首乌叶中黄酮类化合物成分的研究
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摘要
蓼科植物何首乌(Polygonum multiflorum Thunb.)为多年生缠绕性草本，通常根据何首乌的部位不同，块根称为“何首乌”，藤茎称为“夜交藤”，叶称为“首乌叶”。已从何首乌、夜交藤中得到多种黄酮类化合物[1-4]，而首乌叶的研究应用相对较少[5,6]。黄酮类化合物作为优良的自由基清除剂和抗氧化剂，已广泛应用于食品和营养保健品中[7]。
本课题的研究包括四个部分：
（1）利用微波辅助提取法提取了首乌叶黄酮，对影响首乌叶总黄酮提取的因素进行探讨。实验结果表明，最佳条件是：乙醇浓度70%，功率400W，时间5min，温度70℃，料液比（w/v）为1:20，在此条件下得到首乌叶总黄酮的提取率为3.86%。
（2）以吸附量和解析率为参考指标，比较了AB-8、D4006、S-8、D101、NKA-Ⅱ、NKA-9六种大孔树脂对首乌叶总黄酮的吸附和解析效果，并通过HPLC分析证明大孔树脂对首乌叶黄酮起到分离纯化的效果。结果表明：D101大孔树脂对首乌叶总黄酮具有较好的吸附和解析性能，吸附容量为29.0mg·g~(-10，解析率达到98.8%，选择浓度为1.0mg·g-1的样液上样，用pH值为6.0、浓度为70%的乙醇溶液洗脱。该方法简单易行，分离效果好，适于首乌叶总黄酮的分离纯化。
（3）建立高效液相色谱—质谱法（HPLC-LC/MS）分析首乌叶中活性成分,其中有七种黄酮类成分：木樨草苷、牡荆素、芹菜素-7-葡萄糖苷、金丝桃苷、芦丁、紫云英苷和槲皮苷。所建立的高效液相色谱—质谱法高效有效，可以用于快速鉴定首乌叶中黄酮类成分，并首次在该植物中鉴定出芹菜素-7-葡萄糖苷和紫云英苷。
（4）测定首乌叶黄酮溶液的总体抗氧化能力、清除超氧阴离子自由基能力、清除羟基自由基能力、清除DPPH自由基能力和对卵黄脂蛋白脂质抗氧化能力，并与合成抗氧化剂Vc、PG比较，对首乌叶黄酮溶液抗氧化和清除游离基活性进行评价。结果表明：首乌叶黄酮溶液具有良好的抗氧化效果，在一定浓度范围内，首乌叶黄酮溶液对卵黄脂蛋白脂质抗氧化能力优于Vc和PG，其总体抗氧化能力、清除超氧阴离子自由基能力优于PG略低于Vc，清除羟自由基能力优于Vc低于PG，但清除DPPH自由基能力比Vc和PG略弱。
Polygonum multiflorum Thunb., which is a traditional Chinese medicinal plant wasofficially listed in the Chinese Pharmacopoeia(China Pharmacopoeia Committee,2010), is one ofthe most important and commonly used traditional Chinese medicine (TCM). It’s dried roots andstems are well known as “He shou wu” and “Shou wu teng” and it’s leaves are called “Shou wuye” respectively. P. multiflorum has been used in many remedies of traditional medicine inChina such as “Shouwu Yanshou Pian” and other countries for its possessing many activitiessuch as antioxidation, anti-aging, reducing blood lipid and improve memory.
The leaves of Polygonum multiflorum Thunb flavonoids were extracted bymicrowave-assisted extraction technique. The effects of ethanol concentration, microwave power,microwave treatment time, microwave treatment temperature and ratio of material to solvent onthe extraction yields were investigated. The experimental results showed that the optimumconditions of microwave extraction were as follows: ethanol concentration70%, microwavepower400W, microwave treatment time5min, microwave treatment temperature70oC and theratio of material to solvent1:20(w/v). Under the above–mentioned conditions, the extract rate oftotal flavonoids was3.86%.
Adsorption and desorption properties of six kinds of resins, including AB-8, D4006, S-8,D101, NKA-II and NKA-9, for the leaves of P. multiflorum flavonoids were studied in this paperusing total flavonoids content. Furthermore, with the method of HPLC, we proved that themacroporous resins had a separation and purification effect on flavonoids from the leaves of P.multiflorum. It was showed that the macroporous resins D101was selected with high desorptioncapacity (28.99mg·g~(-1)) and desorption rate (98.84%). The concentration of the sample solutionwas1.0mg·mL-1, and pH6.0,70%of ethanol could accomplish desorption completely. Thismethod is simple and feasible and fit for separation and purification of flavonoids from theleaves of P. multiflorum.
