技术领域：Technical field:
本发明属于药物技术领域，具体地说，涉及具有抗肝纤维化活性的呋喃酮聚酮类活性化合物Penicilfuranone A及其制备方法，以该化合物为活性成分的药物组合物，以及其在制备治疗肝纤维化疾病的药物中的应用。The invention belongs to the technical field of medicines, in particular, it relates to a furanone polyketide active compound Penicilfuranone A with anti-hepatic fibrosis activity and a preparation method thereof, a pharmaceutical composition using the compound as an active ingredient, and its preparation for treating liver disease. Drug use in fibrotic diseases.
背景技术：Background technique:
呋喃酮类聚酮类化合物是一类有真菌产生的次生代谢产物，目前见报道于真菌Aspergillus rugulosus,Cephalosporium gregatum,以及Penicillium daleae的次生代谢产物中。迄今为止，文献报道的呋喃聚酮类化合物不足20个化合物，且其分子量皆为300左右。前人研究表明该类化合物具有抗菌，致植物枯萎，抑制哺乳动物Y族DNA酶等作用。本发明化合物结构新颖，分子量为486，与之前报道的该类化合物，结构完全不一样。迄今为止，现有技术中尚未见有本发明的结构新颖的呋喃酮聚酮类化合物被报道，同时也未见任何文献报道该类呋喃酮类化合物具有抗肝纤维化活性的研究。Furanone polyketides are a class of secondary metabolites produced by fungi, which have been reported in the secondary metabolites of fungi Aspergillus rugulosus, Cephalosporium gregatum, and Penicillium daleae. So far, there are less than 20 furan polyketide compounds reported in the literature, and their molecular weights are all about 300. Previous studies have shown that this type of compound has antibacterial effects, causes plant withering, and inhibits mammalian Y family DNA enzymes. The compound of the present invention has a novel structure with a molecular weight of 486, which is completely different from the previously reported compounds. So far, no furanone polyketide compound with novel structure of the present invention has been reported in the prior art, and there is no literature report on the research on the anti-hepatic fibrosis activity of this furanone polyketide compound.
发明内容：Invention content:
本发明目的在于提供从内生真菌Penicillium daleae中分离得到的呋喃聚酮类化合物A，其制备方法，在药物中特别是在抗肝纤维化药物中的应用。The object of the present invention is to provide furanopolyketone compound A isolated from endophytic fungus Penicillium daleae, its preparation method, and its application in medicine, especially in anti-hepatic fibrosis medicine.
为了实现本明的上述目的，本发明提供了如下的技术方案：In order to realize the above-mentioned purpose of the present invention, the present invention provides following technical scheme:
下述结构式(I)所示的青霉呋喃酮A化合物或其药学上可接受的盐，Penicillofuranone A compound represented by the following structural formula (I) or a pharmaceutically acceptable salt thereof,
药物组合物，其包含所述的青霉呋喃酮A化合物或其药学上可接受的盐和至少一种药学上可接受的载体和/或稀释剂。A pharmaceutical composition, which comprises the penicillofuranone A compound or a pharmaceutically acceptable salt thereof and at least one pharmaceutically acceptable carrier and/or diluent.
根据所述的青霉呋喃酮A化合物或其药学上可接受的盐，其中所述的药学上可接受的盐为盐酸盐、氢溴酸盐、硝酸盐、硫酸盐、磷酸盐、酒石酸盐、柠檬酸盐、甲酸盐、乙酸盐、乙二酸盐。According to the penicillin furanone A compound or a pharmaceutically acceptable salt thereof, wherein the pharmaceutically acceptable salt is hydrochloride, hydrobromide, nitrate, sulfate, phosphate, tartrate , citrate, formate, acetate, oxalate.
本发明同时提供了青霉呋喃酮A化合物或其药学上可接受的盐的制备方法，取青霉呋喃酮A Penicillium daleae发酵物，用有机溶剂提取，浓缩得浸膏，分别用乙酸乙酯萃取，硅胶层析柱，制备型高效液相色谱等方法得结构式(I)化合物。The present invention also provides a preparation method of the penicillium furanone A compound or a pharmaceutically acceptable salt thereof. The penicillium furanone A Penicillium daleae fermentation product is extracted with an organic solvent, concentrated to obtain an extract, and extracted with ethyl acetate respectively. , silica gel chromatography column, preparative high performance liquid chromatography and other methods to obtain the compound of structural formula (I).
