滤光膜对黄檗(Phellodendron amuranse)幼苗三种生物碱含量的影响

以滤光膜遮光处理1年生黄檗(Phellodendron amuranse)幼苗,测定了幼苗生物量及根、茎外皮中的小檗碱、药根碱和掌叶防己碱含量.红色、黄色、蓝色和绿色滤光膜对黄檗幼苗的根和茎生物量都有不同程度的抑制作用,其中黄膜的抑制作用最小,而蓝膜和绿膜的抑制作用最强.滤光膜处理也不同程度地抑制了小檗碱、药根碱和掌叶防己碱的合成和积累,红膜的抑制作用最小,蓝膜和绿膜的抑制作用最强.滤光膜处理后,3种生物碱的单株产量都低于对照,红膜处理小檗碱的产量显著高于黄膜处理,但红膜和黄膜处理下的药根碱和掌叶防己碱的产量差异不显著,蓝膜和绿膜处理的3种生物碱的产量都始终最低.滤光膜处理不利于黄檗幼苗的生长和上述3种生物碱的积累.     

氮素形态对黄檗幼苗三种生物碱含量的影响

通过改变水培溶液中NH 4-N和NO3--N的比例,探讨了氮素形态对黄檗幼苗主要药用成分小檗碱、药根碱及掌叶防己碱含量的影响。结果表明,小檗碱的含量与氮素形态有明显关系,在根和茎中都表现为NO3--N比例高更有利于小檗碱的积累。单纯的NO3--N供给最有利于药根碱在黄檗幼苗根中的积累,而处理46d后,NH4 -N/NO3--N为75/25和25/75时黄檗幼苗茎中的药根碱含量最高。黄檗幼苗根和茎中的掌叶防己碱含量对营养液中NH4 -N和NO3--N的比例反应类似,在处理46d后,掌叶防己碱含量由高到低所对应的NH4 -N/NO-3-N为25/75、50/50、75/25、0/100、100/0。处理33d后,黄檗幼苗根中的小檗碱、药根碱、掌叶防己碱含量随处理天数的增加呈下降趋势,而在茎中呈上升趋势。如果同时考虑氮素形态对黄檗幼苗生物量的影响,那么NH4 -N/NO3--N为0/100时其小檗碱产量(含量与生物量乘积)最高,而NH4 -N/NO-3-N为25/75时药根碱和掌叶防已碱的产量最高。     

水分胁迫对黄檗幼苗三种生物碱含量的影响

黄檗（Phellodendron amurense）是东北阔叶红松林的重要伴生树种,也是我国名贵中药关黄柏的药源植物,小檗碱（berberine）、药根碱（jatrorrhizine）、掌叶防己碱（palmatine）是研究者通常关注的主要黄檗生物碱。通过池栽法和渗灌控水方式设置了对照、轻度干旱、重度干旱和水涝（土壤水势分别控制在-40～-20KPa、-80～-60KPa、〈-80KPa和-20～0KPa）4种水分处理。比较了不同水分状况下黄檗幼苗小檗碱、药根碱及掌叶防己碱含量的变化。对于当年生的黄檗幼苗,总体上轻度干旱有利于上述3种生物碱的合成与积累,重度干旱处理下幼苗3种生物碱的含量与对照差异不大。而水涝处理则导致幼苗3种生物碱的含量显著降低。同时,干旱或水涝处理都导致黄檗幼苗生长受抑,幼苗的株高、茎径和生物量都显著低于对照幼苗,因此3种生物碱的单株产量仍是对照的黄檗幼苗最高。茎外皮是中药黄柏的药用部位,也是黄檗幼苗中3种生物碱含号最丰的部位。实验结果预示短期的轻度干旱处理可以提高茎外皮的小檗碱含量。这对以获取生物碱为目的的黄檗幼苗培育可能有参考价值。     