Leaves of P. multiflorum were analysed with a high performance liquidchromatography-liquid chromatography/mass spectrometry (HPLC-LC/MS) system equipped inpositive mode. In this study, an HPLC-LC/MS was successfully developed to identify andcharacterize seven flavonoids (Luteoloside, Vitexin, Apigenin-7-glucoside, Hyperoside, Rutin, Astragalin, Quercitrin), one stilbene glycoside（2,3,5,4’-tetrahydroxystilbane-2-O-β-D-glucoside）and one sterol (Stigmasterol) of the leavesof P. multiflorum. Among these compounds, Apigenin-7-glucoside and Astragalin are isolatedfrom leaves of P. multiflorum for the first time. The proposed method is rapid and reproducibleand is useful for characterization and evaluation of leaves of P. multiflorum flavonoids andstilbene glycoside.
In the present research, the total antioxidant activities,superoxide anion,hydroxyl radicalscavenging activities, scavenging DPPH radical of nutmeg and antioxidant activities onlipoprotein and lipid of yolk （Myristica fragrans Houtt）flavonoids from leaves of P. multiflorumwere determined and the results were compared with those of synthetic antioxidants such asascorbic acid（Vc）and propyl gallate（PG）.The results revealed clearly demonstrated thatflavonoids from leaves of P. multiflorum possesses marked antioxidant activities on on lipoproteinand lipid of yolk was concentration dependent and was higher than those of Vc and PG.Totalantioxidant activities and superoxide anion, hydroxyl radical scavenging activities of flavonoidsfrom leaves of P. multiflorum were higher than PG and weaker than Vc. Hydroxyl radicalscavenging was higher than Vc and weaker than PG. DPPH radical scavenging activities was alittle weaker than those of two synthetic antioxidants.
本课题的研究包括四个部分：
（1）利用微波辅助提取法提取了首乌叶黄酮，对影响首乌叶总黄酮提取的因素进行探讨。实验结果表明，最佳条件是：乙醇浓度70%，功率400W，时间5min，温度70℃，料液比（w/v）为1:20，在此条件下得到首乌叶总黄酮的提取率为3.86%。
（2）以吸附量和解析率为参考指标，比较了AB-8、D4006、S-8、D101、NKA-Ⅱ、NKA-9六种大孔树脂对首乌叶总黄酮的吸附和解析效果，并通过HPLC分析证明大孔树脂对首乌叶黄酮起到分离纯化的效果。结果表明：D101大孔树脂对首乌叶总黄酮具有较好的吸附和解析性能，吸附容量为29.0mg·g~(-10，解析率达到98.8%，选择浓度为1.0mg·g-1的样液上样，用pH值为6.0、浓度为70%的乙醇溶液洗脱。该方法简单易行，分离效果好，适于首乌叶总黄酮的分离纯化。
（3）建立高效液相色谱—质谱法（HPLC-LC/MS）分析首乌叶中活性成分,其中有七种黄酮类成分：木樨草苷、牡荆素、芹菜素-7-葡萄糖苷、金丝桃苷、芦丁、紫云英苷和槲皮苷。所建立的高效液相色谱—质谱法高效有效，可以用于快速鉴定首乌叶中黄酮类成分，并首次在该植物中鉴定出芹菜素-7-葡萄糖苷和紫云英苷。
（4）测定首乌叶黄酮溶液的总体抗氧化能力、清除超氧阴离子自由基能力、清除羟基自由基能力、清除DPPH自由基能力和对卵黄脂蛋白脂质抗氧化能力，并与合成抗氧化剂Vc、PG比较，对首乌叶黄酮溶液抗氧化和清除游离基活性进行评价。结果表明：首乌叶黄酮溶液具有良好的抗氧化效果，在一定浓度范围内，首乌叶黄酮溶液对卵黄脂蛋白脂质抗氧化能力优于Vc和PG，其总体抗氧化能力、清除超氧阴离子自由基能力优于PG略低于Vc，清除羟自由基能力优于Vc低于PG，但清除DPPH自由基能力比Vc和PG略弱。
Polygonum multiflorum Thunb., which is a traditional Chinese medicinal plant wasofficially listed in the Chinese Pharmacopoeia(China Pharmacopoeia Committee,2010), is one ofthe most important and commonly used traditional Chinese medicine (TCM). It’s dried roots andstems are well known as “He shou wu” and “Shou wu teng” and it’s leaves are called “Shou wuye” respectively. P. multiflorum has been used in many remedies of traditional medicine inChina such as “Shouwu Yanshou Pian” and other countries for its possessing many activitiessuch as antioxidation, anti-aging, reducing blood lipid and improve memory.
