牛樟芝滴丸保肝功效及药代动力学研究
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摘要
目的：1.将牛樟芝子实体的水及醇的提取物制备成滴丸剂型,建立滴丸质量控制标准,确定最佳牛樟芝滴丸处方,并完成可制备工艺研究。2.比较牛樟芝子实体滴丸及牛樟芝子实体粉末口服后,于小鼠体内药物吸收度及血中浓度变化之药代动力学关系。3.观察牛樟芝子实体滴丸对腹腔注射APAP的大白鼠肝功能指数之影响与变化情形,以及SOD及GPx等相关抗氧化酶酵素的活性表现,探讨其对于肝脏损伤的保护功效。
方法：1.采用固体分散技术,将牛樟芝子实体的水及醇的提取物制备成滴丸剂型,并以实验设计法找出最佳处方。其中以管拄法分离纯化出已知保肝成份Antcin B,以做为牛樟芝滴丸成分分析的指标成份。再利用液相层析仪、傅立叶变换红外光谱仪、电子显微镜检查及体外溶出试验等建立滴丸质量控制标准。2.由体内血清药代动力学试验,进行牛樟芝滴丸口服生体可用率药理活性研究,依给药方式分为静脉注射及口服两组,以成年雄性Sprague-Dawley大白鼠(230-280g)进行实验,每组六只大白鼠,共十二只。并于颈静脉插管,作为采血途径。以乙醇溶解牛樟芝滴丸,剂量为50mg/kg,牛樟芝粉末,剂量为1g/kg,手术后十二小时给药,由颈静脉抽取0.2mL之血液,去除蛋白质,取上层澄清液以HPLC进行AntcinB含量分析。3.护肝功能药理活性研究将研究组别分为A.正常对照组：生理食盐水(I.P.)+d.d.H20(P.0.)B.负对照组(肝损伤组)：400mg APAP/kg bw(I.P.)+d.d.H20(P.0.)C正对照组(N-acetylcysteine(NAC)治疗对照组)：400mg APAP/kg bw(I.P.)+600mg NAC/kg bw(P.0.)D牛樟芝滴丸一倍剂量(1X)实验组：400mg APAP/kg bw(I.P.)+牛樟芝滴丸(20颗/kg bw)(P.O.)E.牛樟芝滴丸3倍剂量(3X)实验组：400mg APAP/kg bw(I.P.)+牛樟芝滴丸(60颗/kg bw)(P.O.)F.牛樟芝滴丸5倍剂量(5X)实验组：400mg APAP/kg bw(I.P.)+牛樟芝滴丸(100颗/kgbw)(P.O.)。A组腹腔注射生理食盐水(0.6ml/30g bw for mice),B-F组腹腔注射400mgAPAP/kg bw(0.6ml/30g bw for mice),每周两次(星期一及四)于样品灌喂前一小时进行肝炎诱导,所有实验动物每周投予试验样品(d.d.H2O或NAC)后4小时,以尾部采血方式检测肝脏的生化功能：GOT(AST)、GPT(ALT)。最后于第八周结束时全部牺牲,并剖腹取肝脏标本,进行病理学观察并做胶原蛋白的特殊染色(Masson's trichrome stain)。另外将其余肝脏分别进行检测抗氧化成分glutathione (GSH)、 glutathione peroxidase (GSH Px)、glutathione reductase (GSH Rd)、glutathione S-transferase (GST)、superoxide dismutase (SOD)与catalase等酵素活性。
结果：
1.通过对牛樟芝提取分离纯化工艺及HPLC结果,建立了保肝成份Antcin B标准品的质量控制标准及确效。使用固体分散技术将牛樟芝水及醇提取物浓缩乳化后与固体基质(PEG6000)熔融成固体分散溶液,以滴制法制备牛樟芝滴丸。利用实验设计法找出最佳处方,得到三萜类组分：载体：乳化剂=1：2.5：0.5为牛樟芝滴丸最佳条件。建立完成了牛樟芝滴丸之优化条件研究方法、质量标准及可制备工艺的研究。使得本研究所使用之样品质量均一。
2.生物利用度实验结果,于大白鼠体内测得其Antcin B之最高血中(峰)浓度,牛樟芝滴丸0.0310±0.004μg/ml,牛樟芝粉末0.0291±0.004μg/m1；达最高血中浓度(峰)时间牛樟芝滴丸26min,牛樟芝粉末45mim。AUC/Dose牛樟芝滴丸0.0478,牛樟芝粉末0.00721。由本药代动力学实验得知,牛樟芝滴丸之口服生物利用度,比粉末高6.6倍。
3.治疗组的GOT及GPT指数,与负对照组相较可随着牛樟芝滴丸剂量的增加,呈现剂量相关性的降低。GSH浓度则较负对照组显着的提高；并具剂量相关性。GPx活性,可随着牛樟芝滴丸剂量的增加,逐渐地恢复；Mn-SOD的活性或是Cu/Zn-SOD的活性皆会随着牛樟芝滴丸剂量的增加而逐渐的提高。病理切片观察显示：五倍剂量(5X)牛樟芝滴丸的作用下,肝脏组织内伪小叶形成及发炎细胞浸润的现象由原来的中等程度转为少量程度；同时,中央静脉和门脉区间中隔的形成较不完全,显示肝纤维化的程度较负对照组轻微。
结论：
1.分离并建立保肝成份Antcin B的质量控制标准;
2.确定最佳牛樟芝滴丸处方,并完成可制备工艺研究;
3.牛樟芝滴丸对于肝脏细胞具有保护作用。
4.牛樟芝滴丸较粉末吸收快、较高吸收度：滴丸能增加三萜类的口服吸收,作用迅速、生物利用度高。
5.牛樟芝滴丸能使因被APAP损坏之肝细胞其粒腺体及网状内皮组织系统恢复正常,对于肝功能指数或是抗氧化酶具有回复作用。提供了牛樟芝滴丸对于肝脏细胞具有保护作用的科学证据。
Objects:
1.Antrodia cinnamomea fruiting bodies of water and alcohol extract preparation into dripping pills dosage forms, the establishment of dripping pills quality control standards, to determine the best Antrodia cinnamomea dripping pills (ACDP)prescription and completed can be prepared by technology research.
