不同施肥处理对曼地亚红豆杉‘Hicksii’生长和紫杉醇含量的影响

采用L9（3^4）正交实验设计,研究了温室盆栽和田间栽培条件下,不同氮、磷、钾施用水平对曼地亚红豆杉‘Hicksii’（Taxus media‘Hicksii’）生长及紫杉醇含量的影响。结果表明,不同的氮、磷、钾施用水平对曼地亚红豆杉‘Hicksii’盆栽苗和田间栽培苗的生长量、枝叶产量和紫杉醇含量有显著影响,影响效应从大至小依次为氮、磷、钾。根据生长综合指标确定最有利于曼地亚红豆杉幼苗生长的氮、磷、钾施用量分别为400、66和200mg·L^-1;最有利于紫杉醇积累的氮、磷、钾施用量分别为200、66和133mg·L^-1。此外,曼地亚红豆杉‘Hicksii’田间栽培苗的生长量、枝叶产量和紫杉醇含量显著高于盆栽苗。     

南方红豆杉枝叶中6种紫杉烷类化合物含量季节变化

紫杉醇(taxol)是主要来源于红豆杉属植物的一种天然抗肿瘤药物,紫杉烷(taxane)是紫杉醇的代谢前体或支路代谢产物,同样具有开发成为抗肿瘤新药的潜质。本文采用高效液相色谱法研究了紫杉醇(Taxol)、10-去乙酰巴卡亭Ⅲ(10-DAB)、7-木-10-去乙酰紫杉醇(7-xyl-10-DAT)、10-去乙酰紫杉醇(10-DAT)、三尖杉宁碱(CE)和7-表-10-去乙酰紫杉醇(7-epi-10-DAT)6种紫杉烷类化合物在南方红豆杉枝叶中含量的季节变化,结果显示Taxol和10-DAT在8、9月份含量最低,10-DAB和7-xyl-10-DAT在8、9月份含量相对最高,Taxol含量季节变化和10-DAB呈负相关,与CE呈显著正相关。7-xyl-10-DAT含量季节变化和10-DAB、10-DAT分别呈正相关。本文为研究南方红豆杉中紫杉醇及相关紫杉烷的代谢、积累规律提供了依据,不但有助于阐明紫杉醇的生物合成的关键步骤及调控的生理机制,而且对红豆杉资源的深度开发具有指导意义。     

南方红豆杉枝叶中药用抗癌活性物质含量

通过高效液相色谱（HPLC）,测定了人工栽培南方红豆杉枝叶中药用抗癌活性物质紫杉醇（taxol）、三尖杉宁碱（cephalomannine）和人工半合成紫杉醇的主要原料巴卡亭Ⅲ（baccatin Ⅲ）、10 去乙酰基巴卡亭Ⅲ（10-deacetylbaccatin Ⅲ,10-DAB）在一个生长季的含量变化.研究表明：从4月新枝叶萌发至11月枝叶基本停止生长,南方红豆杉枝叶中紫杉醇等药用活性物质含量季节性变化明显.其中紫杉醇和三尖杉宁碱含量的最高值都出现在5月;巴卡亭Ⅲ和10-去乙酰基巴卡亭Ⅲ含量的最高峰值分别出现在9月和4月.相关分析发现,南方红豆杉枝叶中4种药用活性物质之间有很好的相关性,其中紫杉醇与三尖杉宁碱含量呈显著正相关(P<0.05),三尖杉宁碱含量与10-去乙酰巴卡亭Ⅲ含量呈显著负相关(P<0.05).     