更具体的制备方法如下：取Penicillium daleae菌丝体接种于灭菌马铃薯葡萄糖培养液中PDA，于室温下以170转每分钟在摇床中震荡五天，做成种子液，将种子液转接到灭菌的大米培养基中，28℃条件下无菌培养40天，用乙酸乙酯提取利用大米培养基培养40天后的菌丝体，共提取四次，得到乙酸乙酯提取物，利用硅胶柱色谱，高效液相色谱质谱联用，制备型高效液相色谱处理乙酸乙酯提取物，最后得到青霉呋喃酮A，其制备液相色谱条件为：12mL/min,UVλmax 230nm,MeCN-H2O 45:55，保留时间15.0min。The more specific preparation method is as follows: take Penicillium daleae mycelium and inoculate PDA in sterilized potato glucose culture solution, shake in a shaking table at 170 rpm at room temperature for five days, make seed solution, and transfer the seed solution to Into the sterilized rice culture medium, aseptic culture at 28°C for 40 days, extract the mycelium after 40 days of culture in rice culture medium with ethyl acetate, extract four times in total to obtain ethyl acetate extract, use silica gel Column chromatography, high performance liquid chromatography mass spectrometry, preparative high performance liquid chromatography to process the ethyl acetate extract, and finally obtain penicillium furanone A, and its preparative liquid chromatography conditions are: 12mL/min, UVλmax 230nm, MeCN- H2 O 45:55, retention time 15.0min.
上述制备方法中，青霉呋喃酮A化合物可进一步与适当的酸成盐，适当的酸选自盐酸、氢溴酸、硝酸、硫酸、磷酸、酒石酸、柠檬酸、甲酸、乙酸、乙二酸或其他适合的有机酸或无机酸。In the above-mentioned preparation method, the penicillium furanone A compound can be further salted with an appropriate acid, and the appropriate acid is selected from hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, phosphoric acid, tartaric acid, citric acid, formic acid, acetic acid, oxalic acid or Other suitable organic or inorganic acids.
本发明还提供了所述的青霉呋喃酮A化合物或药物组合物在制备抗肝纤维化的药物中的应用。The present invention also provides the application of the penicillofuranone A compound or the pharmaceutical composition in the preparation of anti-hepatic fibrosis medicine.
本发明的药物组合物，能作为适宜口服或注射给药的制剂形式。The pharmaceutical composition of the present invention can be in the form of a preparation suitable for oral or injection administration.
其中所述的口服给药的制剂形式为片剂、缓释片、控释片、锭剂、硬或软胶囊、滴丸、微丸、水性或油混悬剂、乳剂、可分散的散剂或颗粒剂、口服溶液、糖浆剂或酏剂，所述的注射给药的制剂形式为灭菌的水性或油性溶液、无菌粉末、脂质体、乳剂、微乳剂、纳米乳剂或微囊。The formulations for oral administration described herein are tablets, sustained-release tablets, controlled-release tablets, lozenges, hard or soft capsules, dropping pills, pellets, aqueous or oil suspensions, emulsions, dispersible powders or Granules, oral solutions, syrups or elixirs, and the preparations for injection are in the form of sterile aqueous or oily solutions, sterile powders, liposomes, emulsions, microemulsions, nanoemulsions or microcapsules.
本发明化合物青霉呋喃酮A结构新颖，分子量为486，与之前报道的该类化合物结构完全不一样，且具有明显的抗肝纤维化活性，其机制与阻断TGF-β1刺激肝星状细胞的过度激活和细胞外基质的形成能力有关，在低于32μM的浓度下，Penicilfuranone A未见明显的肝细胞毒性。The compound penicillin furanone A of the present invention has a novel structure and a molecular weight of 486, which is completely different from the structure of the previously reported compounds, and has obvious anti-hepatic fibrosis activity, and its mechanism is related to blocking TGF-β1 to stimulate hepatic stellate cells The overactivation of Penicilfuranone A is related to the ability to form extracellular matrix. Penicilfuranone A has no obvious liver cytotoxicity at a concentration lower than 32 μM.