2. Comparison of Antrodia cinnamomea fruiting bodies dripping pills and the powder of Antrodia cinnamomea fruiting bodies after oral administration in mice in vivo drug absorption pharmacokinetics between degree of blood concentration.
3. Observation of Antrodia cinnamomea fruiting bodies of dropping pills on the intraperitoneal injection of APAP the rat liver function indexand changes in circumstances, as well as antioxidant enzymes suchas SOD and GPx activity performance, and its efficacy for the protection of the liver damage.
Methods:
1. Using solid dispersion technique, Antrodia cinnamomea fruiting bodies of water and alcohol extract preparation into Dripping Pill formulations, and experimental design to find the best prescription. Which the column were isolated and purified a known hepatoprotectivc the composition Antcin R, as the index composition analysis of ACDP ingredients. And use of liquid chromatography, Fourier transform infrared spectroscopy, electron microscopy and in vitro dissolution tests to establish the dripping pills quality control standards.
2. By the serum pharmacokinetics testing, pharmacological studies of the oral bioavailability of ACDP, Intravenous and oral administration of two groups, according to themode of administration is divided into adult male surname Sprague-Dawley rats (230-280g) experiment, groups of six rats, a total of twelve. And jugular vein cannulation, as the blood taken pathway. ACDP dissolved in ethanol.a dose of50mg/kg, Antrodia cinnamomea powde at a dose of lg/kg and12hours after surgery, administration of0.2mL of blood taken from the jugular vein,to remove the protein, from the upper to clarifyliquid Antcin B content by HPLC analysis.
3. Hepatoprotective pharmacological studios on the research group is divided into A. the normal control group:saline (IP)+ddH2O (PO) B. The negative control group (liver injury group):400mg APAP/kgbw (IP)+d. d. H20(PO) C. The control group-acetylcysteine (NAC) treatment control group):400mg APAP/kgbw(IP)+600mg of NAC/kgbw(PO) D. ACDP single the dose (1X) experimental group:400mg APAPAgbw(IP)+ACDP (20/kgbw)(PO) E. ACDP three times the dose(3X) experimental group:400mg of APAP/kgbw(IP)+ACDP (60/kgbw)(PO) F. ACDP five times the dose (5X) the experimental group:400mg of APAP/kgbw(IP)+ACDP (100/kgbw)(PO). A group of intraperitoneal injection of saline(0.6ml/30g bw formice.), The B-F group by intraperitoneal injection of400mg APAP/kg bw (0.6ml/30g bw for mice.)Twice a week (Monday and Thursday) in the samples were fed one hour before induction ofhcpatitis, all animals weekly administration of the test sample(d. d.1120or NAC),four hours after tail blood biochemical detection of liver function:GOT (AST), GPT (ALT). Finally, at the end of the eighth week, all the expense of laparotomy and liver specimens for pathological examination and do special staining of Masson'strichrome. In addition to the remaining liver were the detection of anti-oxidant glulathione (GSH), glutathione peroxidase (GSHPx) and glutathione reductase,(GSHRd), glutathione S-transferase (GST) of superoxide dismutase(SOD) and catalase enzyme activity.
Results:
1. According to the separation and purification of Antrodia cinnamomea extract and HPLC results. Established hepatoprotective the ingredients Antcin B, for the quality control standard and validation. Antrodia cinnamomea after the emulsification of water and alcohol extract of the solid matrix (PEG6000) melt into the solid dispersion solution using solid dispersion technology. The use of experimental design method to find the best conditions of ACDP is triterpenoid components:carrier:emulsifier=1:2.5:0.5. Established and completed ACDP of optimal conditions, quality standards and the preparation process. Uniform quality of the sample used in this study.
