大纬度引种青蒿药效、产量影响因子及药效分析方法学研究
|
摘要
本文改进并建立了青蒿素提取、分析方法;以此为基础评价了黑龙江省青蒿野生资源的利用价值;并分别于2006、2007和2008年从北纬23-30°间的云南、广西、四川、湖南等地,引进了若干优质种源,在北纬46°以上的黑龙江省不同地区栽培,筛选出适合本地生长、含量高的青蒿种源,建立了青蒿引种栽培技术;研究了影响青蒿素含量、药用生物量的主要因子,选择对青蒿素产量影响大的因子进行正交设计试验,确定了高产青蒿的施肥技术。为黑龙江引种青蒿,开展大规模栽培生产提供了理论依据。主要研究结果如下:
1、改进建立了恒温振荡浸提法作为青蒿素的提取方法:精确称取青蒿粗粉1.000g,40℃干燥3 h,以20 mL石油醚(60～90℃)为溶剂,30℃恒温振荡提取24 h,频率120 r/min,连续提取4次,每次将前一次的提取液倾出加入10 mL提取溶剂,共提取4d,合并提取液,总量为50 mL。该方法应用设备简单、可操作性强;提取溶剂经济、毒性小、回收方便、可重复利用;提取物杂质量小、适合工业生产操作。
2、改进建立了新的青蒿素柱前衍生HPLC-UV测定方法。按文献方法进行青蒿素测定,发现结果不稳定、操作繁琐。研究表明影响柱前衍生HPLC-UV法测定青蒿素含量结果的主要因素是溶解青蒿素标准品(或青蒿提取浸膏)的95%乙醇用量和碱、酸化过程的碱、酸用量。以此为基础改进建立了新的青蒿素柱前衍生HPLC-UV测定方法。
3、评价了黑龙江省野生青蒿资源利用价值。对黑龙江省内不同地区野生青蒿中青蒿素含量进行测定表明,青蒿素含量均低于0.1%,远远低于青蒿素含量≥0.5%的药用标准,即黑龙江省野生青蒿资源无利用价值。
4、建立了青蒿引种栽培技术。对比室内和野外育苗结果及育苗方法表明:黑龙江省引种青蒿,采用温室育苗,可延长青蒿的生长期45 d左右,集成板式纸筒育苗移栽方便、用土量小,成本低。不同地区引种青蒿相同时间收获,育苗和移栽时间越早青蒿素产量最高。引种的青蒿,营养生长可持续到9月初,生长期长近170 d,可获得巨大药用部位生物量;8月20日后陆续转入生殖生长,本地气候高温、光照强,此时可获得高含量的青蒿素。即提早育苗移栽,可延长其生长时间,在气温较高的月份积累最高的青蒿素含量并达到大的生物量,从而提高青蒿素产量。
5、通过大田栽培和露天盆栽方法,研究了植物生长必需的多数营养因子、逆境胁迫、辐射青蒿种子、日光、昼夜温差、栽培密度、育苗和移栽时间对青蒿药用生物量和青蒿素含量的影响,结果如下:
营养元素单因子对青蒿素产量的影响大小顺序为复合肥(NPK)＞K＞Ca＞Fe＞Mg＞B＞P＞Mn＞Cu＞N＞Mo＞Zn＞ck(对照)。最大可使青蒿素含量提高对照的49.3%、青蒿药用生物量提高对照的55.4%、青蒿素产量提高对照的111.8%。
植物生长调节剂单因素对青蒿素产量影响大小顺序为:GA_3＞NAA＞氨基酸微肥＞B9＞KT。最大可使青蒿素含量提高对照的36.9%、青蒿药用生物量提高对照的93.3%、青蒿素产量提高对照的125.5%。
栽培密度大小对青蒿素含量虽有影响但幅度不大,但种植密度大可提高青蒿素产量(源于生物量产量的贡献)。
大田栽培施肥技术:氮肥720 kg/ha、磷肥1080 kg/ha、钾肥1080 kg/ha及钙肥360kg/ha,赤霉素、萘乙酸及氨基酸微肥喷施浓度分别为40 mg/L、5 mg/L及225倍液。青蒿素含量最高可达到:1.0484%,最低为:0.5598%。
6、引种青蒿中青蒿素含量累积趋势与本地野生青蒿一致,但含量高低差异极大,说明青蒿素含量主要受遗传背景控制,其次受环境条件影响。青蒿素含量最高达到:1.0484%,最低为:0.5598%,远大于0.5%的药用标准,引种青蒿中青蒿素含量可达到药用标准,且药用部位生物量超过原产地。引种栽培的青蒿对本地的各种立地条件具有很强的适应性,说明引种青蒿栽培获得成功。因此黑龙江省可充分利用青蒿适应性强的特点,在荒地、废弃地、闲置地、石砾地,开展大规模栽培生产,是个很好的惠农惠林项目。
In this dissertation,the modified extraction and analytical method of artemisinin is proposed,evaluating the value of wild Artemisia annua resource in Heilongjiang Province on the basis.Some high quality provenances of A.annua from Yunnan,Guangxi,Sichuan and Hunanmain in northern latitude 23～30~0 were introduced in the different areas over northern latitude 46~0 of Heilongjiang Province in 2006,2007and 2008,for selecting the one which adapted to the environment and climate of the cultivated area and has high artemisinin content. The technique of introduced A.annua cultivation is established.The main factors which affect A.annua's artemisinin content and medicinal biomass yield are studied in detail and major of them are researched to establish fertilization technique of high A.annua yield by orthogonal design test.It provides theoretical basis for introducing A.annua and promoting a large-scale planting production in Heilongjiang Province,of which the concrete results are as below:
1.The modified extraction method of constant-temperature oscillating digestion is proposed,of which the appropriate technical conditions are established as follows:drying the powder sample in a baker(40℃) to constant weight,weighing 1.000 g and puting it in a reactor,adding petroleum ether(60～90℃) 20 mL,oscillating digestion(120 r/min) at constant temperature of 30℃for 24 h.Do the same thing but using petroleum ether(60～90℃) 10mL to the residue,repeatedly 3 times.Put the extract of the four times together(50mL in total) as the artemisinin extract.This extraction,which performed with simple apparatus under easy operation,uses economical,low-toxicity,recyclable and easy-recovering solvent.It also provides extraction of a relatively high pureness,with which all the above makes it fit for mass production.
2.The artemisinin content detecting method of the newly modified HPLC-UV with Postcolumn Derivation is proposed.Doing the test according to the references showed unstable results and very complex operation.According to our researches and experiments,the main factors that affect the results of HPLC-UV with postcolumn derivation are the amount of 95% ethanol,which is used as solvent for artemisinin standard sample(or epurated artemisinin),and that of NaOH,HAc used during alkali-treating and acid-treating process,by which the newly modified method was proposed.
3.Evaluated the developing value of wild A.annua resource in Heilongjiang Province. After detecting the artemisinin content of A.annua in different areas of Heilongjiang Province, we found that the artemisinin contents are lower than 0.1%,which is far lower than the medicinal criterion of artemisinin content≥0.5%.It means that the wild A.annua in Heilongjiang is useless.
4.The technique of introducing A.annua cultivation is established.Comparing with the methods and results of sowing seedling in fields and greenhouse,the later,can prolong the growth period about 45 days.What's more,the integrated plate-type paper containers,which require very little soil and enable easy transplanting,is quite economical.If we harvest A. annua in different areas at the same time,the earlier we grow,the higher yield of artemisinin we have.The nutritious growth period can prolong to the beginning of September for introduced A.annua,which is nearly 170 days.Above all,great medicinal biomass can be obtained.After 20~(th) of August,the nutritious growth period of A.annua transfers to breeding growth period in succession,and in this period,high artemisinin content can be obtained because the local climate is at high temperature and strong illumination.It is that the growth period can be lengthened if growing seedlings and transplanting are done earlier.Therefore, not only high artemisinin content can be accumulated,but also great medicinal biomass can be obtained at the month of high temperature.Thus,artemisinin production is increased.
5.Through field planting and outdoor pot experiments,we studied most of the essential nutritious factors for plants growth,adversity stress,irradiating the seeds of A.annua,sunshine, temperature difference between day and night,planting density,sowing seedling and transplanting time,which affect A.annua's artemisinin content and medicinal biomass.The results are as below:
Each factor of mineral nutrition has its own influence on the artemisinin production, generalizing the results,the sequence ranked:NPK > K > Ca > Fe > Mg > B > P > Mn > Cu > N > Mo > Zn > ck(control).With proper fertilization,artemisinin content can be increased by 49.3%,medicinal biomass by 55.4%,artemisinin production by 111.8%as a maximum, compared with the control.
Each factor of hormone analogue can influence artemisinin production differently.As for the effect of each factor,the sequence ranked:gibberellins(GA_3)> naphthlcetic acid (NAA)>animo acid micro fertilizer>B_9>6-furifuryl-aminopurine(KT).When used properly, artemisinin content can be increased by 36.9%,medicinal biomass by 93.3%,artemisinin production by 125.5%as a maximum,compared with the control.
Planting density also can influence artemisinin content but not greatly.However, relatively high planting density leads to high artimisinin production(mainly due to high medicinal biomass).