不同植被地区天然黄檗生物碱含量对季节的响应

黄檗(Phellodendron amurense Rupr.)不仅为中国特有的濒危植物,还是我国著名中药材"关黄柏"的药源植物,由于其珍贵的药用价值而被过度开采。本研究采用HPLC检测不同植被地区天然黄檗种群中药根碱、掌叶防己碱和小檗碱等生物碱的含量,分析不同部位,不同季节对生物碱分布的影响。研究表明,药根碱在黄檗各部位的分布为:根皮茎皮多年生枝一年生枝叶片。掌叶防己碱含量分布为:茎皮多年生枝皮根皮一年生枝叶片。小檗碱在不同部位的分布规律表现为:根皮茎皮多年生枝一年生枝叶片。根皮和茎皮中的药根碱含量随着春、夏、秋季节的推移呈逐渐上升的趋势,表现为秋季夏季春季,而多年生枝皮、一年生枝、叶片中的药根碱则是秋季春季夏季。各部位的掌叶防己碱均随着季节推移而升高,表现为秋季夏季春季。根皮、茎皮和多年生枝皮中小檗碱含量随着季节变化表现为夏季秋季春季。一年生枝中小檗碱在夏季降低,而到了秋季再升高。     

季节和地理差异对天然黄檗小檗碱和药根碱含量的影响

用高效液相色谱法测定了黄檗根皮、茎皮、多年生枝皮、1年生枝和叶片中小檗碱和药根碱的含量,研究黄檗不同部位两种生物碱的季节和地理差异。结果表明:不同季节,天然黄檗各部位的小檗碱、药根碱含量均表现为:根皮茎皮多年生枝皮1年生枝叶片。小檗碱含量随着季节变化在各部位呈现夏季升高,到秋季下降的趋势。而药根碱在不同部位中的含量随着季节变化的规律不同。根皮、茎皮中的药根碱含量随着季节推移逐渐增多,在夏季升高不明显,而到秋季大量积累。多年生枝皮、1年生枝、叶片中的药根碱在3个季节表现则为先降后升。从纬度水平上看,不同地域间黄檗各部位的生物碱差异显著,在调查的40°—50°N范围内,黄檗两种生物碱的含量普遍在纬度低的地区较高,随着纬度增大先逐渐降低,到N44°左右达到最低值,到了高纬度地区略有回升。     

安徽小檗生物碱成分的研究

安徽小檗(Berberis anhweiensis Ahrendt)的酸水提取部分进行硅胶柱色谱分离纯化,并通过波谱学方法,鉴定出6个原小檗碱型生物碱,分别为小檗碱(1)、巴马汀(2)、dehydroeapaurinine(3)、药根碱(4)、非洲防己碱(5)和jatrorubine(6).这6种生物碱均为首次从安徽小檗中分得.其中巴马汀的含量比大多数小檗科其他植物高.     

产小檗碱诱变菌株培养条件的优化

S-NU-3-2菌株是对源自药用植物黄檗的内生真菌S6进行诱变获得的小檗碱产量比出发菌株有明显提高的突变株,本研究对其培养条件进行了优化,以期进一步提高其产量,为开发利用真菌发酵生产植物活性成分的新途径奠定基础。以菌丝生长和小檗碱产量为指标,筛选出了适宜该菌株发酵的基本培养基、碳源、氮源、光照和培养温度,并经进一步的正交试验优化,得出该高产菌株的最佳培养条件为豆芽汁基本培养基含蔗糖3%,酵母膏0.2%,pH7.0,温度26℃,全光照培养。与初始培养条件相比,在优化后的条件下,该菌株的小檗碱得率提高了47.2%,菌体生物量提高了24%。     

不同海拔云南黄连生物量和主要有效成分变化

研究了不同海拔(2 100～2 700 m)下,野生和人工栽培云南黄连的生物量、主要有效成分含量及产量.结果表明:野生云南黄连根茎和根生物量沿海拔梯度呈上升趋势,但无显著性差异(P>0.05);人工栽培云南黄连根茎生物量平均值在海拔2 600 m和2 700 m处分别为87.5 kg·hm^-2和97.0 kg·hm^-2,显著高于海拔2 300 m处(34.8 kg·hm^-2,P<0.05),且海拔2 300、2 600和2 700 m的人工栽培云南黄连根茎和根生物量均大于野生云南黄连,但无显著性差异(P>0.05). 野生云南黄连的根茎和根生物量均与全株生物量呈显著正相关. 野生云南黄连根茎和根小檗碱含量在海拔2 700 m处最高,分别为4.60%和1.93%; 根茎巴马汀和药根碱含量、根药根碱含量在海拔2 600～2 700 m处最高;根巴马汀含量在2 300 m处最高.人工云南黄连根茎和根小檗碱含量在海拔2 600 m处最高,分别为4.41%和1.90%; 根茎巴马汀含量,根小檗碱、巴马汀和药根碱含量在海拔2 600～2 700 m处最高;根茎药根碱含量在海拔2 300 m处最高.海拔2 600～2 700 m处野生云南黄连根茎和根中各有效成分产量显著高于海拔2 100和2 300 m处(P<0.05). 野生云南黄连分株的根茎生物量、根生物量、叶生物量、总生物量、高度和冠幅沿海拔梯度呈先升后降趋势.增大种植密度和加强人工管理可以提高云南黄连生物量和主要有效成分产量.     