2. The bioavailability pharmacokinetics experimental results obtained, the concentration in the blood of the rat body measured of obtaining Antcin B, ACDP0.0470μg/ml Antrodia cinnamomea powder0.0200μg/ml;reached the highest the blood concentration time ACDP0.50hr faster than powder0.75hr Antrodia cinnamomea.
3. GOT and GPT index of the treatment groups, compared with negative control group. With the increase of ACDP dose, a dose-related reduce. GSH concentration than the negative control group was significantly improved; and a dose-related. GPx activity with increasing doses of ACDP, the gradual recovery;Mn-SOD activity, or of Cu/Zn-SOD activity were gradually increased with increasing Dripping Pills dose of Antrodia cinnamomea. Pathological observation:five times the dose (5X) ACDP under pseudo-lobule within the liver tissue formation and inflammatory cell infiltration from moderate into a small amount of the degree; the same time, the central vein andportal interval septum formation is less complete, show the degree of hepatic fibrosis is minor than the negative control group.
Conclusion:
1. separation and the establishment of the hepatoprotective ingredients Antcin B, quality control standards.
2. To determine the best ACDP prescription, and completed research can be prepared by process.
3. ACDP for the liver cells have a protective effect.
4. ACDP than the powder to absorb quickly, a higher absorption rate. Dripping Pills can increase the the triterpenoids oral, absorption, the role of rapid inc.reasebioavailability.
5. ACDP can make because of the APAP damage of liver cell mitochondria and the reticulo endothelial system of organizations back to normal,for liver function or antioxidant enzymes reply. For the liver cells ACDP have a protective effect of scientific evidence.
方法：1.采用固体分散技术,将牛樟芝子实体的水及醇的提取物制备成滴丸剂型,并以实验设计法找出最佳处方。其中以管拄法分离纯化出已知保肝成份Antcin B,以做为牛樟芝滴丸成分分析的指标成份。再利用液相层析仪、傅立叶变换红外光谱仪、电子显微镜检查及体外溶出试验等建立滴丸质量控制标准。2.由体内血清药代动力学试验,进行牛樟芝滴丸口服生体可用率药理活性研究,依给药方式分为静脉注射及口服两组,以成年雄性Sprague-Dawley大白鼠(230-280g)进行实验,每组六只大白鼠,共十二只。并于颈静脉插管,作为采血途径。以乙醇溶解牛樟芝滴丸,剂量为50mg/kg,牛樟芝粉末,剂量为1g/kg,手术后十二小时给药,由颈静脉抽取0.2mL之血液,去除蛋白质,取上层澄清液以HPLC进行AntcinB含量分析。3.护肝功能药理活性研究将研究组别分为A.正常对照组：生理食盐水(I.P.)+d.d.H20(P.0.)B.负对照组(肝损伤组)：400mg APAP/kg bw(I.P.)+d.d.H20(P.0.)C正对照组(N-acetylcysteine(NAC)治疗对照组)：400mg APAP/kg bw(I.P.)+600mg NAC/kg bw(P.0.)D牛樟芝滴丸一倍剂量(1X)实验组：400mg APAP/kg bw(I.P.)+牛樟芝滴丸(20颗/kg bw)(P.O.)E.牛樟芝滴丸3倍剂量(3X)实验组：400mg APAP/kg bw(I.P.)+牛樟芝滴丸(60颗/kg bw)(P.O.)F.牛樟芝滴丸5倍剂量(5X)实验组：400mg APAP/kg bw(I.P.)+牛樟芝滴丸(100颗/kgbw)(P.O.)。A组腹腔注射生理食盐水(0.6ml/30g bw for mice),B-F组腹腔注射400mgAPAP/kg bw(0.6ml/30g bw for mice),每周两次(星期一及四)于样品灌喂前一小时进行肝炎诱导,所有实验动物每周投予试验样品(d.d.H2O或NAC)后4小时,以尾部采血方式检测肝脏的生化功能：GOT(AST)、GPT(ALT)。最后于第八周结束时全部牺牲,并剖腹取肝脏标本,进行病理学观察并做胶原蛋白的特殊染色(Masson's trichrome stain)。另外将其余肝脏分别进行检测抗氧化成分glutathione (GSH)、 glutathione peroxidase (GSH Px)、glutathione reductase (GSH Rd)、glutathione S-transferase (GST)、superoxide dismutase (SOD)与catalase等酵素活性。
结果：
1.通过对牛樟芝提取分离纯化工艺及HPLC结果,建立了保肝成份Antcin B标准品的质量控制标准及确效。使用固体分散技术将牛樟芝水及醇提取物浓缩乳化后与固体基质(PEG6000)熔融成固体分散溶液,以滴制法制备牛樟芝滴丸。利用实验设计法找出最佳处方,得到三萜类组分：载体：乳化剂=1：2.5：0.5为牛樟芝滴丸最佳条件。建立完成了牛樟芝滴丸之优化条件研究方法、质量标准及可制备工艺的研究。使得本研究所使用之样品质量均一。
2.生物利用度实验结果,于大白鼠体内测得其Antcin B之最高血中(峰)浓度,牛樟芝滴丸0.0310±0.004μg/ml,牛樟芝粉末0.0291±0.004μg/m1；达最高血中浓度(峰)时间牛樟芝滴丸26min,牛樟芝粉末45mim。AUC/Dose牛樟芝滴丸0.0478,牛樟芝粉末0.00721。由本药代动力学实验得知,牛樟芝滴丸之口服生物利用度,比粉末高6.6倍。
3.治疗组的GOT及GPT指数,与负对照组相较可随着牛樟芝滴丸剂量的增加,呈现剂量相关性的降低。GSH浓度则较负对照组显着的提高；并具剂量相关性。GPx活性,可随着牛樟芝滴丸剂量的增加,逐渐地恢复；Mn-SOD的活性或是Cu/Zn-SOD的活性皆会随着牛樟芝滴丸剂量的增加而逐渐的提高。病理切片观察显示：五倍剂量(5X)牛樟芝滴丸的作用下,肝脏组织内伪小叶形成及发炎细胞浸润的现象由原来的中等程度转为少量程度；同时,中央静脉和门脉区间中隔的形成较不完全,显示肝纤维化的程度较负对照组轻微。
结论：
1.分离并建立保肝成份Antcin B的质量控制标准;
