大乌头及藤乌头类乌头的调查鉴定与成分分析

本文报道了民间药大乌头及藤乌头类乌头的植物来源,前者属于乌头属乌头亚属乌头组中的显柱乌头系(Ser Stylosa)类植物,后者系同组的蔓乌头系(Ser Volubulia)类植物,药用部分为其块根。研究了生药中生物碱的高效液相色谱测定方法,用十八烷基键合相柱,以甲醇—水—三乙胺(75:25:0.2)作流动相,各生物碱均能基线分离。分析了属于这类乌头的十二种不同植物品种的生药块根,均含有滇乌碱类生物碱,从化学分类学上证明这两群乌头植物在亲缘关系上有较密切的联系。     

宾川乌头中的生物碱及其结构的研究

从云南药用草乌之一的宾川乌头(Aconitum duclouxii Levl)根中分得两种结晶性生物碱,称为碱A及碱B。经鉴定,碱B为乌头碱(aconitine)。碱A分子式为C₃₄H₄₇O₁₀N,熔点168～169℃,[α]_D^28.1+28.3(C,1.43,氯仿),此系一新生物碱,命名为宾乌碱(duclouxine),经光谱分析,初步推定其化学结构式为12-β-羟基查斯乌头碱(12-β-hydroxy chasmaconitine)。宾乌碱经初步药理试验说明有镇痛作用。     

人参四逆汤质量控制方法的研究——乌头类生物碱的鉴别、含量测定及乌头碱的限量检查

本文以人参四逆汤中附片的有效成分为指标。用薄层色谱法作乌头类生物碱鉴别及乌头碱的限量检查。用离子对萃取—分光光度法测定乌头类生物碱含量,乌头碱的回收率为98.33%±4.72%～103.6±3.09%,加样回收率为95.09±2.09%。方法准确、重现性好。     

反相离子对色谱法测定附子中生物碱成分

目的测定附子中乌头碱、新乌头碱、次乌头碱、北乌碱、苯甲酰乌头原碱和苯甲酰新乌头原碱等6种生物碱的含量。方法用反相离子对HPLC法，使用AichromBond-1 C₁₈柱(250 mm×4.6 mm ID)；流动相：乙腈-5 mmol·L^-1 NaH₂PO₄溶液，磷酸调至pH 4.5(50∶50)，内含7 mmol·L^-1 十二烷基硫酸钠(SDS)；检测波长：235 nm；流速：1.0 mL·min^-1；柱温：35 ℃。结果以上6种生物碱可以完全分离，准确测定。结论该方法准确度高，可应用于附子中生物碱的含量测定。     

小活络丸中乌头类生物碱成分含量测定方法的建立

目的：建立小活络丸中生物碱成分乌头碱、次乌头碱、新乌头碱、苯甲酰乌头原碱、苯甲酰次乌头原碱与苯甲酰新乌头原碱的含量测定方法，以完善其质量标准。方法：采用高效液相色谱法， 选择PICKERING C₁₈（4.6 mm×250 mm，5μm）色谱柱，以甲醇、乙腈和0.1%磷酸水为流动相，梯度洗脱，流速1.0 mL·min^-1，柱温25 ℃，检测波长232 nm，进样量15 μL。结果：乌头碱、次乌头碱、新乌头碱、苯甲酰乌头原碱、苯甲酰次乌头原碱及苯甲酰新乌头原碱分别在各自范围内线性关系良好（r=0.9999），平均加样回收率分别为99.8%（RSD=0.8%）、99.9%（RSD=1.0%）、100.6% （RSD=1.4%）、100.8%（RSD=1.5%）、100.9%（RSD=1.5%）、100.6%（RSD=0.8%）。5批样品中上述6个成分含量范围分别为0.5~2.9、5.4~27.4、0.5~10.5、18.8~24.6、18.8~30.2及142.4~181.6 μg·g^-1。 结论：所建立的同时测定6个生物碱含量的测定方法可准确测定小活络丸中生物碱成分的含量，有助于提高小活络丸的质量标准。     

中国乌头之研究 ⅩⅨ 四川雪上一枝蒿中生物碱及其结构

从四川金沙江畔雪上一枝蒿(Aconitum pendulum)中分出四个结晶性的生物碱,证明其中三个为已知生物碱,即次乌头碱(hypaconitine),乌头碱(aconitine)和3-乙酰乌头碱(3-acetylaconitine);另一种为新生物碱,暂称为雪乌碱(penduline)熔点166～167℃,分子式C₃₄H₄₇NO₉,结构式初步推定如下式所示。雪上一枝蒿中主要生物碱为乌头碱,得率约为0.14%;雪乌碱对动物具有镇痛局麻等作用。     

中国乌头之研究——ⅩⅪ.赣皖乌头的研究

从赣皖乌头(Aconitum finetianum Hand-Mazz)中又分得五个生物碱,经光谱及化学方法鉴定,其中二个为新的二萜类生物碱,即脱乙酰冉乌头碱(N-deacetylranconitine),C₃₀H₄₂N₂O₆,熔点125～127℃,和脱乙酰赣乌碱(N-deacetylfinaconitine),C₃₀H₄₂N₂O₉,熔点121～123℃,及三个已知生物碱,即阿娃乌头碱(avadharidine)、牛扁次碱(lycoctonine)和脱乙酰刺乌头碱(N-deacetyllappaconitine)。经药理试验表明,脱乙酰冉乌头碱,脱乙酰赣乌碱和脱乙酰刺乌头碱均具有良好的镇痛效果。     

