得到了蛋白质印迹法的验证。对脑组织氧化应激水平的研究发现机体的总抗氧化能力增强[T-AOC:(66.48±16.17)U/g vs(124.75±28.43)U/g,P<0.05),表现在GST和SOD的酶活性增加[GST:(40.33±5.24)U/mg vs(70.50±6.24)U/mg,P<0.05;SOD:(109.25±23.61)U/mg vs(192.60±23.95)U/mg,P<0.05],而氧化应激产物丙二醛含量减少[(3.96±0.45)nmol/mg vs(2.04±0.31)nmol/mg,P<0.05]。结论长期服用牛樟芝能延长易卒中的自发性高血压大鼠的生存时间,这可能与其增加脑组织抗氧化酶的表达、减轻氧化应激的损伤有关。

牛樟芝固态发酵产安卓奎诺尔工艺研究

对牛樟芝Antrodia cinnamomea固态发酵生产安卓奎诺尔的工艺条件进行优化。先通过单因素试验,筛选出小麦Triticum aestivum质量、相对湿度、培养温度3个主要影响参数,再应用响应曲面分析法优化。结果表明,牛樟芝产安卓奎诺尔固态发酵最佳条件为:小麦80.82 g,相对湿度58.01%,培养温度28.48℃,安卓奎诺尔的平均产量可达259.79 mg·kg^(-1)。

牛樟芝和沉水樟芝的物种同源性及质量控制研究

目的比较研究牛樟芝和沉水樟芝药材间的物种同源性,进行质量控制研究。方法建立牛樟芝和沉水樟芝药材的HPLC指纹图谱,采用Diamonsil C18(2)色谱柱(250 mm×4.6 mm,5μm),乙腈-1%甲酸水为流动相梯度洗脱,流速0.8 m L/min,检测波长254 nm,柱温30℃。结果牛樟芝和沉水樟芝的HPLC指纹图谱中标定了9个共有峰,11批牛樟芝相似度在0.906~0.995之间,10批沉水樟芝相似度在0.956~0.998之间。结论本法简便、重复性好,牛樟芝和沉水樟芝的HPLC指纹图谱和红外光谱具有一定相似性。