2.确定最佳牛樟芝滴丸处方,并完成可制备工艺研究;
3.牛樟芝滴丸对于肝脏细胞具有保护作用。
4.牛樟芝滴丸较粉末吸收快、较高吸收度：滴丸能增加三萜类的口服吸收,作用迅速、生物利用度高。
5.牛樟芝滴丸能使因被APAP损坏之肝细胞其粒腺体及网状内皮组织系统恢复正常,对于肝功能指数或是抗氧化酶具有回复作用。提供了牛樟芝滴丸对于肝脏细胞具有保护作用的科学证据。
Objects:
1.Antrodia cinnamomea fruiting bodies of water and alcohol extract preparation into dripping pills dosage forms, the establishment of dripping pills quality control standards, to determine the best Antrodia cinnamomea dripping pills (ACDP)prescription and completed can be prepared by technology research.
2. Comparison of Antrodia cinnamomea fruiting bodies dripping pills and the powder of Antrodia cinnamomea fruiting bodies after oral administration in mice in vivo drug absorption pharmacokinetics between degree of blood concentration.
3. Observation of Antrodia cinnamomea fruiting bodies of dropping pills on the intraperitoneal injection of APAP the rat liver function indexand changes in circumstances, as well as antioxidant enzymes suchas SOD and GPx activity performance, and its efficacy for the protection of the liver damage.
Methods:
1. Using solid dispersion technique, Antrodia cinnamomea fruiting bodies of water and alcohol extract preparation into Dripping Pill formulations, and experimental design to find the best prescription. Which the column were isolated and purified a known hepatoprotectivc the composition Antcin R, as the index composition analysis of ACDP ingredients. And use of liquid chromatography, Fourier transform infrared spectroscopy, electron microscopy and in vitro dissolution tests to establish the dripping pills quality control standards.
2. By the serum pharmacokinetics testing, pharmacological studies of the oral bioavailability of ACDP, Intravenous and oral administration of two groups, according to themode of administration is divided into adult male surname Sprague-Dawley rats (230-280g) experiment, groups of six rats, a total of twelve. And jugular vein cannulation, as the blood taken pathway. ACDP dissolved in ethanol.a dose of50mg/kg, Antrodia cinnamomea powde at a dose of lg/kg and12hours after surgery, administration of0.2mL of blood taken from the jugular vein,to remove the protein, from the upper to clarifyliquid Antcin B content by HPLC analysis.
3. Hepatoprotective pharmacological studios on the research group is divided into A. the normal control group:saline (IP)+ddH2O (PO) B. The negative control group (liver injury group):400mg APAP/kgbw (IP)+d. d. H20(PO) C. The control group-acetylcysteine (NAC) treatment control group):400mg APAP/kgbw(IP)+600mg of NAC/kgbw(PO) D. ACDP single the dose (1X) experimental group:400mg APAPAgbw(IP)+ACDP (20/kgbw)(PO) E. ACDP three times the dose(3X) experimental group:400mg of APAP/kgbw(IP)+ACDP (60/kgbw)(PO) F. ACDP five times the dose (5X) the experimental group:400mg of APAP/kgbw(IP)+ACDP (100/kgbw)(PO). A group of intraperitoneal injection of saline(0.6ml/30g bw formice.), The B-F group by intraperitoneal injection of400mg APAP/kg bw (0.6ml/30g bw for mice.)Twice a week (Monday and Thursday) in the samples were fed one hour before induction ofhcpatitis, all animals weekly administration of the test sample(d. d.1120or NAC),four hours after tail blood biochemical detection of liver function:GOT (AST), GPT (ALT). Finally, at the end of the eighth week, all the expense of laparotomy and liver specimens for pathological examination and do special staining of Masson'strichrome. In addition to the remaining liver were the detection of anti-oxidant glulathione (GSH), glutathione peroxidase (GSHPx) and glutathione reductase,(GSHRd), glutathione S-transferase (GST) of superoxide dismutase(SOD) and catalase enzyme activity.