附图说明：Description of drawings:
图1为本发明Penicilfuranone A的结构示意图；Fig. 1 is the structural representation of Penicilfuranone A of the present invention;
图2为本发明Penicilfuranone A的X-单晶衍射结构示意图；Fig. 2 is the X-single crystal diffraction structure schematic diagram of Penicilfuranone A of the present invention;
图3为本发明Penicilfuranone A对正常人肝脏细胞生长活力的影响(MTT法；处理48h)；Fig. 3 is the effect of Penicilfuranone A of the present invention on the growth activity of normal human liver cells (MTT method; processing 48h);
图4为本发明Penicilfuranone A抑制TGF-β1诱导下，肝星状细胞纤维化形成相关蛋白的表达(western blot法；A为大鼠T6肝星状细胞株，B为人LX-2肝星状细胞株)。Fig. 4 is that Penicilfuranone A of the present invention inhibits the expression of proteins related to the formation of hepatic stellate cell fibrosis under the induction of TGF-β1 (western blot method; A is rat T6 hepatic stellate cell strain, and B is human LX-2 hepatic stellate cell strain).
具体实施方式：detailed description:
下面结合附图，用本发明的实施例来进一步说明本发明的实质性内容，但并不以此来限定本发明。The substantive content of the present invention will be further described below with reference to the accompanying drawings, but the present invention is not limited thereto.
实施例1：Example 1:
化合物Penicilfuranone A的制备：Preparation of compound Penicilfuranone A:
分离流程：取少量Penicillium daleae菌丝体接种于1000ml灭菌马铃薯葡萄糖培养液中(PDA)，于室温下以170转每分钟在摇床中震荡五天，做成种子液。将种子液转接到2.0公斤灭菌的大米培养基中，25度条件下，无菌培养35天。用乙酸乙酯5L提取利用大米培养基培养35天后的菌丝体，共提取五次，每次5L，得到乙酸乙酯提取物18g。利用硅胶柱色谱，高效液相色谱质谱联用，制备型高效液相色谱处理18g的乙酸乙酯提取物，最后得到Penicilfuranone A 12mg，其制备液相色谱条件为：12mL/min,UVλmax 230nm,MeCN-H2O45:55，保留时间15.0min。Separation process: Inoculate a small amount of Penicillium daleae mycelia into 1000ml sterilized potato dextrose culture solution (PDA), shake in a shaker at 170 rpm for five days at room temperature, and make a seed solution. The seed solution was transferred to 2.0 kg of sterilized rice medium, and cultured aseptically for 35 days at 25°C. The mycelium after 35 days of cultivation in rice culture medium was extracted with 5 L of ethyl acetate, extracted five times in total, 5 L each time, to obtain 18 g of ethyl acetate extract. Using silica gel column chromatography, high performance liquid chromatography mass spectrometry, preparative high performance liquid chromatography to process 18g of ethyl acetate extract, finally obtain Penicilfuranone A 12mg, its preparative liquid chromatography conditions are: 12mL/min, UVλmax 230nm, MeCN-H2 O 45:55, retention time 15.0min.