The fertilization technique of planting A.annua in field is as follow:N is 720 kg/ha,P is 1080 kg/ha,K is 1080 kg/ha,Ca is 360 kg/ha.Besides,animo acid micro fertilizer is diluted to 225 times,and GA_3,NAA is 40 mg/L and 5 mg/L,respectively.The highest artemisinin content is 1.0484%,and the lowest one is 0.5598%.
6.The drifts of artemisinin content of introduced A.annua and local wild ones are the same,but their levels differ greatly.It means artemisinin content is mainly controlled by their genetic backgrounds and secondly controlled by the natural circumstance.The highest artemisinin content is 1.0484%,and the lowest one is 0.5598%of introduced A.annua,which are far higher than the medicinal criterion of artemisinin content≥0.5%.While,with artemisinin content reaching industrial standard and medicinal biomass exceeding that of their origin,introduced A.annua has showed great adaptation to the local natural conditions.It means introduced A.annua get great succeed.So considering A.annua's advantage of great adaptation,starting mass planting production in wasteland,abandoned land,uncultivated land and wasteland with some debris will be a nice forestry and agriculture-benifiting project.
1、改进建立了恒温振荡浸提法作为青蒿素的提取方法:精确称取青蒿粗粉1.000g,40℃干燥3 h,以20 mL石油醚(60～90℃)为溶剂,30℃恒温振荡提取24 h,频率120 r/min,连续提取4次,每次将前一次的提取液倾出加入10 mL提取溶剂,共提取4d,合并提取液,总量为50 mL。该方法应用设备简单、可操作性强;提取溶剂经济、毒性小、回收方便、可重复利用;提取物杂质量小、适合工业生产操作。
2、改进建立了新的青蒿素柱前衍生HPLC-UV测定方法。按文献方法进行青蒿素测定,发现结果不稳定、操作繁琐。研究表明影响柱前衍生HPLC-UV法测定青蒿素含量结果的主要因素是溶解青蒿素标准品(或青蒿提取浸膏)的95%乙醇用量和碱、酸化过程的碱、酸用量。以此为基础改进建立了新的青蒿素柱前衍生HPLC-UV测定方法。
3、评价了黑龙江省野生青蒿资源利用价值。对黑龙江省内不同地区野生青蒿中青蒿素含量进行测定表明,青蒿素含量均低于0.1%,远远低于青蒿素含量≥0.5%的药用标准,即黑龙江省野生青蒿资源无利用价值。
4、建立了青蒿引种栽培技术。对比室内和野外育苗结果及育苗方法表明:黑龙江省引种青蒿,采用温室育苗,可延长青蒿的生长期45 d左右,集成板式纸筒育苗移栽方便、用土量小,成本低。不同地区引种青蒿相同时间收获,育苗和移栽时间越早青蒿素产量最高。引种的青蒿,营养生长可持续到9月初,生长期长近170 d,可获得巨大药用部位生物量;8月20日后陆续转入生殖生长,本地气候高温、光照强,此时可获得高含量的青蒿素。即提早育苗移栽,可延长其生长时间,在气温较高的月份积累最高的青蒿素含量并达到大的生物量,从而提高青蒿素产量。
5、通过大田栽培和露天盆栽方法,研究了植物生长必需的多数营养因子、逆境胁迫、辐射青蒿种子、日光、昼夜温差、栽培密度、育苗和移栽时间对青蒿药用生物量和青蒿素含量的影响,结果如下:
营养元素单因子对青蒿素产量的影响大小顺序为复合肥(NPK)＞K＞Ca＞Fe＞Mg＞B＞P＞Mn＞Cu＞N＞Mo＞Zn＞ck(对照)。最大可使青蒿素含量提高对照的49.3%、青蒿药用生物量提高对照的55.4%、青蒿素产量提高对照的111.8%。
植物生长调节剂单因素对青蒿素产量影响大小顺序为:GA_3＞NAA＞氨基酸微肥＞B9＞KT。最大可使青蒿素含量提高对照的36.9%、青蒿药用生物量提高对照的93.3%、青蒿素产量提高对照的125.5%。
栽培密度大小对青蒿素含量虽有影响但幅度不大,但种植密度大可提高青蒿素产量(源于生物量产量的贡献)。
大田栽培施肥技术:氮肥720 kg/ha、磷肥1080 kg/ha、钾肥1080 kg/ha及钙肥360kg/ha,赤霉素、萘乙酸及氨基酸微肥喷施浓度分别为40 mg/L、5 mg/L及225倍液。青蒿素含量最高可达到:1.0484%,最低为:0.5598%。
6、引种青蒿中青蒿素含量累积趋势与本地野生青蒿一致,但含量高低差异极大,说明青蒿素含量主要受遗传背景控制,其次受环境条件影响。青蒿素含量最高达到:1.0484%,最低为:0.5598%,远大于0.5%的药用标准,引种青蒿中青蒿素含量可达到药用标准,且药用部位生物量超过原产地。引种栽培的青蒿对本地的各种立地条件具有很强的适应性,说明引种青蒿栽培获得成功。因此黑龙江省可充分利用青蒿适应性强的特点,在荒地、废弃地、闲置地、石砾地,开展大规模栽培生产,是个很好的惠农惠林项目。
In this dissertation,the modified extraction and analytical method of artemisinin is proposed,evaluating the value of wild Artemisia annua resource in Heilongjiang Province on the basis.Some high quality provenances of A.annua from Yunnan,Guangxi,Sichuan and Hunanmain in northern latitude 23～30~0 were introduced in the different areas over northern latitude 46~0 of Heilongjiang Province in 2006,2007and 2008,for selecting the one which adapted to the environment and climate of the cultivated area and has high artemisinin content. The technique of introduced A.annua cultivation is established.The main factors which affect A.annua's artemisinin content and medicinal biomass yield are studied in detail and major of them are researched to establish fertilization technique of high A.annua yield by orthogonal design test.It provides theoretical basis for introducing A.annua and promoting a large-scale planting production in Heilongjiang Province,of which the concrete results are as below:
1.The modified extraction method of constant-temperature oscillating digestion is proposed,of which the appropriate technical conditions are established as follows:drying the powder sample in a baker(40℃) to constant weight,weighing 1.000 g and puting it in a reactor,adding petroleum ether(60～90℃) 20 mL,oscillating digestion(120 r/min) at constant temperature of 30℃for 24 h.Do the same thing but using petroleum ether(60～90℃) 10mL to the residue,repeatedly 3 times.Put the extract of the four times together(50mL in total) as the artemisinin extract.This extraction,which performed with simple apparatus under easy operation,uses economical,low-toxicity,recyclable and easy-recovering solvent.It also provides extraction of a relatively high pureness,with which all the above makes it fit for mass production.
2.The artemisinin content detecting method of the newly modified HPLC-UV with Postcolumn Derivation is proposed.Doing the test according to the references showed unstable results and very complex operation.According to our researches and experiments,the main factors that affect the results of HPLC-UV with postcolumn derivation are the amount of 95% ethanol,which is used as solvent for artemisinin standard sample(or epurated artemisinin),and that of NaOH,HAc used during alkali-treating and acid-treating process,by which the newly modified method was proposed.
3.Evaluated the developing value of wild A.annua resource in Heilongjiang Province. After detecting the artemisinin content of A.annua in different areas of Heilongjiang Province, we found that the artemisinin contents are lower than 0.1%,which is far lower than the medicinal criterion of artemisinin content≥0.5%.It means that the wild A.annua in Heilongjiang is useless.
4.The technique of introducing A.annua cultivation is established.Comparing with the methods and results of sowing seedling in fields and greenhouse,the later,can prolong the growth period about 45 days.What's more,the integrated plate-type paper containers,which require very little soil and enable easy transplanting,is quite economical.If we harvest A. annua in different areas at the same time,the earlier we grow,the higher yield of artemisinin we have.The nutritious growth period can prolong to the beginning of September for introduced A.annua,which is nearly 170 days.Above all,great medicinal biomass can be obtained.After 20~(th) of August,the nutritious growth period of A.annua transfers to breeding growth period in succession,and in this period,high artemisinin content can be obtained because the local climate is at high temperature and strong illumination.It is that the growth period can be lengthened if growing seedlings and transplanting are done earlier.Therefore, not only high artemisinin content can be accumulated,but also great medicinal biomass can be obtained at the month of high temperature.Thus,artemisinin production is increased.
5.Through field planting and outdoor pot experiments,we studied most of the essential nutritious factors for plants growth,adversity stress,irradiating the seeds of A.annua,sunshine, temperature difference between day and night,planting density,sowing seedling and transplanting time,which affect A.annua's artemisinin content and medicinal biomass.The results are as below:
Each factor of mineral nutrition has its own influence on the artemisinin production, generalizing the results,the sequence ranked:NPK > K > Ca > Fe > Mg > B > P > Mn > Cu > N > Mo > Zn > ck(control).With proper fertilization,artemisinin content can be increased by 49.3%,medicinal biomass by 55.4%,artemisinin production by 111.8%as a maximum, compared with the control.
Each factor of hormone analogue can influence artemisinin production differently.As for the effect of each factor,the sequence ranked:gibberellins(GA_3)> naphthlcetic acid (NAA)>animo acid micro fertilizer>B_9>6-furifuryl-aminopurine(KT).When used properly, artemisinin content can be increased by 36.9%,medicinal biomass by 93.3%,artemisinin production by 125.5%as a maximum,compared with the control.