Results:
1. According to the separation and purification of Antrodia cinnamomea extract and HPLC results. Established hepatoprotective the ingredients Antcin B, for the quality control standard and validation. Antrodia cinnamomea after the emulsification of water and alcohol extract of the solid matrix (PEG6000) melt into the solid dispersion solution using solid dispersion technology. The use of experimental design method to find the best conditions of ACDP is triterpenoid components:carrier:emulsifier=1:2.5:0.5. Established and completed ACDP of optimal conditions, quality standards and the preparation process. Uniform quality of the sample used in this study.
2. The bioavailability pharmacokinetics experimental results obtained, the concentration in the blood of the rat body measured of obtaining Antcin B, ACDP0.0470μg/ml Antrodia cinnamomea powder0.0200μg/ml;reached the highest the blood concentration time ACDP0.50hr faster than powder0.75hr Antrodia cinnamomea.
3. GOT and GPT index of the treatment groups, compared with negative control group. With the increase of ACDP dose, a dose-related reduce. GSH concentration than the negative control group was significantly improved; and a dose-related. GPx activity with increasing doses of ACDP, the gradual recovery;Mn-SOD activity, or of Cu/Zn-SOD activity were gradually increased with increasing Dripping Pills dose of Antrodia cinnamomea. Pathological observation:five times the dose (5X) ACDP under pseudo-lobule within the liver tissue formation and inflammatory cell infiltration from moderate into a small amount of the degree; the same time, the central vein andportal interval septum formation is less complete, show the degree of hepatic fibrosis is minor than the negative control group.
Conclusion:
1. separation and the establishment of the hepatoprotective ingredients Antcin B, quality control standards.
2. To determine the best ACDP prescription, and completed research can be prepared by process.
3. ACDP for the liver cells have a protective effect.
4. ACDP than the powder to absorb quickly, a higher absorption rate. Dripping Pills can increase the the triterpenoids oral, absorption, the role of rapid inc.reasebioavailability.
5. ACDP can make because of the APAP damage of liver cell mitochondria and the reticulo endothelial system of organizations back to normal,for liver function or antioxidant enzymes reply. For the liver cells ACDP have a protective effect of scientific evidence.
引文
1. Anti-fatigue cyclohexenone compounds from Antrodia camphorata. Ao ZH, Xu ZH, Lu ZM, Xu HY, Zhang XM, Dou WF. Niuchangchih (Antrodia camphorata) and its potential in treating liver diseases. J Ethnopharmacol.12009,121 (2):194-212 [2009-02-03]. doi:10.1016/j. jep.2008.10.039. PMID 19061947.
2. Hseu Y-C, Chang W-C, Hseu Y-T, Lee C-Y, Yech Y-J, Chen P-C, Chen J-Y, Yang H-L. (2002). "Protection of oxidative damage by aqueous extract from Antrodia camphorata mycelia in normal human erythrocytes". Life Sciences 71(4):469-482.
3. S.-H. Wu, Z.-H. Yu.'Y.-C. Dai, C.-H. Su, L.-C. Chen, W.-C. Hsu, and G.-Y. Hwang. "Taiwanofungus, a polypore new genus". Fung. Sci.19 (3-4) 109-116,2004
4. Z.-H. Yu, S.-H. Wu, D.-M. Wang, C.-T. Chen. Phylogenetic Relationships of Antrodia Species and Related Taxa Based on Analyses of Nuclear Large Subunit Ribosomal DNA Sequences. Botanical Studies.2010,51 (1):53-60
5. Arthur JR.2000, The glutathione peroxidases. Cell Mol Life Sci 57:1825-1835
6. Gabriele B.1997, Silymarin retards collagen accumulation in early and advanced biliary fibrosis secondary to complete bile duct obliteration in rats. Hepatology 26:643-649
7. Gutteridge JM. Halliwell B.2000, Free radicals and antioxidants in the year 2000. A historical look to the future. Ann NY Acad Sci 899:136-147
8. Hase K. Kadota S. Basnet P. Takahashi T. Namba T.1996, Protective effect of celosian, an acidic polysaccharide, on chemically and immunologically induced liver injuries. Biol Pharma Bulletin 19:567-572
9. Jonker AM. Dijkhuis FWJ. Boes A. Hardonk MJ. Grond J.1992, Immunohistochemical study of extracellular matrix in acute galactosamine hepatitis in rats. Hepatology 15:423-431
10. Ketterer B.1998, Glutathione S-transferases and prevention of cellular free radical damage. Free Radic Res 28:647-658
11.Lowry OH. Rosebrough NJ. Farr AL. Randall RJ.1951, Protein measurement with the Folin phenol reagent. J Biol Chem 193:265-275
12. Ruwart MJ. Wilkinson KF. Rush BD. Vidmar TJ. Peters KM. Henley KS. Appelman HD. Kim KY. Schuppan D. Hahn EG.1989, The integrated value of serum procollagen III peptide over time predicts hepatic hydroxyproline content and stainable collagen in a model of dietary cirrhosis in the rat. Hepatology 10:801-806
13. Wells PG. Kim PM. Laposa RR. Nicol CJ. Parman T. Winn LM.1997, Oxidative damage in chemical teratogenesis. Mutat Res 396:65-78
14.Wong CK. Ooi VE. Ang PO.2000, Protective effects of seaweeds against liver injury caused by carbon tetrachloride in rats. Chemosphere 41:173-176
15. Chang, T. T. and W. N. Chou.1995. Antrodia cinnamomea sp. nov. on Cinnamomum kanehirai in Taiwan. Mycol. Res.99:756-758.