实施例2：Example 2:
Penicilfuranone A结构解析：Penicilfuranone A structure analysis:
化合物Penicilfuranone A，金色针状晶体，[α]22D–6(c 0.2,MeOH)。ESI-MS谱给出准分子离子峰为m/z 509[M+Na]+，结合高分辨正离子HR-ESI-MS(m/z[M+Na]+509.1792，计算值为C26H30O9Na,509.1782)和13C NMR、DEPT谱提供的信息，确定其分子式为C26H30O9，不饱和度为12。UV光谱在196(4.5),222(4.6),359(4.0),495(2.6)nm处有吸收，说明分子中存在大共轭基团。分析1H和13C NMR谱(表1)，再结合HSQC，HMBC，1H-1H COSY，以及ROESY谱表明得到化合物Penicilfuranone A的结构。最后，通过X-单晶衍射分析进一步证实了以上分析，并确定了其立体构型(图2)。Penicilfuranone A的X-单晶衍射数据已经上传到应该剑桥大学单晶数据库www.ccdc.cam.ac.uk，其登记号为：CCDC 1042018。Compound Penicilfuranone A, golden needle-like crystals, [α]22 D–6 (c 0.2, MeOH). The ESI-MS spectrum gives the quasi-molecular ion peak as m/z 509[M+Na]+ , combined with high-resolution positive ion HR-ESI-MS (m/z[M+Na]+ 509.1792, the calculated value is C26 H30 O9 Na, 509.1782) and the information provided by13 C NMR and DEPT spectrum, it is determined that its molecular formula is C26 H30 O9 , and its degree of unsaturation is 12. The UV spectrum has absorption at 196 (4.5), 222 (4.6), 359 (4.0), 495 (2.6) nm, indicating that there are large conjugated groups in the molecule. Analysis of1 H and13 C NMR spectra (Table 1), combined with HSQC, HMBC,1 H-1 H COZY, and ROESY spectra showed that the structure of the compound Penicilfuranone A was obtained. Finally, the above analysis was further confirmed by X-single crystal diffraction analysis, and its stereo configuration was confirmed (Fig. 2). The X-single crystal diffraction data of Penicilfuranone A have been uploaded to the Cambridge University single crystal databasewww.ccdc.cam.ac.uk , and its accession number is: CCDC 1042018.
表1.Penicilfuranone A的NMR波谱数据(δin ppm,Jin Hz)Table 1. NMR spectral data of Penicilfuranone A (δin ppm, Jin Hz)
a测试于600MHz仪器，溶剂为DMSO-d6.b测试于600MHz仪器，溶剂为acetone-d6.a Tested on 600MHz instrument, the solvent is DMSO-d6 .b Tested on 600MHz instrument, the solvent is acetone-d6 .
实施例3：Example 3:
Penicilfuranone A对正常人肝细胞的毒性检测：Toxicity test of Penicilfuranone A to normal human liver cells:
正常人肝脏细胞(L-02细胞株)购自中国科学院上海生命科学研究院细胞资源中心；RPMI 1640培养基为Hyclone公司产品；胎牛血清为Gibco公司产品；二甲基亚砜(DMSO)和噻唑蓝(MTT)为sigma公司产品；全自动酶标仪(美国，Thermo公司)；恒温CO2培养箱(美国，Thermo公司)；正常人肝细胞L-02细胞株用RPMI1640培养基(含10％灭活胎牛血清，100U/mL青霉素和100μg/mL链霉素)于37℃、饱和空气湿度、含5％CO2的培养箱内常规传代培养。取指数生长期L-02细胞株，1000×g离心5min，弃去上清液，用相应的培养基打匀，制成细胞悬液，计数后稀释成浓度为3×104个细胞/mL的细胞悬液，接种于96孔板，每孔200μL，置于细胞培养箱24h后给药，分别设置细胞对照组和11个浓度的Penicilfuranone A样品组(0.125、0.25、0.5、1、2、4、8、16、32、64、128μM)，每个浓度设置3个复孔，加药后将96孔板分别培养于细胞培养箱中48h后，每孔加20uL MTT溶液(用PBS将MTT粉末配成5mg/mL的储存液)，在细胞培养箱中继续孵育4h后离心弃去上清液，每孔加入100uL DMSO，轻微振荡使紫色晶体完全溶解，用酶标仪于570nm处测定每孔吸光度(OD值),计算3个复孔OD值平均值，并计算细胞存活率：细胞存活率(％)＝(给药组细胞OD均值－背景OD均值)/(对照组细胞OD均值－背景OD均值)×100％。