Planting density also can influence artemisinin content but not greatly.However, relatively high planting density leads to high artimisinin production(mainly due to high medicinal biomass).
The fertilization technique of planting A.annua in field is as follow:N is 720 kg/ha,P is 1080 kg/ha,K is 1080 kg/ha,Ca is 360 kg/ha.Besides,animo acid micro fertilizer is diluted to 225 times,and GA_3,NAA is 40 mg/L and 5 mg/L,respectively.The highest artemisinin content is 1.0484%,and the lowest one is 0.5598%.
6.The drifts of artemisinin content of introduced A.annua and local wild ones are the same,but their levels differ greatly.It means artemisinin content is mainly controlled by their genetic backgrounds and secondly controlled by the natural circumstance.The highest artemisinin content is 1.0484%,and the lowest one is 0.5598%of introduced A.annua,which are far higher than the medicinal criterion of artemisinin content≥0.5%.While,with artemisinin content reaching industrial standard and medicinal biomass exceeding that of their origin,introduced A.annua has showed great adaptation to the local natural conditions.It means introduced A.annua get great succeed.So considering A.annua's advantage of great adaptation,starting mass planting production in wasteland,abandoned land,uncultivated land and wasteland with some debris will be a nice forestry and agriculture-benifiting project.
引文
[1]屠呦呦,倪慕云,钟裕亮,等.中药青蒿化学成分研究Ⅰ.药学学报,1981,16(5):366-370
[2]青蒿素结构研究协作组.一种新型的倍半萜内酯--青蒿素.科学通报,1977,22(3):142
[3]Sipahimal A T,Fulzele D P,Heble M R.Rapid method for the detection and determination of artemisinin by gas chromatography.J Chromatogr,1991,538(2):425-455
[4]Klayman D L,Lin A J,Acton N,et al.Isolation of artemisinin(Qinghaosu) from Artemisia annua growing in the United States.J Nat Prod,1984,47(40):715-717
[5]Klayman D L.Qinghaosu(Artemisinin):An antimalarial drug from China.Science,1985,228(4703):1049-1055
[6]Avery M A,Jennings W C,Chong W K M.Simplified anelogues of the antimalarial artemisinin:Synthesis of 6,9-Desmethyl artmisinin.J Org Chem,1989,54(8):1792-1795
[7]Posner G H,Chong H O.A regiospecifieally oxygen-18 labeled 1,2,4-Trioxane;a single chemical model system to probe the mechanison(s) for the syntimal activity of artemisinin.J Am Chem Soc,1992,114(21):8328-8329
[8]王明学.青蒿素及其衍生物的开发和产业化.中国医药技术与市场,2006,6(2):31-33
[9]P.Christen,J.L.Veuthey.New Trends in Extraction,Identification and Quantification of Artemisinin and its Derivatives.Current Medicinal Chemistry,2001,(8):1827-1839
[10]王宗德,孙芳华.青蒿素理化性质及其测定方法的研究进展.江西农业大学学报,1999,21(4):606-611
[11]国家药典委员会.中华人民共和国药典(二部).北京:化学工业出版社,2005:309
[12]赵世善,曾美怡.高效液相色谱法测定青蒿植物中的青蒿素.药物分析杂志,1986,6(1):3-4
[13]杜小英,张玲,石红,等.青蒿素最佳提取工艺研究.河北中医药学报.2005,20(3):31,41
[14]赵兵,王玉春,吴江,等.青蒿素提取条件研究.中草药,2000,31(6):421-423
[15]韦国锋,覃特营,莫少泽.提取青蒿素实验条件的研究.右江民族医学院学报,1995,17(2):137-139
[16]邓启华,李子成,杨宗伟.超声波连续逆流提取青蒿素的新工艺.四川化工,2008,11(3):17-19
[17]Ganzler K,Salgo A,Valko K.Microwave extraction-a novel simple preparation method for chromatography.J Chromatogr,1986,371:299-306
[18]Craveriro A A,Matos F J.Microwave oven extraction of essential oil.Flavour Fragrance J, 1989,4(1):43-46
[19]Jean F I,Collin G J,Lord D.Essential oils and microwave extracts of cultivated plants.Perfum Flavor,1992,17(3):35-41
[20]梁忠生.青蒿微波预处理对青蒿素提取产率的影响研究.中南药学,2004,2(6):342-344
[21]喻凌寒,宋之光,陈江韩,等.快速溶剂萃取反相高效液相色谱法测定青蒿中的青蒿素.分析实验室,2006,25(8):68-71
[22]周毅峰,石开明,艾训儒,等.快速溶剂萃取法提取青蒿素条件研究.中药材,2008,31(2):296-298
[23]阮栋梁,王丰玲,张英锋,等.青蒿素的制备、用途和展望.渤海大学学报(自然科学版),2007,28(2):108-112
[24]Socrates Quispe-Condori,Deny Sanchez,Mary A.Foglio,et al.Global yield isotherms and kinetic of artemisinin extraction from Artemisia annua L.leaves using supercritical carbon dioxide.J.of Supercritical Fluids 2005,36:40-48
[25]钱国平,杨亦文,吴彩娟,等.超临界CO_2从青蒿中提取青蒿素的研究.化工进展,2005,24(3):286-290 302
[26]韦国锋,黄祖良,何有成.大孔吸附树脂提取青蒿素的研究.离子交换与吸附,2007,23(4):373-377
[27]张玲,杜小英,姜锦花,等.青蒿素提取分离工艺研究.现代中药研究与实践,2005,19(6):57-58
[28]胡淼,钱国平,苏宝根,等.硅胶柱层析纯化青蒿素.华西药学杂志,2005,20(4):283-285.
[29]金玲,居明秋.引种青蒿中青蒿素含量的测定.基层中药杂志,1998,12(3):34
[30]沈旋坤,严克东.紫外分光光度法测定青蒿素含量.药物分析杂志,1983,3(1):24
[31]李春莉,王莎莉,王亚平,等.紫外分光光度法测定青蒿素的含量.重庆医科大学学报,2007,32(4):413-415
[32]周浓,段意梅,陈强.青蒿素含量测定方法改进及青蒿植物质量评价.时珍国医国药,2008,19(4):911-912
[33]赵永成,刘荣,赵正荣,等.昭通产青蒿植物中青蒿素的分析研究.基层中药杂志,2001,15(2):7-8
[34]刘炳玉,田惠君,崔进,等.青蒿素的红外光谱定量测定.药物分析杂志,1994,14(4):47-48
[35]刘浩,黄艳萍,相秉仁,等.青蒿中青蒿素的近红外光谱法测定.中国医药工业杂志,2007,38(8):584-587
[36]张荣沭,赵敏,韩颂.引种的不同种源黄花蒿青蒿素的含量研究.林产化学与工业,2008,28(6):贝码待定
[37]Congyue A.Peng,Jorge F.S.Ferreira,Andrew J.Wood.Direct analysis of artemisinin from Artemisia annua L.using high-performance liquid chromatography with evaporative light scattering detector,and gas chromatography with flame ionization detector.Journal of Chromatography A,2006,1133:254-258
[38]赵兵,王玉春,吴江,等.超声波用于强化石油醚提取青蒿素.化工冶金,2000,21(3):310-313
[39]王剑文,郑丽屏,谭仁祥.促进青蒿发根青蒿素合成的内生真菌诱导子的制备.生物工程学报,2006,22(5):829-834
[40]Filip C.W.,Sofie R.F.,Lies Maes,et al.Quantitation of artemisinin and its biosynthetic precursors in Artemisia annua L.by high performance liquid chromatography-electrospray quadrupole time-of-flight tandem mass spectrometry.Journal of Chromatography A,1118(2006) 180-187
[41]刘丽芳,王茜,李海燕,等.柱前衍生-RP-HPLC法测定青蒿中青蒿素的含量.中国野生植物资源,2004,23(6):60-62
[42]申薇薇,王锐利,胡爽,等.高效液相-柱后衍生法测定青蒿素含量.中国药物与临床,2007,7(9):657-659
[43]陈迪钊,刘坚.青蒿素的热稳定行为及气相色谱分析.怀化学院学报,2006,25(8):62-64
[44]郭燕,王俊,陈正堂.青蒿素类药物的药理作用新进展.中国临床药理学与治疗学,2006,11(6):615-620
[45]徐振海.青蒿素药物作用研究简介.安徽预防医学杂志,2004,10(3):181-182
[46]Pandey A V,Tekwani B L,Singh R L,Chauhan V S.Artemisinin,an endoperoxide antimalarial,disrupts the hemoglobin catabolism and heme- detoxification systems in malarial parasite.J Biol Chem,1999,274:19383-8
[47]Dorn A,Vippagunta S R,Matile H,et al.An assessment of drug-haematin binding as a mechanism for inhibition of haematin polymerisation by quinoline antimalarials.Biochem Pharmacol,1998,55:727-736
[48]Hoppe H C,Van Schalkwyk D A,Wiehart U I,et al.Antimalarial quinolines and artemisinin inhibit endocytosis in Plasmodium falciparum.Antimicrob Agents Chemother,2004,48:2370-8
[49]Aldieri E,Atragene D,Bergandi L,et al.Artemisinin inhibits inducible nitric- oxide synthase and nuclear factor NF-kB activation.FEBS Lett,2003,552:141-144