The leaves of Polygonum multiflorum Thunb flavonoids were extracted bymicrowave-assisted extraction technique. The effects of ethanol concentration, microwave power,microwave treatment time, microwave treatment temperature and ratio of material to solvent onthe extraction yields were investigated. The experimental results showed that the optimumconditions of microwave extraction were as follows: ethanol concentration70%, microwavepower400W, microwave treatment time5min, microwave treatment temperature70oC and theratio of material to solvent1:20(w/v). Under the above–mentioned conditions, the extract rate oftotal flavonoids was3.86%.
Adsorption and desorption properties of six kinds of resins, including AB-8, D4006, S-8,D101, NKA-II and NKA-9, for the leaves of P. multiflorum flavonoids were studied in this paperusing total flavonoids content. Furthermore, with the method of HPLC, we proved that themacroporous resins had a separation and purification effect on flavonoids from the leaves of P.multiflorum. It was showed that the macroporous resins D101was selected with high desorptioncapacity (28.99mg·g~(-1)) and desorption rate (98.84%). The concentration of the sample solutionwas1.0mg·mL-1, and pH6.0,70%of ethanol could accomplish desorption completely. Thismethod is simple and feasible and fit for separation and purification of flavonoids from theleaves of P. multiflorum.
Leaves of P. multiflorum were analysed with a high performance liquidchromatography-liquid chromatography/mass spectrometry (HPLC-LC/MS) system equipped inpositive mode. In this study, an HPLC-LC/MS was successfully developed to identify andcharacterize seven flavonoids (Luteoloside, Vitexin, Apigenin-7-glucoside, Hyperoside, Rutin, Astragalin, Quercitrin), one stilbene glycoside（2,3,5,4’-tetrahydroxystilbane-2-O-β-D-glucoside）and one sterol (Stigmasterol) of the leavesof P. multiflorum. Among these compounds, Apigenin-7-glucoside and Astragalin are isolatedfrom leaves of P. multiflorum for the first time. The proposed method is rapid and reproducibleand is useful for characterization and evaluation of leaves of P. multiflorum flavonoids andstilbene glycoside.
In the present research, the total antioxidant activities,superoxide anion,hydroxyl radicalscavenging activities, scavenging DPPH radical of nutmeg and antioxidant activities onlipoprotein and lipid of yolk （Myristica fragrans Houtt）flavonoids from leaves of P. multiflorumwere determined and the results were compared with those of synthetic antioxidants such asascorbic acid（Vc）and propyl gallate（PG）.The results revealed clearly demonstrated thatflavonoids from leaves of P. multiflorum possesses marked antioxidant activities on on lipoproteinand lipid of yolk was concentration dependent and was higher than those of Vc and PG.Totalantioxidant activities and superoxide anion, hydroxyl radical scavenging activities of flavonoidsfrom leaves of P. multiflorum were higher than PG and weaker than Vc. Hydroxyl radicalscavenging was higher than Vc and weaker than PG. DPPH radical scavenging activities was alittle weaker than those of two synthetic antioxidants.
引文
[1]周立新,林茂,李建北等.何首乌乙酸乙酯不溶部分化学成分的研究[J].药学学报,1994,29(2):107.
[2]张志国,吕泰省,姚庆强.何首乌的研究进展[J].解放军药学学报,2008,24(1):62-64.
[3]夏扎旦,萨提瓦力地·赫力力,买合布白·阿不都热依木等.分光光度法测定维吾尔传统药用植物何首乌的总黄酮含量[J].食品科学,2006,27(12):704-705.
[4]王自军.夜交藤中总黄酮的超声提取及含量测定[J].时珍国医国药,2010,21(12):3211-3212.
[5] Xu YL, Dong Q, Hu FZ. Simultaneous quantitative determination of eight active componentsin Polygonum multiflorum Thumb by RP-HPLC [J]. J Chin Pharm Soc,2009,18:358-361.
[6]饶高雄,薛咏梅,惠婷婷等.首乌叶化学成分研究[J].中药材,2009,32(6):891-893.
[7]延玺,刘会青,邹永青等.黄酮类化合物生理活性及合成研究进展[J].有机化学,2008,28(9):1534-1544.
[8] Duan L, Carrier DL, Clausen EC. Silymarin extraction from milk thistle using hotwater [J].Appl Biochem Biotechnol.,2004,(113-116):559-568.
[9] Rogerio AP, Kanashiro A, Fontanar C, et al. Anti-inflammatory activity ofquercetin andisoquercitrin in experimental murine allergic asthma [J]. Inflamm Res,2007,56(10):402-408.
[10]徐美奕,韩雅莉,东野广智等.紫荆花总黄酮的分离纯化与光谱分析[J].中药材,2007,30(10):1252-1255.
[11]姚新生.天然药物化学[M].北京:人民卫生出版社,2001.
[12] Paniwnyk L, Beaufoy E, Lorimer JP, et al. The extraction of rutin from flowerbuds ofSophora japonica [J]. Ultrason Sonochem,2001,8(3):299-301.