Normal human liver cells (L-02 cell line) were purchased from the Cell Resource Center of Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences; RPMI 1640 medium was produced by Hyclone; fetal bovine serum was produced by Gibco; dimethyl sulfoxide (DMSO) and Thiazolium blue (MTT) is the product of sigma company; automatic microplate reader (U.S., Thermo company); constant temperatureCO incubator (U.S., Thermo company); normal human liver cell L-02 cell line RPMI1640 medium (containing 10 % inactivated fetal bovine serum, 100 U/mL penicillin and 100 μg/mL streptomycin) were routinely subcultured in an incubator at 37° C., saturated air humidity, and containing 5% CO2 . Take the L-02 cell line in the exponential growth phase, centrifuge at 1000×g for 5 minutes, discard the supernatant, mix well with the corresponding medium to make a cell suspension, count and dilute to a concentration of3 ×104 cells/mL The cell suspension was inoculated in a 96-well plate, 200 μL per well, placed in a cell culture incubator for 24 hours and administered, and the cell control group and 11 concentrations of Penicilfuranone A sample groups (0.125, 0.25, 0.5, 1, 2, 4, 8, 16, 32, 64, 128μM), each concentration set 3 duplicate wells, after the addition of drugs, the 96-well plate was cultured in the cell culture incubator for 48h, and 20uL MTT solution was added to each well (MTT was mixed with PBS 5 mg/mL storage solution), continue to incubate in the cell culture incubator for 4 hours, centrifuge and discard the supernatant, add 100uL DMSO to each well, shake slightly to dissolve the purple crystals completely, and measure each well at 570nm with a microplate reader. Well absorbance (OD value), calculate the average OD value of 3 duplicate wells, and calculate the cell survival rate: cell survival rate (%) = (average OD value of cells in the administration group - mean OD value of the background) / (average OD value of cells in the control group - Background OD mean) x 100%.
肝细胞损伤往往被认为是发生肝脏纤维化的诱发因子，因此，药物可以通过保护肝细胞的方式来阻止肝脏纤维化进程，至少其本身应没有肝细胞毒性。MTT检测结果发现，经Penicilfuranone A各浓度处理48h后，在非常高的浓度(32μM)下，人正常肝脏细胞也没有出现明显的活力抑制现象，这意味着，在低于32μM的浓度下，Penicilfuranone A未见明显的肝细胞毒性，结果见图3。Hepatic cell injury is often considered to be an inducing factor for liver fibrosis. Therefore, drugs can prevent the process of liver fibrosis by protecting liver cells, at least by themselves, they should not be toxic to liver cells. The results of MTT test found that after being treated with various concentrations of Penicilfuranone A for 48 hours, at a very high concentration (32μM), human normal liver cells did not appear to significantly inhibit the activity, which means that at a concentration lower than 32μM, Penicilfuranone A There was no obvious liver cytotoxicity, the results are shown in Figure 3.
实施例4：Example 4:
采用western blot方法检测Penicilfuranone A对TGF-β1刺激肝星状细胞激活及细胞外基质成分表达的影响The effect of penicilfuranone A on the activation of hepatic stellate cells stimulated by TGF-β1 and the expression of extracellular matrix components were detected by western blot