指数施肥对云曼红豆杉幼苗活性成分10-DAB及养分累积的影响北大核心CSCD

以1年生云曼红豆杉实生容器苗为材料,设置不施肥对照(CK)以及不同施肥量的平均施肥(AF1、AF2)、指数施肥(EF1、EF2、EF3和EF4)等7个处理,测定不同施肥方式和施肥量下苗木各时期活性成分10-DAB含量、生长量、枝叶生物量和10-DAB累积量,以及各部位N、P和K养分累积情况,探究施肥方式及施肥量对云曼红豆杉幼苗活性成分10-DAB及养分累积的影响,为云曼红豆杉苗期养分精准调控与高效培育提供理论和实践依据。结果表明:(1)施肥显著提高了幼苗枝叶中10-DAB含量,促进了生长及枝叶生物量累积(P<0.05);在相同施氮量下,指数施肥处理枝叶生物量显著大于平均施肥(P<0.05),枝叶中10-DAB含量以EF2、EF3处理较高,生长和枝叶生物量以EF2处理最大。(2)EF2处理的枝叶10-DAB累积量显著大于其他处理(P<0.05),并表现为:EF2>EF1>AF2>EF3>AF1>EF4>CK,EF2处理分别较AF1和AF2处理提高30.61%~41.94%和18.14%~25.00%。(3)施肥对幼苗各部位N、P和K含量的影响显著(P<0.05);在相同施氮量下,指数施肥各部位的N、P和K含量均优于平均施肥;指数施肥处理中,根系中的N含量以及各器官K含量随施肥量增加而上升,各部位P含量以及茎和叶中N含量随施肥量增加先上升后下降;各处理N、P和K分配比例呈现出叶>根>茎规律。幼苗各器官中N与P、P与K以及根、叶中的N与K之间均表现出显著(P<0.05)正相关,呈现出协同关系。(4)云曼红豆杉幼苗的10-DAB累积量在9~10月份最高,此时采收可获得的10-DAB累积量最高,收益也最大。研究发现,指数施肥与平均施肥相比有效提高了云曼红豆杉苗木活性成分10-DAB及养分累积,并以施肥量为1600 mg·株^(-1)的苗木各指标表现较优。     

叶面施肥对南方红豆杉针叶中紫杉醇和10-DAB含量的影响

人工栽培红豆杉已成为目前解决紫杉醇市场需求的主要途径,如何提高人工栽培的红豆杉中紫杉醇及其化学半合成前体10-脱酰基巴卡丁Ⅲ(10-DAB)的含量是一个十分关键的问题.对人工栽培南方红豆杉叶面喷施不同浓度的N、K、Ca、Mg 肥和自行配置的稀土混合肥(含La、Ce、Sm),连续4次跟踪采样,超高效液相色谱(UPLC)测定样品中紫杉醇和10-DAB的含量.结果表明,叶面喷施不同浓度的N元素均引起红豆杉针叶中紫杉醇和10-DAB含量的降低;喷施K元素,前期引起10-DAB含量的增加但随之降低,紫杉醇的含量则持续降低;喷施不同浓度的Ca、Mg提高了红豆杉中紫杉醇和10-DAB的含量,其作用可持续一段时间;自行配置的稀土混合肥明显地提高了紫杉醇和10-DAB的含量且其持续作用时间较长,紫杉醇和10-DAB的含量分别最高可达0.261mg g-1和0.641mg g-1.     

不同地理种源南方红豆杉中紫杉醇和10-DAB含量及影响因子

选择福建省梁野山、龙栖山、闽侯旗山、南靖书洋乡和南平大坪村南方红豆杉（Taxus chinensis var. marei）天然群体分布相对集中区,采集胸径相同的南方红豆杉的当年生针叶和树皮并取树下土壤样品。利用超高效液相色谱分析测定南方红豆杉当年生针叶及树皮中紫杉醇及10-去乙酰基巴卡亭Ⅲ（10-deacetylbaccatin Ⅲ, 10-DAB）含量,另测定土壤pH值、速效磷、速效氮、铁、钙、镁、锌含量。结果表明,不论是紫杉醇还是10-DAB含量,不同地理种源南方红豆杉间差异均较大,说明地理位置对于红豆杉中紫杉醇和10-DAB含量具有明显的影响;树皮中紫杉醇含量与土壤中铁含量显著相关（P＜0.01）;南方红豆杉当年生针叶中及树皮中紫杉醇含量与各种源地气象因子的相关性不显著（P＞0.05）。     