[13] Yang Y, Zhang F. Ultrasound-assisted extraction of rutin and quercetin from Euonymusalatus(Thunb.) Sieb [J]. Ultrason Sonochem,2008,15(4):308-313.
[14] Wang X, Ding L, Zhang H, et al. Development of ananalytical method for organotincompounds in fortified flour samples using microwave-assisted extraction andnormal-phase HPLC with UV detection [J]. J Chromatogr B,2006,843(2):268-274.
[15] Chen L, Ding L, Yu A, et al. Continuous determination of total flavonoids in Platycladusorientalis(L.) Franco by dynamic microwave-assisted extraction coupled with on-linederivatization and ultraviolet-visible detection [J]. Anal Chim Acta,2007,596(1):164-170.
[16] Song JZ, Mo SF, Yip YK, et al. Development of microwave assisted extraction for thesimultaneous determination of isoflavonoids and saponins in radix astragali by highperformance liquid chromatography [J]. J Sep Sci,2007,30(6):819-824.
[17] Safaralie A, Fatem S, Sefidkon F..Essentialoil composition ofValeriana officinalis L.rootscultivated in Iran.Comparative analysis between supercritical CO2extraction andhydrodistillation [J]. J Chromatogr A,2008,1180(1-2):159-164.
[18]万茵,谢明勇,董欢欢等.超临界CO2流体提取车前子总黄酮的工艺优化[J].南昌大学学报(理科版),2007,31(2):160-163.
[19]余洪波,张晓昱.酶法在中药提取中的研究进展[J].中成药,2005,27(5):591-593.
[20]庞允,杨建秀,刘子兰.银杏叶中黄酮的提取工艺比较[J].制剂技术,2007,16(3):38-39.
[21]张兆旺,孙秀梅.“半仿生提取法”的特点与应用[J].世界科学技术,2000,2(1):35-38.
[22]赵雪梅,孙秀梅,张兆旺等.杜仲降压方药的半仿生提取工艺研究[J].中国中药杂志,2007,32(21):2247-2252.
[23]陈丛瑾,黄克瀛,李德良.正交试验法优选香椿叶总黄酮的半仿生提取工艺[J].食品科技,2007,(6):119-121.
[24]郭文晶,张守勤,吴华等.均匀设计优化高压提取刺五加叶中总黄酮[J].中药材,2007,30(6):718-720.
[25] Alvarez-Munoz D, Saez M, Lara-Martin PA, et al. New extraction method for the analysisof linear alkylbenzene sulfonates inmarine organisms Pressurized liquid extraction versusSoxhlet extraction [J]. J Chromatogr A,1052(1-2):33-38.
[26] Zhang S, Liu C, Bi H, et al. Extraction of flavonoids from Rhodiola sachlinesisA.Bor byUPE and the antioxidant activity of its extract [J]. Nat Prod Res,2008,22(2):178-187.
[27]田庆来,谢渝春,张波等.溶剂萃取法分离水溶性甘草黄酮[J].过程工程学报,2007,7(3):496-500.
[28] Zheng Y, Haworth IS, Zuo Z, et al. Physicochemical and structural characterization ofquercetin-beta-cyclodex trin complexes [J]. J Pharm Sci,2005,94(5):1079-1089.
[29]屠鹏飞,贾存勤,张洪全.大孔吸附树脂在中药新药研究和生产中的应用[J].世界科学技术,2004,6(3):22-29.
[30] Sun A, Sun Q, Liu R. Preparative isolation and purification of flavone compoundsfromsophora japonica L. by high-speed counter-current chromatography combined withmacroporous resin column separation [J]. J Sep Sci,2007,30(7):1013-1018.
[31] Yao S, Luo JG, Huang XF, et al. Application of preparative high-speed counterchromatography/preparative high-performance liquid chromatographymode in rapidseparation of saponins [J]. J Chromatogr B,2008,864:69-77.
[32] Kohler N, Wray V, Winterhalter P. Preparative isolation of procyanidins from grape seedextracts by high speed counter-current chromatography [J]. J Chromatogr A,2008,1177(1):114-125.
[33] Zhang Q, Chen LJ, Hao YY, et al. Isolation and purification of flavonol glycosides fromGinkgo biloba leaves by high-speed counter-current chromatography [J]. J Sep Sci,2007,30(13):2153-2159.
[34] Arvind R. Equilibrium theory-based design of simulated moving bed proceses underreduced purity requitements Linear isotherms [J]. J Chromatogr A,2008,1185(2):216-222.
[35] Cong JX, Lin BC. Separation of liquiritin by simulated moving bed chromatography. JChromatogr A,2007,1145(1-2):190-194.
[36]李瑞萍,黄骏雄.高效制备液相色谱柱技术的研究进展[J].化学进展,2004,16(2):273-283.
[37]罗艺萍.黄酮类化合物的药理活性研究进展[J].亚太传统医药,2010,6(4):126-128.