人肝星状细胞LX-2细胞株引自中国典型培养物保藏中心；大鼠肝星状细胞T6细胞株为引自中国科学院上海生命科学研究院细胞资源中心；分别用DMEM培养基和RPMI1640培养基(含10％灭活胎牛血清，100U/mL青霉素和100μg/mL链霉素)，于37℃，含5％CO2的培养箱内常规传代培养。取对数生长期的肝星状细胞株LX-2和T6，在6孔培养板以5×104个细胞/孔进行接种，细胞培养过夜后，分别以相应的无血清培养基同步化培养24h，此后，分组换液，分别加入终浓度10ng/ml的重组人TGF-β1(美国Peprotech公司)和不同浓度的Penicilfuranone A(1,2和4μM)，同时分别设溶剂对照组(0.5％DMSO)和浓度为10μM的SB431542(TGF-β1Ⅰ型受体激酶抑制剂，美国Cayman公司)阳性对照组。药物作用48h后收集细胞，蛋白裂解液裂解细胞，4℃、12000×g低温离心30min，收集上清。上清经BCA法测定其蛋白浓度。加入等体积2×SDS加样缓冲液(125mmol/L Tris-HCl、20％甘油，0.01％溴酚蓝、4％SDS、200mmol/L DTT)，调整各组上样蛋白浓度一致后，沸水浴中加热5min，SDS-聚丙烯酰胺凝胶电泳，转膜，含5％脱脂奶粉的TBST封闭1h后，分别加入α-平滑肌肌动蛋白(α-SMA),波形蛋白(vimentin)、I型胶原(type I collagen)、结缔组织生长因子(CTGF)和内参β-actin抗体，4℃孵育过夜。TBST洗涤3次，加入辣根酶标记羊抗兔IgG二抗，37℃下与膜反应1h，TBST洗膜3次，按ECL试剂盒说明书进行化学发光，分析相关蛋白的表达情况。The human hepatic stellate cell LX-2 cell line was quoted from the China Center for Type Culture Collection; the rat hepatic stellate cell T6 cell line was cited from the Cell Resource Center of Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences; cultured in DMEM medium and RPMI1640 respectively Substratum (containing 10% inactivated fetal bovine serum, 100 U/mL penicillin and 100 μg/mL streptomycin) was routinely subcultured in an incubator containing 5% CO2 at 37°C. Hepatic stellate cell lines LX-2 and T6 in the logarithmic growth phase were seeded in 6-well culture plates with5 ×104 cells/well. After the cells were cultured overnight, they were cultured synchronously with corresponding serum-free medium. After 24 hours, the medium was changed in groups, and recombinant human TGF-β1 (Peprotech, USA) with a final concentration of 10 ng/ml and Penicilfuranone A (1, 2 and 4 μM) of different concentrations were added respectively, and a solvent control group (0.5% DMSO ) and a positive control group of SB431542 (TGF-β1 type I receptor kinase inhibitor, Cayman, USA) at a concentration of 10 μM. Cells were collected after 48 hours of drug action, lysed with protein lysate, centrifuged at 12000×g at 4°C for 30 minutes at low temperature, and the supernatant was collected. The protein concentration of supernatant was determined by BCA method. Add an equal volume of 2×SDS loading buffer (125mmol/L Tris-HCl, 20% glycerol, 0.01% bromophenol blue, 4% SDS, 200mmol/L DTT), adjust the protein concentration of each group to be the same, and place in a boiling water bath Heated at medium temperature for 5 minutes, electrophoresed on SDS-polyacrylamide gel, transferred to membrane, blocked with TBST containing 5% skimmed milk powder for 1 hour, then added α-smooth muscle actin (α-SMA), vimentin and type I collagen respectively (type I collagen), connective tissue growth factor (CTGF) and internal control β-actin antibody, incubated overnight at 4°C. Wash 3 times with TBST, add horseradish enzyme-labeled goat anti-rabbit IgG secondary antibody, react with the membrane for 1 h at 37°C, wash the membrane 3 times with TBST, perform chemiluminescence according to the instructions of the ECL kit, and analyze the expression of related proteins.