红豆杉低温半致死温度和低温胁迫下紫杉烷含量

通过测定低温胁迫下红豆杉电阻抗参数和紫杉烷含量变化,确定适合红豆杉低温半致死温度测定的电阻抗参数,探讨低温对红豆杉紫杉烷生物合成的影响,为扩大红豆杉引种范围及利用低温条件提高红豆杉中紫杉醇含量奠定了理论基础。以云南红豆杉、南方红豆杉(山西长治)、中国红豆杉、曼地亚红豆杉和东北红豆杉为材料,分别采用电导率法和电阻抗法测定上述树种低温半致死温度,并利用LC-MS法测定低温胁迫下红豆杉叶片中紫杉烷含量和脱落酸(abscisic acid, ABA)含量。结果表明,5种红豆杉中,中国红豆杉、南方红豆杉(山西长治)、东北红豆杉和曼地亚红豆杉均能耐受-14.668--9.106℃低温;云南红豆杉能耐受-8.802--3.521℃低温,高于其他4种红豆杉。电阻抗τ_m参数与电导法测定的5种红豆杉一年生枝低温半致死温度显著相关,相关系数为0.912。低温胁迫下,红豆杉一年生叶片中紫杉醇含量高于当年生叶片。随着处理温度的下降,云南红豆杉和东北红豆杉一年生叶片中紫杉醇含量显著增加,中国红豆杉当年生叶片中巴卡亭Ⅲ含量和10-去乙酰基紫杉醇含量显著增加。此外,红豆杉叶片中ABA含量随温度的...     

赤霉素和烯效唑对东北红豆杉紫杉醇代谢的影响

以一年生东北红豆杉扦插苗为材料,采用Hoagland营养液添加赤霉素及其合成抑制剂烯效唑的方法,研究了不同处理对东北红豆杉紫杉醇(taxol)及其前体巴卡亭Ⅲ(baccatinⅢ)与10-去乙酰基巴卡亭Ⅲ(10-DAB)含量变化的影响,同时比较了苯丙氨酸解氨酶(PAL)、过氧化物酶(POD)和超氧化物酶(SOD)活性变化,分析了紫杉醇合成旁路途径物质及抑制剂对紫杉醇代谢的作用效应.结果表明,不同浓度赤霉素(GA3)和烯效唑(S3307)处理对东北红豆杉POD、SOD及PAL活性的影响不显著;但整个处理过程中叶片中紫杉醇含量均为对照的1.28～6.44倍,茎中紫杉醇、baccatinⅢ及叶片中10-DAB含量在第6天均高于相应对照;赤霉素与烯效唑对紫杉醇及其前体合成的作用途径存在一致性.     

不同地理种源南方红豆杉中紫杉醇和10-DAB含量及影响因子

选择福建省梁野山、龙栖山、闽侯旗山、南靖书洋乡和南平大坪村南方红豆杉(Taxus chinensis var.marei)天然群体分布相对集中区,采集胸径相同的南方红豆杉的当年生针叶和树皮并取树下土壤样品.利用超高效液相色谱分析测定南方红豆杉当年生针叶及树皮中紫杉醇及10-去乙酰基巴卡亭Ⅲ(10-deacetylbaccatin Ⅲ,10-DAB)含量,另测定土壤pH值、速效磷、速效氮、铁、钙、镁、锌含量.结果表明,不论是紫杉醇还是10-DAB含量,不同地理种源南方红豆杉间差异均较大,说明地理位置对于红豆杉中紫杉醇和10-DAB含量具有明显的影响;树皮中紫杉醇含量与土壤中铁含量显著相关(P<0.01);南方红豆杉当年生针叶中及树皮中紫杉醇含量与各种源地气象因子的相关性不显著(P>0.05).     

胸径、构件和季节对南方红豆杉中紫杉醇和10-DAB含量的影响

系统采集了12个不同胸径的南方红豆杉当年生针叶和树皮样品,用超高效液相色谱(UPLC)测定其紫杉醇(taxol)和10-去乙酰基巴卡亭Ⅲ(10-deacetylbaccatin Ⅲ,10-DAB)含量,结果显示针叶的两种被测试物质含量的平均值分别为0.0127mg·g-1和0.0805mg·g-1,树皮的这两种物质含量平均值分别为0.1164mg·g-1和0.4842mg·g-1。当年生针叶、树皮中紫杉醇含量与胸径相关性不显著。对植株不同构件(枝条、针叶、树皮和根)中紫杉醇和10-DAB含量的测定表明,根中紫杉醇和10-DAB含量最高,当年生针叶紫杉醇含量最低。天然和人工栽培南方红豆杉当年生针叶中紫杉醇和10-DAB含量呈明显的季节变化,各月份间含量差异显著,人工栽培和天然植株针叶在生长速度明显减缓的10月份,紫杉醇和10-DAB含量均出现一个明显的峰值。     