[38] Nkenqfack AE, Vouffo TW, Fomum ZT, et al. Prenylated isoflavanone from the roots ofErythrina sigmoidea [J]. Phytochem,1994,36(4):1074-1077.
[39]吉琦正雄.何首乌的成分研究[J]．国外医学·中医药分册，1985,7(1):55.
[40] Zhu HB, Wang YZ, Liu YX, et al. Analysis of flavonoids in Portulaca oleracea L. byUV–Vis spectrophotometry with comparative study on different extraction technologies [J].Food Anal Methods,2010,3:90–97.
[41]仇燕.菜芙蓉花中总黄酮微波提取的研究[J]．中药材,2006,29(4):387-389
[42]陈伟,刘青梅.微波技术在杜仲黄酮提取工艺中的应用研究[J]．食品科学,2006,27(10):285-288.
[43]张琪,刘慧灵,朱瑞等.苦荞麦中总黄酮和芦丁的含量测定方法的研究[J].食品科学,2003,24(7):113-116.
[44]毕洁,杨庆利,于丽娜等.花生壳黄酮乙醇提取工艺探讨[J].现代化工,2008,10(28):316-319.
[45]陈恩赞,苏海佳,谭天伟.微波提取酱渣中总异黄酮的实验研究[J].环境工程学报,2010,4(2):427-430.
[46]周桃英,罗登宏,李国庆等. AB-8大孔树脂纯化荷叶总黄酮的工艺研究[J].中国食品添加剂,20095:113-119.
[47]陶海腾,陈晓明,吕飞杰等.大孔吸附树脂法纯化木薯叶黄酮的初步研究[J].食品工业科技,2009,30(8):192-194.
[48]李旭洲,韩立炜,张岩峰等.葛根中葛根黄酮的水提取及大孔树脂纯化工艺研究[J].北京中医药大学学报,2009,32(9):626-629.
[49] Fu BQ, Liu J, Li H, et al. The application of macroporous resins in the separation of licoriceflavonoids and glycyrrhizic acid[J]. J Chromatogr A,2005,1089:18–24.
[50]李颖畅,孟宪军,李小红.大孔树脂纯化蓝萄叶总黄酮的工艺研究[J].食品工业与发酵,2008,34(1):133-137.
[51]王俊,赵辉,吴福安.树脂吸附法分离纯化桑叶总黄酮[J].离子交换与吸附,2008,24(2):139-147.
[52]刘志祥,曾超珍.大孔树脂法纯化苦丁茶总黄酮的研究[J].时珍国医国药,2009,20(9):2183-2184.
[49] Fu BQ, Liu J, Li H, et al. The application of macroporous resins in the separation of licoriceflavonoids and glycyrrhizic acid[J]. J Chromatogr A,2005,1089:18–24.
[50]李颖畅,孟宪军,李小红.大孔树脂纯化蓝萄叶总黄酮的工艺研究[J].食品工业与发酵,2008,34(1):133-137.
[51]王俊,赵辉,吴福安.树脂吸附法分离纯化桑叶总黄酮[J].离子交换与吸附,2008,24(2):139-147.
[52]刘志祥,曾超珍.大孔树脂法纯化苦丁茶总黄酮的研究[J].时珍国医国药,2009,20(9):2183-2184.
[53]廖琼峰.陆英药效物质基础研究[D].2005.05.
[54]冯广卫,李翠兵,廖尚高等.荭草素和异荭草素、牡荆素和异牡荆素2对碳苷化合物的快速检测与鉴定[J].药物分析杂志,2011,31(7):1263-1268.
[55]于澎,张虹. HPLC-ESI-MS法同时测定虎杖提取物中的17种成分[J].沈阳药科大学学报,2011,28(12):963-968.
[56]张语迟,刘春明,刘志强.罗布白麻与罗布红麻的液相色谱-质谱联用分析[J].分析测试学报,2009,28(10):1148-1154.
[57]梁艳.蜂蜜酚类化合物的色谱分析及其电喷雾质谱断裂途径的研究.(博士论文).2009.08
[58] Ozkan G, Simsek B, Kuleasan H. Antioxidant activities of Satureja cilicica essential oil inbutter and in vitro [J]. J Food Engn,2007,79:1391-1396.
[59]刘骏.结晶紫分光光度法测定Fenton反应产生的羟自由基[J].武汉工业学院学报,2005,24(2):53-55.
[60]程丽华,黄君礼,倪福祥. Fenton试剂生成·OH的动力学研究[J].环境污染治理技术与设备,2003,4(5):12-14.
[61]罗刚,黄君礼. Fe(phen)32+光度法研究Fenton试剂体系中·OH生成率的影响因素[J].化学工程师,2001,87(6):23-25.