肝星状细胞激活后可以产生大量的细胞外基质成分，因此在肝脏纤维化的发生、发展过程中起到关键作用。TGF-β1是目前已知诱导肝星状细胞激活的最强因子，它不仅诱导肝星状细胞激活，上调细胞标志物α-SMA和vimentin的表达水平，同时还提高肝脏内typeI collagen为主的细胞外基质合成。Western blot结果显示，Penicilfuranone A处理肝星状细胞48h后，可以明显降低TGF-β1诱导下细胞激活的标志物蛋白α-SMA和vimentin表达，同时显著降低肝星状细胞合成type I collagen的能力，其降调vimentin表达的能力，甚至强于阳性对照药SB431542。此外，CTGF作为TGF-β1下游最重要的调控蛋白，可协同TGF-β1，促进肝组织内大量生成细胞外基质。Western blot的结果同时发现，Penicilfuranone A各剂量组处理后，与TGF-β1诱导组比较，肝星状细胞内CTGF蛋白表达水平显著降低。上述结果提示，Penicilfuranone A具有明显的抗肝纤维化作用，其机制与阻断TGF-β1刺激肝星状细胞的过度激活和细胞外基质的形成能力有关，结果见图4。After activation, hepatic stellate cells can produce a large amount of extracellular matrix components, so they play a key role in the occurrence and development of liver fibrosis. TGF-β1 is the strongest factor known to induce the activation of hepatic stellate cells. It not only induces the activation of hepatic stellate cells, but also up-regulates the expression levels of cell markers α-SMA and vimentin. Synthesis of extracellular matrix. Western blot results showed that after Penicilfuranone A treated hepatic stellate cells for 48 hours, it could significantly reduce the expression of TGF-β1-induced cell activation marker proteins α-SMA and vimentin, and significantly reduce the ability of hepatic stellate cells to synthesize type I collagen. Its ability to down-regulate the expression of vimentin is even stronger than that of the positive control drug SB431542. In addition, CTGF, as the most important regulatory protein downstream of TGF-β1, can cooperate with TGF-β1 to promote a large amount of extracellular matrix in liver tissue. The results of Western blot also found that the expression level of CTGF protein in hepatic stellate cells was significantly reduced after treatment with each dose of Penicilfuranone A compared with the TGF-β1 induction group. The above results suggest that Penicilfuranone A has an obvious anti-hepatic fibrosis effect, and its mechanism is related to blocking the excessive activation of hepatic stellate cells and the formation of extracellular matrix stimulated by TGF-β1. The results are shown in Figure 4.
实施例5：Example 5:
按实施例1制得Penicilfuranone A，按其与赋形剂重量比1:1的比例加入赋形剂，制粒压片。Penicilfuranone A was prepared according to Example 1, and the excipient was added according to the weight ratio of the penicilfuranone A to the excipient at a ratio of 1:1, and then granulated and compressed into tablets.
实施例6：Embodiment 6:
按实施例1制得Penicilfuranone A，按其与赋形剂重量比1:2的比例加入赋形剂，制粒压片。Penicilfuranone A was prepared according to Example 1, and excipients were added at a weight ratio of 1:2 to the excipients, granulated and compressed into tablets.
实施例7：Embodiment 7:
按实施例1制得Penicilfuranone A，按常规胶囊制剂方法制成胶囊。Prepare Penicilfuranone A according to Example 1, and make capsules according to conventional capsule preparation methods.
实施例8：Embodiment 8:
按实施例1制得Penicilfuranone A，再按下述方法制成片剂：Obtain Penicilfuranone A according to embodiment 1, then make tablet according to the following method:
实施例9：Embodiment 9:
胶囊剂：Penicilfuranone A， 100mgCapsules: Penicilfuranone A, 100mg
淀粉 适量 Appropriate amount of starch
硬脂酸镁 适量 Magnesium Stearate Appropriate amount
制备方法：将Penicilfuranone A，与助剂混合，过筛，在合适的容器中均匀混合，把得到的混合物装入硬明胶胶囊。Preparation method: mix Penicilfuranone A with auxiliary agents, sieve, mix evenly in a suitable container, and put the obtained mixture into hard gelatin capsules.
实施例10：Example 10:
制备方法：搅拌下于适当体积的重蒸馏水中每次加入一种成分，直至完全深解，然后再加入另一种成分。加水至2ml后，将该溶液在无菌过滤器上过滤，装入瓶中并按照适当的剂量分隔。Preparation method: Add one ingredient at a time to an appropriate volume of double-distilled water with stirring until completely decomposed, and then add another ingredient. After making up to 2 ml with water, the solution is filtered on a sterile filter, filled into bottles and divided according to the appropriate doses.