[50]Eckstein-Ludwig U,Webb R J,Van Goethem I D,et al.Artemisinin target the SERCA of Plasmodium falciparum.Nature,2003;424:957-964
[51]Ridley R G.To kill a parasite.Nature,2003;424:887-893
[52]梅林,石开云,苏建华,等.青蒿素国内研究进展.激光杂志,2008,29(2):95-96
[53]张剑锋,闻礼永.抗日本血吸虫药物作用机制的研究进展.国外医学·流行病学传染病学分册,2004,31(6):368-370
[54]崔金凤.治疗血吸虫病药物综述.安徽预防医学杂志,2005,11(3):172-173
[55]杨翠萍.青蒿素及其衍生物抗弓形虫的研究进展.国际医学寄生虫病杂志,2006,33(3):160-162
[56]邓定安.有抗肿瘤活性的青蒿素衍生物.药学学报,1992,27(4):317-320
[57]Efferth T,Dunstan H,Sauerbrey A,et al.The antimalarial artemisinin is also active against cancer.Int J Oncol,2001,18(4):767-773
[58]Singh N P,Lai H.Selective toxicity of dihydroartemisinin and holotransferrin toward human breast cancer cells.Life Sciences,2001,70:49-56
[59]温悦,孟德胜.青蒿素类药物药理作用研究进展.医药导报,2007,26(10):1193-1195
[60]易剑峰,王珏,林色奇,等.青蒿素及其衍生物免疫调节研究概况.江西中医学院学报,2006,18(2):69-71
[61]郑全芳,杨志刚,孙红祥.青蒿素及其衍生物药理作用研究进展.中国兽药杂志,2006,40(1):40-44
[62]贺小青,方鹏飞.青蒿素及其衍生物的药理作用.医药导报,2006,25(6):528-530
[63]胡世林,许有玲.纪念青蒿素30年.世界科学技术-中医药现代化,2005,7(2):1-2
[64]韦霄,李锋,许成琼,等.不同栽培措施对青蒿产量和青蒿素含量的影响.广西科学院学报,1999,15(3):132-136
[65]岑丽华,徐良,黄茱刚,等.不同纬区及不同栽培立地条件对青蒿青蒿素含量的影响.安徽农学通报,2007,13(13):46-47
[66]钟国跃,周华蓉,凌云,等.青蒿优质种质资源的研究.中草药,1998,29(4):264-266
[67]杨水平,杨宪,黄建国,等.青蒿素生产研究进展.热带亚热带植物学报,2004,12(2):189-194
[68]韦美丽,崔秀明,陈中坚,等.青蒿栽培研究进展.现代中药研究与实践,2005,19(5):60-64
[69]钟国跃,周华蓉,凌云,等.青蒿优质种植资源的研究.中草药,1998,29(4):264-267
[70]钟凤林,陈和荣,陈敏,等.青蒿最佳采收时期、采收部位和干燥方式的实验研究.中国中药杂志,1997,22(7):405-406
[71]齐向娟,李士雨,何玉娟.天津地区青蒿中青蒿素测定.中草药,2006,37(3):449-450
[72]陈强,张积强.青蒿素的提取、分离和测定.宝鸡文理学院学报(自然科学版),1999,19(1):47-48
[73]李吉和,张慧灵,李小玲,等.内蒙古地区青蒿中青蒿素的SFEHPLC测定.中药材,2000,23(12):756-758
[74]Woerdenbag H J,Pras N,Chan N G,et al.Artemisinin Related Sesquiterpenes and Essential oil in Artemisia annua During a Vegtation Period in Vietem.Plants Medics,1994, 60(3):272-275
[75]Chan Kit L.Selection of high artemisinin Melding Artemisia annua.Plants Medics,1995,61(3):285-287
[76]Charles D J,Simon J E,Wood K V,Heinstein P.Germplasm variation in artemisinin content of Artemisia annua using an alternative method of artemisinin analysis from crude plant extracts.J Nat Prod,1990,53:157-160
[77]Trigg P I.Qinghaosu(Artemisinin) as an antimalarial drug.Econ Med Plant Res,1990,3:20-55
[78]Klayman D L.Qinghaosu(artemisinin):an antimalarial drug from China.Science,1985,228:1049-1055
[79]耿飒,叶和春,李国凤,等.中药青蒿的生理生化特征及其研究进展.应用与环境生物学报,2002,8(1):90-97
[80]张萍,张子忠.山东引种青蒿青蒿素含量分析.山东中医药大学学报,2001,25(3):229-231
[81]韦霄,李锋,许成琼,等.青蒿生物学特性研究.广西植物,1997,17(2):166-168
[82]韦记青,韦霄,蒋运生,等.青蒿高产栽培技术.广西农业科学,2005,36(5):472-743
[83]李典鹏,梁小燕,陈秀珍,等.采用薄层层析-紫外分光光度法测定广西不同产地青蒿中青蒿素含量.广西植物,1995,15(3):254-255
[84]傅德明,蔡正江,毛禄国.青蒿生物学特性及规范化栽培技术.中国农技推广,2005,(12):34-35
[85]胡四化,胡家华,李让平.青蒿菊花瘿蚊的特征性与综合防治.广东农业科学,2006,(8):66
[86]陈海梅.青蒿栽培技术.特种经济动植物,2006,(3):26
[87]Singh A,Vishwakarma R A,Husssin A.Evaluation of Artemisia annua strains for higher artemisinin production.Planta Med,1988,54:475
[88]Ferreira J F S,Janick J.Production and detection of artemisinin form Artemisia annua.Acta Hort,1995,390:41-49
[89]Ferreira J F S,Charles D,Simon J E,Janick J.Effect of drying methods on the recovery and yield of artemisinin from Artemisia annua L..Hort Science,1992,27:650(Abstr.565)
[90]陈平.黄花资源及开发利用概述.广西林业科学,1998,27(1):35-36
[91]陈和荣.青蒿新品系“京厦Ⅰ号”的选育.中国技术成果大全,1992,1:186-187
[92]李锋,韦霄,许成琼,等.广西青蒿类型调查研究.广西植物,1997,7(3):231-234
[93]黄正方,郑贵华,李成东,等.影响青蒿成分(青蒿素)含量的因素研究.西南农业大学学报,1997,19(1):93-94
[94]Delabays N,Benakis A,Coll J.et al.Selection and breeding for high arteminin (qinghaosu)yielding strains of Artemisia annua.Acta Hort,1993,330:203-207
[95]Pras N,Visser J F,Batterman S,Woerdenbag H J,Malingre T M,Lugt C B.Laboratory selection of Artemisia annua L.for high artemisinin yielding types.Phytochem Anal,1991,2:80-83
[96]Geldre E U,Vergauwe A,Vanden Eeckout E.State of the art of the production of the antimalarial compound artemisinin in plants.Plant Mol Biol.1997,33:199-209
[97]Elhag H M,EL-Domiaty M M,El-Feraly F S,Mossa J S,El-Olemy M M.Selection and micropropagation of high artemisinin producing clones of Artemisia annua L..Phytother Res.1992,6:20-24
[98]Ferreira J F S,Simon J E,Janick J.Developmental studies of Artemisia annua flowering and artemisinin production under greenhouse and field conditions.Plants Med,1995,61:167-170
[99]Sangwan R S,Sangwan N S,Jain D C,et al.RAPD profile based genetic characterization of chemotypic variants of Artemisia annua L..Biochem Mol Biol Int,1999,47:935-944
[100]Wallaart T E,Pras N,Quax W J.Seasonal variation of artemisinin and its biosynthetic precursors in tetraploid Artemisia annua plants compared with the diploid wild-type[J].Planta Med,1999,65:723-728
[101]Magalhaes Redro M De.New hybrid lines of the antinalarial species Artemisia annua L.guarantee its growth in Brazil.Cienciae Culura(Sao Paulo),1997,49(5):413-415
[102]Enserink M.Infectious diseases:source of new hope against malaria is in short supply.Science,2005,307(5706):33
[103]Bouwmeester H J,Wallarrt T E,Janssen M H A,et al.Amorpha-4-11-diene synthase catalyses the first probable step in artemisinin biosynthesis.Phytochemistry,1999,52:843-854
[104]Picausd S,Mercke P,He X,et al.Amorpha-4,11-diene synthase:mechanism and stereochemistry of the enzymatic cyclization of farnesyl diphosphate.A rchies Biochem Biophy,2006,448:150-155
[105]Kraker J W,Franssen M C R,Dalm M C F,et al.Biosynthesis of germacrene carbosylic acid in chicory roots:demonstracion of a cytochrome P450(+)germacrene a hydroxyase and NADP+-denpendent sesquiterpenoid dehy-drogenase(s)involved in sesquiterpene lactone biosynthesis.Plant Physiol,2001,125(4):1930-1940
[106]Bertea C M,Freije J R,DerWoude H,et al.Identification of intermediates and enzymes involed in the early steps of artemisinin biosynthesis in Artemisia annua.Planta Med,2005,71:40-47
[107]Bogers R J,Craker L E,Langer D.Medicinal and Aromatic Plants.Amsterdam:Springer,2006:162
[108]Teoh K H,Polichuk D R,Reed D W,et al.Artemisia annua(Asteraceae)trichome- specific cDNAs reveal cyphavl,a cytochrome P450 with a key role in the biosynthesis of the antimalarial sequiterpene lactone artemisinin.FEB SLett,2006,580:1411-1416