[62]刘骏.结晶紫分光光度法测定Fenton反应产生的羟自由基[N].武汉工业学院学报,2005,24(2):53-55.
[63]冯雪.草果精油的抗氧化与分子微胶囊化研究[D].2010.05.
[64]李荣,姜子涛,马丽.月桂精油抗氧化性能及清除自由基能力的研究[J].中国调味品,2009,34(11):58-62.
[65]杨欣,姜子涛,李荣等.分段提取的爪哇香茅挥发油抗氧化性能及清除自由基能力的比较[J].中国食品学报,2011,11(1):34-39.
[66]赫春香,卢丽男,曹璨.荧光动力学分析新方法测定超氧自由基抑制剂的抗氧化活性[J].辽宁师范大学学报(自然科学版),2005,28:73-75.
[67]杨欣,姜子涛,李荣等.柠檬草精油的成分分析和抗氧化能力比较[J].食品科技,2010,35(8):311-316.
[68]杨欣,姜子涛,李荣等.天然香料锡兰香茅挥发油抗氧化活性研究[J].粮食与油脂,2009(11):19-21.
[69]陆占国,郭红转,封丹.芜荽茎叶精油成分及清除DPPH自由基能力研究[J].食品与发酵工业,2006,32(8):24-27.
[70]金鸣,蔡亚欣,李金荣.邻二氮菲-Fe2+氧化法检测H2O2/Fe2+产生的羟自由基[J].生物
化学与生物物理进展,1996,23:553-555.
[2]张志国,吕泰省,姚庆强.何首乌的研究进展[J].解放军药学学报,2008,24(1):62-64.
[3]夏扎旦,萨提瓦力地·赫力力,买合布白·阿不都热依木等.分光光度法测定维吾尔传统药用植物何首乌的总黄酮含量[J].食品科学,2006,27(12):704-705.
[4]王自军.夜交藤中总黄酮的超声提取及含量测定[J].时珍国医国药,2010,21(12):3211-3212.
[5] Xu YL, Dong Q, Hu FZ. Simultaneous quantitative determination of eight active componentsin Polygonum multiflorum Thumb by RP-HPLC [J]. J Chin Pharm Soc,2009,18:358-361.
[6]饶高雄,薛咏梅,惠婷婷等.首乌叶化学成分研究[J].中药材,2009,32(6):891-893.
[7]延玺,刘会青,邹永青等.黄酮类化合物生理活性及合成研究进展[J].有机化学,2008,28(9):1534-1544.
[8] Duan L, Carrier DL, Clausen EC. Silymarin extraction from milk thistle using hotwater [J].Appl Biochem Biotechnol.,2004,(113-116):559-568.
[9] Rogerio AP, Kanashiro A, Fontanar C, et al. Anti-inflammatory activity ofquercetin andisoquercitrin in experimental murine allergic asthma [J]. Inflamm Res,2007,56(10):402-408.
[10]徐美奕,韩雅莉,东野广智等.紫荆花总黄酮的分离纯化与光谱分析[J].中药材,2007,30(10):1252-1255.
[11]姚新生.天然药物化学[M].北京:人民卫生出版社,2001.
[12] Paniwnyk L, Beaufoy E, Lorimer JP, et al. The extraction of rutin from flowerbuds ofSophora japonica [J]. Ultrason Sonochem,2001,8(3):299-301.
[13] Yang Y, Zhang F. Ultrasound-assisted extraction of rutin and quercetin from Euonymusalatus(Thunb.) Sieb [J]. Ultrason Sonochem,2008,15(4):308-313.
[14] Wang X, Ding L, Zhang H, et al. Development of ananalytical method for organotincompounds in fortified flour samples using microwave-assisted extraction andnormal-phase HPLC with UV detection [J]. J Chromatogr B,2006,843(2):268-274.
[15] Chen L, Ding L, Yu A, et al. Continuous determination of total flavonoids in Platycladusorientalis(L.) Franco by dynamic microwave-assisted extraction coupled with on-linederivatization and ultraviolet-visible detection [J]. Anal Chim Acta,2007,596(1):164-170.
[16] Song JZ, Mo SF, Yip YK, et al. Development of microwave assisted extraction for thesimultaneous determination of isoflavonoids and saponins in radix astragali by highperformance liquid chromatography [J]. J Sep Sci,2007,30(6):819-824.
[17] Safaralie A, Fatem S, Sefidkon F..Essentialoil composition ofValeriana officinalis L.rootscultivated in Iran.Comparative analysis between supercritical CO2extraction andhydrodistillation [J]. J Chromatogr A,2008,1180(1-2):159-164.
[18]万茵,谢明勇,董欢欢等.超临界CO2流体提取车前子总黄酮的工艺优化[J].南昌大学学报(理科版),2007,31(2):160-163.