悬浮培养南方红豆杉细胞的紫杉醇生物合成路径中诱导子的作用位点

作者研究开发出一种基于分析中间响应模式确定悬浮培养诱导子作用位点的新方法.研究结果表明,一个诱导子的作用位点存在于浓度变化方向相反的相邻两个中间代谢物之间;该方法的有效性在悬浮培养南方红豆杉(Taxus chinensis(Pilg.)Rehd.var.mairei(Lemee et Levl.)Cheng et L. K.Fu)生物合成紫杉醇过程中得以证实;经确定,甲基茉莉酮酸、硝酸根和柠檬酸铵的作用位点在baccatinⅢ至10-去乙酰基紫杉醇之间;水杉酸、花生四烯酸的作用位点存在3种可能性,即增强10-去乙酰基紫杉醇的合成、防止紫杉醇和cephalomannine的降解.该方法对指导诱导子配伍优化紫杉醇生产具有指导意义.     

不同施肥处理对木棉叶片光合特性和幼苗生长的影响

采用L25(53)正交实验设计设置不同氮、磷和钾肥施用量及配比﹝单株施用量分别为CO(NH2)22.0、4.0、6.0、8.0和10.0 g,Ca(H2PO4)24.0、6.0、8.0、10.0和12.0 g,KCl 0.6、1.2、1.8、2.4和3.0 g﹞,并设置不同复合肥施用量(单株施用量分别为10、20、30、40和50 g),比较了施肥后第1至第3个月木棉( Bombax ceiba Linn.)幼苗叶片光合指标﹝包括净光合速率( Pn)、水分利用效率( WUE)、PSⅡ最大光化学效率( Fv/Fm )和叶绿素相对含量( SPAD)﹞和幼苗生长指标(包括株高增长量、地径增长量和叶面积增长量)的变化。结果表明：总体来看,不同施肥处理组木棉幼苗叶片的Pn和WUE值升高,幼苗的株高增长量、地径增长量和叶面积增长量增加,但不同施肥处理对叶片Fv/Fm和SPAD值的影响较小;复合肥对叶片光合生理特性和幼苗生长的影响也较小。在施肥后的第1至第3个月,单株施用量氮肥4.0或6.0 g,磷肥4.0或8.0 g,钾肥1.2、1.8或3.0 g处理组幼苗叶片的Pn和WUE值显著高于对照(不施肥)和大多数处理组;单株施用量氮肥4.0或6.0 g、磷肥4.0~12.0 g、钾肥1.2~3.0 g处理组幼苗的株高增长量、地径增长量和叶面积增长量也均较高。综合分析结果显示：氮肥对木棉幼苗光合生理特性及生长的影响最大,钾肥次之,磷肥最小。综合考虑Pn值、WUE值、株高增长量、地径增长量和叶面积增长量,木棉苗期的适宜单株施肥量为N 1.84或2.76 g、P2 O50.72~2.16 g和K2 O 0.72~1.80 g。     

Seasonal changes in the concentrations of four taxoids in Taxus baccata L. during the autumn-spring period.

The concentrations of four common taxoids: baccatin III, paclitaxel, cephalomannine and 10-deacetylbaccatin III (10-DAB III) were measured in fresh needles and stems of Taxus baccata L. during the late autumn-spring period (November'96-April'97) which has not been investigated to date in this species. Baccatin III, paclitaxel and 10-DAB III were present on the surface of the twigs in concentrations of 8-26 micrograms/1000 g (fresh weight). Changes in the levels of baccatin III and paclitaxel inside the needles and stems showed similar trends over the investigated period. From November to March the total level of taxoids differed between the needles and stems, and were the same only in April. Total levels in fresh needles were stable from December to March. The highest concentrations of 10-DAB III in the whole analysed period in fresh stems were measured, as well as in the fresh needles except for samples collected in November and December when the levels of cephalomannine were higher. The concentrations of paclitaxel were usually the lowest. These results confirm that epigenetic factors--date of collection (and thus phyllogenesis) and kind of plant tissue--determine taxoid levels during the late autumn-spring period in T. baccata. The opposite patterns of changes for 10-DAB III and cephalomannine, especially in the fresh needles, suggest a possible role for 10-DAB III in the biosynthetic pathway to cephalomannine, a less polar taxoid with a side-chain at position C-13. As well, owing to the thermolability of taxoids, the influence of low temperatures in December and January could explain the highest observed concentrations of 10-DAB III in the fresh stems and needles, respectively.     