16. Chang, T. T. and W. N. Chou.2004. Antrodia cinnamomea reconsidered and A. salmonea sp. nov. on Cunninghamia konishii in Taiwan. Bot. Bull. Acad. Sinica 45:347-352.
17.McNeill, J., F. R. Barrie, H. M. Burdet, V. Demoulin, D. L. Hawksworth, K. Marhold, D. H. Nicolson, J. Prado, P. C. Silva, J. E. Skog, J. H. Wiersema and N. J. Turland.2006. International Code of Botanical Nomenclature (Vienna Code).
18. Wu, S.H., L. Ryvarden and, T. T. Chang.1997. Antrodia camphorata ("niu-chang-chih"), new combination of a medicinal fungus in Taiwan. Bot. Bull. Acad. Sinica 38:273-275.
19. Wu, S. H., Z. H.Yu, Y. C. Dai, C. T. Chen, C. H. Su, L. C. Chen, W. C. Hsu and G. Y. Hwang.2004. Taiwano fungus, a poly pore new genus. Fung. Sci.19: 109-116.
20. Yu, Z. H., S. H. Wu, D. M. Wang and C. T. Chen.2010. Phylogenetic relationships of Antrodia species and related taxa based on analyses of nuclear large subunit ribosomal DNA sequences. Bot. Stud.51:53-60.
21. Zang, M. and Q. Su.1990. Ganoderma comphoratum, a new taxon in genus Ganoderma from Taiwan, China. Acta Bot. Yunnanica 12:395-396.
22. Chen YY, Chou PY, Chien YC, Wu CH, Wu TS, Sheu MJ. Chem Pharm Bull (Tokyo). 2006 Apr;Ethanol extracts of fruiting bodies of Antrodia cinnamomea exhibit anti-migration action in human adenocarcinoma CL1-0 cells through the MAPK and PI3K/AKT signaling pathways.54(4):496-500.
23. Han HF, Nakamura N, Zuo F, Hirakawa A, Yokozawa T, Hattori M Evid Based Complement Alternat Med.2012;2012:378415. Epub 2012 Feb
21. Protective effects of a neutral polysaccharide isolated from the mycelium of Antrodia cinnamomea on Propionibacterium acnes and lipopolysaccharide induced hepatic injury in mice.
24. Chen YY, Liu FC, Chou PY, Chien YC, Chang WS, Huang GJ, Wu CH, Sheu MJ. Ethanol Extracts of Fruiting Bodies of Antrodia cinnamomea Suppress CL1-5 Human Lung Adenocarcinoma Cells Migration by Inhibiting Matrix Metalloproteinase-2/9 through ERK, JNK, p38, and PI3K/Akt Signaling Pathways.