明党参幼苗存活与生长对光照强度的响应

以濒危植物明党参低温催芽形成的幼苗和常温正常萌发的幼苗为对象,对其存活及形态、结构、生物量及其分配、生长速率等指标进行测定和分析.结果表明,明党参幼苗在较高光强（65％全光照）存活率最高,有利于幼苗建成,低光强存活率最低.低温催芽形成的幼苗和常温正常萌发的幼苗对光照强度的响应规律类似,明党参幼苗的生长参数（株高除外）叶生物量、根生物量、总生物量、生长速率和总叶面积均在65％全光照条件下达到最大值,表现出最佳生长状况,表明适宜明党参幼苗生长的光照条件是65％全光照;利用低温催芽形成的幼苗生物量约是常温正常萌发幼苗的5倍,能提高其经受不利环境的能力,为明党参的栽培和保护提供一种新的途径.     

朱砂根幼苗在不同光照强度下的形态和生理响应

通过人工遮荫模拟不同生境光强(100%、52%、33%、15%和6%的相对光照)的方法,对朱砂根(Ardisia crenata Sims)幼苗形态、生物量和生理指标的变化特征进行了分析。研究结果显示,朱砂根单株生物量在52%相对光照处理下显著高于其他光照处理,在15%～52%相对光照处理下分配给叶片的生物量比例高于全光照(100%)和极弱(6%)光照处理,但根冠比不受光强影响,其结构可塑性相对较低。硝酸盐含量随遮荫强度的增加而增大,且在6%相对光照处理下显著增加,硝酸盐还原酶的活性同硝酸盐含量变化规律一致,表明朱砂根主要通过改变叶面积和比叶面积等形态指标,以及调整生物量的分配和光合色素含量来适应不同的光环境。叶绿体超微结构分析结果显示,在15%～52%相对光照处理下,叶绿体数量较多且细胞结构较完整,而100%和6%光照处理下的叶绿体数量明显减少,且细胞结构严重受损发生质壁分离现象。因此,朱砂根适宜生长的光照条件为15%～52%,尤其是33%～52%的相对光照条件更佳。     

Effects of water stress on berberine, jatrorrhizine and palmatine contents in amur corktree seedlings

Amur corktree (Phellodendron amurense) is one of the important associated species in broadleaved-Korean pine forests, and in the pharmaceutical resource plants of famous Chinese traditional medicine, named cortex phellodendri. Berberine, jatrorrhizine and palmatine are the main alkaloids to which high attention is paid by the researchers. In the present study, water stress treatments with four conditions, that is, mild drought, severe drought, waterlogging, and control (soil water potentials were controlled in the ranges of-40—-20 KPa,-80—-60 KPa, <-80 KPa and-20—0 KPa), were performed using the technique of root-sphere osmotic irrigation in a soil pond. The changes in the main medicinal compositions of berberine, jatrorrhizine and palmatine contents under different water conditions were discussed. As for the annual growing of amur corktree seedlings, mild drought was generally beneficial to the synthesis and accumulation of the three above-mentioned alkaloid contents. The three alkaloid contents did not show great changes under severe drought whereas those contents had significantly reduced under waterlogging compared with controls. Meanwhile, the growth of amur corktree seedlings was inhibited by the treatments of drought and waterlogging. The height, diameter and biomass of amur corktree seedlings were significantly lower than those of the control seedlings, which meant that the three alkaloid contents in a single seedling still kept the highest in control seedlings. Stem cortex was the medical part in Chinese traditional medicine and also kept the most abundant of the three alkaloid contents in amur corktree. Results indicated that short periods of mild drought could improve the berberine contents in the stem cortex, which might have reference value for the cultivation of amur corktree seedlings to obtain alkaloids.     