[109]Ro D K,Paradise E M,Ouellet M,et al.Production of the antimalarial drug precursor artemisinic acid in engineered yeast.Nature,2006,440(13):940-943
[110]Abdin M Z,Israr M,Srivastava P S,et al.Invitroproduction of artemisinin.a novel antimalarial compound from Artemisia annua L..J Med Arom Plant Sci,2000,22(4A):378-384
[111]Dhingra V,Rajoli C,Narasu M L.Partial purification of proteins involved in the bioconversion of arteannuin B to artemisinin.Bioresour Technol,2000,73:279-282
[112]Dhingra V,Rajoli C,Narasu M L.Purification and characterization of enzyme involved in the biocon version of arteannuin B to artemisinin.Biochem Biophys Res Commun,2001,281:558-561
[113]Tatineni R,Doddapaneni K K,Dalavayi S,et al.Microbacterium trichotecenoly ticum enzymemediated transformation of arteannuin B to artemisinin.Process Biochem,2006,41(12):2464-2467
[114]Brown G D,Sy L K.In vivo transformations of dihydroartemisinin acid in Artemisia annua Plants.Tetrahedron,2004,60:1139-1159
[115]Wallaart T E,Van Uden W,Lubberink H G M,et al.Isolation and identification of dihydroartemisinic acid from Artemisia annua and its possible role in the biosynthesis of artemisinin.J Nat Prod,1999,62:430-433
[116]Wallarrt T E,Pras N,Quax W J.Isolation and identification of dihydroartemisinic acid hydroperoxide from Artemisia annua:a novel biosynthetic precursor of artemisinin.J Nat Prod,1999,62:1160-1162
[117]Sy L K,Brown G D.The mechanism of the spontaneous autoxidation of dihydroartemisinic acid.Tetrahedron,2002,58:879-908
[118]Sy L K,Brown G D.The role of the 12-carboxylic acid group in the spontaneous autoxidation of dihydroartemisinic acid.Tetrahedron,2002,58:909-923
[119]Brown G D,Sy L K.Invivo transformations of dihydroartemisinic acid in Artemisia annua plants.Tetrahedron,2004,60:1139-1159
[120]刘硕谦,田娜,李娟,等．青蒿素组合生物合成的研究进展．中草药,2007,38(9)：1425～1431
[121]Leyes A E,Baker J A,Hahn F M,et al.Biosynthesis of isoprenoids in Escherichia coli:stereochemistry of the reaction catalyzed by isopentenyl diphosphate:dimethylallyl diphosphate isomerase.Chem Commun,1999:717-728
[122]Sandmann G.Combinatorial biosynthesis of carotenoids in a heterologous host:a powerful approach for the biosynthesis of novel structures.Chem BioChem,2002,3:629- 635
[123]Huang Q L,Roessner C A,Croteau R,et al.Engineering E.coli for the synthesis of taxadiene,a key intermediate in the biosynthesis of taxo..Bioorgan Med Chem,2001,9:2237-2242
[124]Cunningham F X,Lafond T P,Gantt E,et al.Evidence of a role for lytb in the nonmevalonate pathway of isoprenoid biosynthesis.J Bacteriol,2000,182:5841-5848
[125]Martin V J J,Pitera D J,Withers S T,et al.Engineering a mevalonate pathway in Escherichia coli for production of terpenoids[J].Nature Biotechnol,2003,21(7):796-802
[126]Zahiri H S,Yoon S H,Keasling J D,et al.Coenzyme Q10 production in recombinant Escherichia coli strains engineered with a heterologous decaprenyl diphosphate synthase gene and foreign mevalonage pathway.Metabolic Eng,2006,8:406-416
[127]Martin V J J,Yoshikuni Y,Keasling J D.The invivo synthesis of plant sesquiterpenes by Escherichia coli.Biotechnol Bioeng,2001,75:497-503
[128]Pitera D J,Paddon C J,Newmanand J D,et al.Balancing a heterologous mevalonate pathway for improved isoprenoid production in Escherichia coli.Metabolic Eng,2007,9(2):193-207
[129]Lindahl A L,Olsson M E,Mercke P,et al.Production of the artemisinin precursor amorpha-4,11-diene by engineered Saccharomyces cerevisiae.Biotechnol Lett,2006,28:571-580
[130]Hampton R,Dimster-Denk D,Rine J.The biology of HM G-CoA reductase:the pros of contra-regulation.Trends Biochem Sci,1996,21:140-145
[131]Donald K,Hampton R,Fritz I.Effects of overproduction of the catalytic domain of 3-hydroxy -3-methylglutaryl coenzyme A reductase on squalene synthesis in Saccharomyces cerevisiae.Appl Environ Microbiol,1997,63:3341-3344
[132]Starari V J,Escalante-Semerana J C.Identification of the protein acetyltransferase(Pat)enzyme that acetylates acetyl-CoA synthetase in Salmonella enterica.J Mol Biol,2004,340:1005-1012
[133]Shiba Y,Paradise E M,Kirby J,et al.Engineering of the pyruvate dehydrogenase bypass in Saccharomyces cerevisiae for high-level production of isoprenoids.Metabolic Eng,2007,9(2):160-168
[134]朱卫平.野生青蒿的引种驯化和高青蒿素含量栽培品种选育目标性状的研究.2003,博士学位论文.15,37
[135]邓素兰,余继宏,毛丽梅.青蒿中青蒿素的提取分离研究.安徽农学通报.2007,13(5):31-34
[136]王轶.青蒿素最佳提取工艺研究.河南农业科学.2007,(7):84-86
[137]黄海滨,岑家铭,奉建芳,等.RP-HPLC测定青蒿中青蒿素的含量.广西大学学报 (自然科学版),1994,19(2):194-196
[138]Ferreira J F S,Simon J E,Janick J.Artemisia annua:botany,horticulture,pharmacology.Hort Re,1997,19:319-371
[139]朱卫平,盛孝邦.青蒿素的组织化学定位及其含量相关性研究.中草药,2003,34(9):852-854
[140]陈立亭,孙玉亭著.黑龙江省气候与农业.北京:气象出版社,2000:1-3
[141]郭家林,姬菊枝编著.哈尔滨市气候预测及其对生态环境的影响.北京:气象出版社,2000:1,2,4,10,12,14,16,104
[142]陈福泰,张桂花.药用青蒿植株中青蒿素合成的若干生理因子的研究.植物生理学通讯.1987,(5):26-30
[143]Ferreira J F S,Simon J E,Janick J.Developtmental studies of Artemisia annua L..Int.J.Plant Sci,1993,154(1):107-118
[144]李国栋,周全,赵长文,等.青蒿素类药物的研究现状.中国药学杂志,1998,33(7):385-389
[145]Ferreira J F S,Simon J E,Janick J.Artemisia annua:botany,horticulture,pharmacology.Hort Re,1997,19:319-371
[146]朱卫平,盛孝邦.青蒿素的组织化学定位及其含量相关性研究.中草药,2003,34(9):852-854
[147]江苏新医学院编.中药大辞典.下册.上海:上海科学技术出版社,1977,2052
[148]Laughlin J C.Effect of agronomic practices on Plant yield and antimalarial constituents of Artemisia annua L..Acta Hort,1993,33:53-61
[149]农业部种植业管理司,全国农业技术推广服务中心编著.测土配方施肥技术.北京:中国农业出版社,2006:3
[150]彭剑涛编.植物生长调节剂和除草剂使用技术.上海:上海科学普及出版社,2000:4-22
[151]刘春朝,王立春,叶和春.青蒿素生物合成研究进展.天然产物研究与开发,2000,12(1):83-86
[152]李弘剑,张毅.青蒿培养细胞中青蒿素生物合成代谢的体外调节.中国生物化学与分子生物学学报,1999,15(3):479-483
[153]Srivastava N K,Sharma S.Influence of micronutrient imbalance on growth and artemisinin content in Artemisia annua.Indian J Pharm Sci,1990,2:225-227
[154]萧浪涛,王少先,彭克勤,等.自然干旱胁迫下配方肥增效剂对水稻内源激素的影响.中国水稻科学,2005,19(5):417-421
[155]Liersch R,Soicke H,Stehr C,Tullner H U.Formation of artemisinin in Artemisia annua during one vegetation period.Planta Med,1986,52:387-390
[156]Shukla A,Farooqi-Abad A H,Shukla Y N,Sharma S.Effect of triacontanol and chlormequat on growth,plant hormones and artemisinin yield in Artemisia annua L..Plant Grow Regul.1992,11:165-171