25.陈经祥(1994)「灵芝菌种接种工具比较与太空包木屑培养基含量对灵芝子实体产量之影响」,中国园艺,
26.程一华(1994)[樟芝之成分研究],国立台湾师范大学化学研究所硕士
27.林文鑫、陈俊宪、陈劲初、吕锋洲,2000,樟芝液态发酵萃取物对肿瘤细胞株之毒杀性分析。
28.陈劲初、林文鑫、陈清农、许胜杰、黄仕政、陈炎炼,2001,台湾特有真菌—樟芝菌丝体之开发,Fungal Science Vol.16, No.1,2 p7-22
29.张中姿、陈俊宪、林文鑫、陈劲初、吕锋洲,2001,深红色樟芝菌丝体之甲醇萃取物对肝癌细胞株之研究。中华民国食品科技学会第三十一次会员大会。
30.陈建志、陈劲初、陈清农、许胜杰、曾虹萍、盛莉莎,2001,樟芝菌丝体对B型肝炎病毒表面抗原及e抗原之活性检测。中华民国食品科技学会第三十一次会员大会
31.蔡雁晖、末祖莹、陈嬿如、颜国钦,2001,樟芝菌丝体液体深层培养液对过氧化氢诱发人类肝细胞氧化伤害之保护作用。中华民国食品科技学会第三十一次会员大会。
32.沈立言、陈怡欣、林文川,2001,牛樟芝菌丝体发酵萃取液对肝脏生理机能性之影响。中华民国食品科技学会第三十一次会员大会。
33.张东柱,1997,牛樟之病害,牛樟生物学及育林技术研讨会论文集,林业丛刊第72号：127—131。
34.简秋源、姜宏哲、陈淑贞,1997,牛樟菇培养性状及其三帖类成分分析之研究,牛樟生物学及育林技术研讨会论文集,林业丛刊第72号：133—137。
35.吴振甫(1992)「洋菇舍微气候控制之模式化与分析」,国立台湾大学农业工程学研究所硕士论文。
36.吴德鹏(1995) 「樟芝微量成分的研究」,国立台湾师范大学化学研究所硕士论文。
37.吴升原(2002)「牛樟抽出物对樟芝生长影响之探讨」,国立台湾大学森林学研究所硕士论文。
38.高郁婷(2004)「牛樟抽出物及精油对樟芝菌丝及子实体生长之影响」, 国立台湾大学环境暨资源学研究所硕士论文。
39.陈清农、陈劲初、张基煌、乔长诚(1999)「樟芝挥发性成分之研究」中华民国食品科学技术学会第二十九次会员大会论文摘要。
40.陈晓娟(2004)「针叶树之牛樟芝菌丝体生长促进因子及牛樟芝菌丝体成分之研究」,台北医学大学生药学研究所硕士论文。
2. Hseu Y-C, Chang W-C, Hseu Y-T, Lee C-Y, Yech Y-J, Chen P-C, Chen J-Y, Yang H-L. (2002). "Protection of oxidative damage by aqueous extract from Antrodia camphorata mycelia in normal human erythrocytes". Life Sciences 71(4):469-482.
3. S.-H. Wu, Z.-H. Yu.'Y.-C. Dai, C.-H. Su, L.-C. Chen, W.-C. Hsu, and G.-Y. Hwang. "Taiwanofungus, a polypore new genus". Fung. Sci.19 (3-4) 109-116,2004
4. Z.-H. Yu, S.-H. Wu, D.-M. Wang, C.-T. Chen. Phylogenetic Relationships of Antrodia Species and Related Taxa Based on Analyses of Nuclear Large Subunit Ribosomal DNA Sequences. Botanical Studies.2010,51 (1):53-60
5. Arthur JR.2000, The glutathione peroxidases. Cell Mol Life Sci 57:1825-1835
6. Gabriele B.1997, Silymarin retards collagen accumulation in early and advanced biliary fibrosis secondary to complete bile duct obliteration in rats. Hepatology 26:643-649
7. Gutteridge JM. Halliwell B.2000, Free radicals and antioxidants in the year 2000. A historical look to the future. Ann NY Acad Sci 899:136-147
8. Hase K. Kadota S. Basnet P. Takahashi T. Namba T.1996, Protective effect of celosian, an acidic polysaccharide, on chemically and immunologically induced liver injuries. Biol Pharma Bulletin 19:567-572
9. Jonker AM. Dijkhuis FWJ. Boes A. Hardonk MJ. Grond J.1992, Immunohistochemical study of extracellular matrix in acute galactosamine hepatitis in rats. Hepatology 15:423-431
10. Ketterer B.1998, Glutathione S-transferases and prevention of cellular free radical damage. Free Radic Res 28:647-658
11.Lowry OH. Rosebrough NJ. Farr AL. Randall RJ.1951, Protein measurement with the Folin phenol reagent. J Biol Chem 193:265-275
12. Ruwart MJ. Wilkinson KF. Rush BD. Vidmar TJ. Peters KM. Henley KS. Appelman HD. Kim KY. Schuppan D. Hahn EG.1989, The integrated value of serum procollagen III peptide over time predicts hepatic hydroxyproline content and stainable collagen in a model of dietary cirrhosis in the rat. Hepatology 10:801-806
13. Wells PG. Kim PM. Laposa RR. Nicol CJ. Parman T. Winn LM.1997, Oxidative damage in chemical teratogenesis. Mutat Res 396:65-78
14.Wong CK. Ooi VE. Ang PO.2000, Protective effects of seaweeds against liver injury caused by carbon tetrachloride in rats. Chemosphere 41:173-176
15. Chang, T. T. and W. N. Chou.1995. Antrodia cinnamomea sp. nov. on Cinnamomum kanehirai in Taiwan. Mycol. Res.99:756-758.
16. Chang, T. T. and W. N. Chou.2004. Antrodia cinnamomea reconsidered and A. salmonea sp. nov. on Cunninghamia konishii in Taiwan. Bot. Bull. Acad. Sinica 45:347-352.
17.McNeill, J., F. R. Barrie, H. M. Burdet, V. Demoulin, D. L. Hawksworth, K. Marhold, D. H. Nicolson, J. Prado, P. C. Silva, J. E. Skog, J. H. Wiersema and N. J. Turland.2006. International Code of Botanical Nomenclature (Vienna Code).