Cultivation of cell cultures of Berberis wilsonae in 20-l airlift bioreactors

Suspension cultures of Berberis wilsonae produce 4 berberine-type alkaloids: berberine, palmatine, columbamine and jatrorrhizine. In particular the formation of the phenolic alkaloids columbamine and jatrorrhizine and of berberine proves to be dependent on the concentration of dissolved oxygen. With higher aeration rates, berberine and jatrorrhizine yields can be increased considerably. Thus we reached an alkaloid yield of more than 3 g × 1^–1 with 50% dissolved oxygen tension in the medium. As far as we know this is one of the best results in fermenting of alkaloid-producing cell cultures.Abbreviations pO₂ dissolved oxygen concentration in % saturation (using air) - HPLC high-performance liquid chromatography - vvm volume air × volume medium^–1 × minute^–1 - rpm revolutions per minute - IAA indole-3-acetic acid - 2,4-D 2,4-dichlorophenoxy acetic acid     

我国黄檗野生种群生存现状及化学表征研究

黄檗(Phellodendron amurense Rupr.)为我国常用中药关黄柏的基原,也是国家二级保护植物,但在20世纪60-90年代其野生资源遭到严重破坏,并且至今种群规模未得到很好的恢复。本文对我国燕山、千山、长白山、张广才岭、小兴安岭5条山脉黄檗野生种群的种群密度、年龄结构(径级结构)、生物碱类成分含量等进行了调查和研究,结果显示:黄檗野生种群年龄结构呈纺锤形,幼龄级、青壮级个体分别占总个体数的13.9%和68.0%,表明幼龄植株储备匮乏,天然更新不足,总体呈严重的衰退趋势,受威胁状况严峻;千山种群的小檗碱、巴马汀、黄柏碱含量及生物碱总量最高,而小兴安岭种群最低,且二者与其他三条山脉黄檗种群的生物碱类成分含量差异显著。本研究结果表明,野生黄檗种群分布区缩小,年龄结构呈衰退趋势,部分种群个体数量极少,即使在保护状态下也难以自我恢复;千山黄檗种群的生物碱类成分含量较高,在开展药材生产时应予以重视。     

Study on noncovalent complexes of cytotoxic protoberberine alkaloids with double-stranded DNA by using electrospray ionization mass spectrometry

The noncovalent complexes of four cytotoxic protoberberine alkaloids that is, berberine, palmatine, jatrorrhizine, and coptisine with double-stranded oligodeoxynucleotides d(AAGAATTCTT)(2) were investigated by electrospray ionization mass spectrometry. These four active components from Chinese herbal medicines showed both 1:1 and 1:2 binding stoichiometries, independent on the alkaloid-to-DNA ratios. Binding affinities in the order of palmatine> or =jatrorrhizine>coptisine>berberine with d(AAGAATTCTT)(2) were obtained. Additionally, the preliminary results indicated that berberine had some sequence selectivities.     

DNA-binding cytotoxic alkaloids: comparative study of the energetics of binding of berberine, palmatine, and coralyne

Deoxyribonucleic acid is the site of storage and retrieval of genetic information through interaction with proteins and other small molecules. In the present study, the interaction of two natural cytotoxic protoberberine plant alkaloids, berberine and palmatine, and a synthetic derivative, coralyne, with mammalian herring testis DNA was investigated using a combination of isothermal titration calorimetry, differential scanning calorimetry, and optical melting experiments to characterize the energetics of their binding. The binding constants of these alkaloids to DNA under identical conditions were evaluated from the UV melting data, and the enthalpy of binding was elucidated from isothermal titration studies. The binding constants of berberine, palmatine, and coralyne to DNA were found to be 1.15 x 10(4), 2.84 x 10(4), and 3.5 x 10(6) M(-1) at 20 degrees C in buffer of 20 mM [Na+]. Parsing of the free energy change of the interaction observed into polyelectrolytic and nonpolyelectrolytic components suggested that although these alkaloids are charged, the major contributor of about 75% of the binding free energy arises from the nonpolyelectrolytic forces. The binding in case of palmatine and coralyne was predominantly enthalpy driven with favoring smaller entropy terms, while that of berberine was favored by both negative enthalpy and positive entropy changes. Temperature dependence of the binding enthalpies determined from ITC studies in the range 20-40 degrees C was used to calculate the binding-induced change in heat capacity (DeltaC(o)(p)) values as -117, -135, and -157 cal/mol K, respectively, for berberine, palmatine, and coralyne. Taken together, the results suggest that the DNA binding of the planar synthetic coralyne is stronger and thermodynamically more favored compared to the buckled natural berberine and palmatine.     