Two New Taxane Diterpenoids from Taxus yunnanensis

Five compounds were isolated from the bark of Taxus yunnanensis Cheng et L. K. Fu. Based on spectral IR, MS, 1H-NMR, 13C-NMR and physico-chemical constant analysis, they were identified as 7-epi-taxol B ( Ⅰ ), 1, 13-diacetoxy-10-deacetyl baccatin Ⅲ (Ⅱ ), taxol C-7-xylose (Ⅲ), taxol (Ⅳ), 3-epi-ursolic acid (Ⅴ). Compounds Ⅰ and Ⅱ were newly identified.     

Effect of the culture medium and biotic stimulation on taxane production in Taxus globosa Schltdl in vitro cultures

Taxus globosa is the only species of the Taxus genus that grows in Mexico. In this study, callus cultures from leaves and young shoots of T. globosa were established in Gamborg’s B5 medium supplemented with 2,4-dichlorophenoxiacetic acid (2 mg/L), kinetin (0.5 mg/L) and gibberellic acid (0.25 mg/L). Callus growth and taxane production were evaluated using two culture media: Woody Plant Medium and Gamborg’s B5 supplemented with picloram (2 mg/L), kinetin (0.1 mg/L) and gibberellic acid (0.5 mg/L). The effect of the inoculum size (50, 100 and 150 g FW/L) and culture media (Woody Plant Medium and Gamborg’s B5) with and without the presence of methyl jasmonate (100 μM) on T. globosa cell suspensions was assessed. Taxane analysis revealed that the calli in Gamborg’s B5 produced taxol (50 μg/g DW), baccatin III, 10-deacetyl baccatin III and 10-deacetyl taxol. Woody Plant Medium also induced the production of taxol, although to a lesser extent. The optimum inoculum size was 50 g FW/L. In cell suspension cultures, both media had a significant effect on taxane production when supplemented with methyl jasmonate. In Woody Plant Medium, at day 14, a total concentration of 197.999 μg/L of taxol, 160.622 μg/L of baccatin III, 633.724 μg/L of 10-deacetyl baccatin III and 229.611 μg/L 10-deacetyl taxol were obtained, with total excretion of baccatin III and 10-deacetyl taxol to the culture medium. In Gamborg’s B5, cephalomanine was obtained at a concentration of 91.428 μg/L without elicitation, and all taxanes were excreted to the medium to a variable extent.     

一株能水解巴卡亭ⅢC-10位乙酰基的成团泛菌的筛选和鉴定

10-脱乙酰巴卡亭Ⅲ是合成紫杉醇和多烯紫杉醇的前体。以巴卡亭Ⅲ为底物,结合TLC、HPLC、HPLC-MS分析方法,通过设计专门的筛选方法筛选产酶菌株,得到一株巴卡亭ⅢC-10位脱乙酰基酶产生菌株Z1-56。以形态特征、生理生化特征、16S rDNA序列分析作为菌株的鉴定手段,Z1-56被鉴定为成团泛菌(Pantoea agglomerans),首次发现成团泛菌产生巴卡亭ⅢC-10-脱乙酰酶。     

High-performance liquid chromatography and micellar electrokinetic chromatography of taxol and related taxanes from bark and needle extracts of Taxus species

High-performance liquid chromatography (HPLC) and micellar electrokinetic chromatography (MEKC) were applied for the separation of taxol, cephalomannine, and baccatin III in crude extracts from the needle and bark of Taxus species. The chromatogram of the bark extract was cleaner than that of the needle allowing a more reliable detection of taxol and cephalomannine in the bark extract. However, HPLC quantitation of taxol in the needle extract would be difficult due to coeluting taxinines. Nevertheless, this was not a problem in the MEKC experiment. In comparison to HPLC, MEKC offered baseline resolution of taxol from taxinines in the needle extract, less solvent waste, a smaller sample requirement, and the simultaneous detection of taxol, cephalomannine and baccatin III in a relatively simpler electrophoretic run.