18. Wu, S.H., L. Ryvarden and, T. T. Chang.1997. Antrodia camphorata ("niu-chang-chih"), new combination of a medicinal fungus in Taiwan. Bot. Bull. Acad. Sinica 38:273-275.
19. Wu, S. H., Z. H.Yu, Y. C. Dai, C. T. Chen, C. H. Su, L. C. Chen, W. C. Hsu and G. Y. Hwang.2004. Taiwano fungus, a poly pore new genus. Fung. Sci.19: 109-116.
20. Yu, Z. H., S. H. Wu, D. M. Wang and C. T. Chen.2010. Phylogenetic relationships of Antrodia species and related taxa based on analyses of nuclear large subunit ribosomal DNA sequences. Bot. Stud.51:53-60.
21. Zang, M. and Q. Su.1990. Ganoderma comphoratum, a new taxon in genus Ganoderma from Taiwan, China. Acta Bot. Yunnanica 12:395-396.
22. Chen YY, Chou PY, Chien YC, Wu CH, Wu TS, Sheu MJ. Chem Pharm Bull (Tokyo). 2006 Apr;Ethanol extracts of fruiting bodies of Antrodia cinnamomea exhibit anti-migration action in human adenocarcinoma CL1-0 cells through the MAPK and PI3K/AKT signaling pathways.54(4):496-500.
23. Han HF, Nakamura N, Zuo F, Hirakawa A, Yokozawa T, Hattori M Evid Based Complement Alternat Med.2012;2012:378415. Epub 2012 Feb
21. Protective effects of a neutral polysaccharide isolated from the mycelium of Antrodia cinnamomea on Propionibacterium acnes and lipopolysaccharide induced hepatic injury in mice.
24. Chen YY, Liu FC, Chou PY, Chien YC, Chang WS, Huang GJ, Wu CH, Sheu MJ. Ethanol Extracts of Fruiting Bodies of Antrodia cinnamomea Suppress CL1-5 Human Lung Adenocarcinoma Cells Migration by Inhibiting Matrix Metalloproteinase-2/9 through ERK, JNK, p38, and PI3K/Akt Signaling Pathways.
25.陈经祥(1994)「灵芝菌种接种工具比较与太空包木屑培养基含量对灵芝子实体产量之影响」,中国园艺,
26.程一华(1994)[樟芝之成分研究],国立台湾师范大学化学研究所硕士
27.林文鑫、陈俊宪、陈劲初、吕锋洲,2000,樟芝液态发酵萃取物对肿瘤细胞株之毒杀性分析。
28.陈劲初、林文鑫、陈清农、许胜杰、黄仕政、陈炎炼,2001,台湾特有真菌—樟芝菌丝体之开发,Fungal Science Vol.16, No.1,2 p7-22
29.张中姿、陈俊宪、林文鑫、陈劲初、吕锋洲,2001,深红色樟芝菌丝体之甲醇萃取物对肝癌细胞株之研究。中华民国食品科技学会第三十一次会员大会。
30.陈建志、陈劲初、陈清农、许胜杰、曾虹萍、盛莉莎,2001,樟芝菌丝体对B型肝炎病毒表面抗原及e抗原之活性检测。中华民国食品科技学会第三十一次会员大会
31.蔡雁晖、末祖莹、陈嬿如、颜国钦,2001,樟芝菌丝体液体深层培养液对过氧化氢诱发人类肝细胞氧化伤害之保护作用。中华民国食品科技学会第三十一次会员大会。
32.沈立言、陈怡欣、林文川,2001,牛樟芝菌丝体发酵萃取液对肝脏生理机能性之影响。中华民国食品科技学会第三十一次会员大会。
33.张东柱,1997,牛樟之病害,牛樟生物学及育林技术研讨会论文集,林业丛刊第72号：127—131。
34.简秋源、姜宏哲、陈淑贞,1997,牛樟菇培养性状及其三帖类成分分析之研究,牛樟生物学及育林技术研讨会论文集,林业丛刊第72号：133—137。
35.吴振甫(1992)「洋菇舍微气候控制之模式化与分析」,国立台湾大学农业工程学研究所硕士论文。
36.吴德鹏(1995) 「樟芝微量成分的研究」,国立台湾师范大学化学研究所硕士论文。
37.吴升原(2002)「牛樟抽出物对樟芝生长影响之探讨」,国立台湾大学森林学研究所硕士论文。
38.高郁婷(2004)「牛樟抽出物及精油对樟芝菌丝及子实体生长之影响」, 国立台湾大学环境暨资源学研究所硕士论文。
39.陈清农、陈劲初、张基煌、乔长诚(1999)「樟芝挥发性成分之研究」中华民国食品科学技术学会第二十九次会员大会论文摘要。
40.陈晓娟(2004)「针叶树之牛樟芝菌丝体生长促进因子及牛樟芝菌丝体成分之研究」,台北医学大学生药学研究所硕士论文。