Determination of protoberberine alkaloids in medicinal plants based on acidic potassium permanganate chemiluminescence system

A simple method was established to determine protoberberine alkaloids in Cortex Phellodendri and Rhizoma Coptidis based on an acidic potassium permanganate chemiluminescence (CL) system. The optimum conditions for the CL reaction between protoberberine alkaloids and potassium permanganate were studied in detail. Under the optimum conditions, the linear response ranges for berberine, palmatine and jatrorrhizine were 0.038–7.27, 0.031–18.1 and 0.012–3.61 μg/mL with detection limits of 0.005, 0.004 and 0.0007 μg/mL, respectively. This method was successfully applied to determine the content of protoberberine alkaloids (calculated using berberine as an index) in Cortex Phellodendri and Rhizoma Coptidis. In addition, a possible mechanism of this CL reaction was proposed on the basis of the investigation of CL, UV and fluorescent spectra of protoberberine alkaloids in acidic solution containing potassium permanganate. Copyright © 2009 John Wiley & Sons, Ltd.     

Simultaneous determination of berberine, palmatine and jatrorrhizine by liquid chromatography-tandem mass spectrometry in rat plasma and its application in a pharmacokinetic study after oral administration of coptis-evodia herb couple

A rapid and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed to simultaneously determine berberine, palmatine and jatrorrhizine in rat plasma. After mixing with the internal standard (IS) tetrahydropalmatine, plasma samples were pretreated by protein precipitation with acetonitrile-methanol (1:2, v/v). Chromatographic separation was carried out on a C18 column using a mixture of water (containing 0.1% formic acid) and acetonitrile (30:70, v/v) as mobile phase. The detection was performed by selected reaction monitoring (SRM) mode via electrospray ionization (ESI) source operating in the positive ionization mode. The method was linear over the concentration range of 1.0-250.0 ng/mL for all components. The intra- and inter-day precision values were less than 14.6% and the deviations were within +/-4.0%. The fully validated LC-MS/MS method has been successfully applied to the pharmacokinetic study of berberine, palmatine and jatrorrhizine in rat plasma after oral administration of coptis-evodia herb couple. Three peaks were observed in both individual and mean plasma-concentration curves of berberine, palmatine and jatrorrhizine, which may be attributed to distribution re-absorption and enterohepatic circulation.     

A comparative study on the anti-inflammatory,antinociceptive and antipyretic effects of isoquinoline alkaloids from the roots of Turkish Berberis species

Roots and barks of various Berberis species are used as folk remedy for the treatment of various inflammatory diseases such as lumbago, rheumatism and to reduce fever. Six isoquinoline alkaloids namely berberine, berbamine, palmatine, oxyacanthine, magnoflorine, and columbamine were isolated as the main components of alkaloidal fraction from the roots of Turkish Berberis species and effects were studied using various in vivo models in mice. All alkaloids inhibited inflammations in varying degrees, among them berberine, berbamine and palmatine were shown to possess significant and dose-dependent inhibitory activity against serotonin-induced hind paw oedema both on oral and topical applications and acetic acid-induced increase in vascular permeability on oral administration. Moreover, these three alkaloids were also shown to possess dose-dependent antinociceptive activity, which assessed by using the model based on the inhibition of p-benzoquinone-induced writhing movements as well as antipyretic activity on FCA-induced increased rectal temperature on subacute administration. However, all alkaloids induced gastric lesions in varying degrees.