[157]杨再强,王立新.观赏植物辐射诱变育种研究进展.四川林业科学,2006,27(3):19-23
[158]冯璐,那日.植物育种的物理学方法.草业科学,2005,22(12):63-66
[159]杨海梅,李明思,谢云.青蒿耗水及生长规律试验研究.石河子大学学报,2005,23(3):339-341
[160]王三根,梁颖.中药青蒿的生态生理及其综合利用中国野生植物资源,2003.8:47-49
[161]Sushil Kumar,S.K.Gupta,Poorinima Singh,et al.High yields of artemisinin by multiharvest of Artemisia annua crops.Industrial Crops and Products,2004,(19):77-90
[162]韦国锋,梁峰,覃特营.青蒿素不同提取工艺的研究.数理医药学杂志,2002,15(2):170-171
[163]郝金玉,韩伟,施超欧,等.青蒿中青蒿素的微波辅助提取.中国医药工业杂志,2002,33(8):385-387
[164]M.Irfan Qureshi,M.Israr,M.Z.Abdin,et al.Responses of Artemisia annua L.and saltinduced oxidative stress.Environmental and Experimental Botany 53(2005) 185-193
[165]雷红松.青蒿组培苗的移栽.特种经济动植物,2004,(2):31
[2]青蒿素结构研究协作组.一种新型的倍半萜内酯--青蒿素.科学通报,1977,22(3):142
[3]Sipahimal A T,Fulzele D P,Heble M R.Rapid method for the detection and determination of artemisinin by gas chromatography.J Chromatogr,1991,538(2):425-455
[4]Klayman D L,Lin A J,Acton N,et al.Isolation of artemisinin(Qinghaosu) from Artemisia annua growing in the United States.J Nat Prod,1984,47(40):715-717
[5]Klayman D L.Qinghaosu(Artemisinin):An antimalarial drug from China.Science,1985,228(4703):1049-1055
[6]Avery M A,Jennings W C,Chong W K M.Simplified anelogues of the antimalarial artemisinin:Synthesis of 6,9-Desmethyl artmisinin.J Org Chem,1989,54(8):1792-1795
[7]Posner G H,Chong H O.A regiospecifieally oxygen-18 labeled 1,2,4-Trioxane;a single chemical model system to probe the mechanison(s) for the syntimal activity of artemisinin.J Am Chem Soc,1992,114(21):8328-8329
[8]王明学.青蒿素及其衍生物的开发和产业化.中国医药技术与市场,2006,6(2):31-33
[9]P.Christen,J.L.Veuthey.New Trends in Extraction,Identification and Quantification of Artemisinin and its Derivatives.Current Medicinal Chemistry,2001,(8):1827-1839
[10]王宗德,孙芳华.青蒿素理化性质及其测定方法的研究进展.江西农业大学学报,1999,21(4):606-611
[11]国家药典委员会.中华人民共和国药典(二部).北京:化学工业出版社,2005:309
[12]赵世善,曾美怡.高效液相色谱法测定青蒿植物中的青蒿素.药物分析杂志,1986,6(1):3-4
[13]杜小英,张玲,石红,等.青蒿素最佳提取工艺研究.河北中医药学报.2005,20(3):31,41
[14]赵兵,王玉春,吴江,等.青蒿素提取条件研究.中草药,2000,31(6):421-423
[15]韦国锋,覃特营,莫少泽.提取青蒿素实验条件的研究.右江民族医学院学报,1995,17(2):137-139
[16]邓启华,李子成,杨宗伟.超声波连续逆流提取青蒿素的新工艺.四川化工,2008,11(3):17-19
[17]Ganzler K,Salgo A,Valko K.Microwave extraction-a novel simple preparation method for chromatography.J Chromatogr,1986,371:299-306
[18]Craveriro A A,Matos F J.Microwave oven extraction of essential oil.Flavour Fragrance J, 1989,4(1):43-46
[19]Jean F I,Collin G J,Lord D.Essential oils and microwave extracts of cultivated plants.Perfum Flavor,1992,17(3):35-41
[20]梁忠生.青蒿微波预处理对青蒿素提取产率的影响研究.中南药学,2004,2(6):342-344
[21]喻凌寒,宋之光,陈江韩,等.快速溶剂萃取反相高效液相色谱法测定青蒿中的青蒿素.分析实验室,2006,25(8):68-71
[22]周毅峰,石开明,艾训儒,等.快速溶剂萃取法提取青蒿素条件研究.中药材,2008,31(2):296-298
[23]阮栋梁,王丰玲,张英锋,等.青蒿素的制备、用途和展望.渤海大学学报(自然科学版),2007,28(2):108-112
[24]Socrates Quispe-Condori,Deny Sanchez,Mary A.Foglio,et al.Global yield isotherms and kinetic of artemisinin extraction from Artemisia annua L.leaves using supercritical carbon dioxide.J.of Supercritical Fluids 2005,36:40-48
[25]钱国平,杨亦文,吴彩娟,等.超临界CO_2从青蒿中提取青蒿素的研究.化工进展,2005,24(3):286-290 302
[26]韦国锋,黄祖良,何有成.大孔吸附树脂提取青蒿素的研究.离子交换与吸附,2007,23(4):373-377
[27]张玲,杜小英,姜锦花,等.青蒿素提取分离工艺研究.现代中药研究与实践,2005,19(6):57-58
[28]胡淼,钱国平,苏宝根,等.硅胶柱层析纯化青蒿素.华西药学杂志,2005,20(4):283-285.
[29]金玲,居明秋.引种青蒿中青蒿素含量的测定.基层中药杂志,1998,12(3):34
[30]沈旋坤,严克东.紫外分光光度法测定青蒿素含量.药物分析杂志,1983,3(1):24
[31]李春莉,王莎莉,王亚平,等.紫外分光光度法测定青蒿素的含量.重庆医科大学学报,2007,32(4):413-415
[32]周浓,段意梅,陈强.青蒿素含量测定方法改进及青蒿植物质量评价.时珍国医国药,2008,19(4):911-912
[33]赵永成,刘荣,赵正荣,等.昭通产青蒿植物中青蒿素的分析研究.基层中药杂志,2001,15(2):7-8
[34]刘炳玉,田惠君,崔进,等.青蒿素的红外光谱定量测定.药物分析杂志,1994,14(4):47-48
[35]刘浩,黄艳萍,相秉仁,等.青蒿中青蒿素的近红外光谱法测定.中国医药工业杂志,2007,38(8):584-587
[36]张荣沭,赵敏,韩颂.引种的不同种源黄花蒿青蒿素的含量研究.林产化学与工业,2008,28(6):贝码待定
[37]Congyue A.Peng,Jorge F.S.Ferreira,Andrew J.Wood.Direct analysis of artemisinin from Artemisia annua L.using high-performance liquid chromatography with evaporative light scattering detector,and gas chromatography with flame ionization detector.Journal of Chromatography A,2006,1133:254-258
[38]赵兵,王玉春,吴江,等.超声波用于强化石油醚提取青蒿素.化工冶金,2000,21(3):310-313
[39]王剑文,郑丽屏,谭仁祥.促进青蒿发根青蒿素合成的内生真菌诱导子的制备.生物工程学报,2006,22(5):829-834
[40]Filip C.W.,Sofie R.F.,Lies Maes,et al.Quantitation of artemisinin and its biosynthetic precursors in Artemisia annua L.by high performance liquid chromatography-electrospray quadrupole time-of-flight tandem mass spectrometry.Journal of Chromatography A,1118(2006) 180-187
[41]刘丽芳,王茜,李海燕,等.柱前衍生-RP-HPLC法测定青蒿中青蒿素的含量.中国野生植物资源,2004,23(6):60-62
[42]申薇薇,王锐利,胡爽,等.高效液相-柱后衍生法测定青蒿素含量.中国药物与临床,2007,7(9):657-659
[43]陈迪钊,刘坚.青蒿素的热稳定行为及气相色谱分析.怀化学院学报,2006,25(8):62-64
[44]郭燕,王俊,陈正堂.青蒿素类药物的药理作用新进展.中国临床药理学与治疗学,2006,11(6):615-620
[45]徐振海.青蒿素药物作用研究简介.安徽预防医学杂志,2004,10(3):181-182
[46]Pandey A V,Tekwani B L,Singh R L,Chauhan V S.Artemisinin,an endoperoxide antimalarial,disrupts the hemoglobin catabolism and heme- detoxification systems in malarial parasite.J Biol Chem,1999,274:19383-8
[47]Dorn A,Vippagunta S R,Matile H,et al.An assessment of drug-haematin binding as a mechanism for inhibition of haematin polymerisation by quinoline antimalarials.Biochem Pharmacol,1998,55:727-736
[48]Hoppe H C,Van Schalkwyk D A,Wiehart U I,et al.Antimalarial quinolines and artemisinin inhibit endocytosis in Plasmodium falciparum.Antimicrob Agents Chemother,2004,48:2370-8
[49]Aldieri E,Atragene D,Bergandi L,et al.Artemisinin inhibits inducible nitric- oxide synthase and nuclear factor NF-kB activation.FEBS Lett,2003,552:141-144
[50]Eckstein-Ludwig U,Webb R J,Van Goethem I D,et al.Artemisinin target the SERCA of Plasmodium falciparum.Nature,2003;424:957-964
[51]Ridley R G.To kill a parasite.Nature,2003;424:887-893
[52]梅林,石开云,苏建华,等.青蒿素国内研究进展.激光杂志,2008,29(2):95-96
[53]张剑锋,闻礼永.抗日本血吸虫药物作用机制的研究进展.国外医学·流行病学传染病学分册,2004,31(6):368-370
[54]崔金凤.治疗血吸虫病药物综述.安徽预防医学杂志,2005,11(3):172-173
[55]杨翠萍.青蒿素及其衍生物抗弓形虫的研究进展.国际医学寄生虫病杂志,2006,33(3):160-162
[56]邓定安.有抗肿瘤活性的青蒿素衍生物.药学学报,1992,27(4):317-320
[57]Efferth T,Dunstan H,Sauerbrey A,et al.The antimalarial artemisinin is also active against cancer.Int J Oncol,2001,18(4):767-773
[58]Singh N P,Lai H.Selective toxicity of dihydroartemisinin and holotransferrin toward human breast cancer cells.Life Sciences,2001,70:49-56