[19]余洪波,张晓昱.酶法在中药提取中的研究进展[J].中成药,2005,27(5):591-593.
[20]庞允,杨建秀,刘子兰.银杏叶中黄酮的提取工艺比较[J].制剂技术,2007,16(3):38-39.
[21]张兆旺,孙秀梅.“半仿生提取法”的特点与应用[J].世界科学技术,2000,2(1):35-38.
[22]赵雪梅,孙秀梅,张兆旺等.杜仲降压方药的半仿生提取工艺研究[J].中国中药杂志,2007,32(21):2247-2252.
[23]陈丛瑾,黄克瀛,李德良.正交试验法优选香椿叶总黄酮的半仿生提取工艺[J].食品科技,2007,(6):119-121.
[24]郭文晶,张守勤,吴华等.均匀设计优化高压提取刺五加叶中总黄酮[J].中药材,2007,30(6):718-720.
[25] Alvarez-Munoz D, Saez M, Lara-Martin PA, et al. New extraction method for the analysisof linear alkylbenzene sulfonates inmarine organisms Pressurized liquid extraction versusSoxhlet extraction [J]. J Chromatogr A,1052(1-2):33-38.
[26] Zhang S, Liu C, Bi H, et al. Extraction of flavonoids from Rhodiola sachlinesisA.Bor byUPE and the antioxidant activity of its extract [J]. Nat Prod Res,2008,22(2):178-187.
[27]田庆来,谢渝春,张波等.溶剂萃取法分离水溶性甘草黄酮[J].过程工程学报,2007,7(3):496-500.
[28] Zheng Y, Haworth IS, Zuo Z, et al. Physicochemical and structural characterization ofquercetin-beta-cyclodex trin complexes [J]. J Pharm Sci,2005,94(5):1079-1089.
[29]屠鹏飞,贾存勤,张洪全.大孔吸附树脂在中药新药研究和生产中的应用[J].世界科学技术,2004,6(3):22-29.
[30] Sun A, Sun Q, Liu R. Preparative isolation and purification of flavone compoundsfromsophora japonica L. by high-speed counter-current chromatography combined withmacroporous resin column separation [J]. J Sep Sci,2007,30(7):1013-1018.
[31] Yao S, Luo JG, Huang XF, et al. Application of preparative high-speed counterchromatography/preparative high-performance liquid chromatographymode in rapidseparation of saponins [J]. J Chromatogr B,2008,864:69-77.
[32] Kohler N, Wray V, Winterhalter P. Preparative isolation of procyanidins from grape seedextracts by high speed counter-current chromatography [J]. J Chromatogr A,2008,1177(1):114-125.
[33] Zhang Q, Chen LJ, Hao YY, et al. Isolation and purification of flavonol glycosides fromGinkgo biloba leaves by high-speed counter-current chromatography [J]. J Sep Sci,2007,30(13):2153-2159.
[34] Arvind R. Equilibrium theory-based design of simulated moving bed proceses underreduced purity requitements Linear isotherms [J]. J Chromatogr A,2008,1185(2):216-222.
[35] Cong JX, Lin BC. Separation of liquiritin by simulated moving bed chromatography. JChromatogr A,2007,1145(1-2):190-194.
[36]李瑞萍,黄骏雄.高效制备液相色谱柱技术的研究进展[J].化学进展,2004,16(2):273-283.
[37]罗艺萍.黄酮类化合物的药理活性研究进展[J].亚太传统医药,2010,6(4):126-128.
[38] Nkenqfack AE, Vouffo TW, Fomum ZT, et al. Prenylated isoflavanone from the roots ofErythrina sigmoidea [J]. Phytochem,1994,36(4):1074-1077.
[39]吉琦正雄.何首乌的成分研究[J]．国外医学·中医药分册，1985,7(1):55.
[40] Zhu HB, Wang YZ, Liu YX, et al. Analysis of flavonoids in Portulaca oleracea L. byUV–Vis spectrophotometry with comparative study on different extraction technologies [J].Food Anal Methods,2010,3:90–97.
[41]仇燕.菜芙蓉花中总黄酮微波提取的研究[J]．中药材,2006,29(4):387-389
[42]陈伟,刘青梅.微波技术在杜仲黄酮提取工艺中的应用研究[J]．食品科学,2006,27(10):285-288.
[43]张琪,刘慧灵,朱瑞等.苦荞麦中总黄酮和芦丁的含量测定方法的研究[J].食品科学,2003,24(7):113-116.
[44]毕洁,杨庆利,于丽娜等.花生壳黄酮乙醇提取工艺探讨[J].现代化工,2008,10(28):316-319.
[45]陈恩赞,苏海佳,谭天伟.微波提取酱渣中总异黄酮的实验研究[J].环境工程学报,2010,4(2):427-430.