实施例11：Example 11:
滴丸：Penicilfuranone A 1gDropping pills: Penicilfuranone A 1g
聚乙二醇6000 9g Macrogol 6000 9g
制法：Penicilfuranone A与聚乙二醇6000熔融液的制备：按上述处方量称取Penicilfuranone A，加入适量无水乙醇，微热溶解后，加入处方量的聚乙二醇熔融液中(60℃水浴保温)，搅拌混合均匀，直至乙醇挥尽为止，静置于60℃水浴中保温30分钟，待气泡除尽，然后将除尽气泡的上述混匀熔融液转入贮液筒内，在保温80-85℃的条件下，控制滴速，一滴滴地滴入冷凝液中，等冷凝完全，倾去冷凝液，收集滴丸，沥净和用滤纸除去丸上的冷凝液，放置硅胶干燥器中或自然干燥即可。Preparation method: Preparation of Penicilfuranone A and Polyethylene Glycol 6000 melt solution: Weigh Penicilfuranone A according to the above prescription amount, add appropriate amount of absolute ethanol, dissolve with slight heat, and then add the prescription amount into the polyethylene glycol melt solution (60°C water bath for heat preservation), stir and mix evenly until the ethanol is evaporated, and keep it in a 60°C water bath for 30 minutes. Under the condition of 80-85℃, control the dripping speed, drip into the condensate drop by drop, wait until the condensation is complete, pour off the condensate, collect the dropping pills, drain and remove the condensate on the pills with filter paper, and place in a silica gel drier Neutral or dry naturally.
| Application Number | Priority Date | Filing Date | Title |
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| CN201510672179.XACN105330629B (en) | 2015-10-16 | 2015-10-16 | Anti-hepatic fibrosis penicillin furanone A compound and its pharmaceutical composition and application |
| Application Number | Priority Date | Filing Date | Title |
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| CN201510672179.XACN105330629B (en) | 2015-10-16 | 2015-10-16 | Anti-hepatic fibrosis penicillin furanone A compound and its pharmaceutical composition and application |
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| CN105330629Btrue CN105330629B (en) | 2017-06-27 |
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| CN201510672179.XAActiveCN105330629B (en) | 2015-10-16 | 2015-10-16 | Anti-hepatic fibrosis penicillin furanone A compound and its pharmaceutical composition and application |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105399708A (en)* | 2015-10-16 | 2016-03-16 | 中国科学院昆明植物研究所 | Anti-liver fibrosis Penicilfuranone A compound as well as pharmaceutical composition and application thereof |
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| CN118324775B (en)* | 2024-03-01 | 2025-03-21 | 中山大学附属第五医院 | A polyketide compound and its application in preparing medicine for treating pulmonary fibrosis |
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| EP2189453A1 (en)* | 2008-11-25 | 2010-05-26 | Université Louis Pasteur | Rocaglaol derivatives as cardioprotectant agents |
| WO2015075165A1 (en)* | 2013-11-22 | 2015-05-28 | Deutsches Krebsforschungszentrum | Translation inhibitors in high-dose chemo- and/or high-dose radiotherapy |
| CN105399708A (en)* | 2015-10-16 | 2016-03-16 | 中国科学院昆明植物研究所 | Anti-liver fibrosis Penicilfuranone A compound as well as pharmaceutical composition and application thereof |
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP2189453A1 (en)* | 2008-11-25 | 2010-05-26 | Université Louis Pasteur | Rocaglaol derivatives as cardioprotectant agents |
| WO2015075165A1 (en)* | 2013-11-22 | 2015-05-28 | Deutsches Krebsforschungszentrum | Translation inhibitors in high-dose chemo- and/or high-dose radiotherapy |
| CN105399708A (en)* | 2015-10-16 | 2016-03-16 | 中国科学院昆明植物研究所 | Anti-liver fibrosis Penicilfuranone A compound as well as pharmaceutical composition and application thereof |
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| LC-MS-Guided Isolation of Penicilfuranone A: A New Antifibrotic Furancarboxylic Acid from the Plant Endophytic Fungus Penicillium sp.sh18;Wei-Guang Wang 等;《J. Nat. Prod.》;20161218;第79卷;149-155* |
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105399708A (en)* | 2015-10-16 | 2016-03-16 | 中国科学院昆明植物研究所 | Anti-liver fibrosis Penicilfuranone A compound as well as pharmaceutical composition and application thereof |
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| CN105330629A (en) | 2016-02-17 |
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