[59]温悦,孟德胜.青蒿素类药物药理作用研究进展.医药导报,2007,26(10):1193-1195
[60]易剑峰,王珏,林色奇,等.青蒿素及其衍生物免疫调节研究概况.江西中医学院学报,2006,18(2):69-71
[61]郑全芳,杨志刚,孙红祥.青蒿素及其衍生物药理作用研究进展.中国兽药杂志,2006,40(1):40-44
[62]贺小青,方鹏飞.青蒿素及其衍生物的药理作用.医药导报,2006,25(6):528-530
[63]胡世林,许有玲.纪念青蒿素30年.世界科学技术-中医药现代化,2005,7(2):1-2
[64]韦霄,李锋,许成琼,等.不同栽培措施对青蒿产量和青蒿素含量的影响.广西科学院学报,1999,15(3):132-136
[65]岑丽华,徐良,黄茱刚,等.不同纬区及不同栽培立地条件对青蒿青蒿素含量的影响.安徽农学通报,2007,13(13):46-47
[66]钟国跃,周华蓉,凌云,等.青蒿优质种质资源的研究.中草药,1998,29(4):264-266
[67]杨水平,杨宪,黄建国,等.青蒿素生产研究进展.热带亚热带植物学报,2004,12(2):189-194
[68]韦美丽,崔秀明,陈中坚,等.青蒿栽培研究进展.现代中药研究与实践,2005,19(5):60-64
[69]钟国跃,周华蓉,凌云,等.青蒿优质种植资源的研究.中草药,1998,29(4):264-267
[70]钟凤林,陈和荣,陈敏,等.青蒿最佳采收时期、采收部位和干燥方式的实验研究.中国中药杂志,1997,22(7):405-406
[71]齐向娟,李士雨,何玉娟.天津地区青蒿中青蒿素测定.中草药,2006,37(3):449-450
[72]陈强,张积强.青蒿素的提取、分离和测定.宝鸡文理学院学报(自然科学版),1999,19(1):47-48
[73]李吉和,张慧灵,李小玲,等.内蒙古地区青蒿中青蒿素的SFEHPLC测定.中药材,2000,23(12):756-758
[74]Woerdenbag H J,Pras N,Chan N G,et al.Artemisinin Related Sesquiterpenes and Essential oil in Artemisia annua During a Vegtation Period in Vietem.Plants Medics,1994, 60(3):272-275
[75]Chan Kit L.Selection of high artemisinin Melding Artemisia annua.Plants Medics,1995,61(3):285-287
[76]Charles D J,Simon J E,Wood K V,Heinstein P.Germplasm variation in artemisinin content of Artemisia annua using an alternative method of artemisinin analysis from crude plant extracts.J Nat Prod,1990,53:157-160
[77]Trigg P I.Qinghaosu(Artemisinin) as an antimalarial drug.Econ Med Plant Res,1990,3:20-55
[78]Klayman D L.Qinghaosu(artemisinin):an antimalarial drug from China.Science,1985,228:1049-1055
[79]耿飒,叶和春,李国凤,等.中药青蒿的生理生化特征及其研究进展.应用与环境生物学报,2002,8(1):90-97
[80]张萍,张子忠.山东引种青蒿青蒿素含量分析.山东中医药大学学报,2001,25(3):229-231
[81]韦霄,李锋,许成琼,等.青蒿生物学特性研究.广西植物,1997,17(2):166-168
[82]韦记青,韦霄,蒋运生,等.青蒿高产栽培技术.广西农业科学,2005,36(5):472-743
[83]李典鹏,梁小燕,陈秀珍,等.采用薄层层析-紫外分光光度法测定广西不同产地青蒿中青蒿素含量.广西植物,1995,15(3):254-255
[84]傅德明,蔡正江,毛禄国.青蒿生物学特性及规范化栽培技术.中国农技推广,2005,(12):34-35
[85]胡四化,胡家华,李让平.青蒿菊花瘿蚊的特征性与综合防治.广东农业科学,2006,(8):66
[86]陈海梅.青蒿栽培技术.特种经济动植物,2006,(3):26
[87]Singh A,Vishwakarma R A,Husssin A.Evaluation of Artemisia annua strains for higher artemisinin production.Planta Med,1988,54:475
[88]Ferreira J F S,Janick J.Production and detection of artemisinin form Artemisia annua.Acta Hort,1995,390:41-49
[89]Ferreira J F S,Charles D,Simon J E,Janick J.Effect of drying methods on the recovery and yield of artemisinin from Artemisia annua L..Hort Science,1992,27:650(Abstr.565)
[90]陈平.黄花资源及开发利用概述.广西林业科学,1998,27(1):35-36
[91]陈和荣.青蒿新品系“京厦Ⅰ号”的选育.中国技术成果大全,1992,1:186-187
[92]李锋,韦霄,许成琼,等.广西青蒿类型调查研究.广西植物,1997,7(3):231-234
[93]黄正方,郑贵华,李成东,等.影响青蒿成分(青蒿素)含量的因素研究.西南农业大学学报,1997,19(1):93-94
[94]Delabays N,Benakis A,Coll J.et al.Selection and breeding for high arteminin (qinghaosu)yielding strains of Artemisia annua.Acta Hort,1993,330:203-207
[95]Pras N,Visser J F,Batterman S,Woerdenbag H J,Malingre T M,Lugt C B.Laboratory selection of Artemisia annua L.for high artemisinin yielding types.Phytochem Anal,1991,2:80-83
[96]Geldre E U,Vergauwe A,Vanden Eeckout E.State of the art of the production of the antimalarial compound artemisinin in plants.Plant Mol Biol.1997,33:199-209
[97]Elhag H M,EL-Domiaty M M,El-Feraly F S,Mossa J S,El-Olemy M M.Selection and micropropagation of high artemisinin producing clones of Artemisia annua L..Phytother Res.1992,6:20-24
[98]Ferreira J F S,Simon J E,Janick J.Developmental studies of Artemisia annua flowering and artemisinin production under greenhouse and field conditions.Plants Med,1995,61:167-170
[99]Sangwan R S,Sangwan N S,Jain D C,et al.RAPD profile based genetic characterization of chemotypic variants of Artemisia annua L..Biochem Mol Biol Int,1999,47:935-944
[100]Wallaart T E,Pras N,Quax W J.Seasonal variation of artemisinin and its biosynthetic precursors in tetraploid Artemisia annua plants compared with the diploid wild-type[J].Planta Med,1999,65:723-728
[101]Magalhaes Redro M De.New hybrid lines of the antinalarial species Artemisia annua L.guarantee its growth in Brazil.Cienciae Culura(Sao Paulo),1997,49(5):413-415
[102]Enserink M.Infectious diseases:source of new hope against malaria is in short supply.Science,2005,307(5706):33
[103]Bouwmeester H J,Wallarrt T E,Janssen M H A,et al.Amorpha-4-11-diene synthase catalyses the first probable step in artemisinin biosynthesis.Phytochemistry,1999,52:843-854
[104]Picausd S,Mercke P,He X,et al.Amorpha-4,11-diene synthase:mechanism and stereochemistry of the enzymatic cyclization of farnesyl diphosphate.A rchies Biochem Biophy,2006,448:150-155
[105]Kraker J W,Franssen M C R,Dalm M C F,et al.Biosynthesis of germacrene carbosylic acid in chicory roots:demonstracion of a cytochrome P450(+)germacrene a hydroxyase and NADP+-denpendent sesquiterpenoid dehy-drogenase(s)involved in sesquiterpene lactone biosynthesis.Plant Physiol,2001,125(4):1930-1940
[106]Bertea C M,Freije J R,DerWoude H,et al.Identification of intermediates and enzymes involed in the early steps of artemisinin biosynthesis in Artemisia annua.Planta Med,2005,71:40-47
[107]Bogers R J,Craker L E,Langer D.Medicinal and Aromatic Plants.Amsterdam:Springer,2006:162
[108]Teoh K H,Polichuk D R,Reed D W,et al.Artemisia annua(Asteraceae)trichome- specific cDNAs reveal cyphavl,a cytochrome P450 with a key role in the biosynthesis of the antimalarial sequiterpene lactone artemisinin.FEB SLett,2006,580:1411-1416
[109]Ro D K,Paradise E M,Ouellet M,et al.Production of the antimalarial drug precursor artemisinic acid in engineered yeast.Nature,2006,440(13):940-943
[110]Abdin M Z,Israr M,Srivastava P S,et al.Invitroproduction of artemisinin.a novel antimalarial compound from Artemisia annua L..J Med Arom Plant Sci,2000,22(4A):378-384
[111]Dhingra V,Rajoli C,Narasu M L.Partial purification of proteins involved in the bioconversion of arteannuin B to artemisinin.Bioresour Technol,2000,73:279-282
[112]Dhingra V,Rajoli C,Narasu M L.Purification and characterization of enzyme involved in the biocon version of arteannuin B to artemisinin.Biochem Biophys Res Commun,2001,281:558-561