[46]周桃英,罗登宏,李国庆等. AB-8大孔树脂纯化荷叶总黄酮的工艺研究[J].中国食品添加剂,20095:113-119.
[47]陶海腾,陈晓明,吕飞杰等.大孔吸附树脂法纯化木薯叶黄酮的初步研究[J].食品工业科技,2009,30(8):192-194.
[48]李旭洲,韩立炜,张岩峰等.葛根中葛根黄酮的水提取及大孔树脂纯化工艺研究[J].北京中医药大学学报,2009,32(9):626-629.
[49] Fu BQ, Liu J, Li H, et al. The application of macroporous resins in the separation of licoriceflavonoids and glycyrrhizic acid[J]. J Chromatogr A,2005,1089:18–24.
[50]李颖畅,孟宪军,李小红.大孔树脂纯化蓝萄叶总黄酮的工艺研究[J].食品工业与发酵,2008,34(1):133-137.
[51]王俊,赵辉,吴福安.树脂吸附法分离纯化桑叶总黄酮[J].离子交换与吸附,2008,24(2):139-147.
[52]刘志祥,曾超珍.大孔树脂法纯化苦丁茶总黄酮的研究[J].时珍国医国药,2009,20(9):2183-2184.
[49] Fu BQ, Liu J, Li H, et al. The application of macroporous resins in the separation of licoriceflavonoids and glycyrrhizic acid[J]. J Chromatogr A,2005,1089:18–24.
[50]李颖畅,孟宪军,李小红.大孔树脂纯化蓝萄叶总黄酮的工艺研究[J].食品工业与发酵,2008,34(1):133-137.
[51]王俊,赵辉,吴福安.树脂吸附法分离纯化桑叶总黄酮[J].离子交换与吸附,2008,24(2):139-147.
[52]刘志祥,曾超珍.大孔树脂法纯化苦丁茶总黄酮的研究[J].时珍国医国药,2009,20(9):2183-2184.
[53]廖琼峰.陆英药效物质基础研究[D].2005.05.
[54]冯广卫,李翠兵,廖尚高等.荭草素和异荭草素、牡荆素和异牡荆素2对碳苷化合物的快速检测与鉴定[J].药物分析杂志,2011,31(7):1263-1268.
[55]于澎,张虹. HPLC-ESI-MS法同时测定虎杖提取物中的17种成分[J].沈阳药科大学学报,2011,28(12):963-968.
[56]张语迟,刘春明,刘志强.罗布白麻与罗布红麻的液相色谱-质谱联用分析[J].分析测试学报,2009,28(10):1148-1154.
[57]梁艳.蜂蜜酚类化合物的色谱分析及其电喷雾质谱断裂途径的研究.(博士论文).2009.08
[58] Ozkan G, Simsek B, Kuleasan H. Antioxidant activities of Satureja cilicica essential oil inbutter and in vitro [J]. J Food Engn,2007,79:1391-1396.
[59]刘骏.结晶紫分光光度法测定Fenton反应产生的羟自由基[J].武汉工业学院学报,2005,24(2):53-55.
[60]程丽华,黄君礼,倪福祥. Fenton试剂生成·OH的动力学研究[J].环境污染治理技术与设备,2003,4(5):12-14.
[61]罗刚,黄君礼. Fe(phen)32+光度法研究Fenton试剂体系中·OH生成率的影响因素[J].化学工程师,2001,87(6):23-25.
[62]刘骏.结晶紫分光光度法测定Fenton反应产生的羟自由基[N].武汉工业学院学报,2005,24(2):53-55.
[63]冯雪.草果精油的抗氧化与分子微胶囊化研究[D].2010.05.
[64]李荣,姜子涛,马丽.月桂精油抗氧化性能及清除自由基能力的研究[J].中国调味品,2009,34(11):58-62.
[65]杨欣,姜子涛,李荣等.分段提取的爪哇香茅挥发油抗氧化性能及清除自由基能力的比较[J].中国食品学报,2011,11(1):34-39.
[66]赫春香,卢丽男,曹璨.荧光动力学分析新方法测定超氧自由基抑制剂的抗氧化活性[J].辽宁师范大学学报(自然科学版),2005,28:73-75.
[67]杨欣,姜子涛,李荣等.柠檬草精油的成分分析和抗氧化能力比较[J].食品科技,2010,35(8):311-316.
[68]杨欣,姜子涛,李荣等.天然香料锡兰香茅挥发油抗氧化活性研究[J].粮食与油脂,2009(11):19-21.
[69]陆占国,郭红转,封丹.芜荽茎叶精油成分及清除DPPH自由基能力研究[J].食品与发酵工业,2006,32(8):24-27.
[70]金鸣,蔡亚欣,李金荣.邻二氮菲-Fe2+氧化法检测H2O2/Fe2+产生的羟自由基[J].生物
化学与生物物理进展,1996,23:553-555.