[113]Tatineni R,Doddapaneni K K,Dalavayi S,et al.Microbacterium trichotecenoly ticum enzymemediated transformation of arteannuin B to artemisinin.Process Biochem,2006,41(12):2464-2467
[114]Brown G D,Sy L K.In vivo transformations of dihydroartemisinin acid in Artemisia annua Plants.Tetrahedron,2004,60:1139-1159
[115]Wallaart T E,Van Uden W,Lubberink H G M,et al.Isolation and identification of dihydroartemisinic acid from Artemisia annua and its possible role in the biosynthesis of artemisinin.J Nat Prod,1999,62:430-433
[116]Wallarrt T E,Pras N,Quax W J.Isolation and identification of dihydroartemisinic acid hydroperoxide from Artemisia annua:a novel biosynthetic precursor of artemisinin.J Nat Prod,1999,62:1160-1162
[117]Sy L K,Brown G D.The mechanism of the spontaneous autoxidation of dihydroartemisinic acid.Tetrahedron,2002,58:879-908
[118]Sy L K,Brown G D.The role of the 12-carboxylic acid group in the spontaneous autoxidation of dihydroartemisinic acid.Tetrahedron,2002,58:909-923
[119]Brown G D,Sy L K.Invivo transformations of dihydroartemisinic acid in Artemisia annua plants.Tetrahedron,2004,60:1139-1159
[120]刘硕谦,田娜,李娟,等．青蒿素组合生物合成的研究进展．中草药,2007,38(9)：1425～1431
[121]Leyes A E,Baker J A,Hahn F M,et al.Biosynthesis of isoprenoids in Escherichia coli:stereochemistry of the reaction catalyzed by isopentenyl diphosphate:dimethylallyl diphosphate isomerase.Chem Commun,1999:717-728
[122]Sandmann G.Combinatorial biosynthesis of carotenoids in a heterologous host:a powerful approach for the biosynthesis of novel structures.Chem BioChem,2002,3:629- 635
[123]Huang Q L,Roessner C A,Croteau R,et al.Engineering E.coli for the synthesis of taxadiene,a key intermediate in the biosynthesis of taxo..Bioorgan Med Chem,2001,9:2237-2242
[124]Cunningham F X,Lafond T P,Gantt E,et al.Evidence of a role for lytb in the nonmevalonate pathway of isoprenoid biosynthesis.J Bacteriol,2000,182:5841-5848
[125]Martin V J J,Pitera D J,Withers S T,et al.Engineering a mevalonate pathway in Escherichia coli for production of terpenoids[J].Nature Biotechnol,2003,21(7):796-802
[126]Zahiri H S,Yoon S H,Keasling J D,et al.Coenzyme Q10 production in recombinant Escherichia coli strains engineered with a heterologous decaprenyl diphosphate synthase gene and foreign mevalonage pathway.Metabolic Eng,2006,8:406-416
[127]Martin V J J,Yoshikuni Y,Keasling J D.The invivo synthesis of plant sesquiterpenes by Escherichia coli.Biotechnol Bioeng,2001,75:497-503
[128]Pitera D J,Paddon C J,Newmanand J D,et al.Balancing a heterologous mevalonate pathway for improved isoprenoid production in Escherichia coli.Metabolic Eng,2007,9(2):193-207
[129]Lindahl A L,Olsson M E,Mercke P,et al.Production of the artemisinin precursor amorpha-4,11-diene by engineered Saccharomyces cerevisiae.Biotechnol Lett,2006,28:571-580
[130]Hampton R,Dimster-Denk D,Rine J.The biology of HM G-CoA reductase:the pros of contra-regulation.Trends Biochem Sci,1996,21:140-145
[131]Donald K,Hampton R,Fritz I.Effects of overproduction of the catalytic domain of 3-hydroxy -3-methylglutaryl coenzyme A reductase on squalene synthesis in Saccharomyces cerevisiae.Appl Environ Microbiol,1997,63:3341-3344
[132]Starari V J,Escalante-Semerana J C.Identification of the protein acetyltransferase(Pat)enzyme that acetylates acetyl-CoA synthetase in Salmonella enterica.J Mol Biol,2004,340:1005-1012
[133]Shiba Y,Paradise E M,Kirby J,et al.Engineering of the pyruvate dehydrogenase bypass in Saccharomyces cerevisiae for high-level production of isoprenoids.Metabolic Eng,2007,9(2):160-168
[134]朱卫平.野生青蒿的引种驯化和高青蒿素含量栽培品种选育目标性状的研究.2003,博士学位论文.15,37
[135]邓素兰,余继宏,毛丽梅.青蒿中青蒿素的提取分离研究.安徽农学通报.2007,13(5):31-34
[136]王轶.青蒿素最佳提取工艺研究.河南农业科学.2007,(7):84-86
[137]黄海滨,岑家铭,奉建芳,等.RP-HPLC测定青蒿中青蒿素的含量.广西大学学报 (自然科学版),1994,19(2):194-196
[138]Ferreira J F S,Simon J E,Janick J.Artemisia annua:botany,horticulture,pharmacology.Hort Re,1997,19:319-371
[139]朱卫平,盛孝邦.青蒿素的组织化学定位及其含量相关性研究.中草药,2003,34(9):852-854
[140]陈立亭,孙玉亭著.黑龙江省气候与农业.北京:气象出版社,2000:1-3
[141]郭家林,姬菊枝编著.哈尔滨市气候预测及其对生态环境的影响.北京:气象出版社,2000:1,2,4,10,12,14,16,104
[142]陈福泰,张桂花.药用青蒿植株中青蒿素合成的若干生理因子的研究.植物生理学通讯.1987,(5):26-30
[143]Ferreira J F S,Simon J E,Janick J.Developtmental studies of Artemisia annua L..Int.J.Plant Sci,1993,154(1):107-118
[144]李国栋,周全,赵长文,等.青蒿素类药物的研究现状.中国药学杂志,1998,33(7):385-389
[145]Ferreira J F S,Simon J E,Janick J.Artemisia annua:botany,horticulture,pharmacology.Hort Re,1997,19:319-371
[146]朱卫平,盛孝邦.青蒿素的组织化学定位及其含量相关性研究.中草药,2003,34(9):852-854
[147]江苏新医学院编.中药大辞典.下册.上海:上海科学技术出版社,1977,2052
[148]Laughlin J C.Effect of agronomic practices on Plant yield and antimalarial constituents of Artemisia annua L..Acta Hort,1993,33:53-61
[149]农业部种植业管理司,全国农业技术推广服务中心编著.测土配方施肥技术.北京:中国农业出版社,2006:3
[150]彭剑涛编.植物生长调节剂和除草剂使用技术.上海:上海科学普及出版社,2000:4-22
[151]刘春朝,王立春,叶和春.青蒿素生物合成研究进展.天然产物研究与开发,2000,12(1):83-86
[152]李弘剑,张毅.青蒿培养细胞中青蒿素生物合成代谢的体外调节.中国生物化学与分子生物学学报,1999,15(3):479-483
[153]Srivastava N K,Sharma S.Influence of micronutrient imbalance on growth and artemisinin content in Artemisia annua.Indian J Pharm Sci,1990,2:225-227
[154]萧浪涛,王少先,彭克勤,等.自然干旱胁迫下配方肥增效剂对水稻内源激素的影响.中国水稻科学,2005,19(5):417-421
[155]Liersch R,Soicke H,Stehr C,Tullner H U.Formation of artemisinin in Artemisia annua during one vegetation period.Planta Med,1986,52:387-390
[156]Shukla A,Farooqi-Abad A H,Shukla Y N,Sharma S.Effect of triacontanol and chlormequat on growth,plant hormones and artemisinin yield in Artemisia annua L..Plant Grow Regul.1992,11:165-171
[157]杨再强,王立新.观赏植物辐射诱变育种研究进展.四川林业科学,2006,27(3):19-23
[158]冯璐,那日.植物育种的物理学方法.草业科学,2005,22(12):63-66
[159]杨海梅,李明思,谢云.青蒿耗水及生长规律试验研究.石河子大学学报,2005,23(3):339-341
[160]王三根,梁颖.中药青蒿的生态生理及其综合利用中国野生植物资源,2003.8:47-49
[161]Sushil Kumar,S.K.Gupta,Poorinima Singh,et al.High yields of artemisinin by multiharvest of Artemisia annua crops.Industrial Crops and Products,2004,(19):77-90
[162]韦国锋,梁峰,覃特营.青蒿素不同提取工艺的研究.数理医药学杂志,2002,15(2):170-171
[163]郝金玉,韩伟,施超欧,等.青蒿中青蒿素的微波辅助提取.中国医药工业杂志,2002,33(8):385-387
[164]M.Irfan Qureshi,M.Israr,M.Z.Abdin,et al.Responses of Artemisia annua L.and saltinduced oxidative stress.Environmental and Experimental Botany 53(2005) 185-193
[165]雷红松.青蒿组培苗的移栽.特种经济动植物,2004,(2):31