口服制川乌提取物后大鼠体内乌头类生物碱的组织分布

目的建立HPLC法测定口服制川乌提取物后大鼠体内乌头类生物碱的方法。方法用Waters 2690@996 PAD系统，Halsil 100 C₁₈柱(250 mm×4.6 mm ID, 5 μm)，水-甲醇-二乙胺(75∶25∶0.1)为流动相，流速0.9 mL·min^-1，检测波长240 nm。结果乌头碱在心脏、脾脏、肺脏、肾脏的线性范围：0.4-100 μg·mL^-1，r分别为0.997 2,0.998 6,0.999 3,0.999 4；在肝脏的线性范围：2-200 μg·mL^-1，r为0.999 0。次乌头碱在心、肝、脾、肺、肾、脑和脊髓的线性范围：5-100 μg·mL^-1，r分别为0.999 4,0.999 7,0.999 8,0.998 4,0.999 8,0.999 8和0.999 7。乌头碱及次乌头碱的检测限(S/N=3)为0.4 μg·mL^-1。各组织中的回收率：乌头碱为88.7%-102.2%，次乌头碱为865%-101.3%。结论本法为确证乌头类生物碱的中毒提供了科学有效的依据。     

中国乌头的研究——ⅩⅩ.赣皖乌头的研究

本文报道了从赣皖乌头Aconitum finetianum Hand-Mazz的根中分到三个二萜类生物碱A.B.c。经鉴定B和C分别为lappaconitine和ranaconitine。碱A,C₃₂H₄₄N₂O₁₀,熔点220～221℃,[α]_D²²+44.7°(甲醇),为一个新生物碱,暂称赣乌碱finaconitine,经光谱推定化学结构为10-β-羟基-冉乌头碱。     

弯喙乌头的化学成分研究

从弯喙乌头(Aconitum campylorrhynchum)中分离出一个微量新生物碱,经光谱和化学方法证明为8-乙酰嘟啦碱-(8-acetytdolaconine,Ⅰ),同时分离得到2个已知生物碱;即嘟啦碱(Ⅱ)和aconosine,(Ⅲ),后者是该植物的主要生物碱成分,在非生物碱部分还分离得到β-谷甾醇,棕榈酸和香豆酸。     

Physicochemical quality evaluation of natural compounds isolated from crude drugs

Aconite root has high toxicity caused by diester alkaloids, thus it was necessary to define the limiting value of diester alkaloids used in medicine formulation. To give the quality of “Processed Aconite Root” and “Powdered Processed Aconite Root” in the Japanese Pharmacopoeia (14th edn, supplement II), we established the official specification and evaluation methods of standard substances. High qualitative grade diester alkaloids, aconitine, hypaconitine, jesaconitine and mesaconitine, which were useful to evaluate the purity of processed aconite root and powdered processed aconite root, were prepared and evaluated for their stability. We studied the physicochemical specification and evaluation methods of these alkaloids. In addition, an “Aconitum diester alkaloids standard solution for purity”, which was used for the purity test, was prepared, and we also studied its physicochemical specification and evaluation methods. In addition, to evaluate the quality of processed aconite root and powdered processed aconite root, a TLC identification test was established. A monoester alkaloid of benzoylmesaconine hydrochloride was used as the reference standard in the latter test, and we also investigated its physicochemical specification and evaluation methods.     

中国乌头的研究Ⅶ.乌头生物碱的薄层层析

本文选择七种已知结构的生物碱,用不加粘合剂氧化铝薄层层析法的桑件进行了研究,结果表明此法完全可应用于乌头生物碱的检定.其较好条件是细度小于150号筛孔,活性Ⅳ级碱性氧化铝为吸附剂;石油醚-乙醚(1:10)为展开剂和以碘蒸气为显色剂.将此薄层层析条件应用到从中国乌头中分出的新生物碱检查时,亦获得良好的结果.     

中国乌头的研究 ⅩⅢ.北草乌中的生物碱

从内蒙古赤峰产的北草乌中共分得五种结晶性的生物碱,并证明其中四种为已知物,即去氧乌头碱(deoxyaconitine),次乌头碱(hypaconitine),乌头碱(aconitine)和新乌头碱(mesaconitine);另一种为新生物碱,暂称北草乌碱(beiwutine),熔点196～198℃,分子式C₃₃H₄₅NO₁₂,结构式推定如图(1)。此草乌中的主要生物碱为新乌头碱。     

Aconitum Alkaloid Poisoning Related to the Culinary Uses of Aconite Roots

Aconite roots (roots or root tubers of the Aconitum species) are eaten as root vegetables and used to prepare herbal soups and meals, mainly for their purported health benefits. Aconite roots contain aconitine and other Aconitum alkaloids, which are well known cardiotoxins and neurotoxins. To better understand why Aconitum alkaloid poisoning related to the culinary uses of aconite roots can occur and characterize the risks posed by these “food supplements”, relevant published reports were reviewed. From 1995 to 2013, there were eight reports of aconite poisoning after consumption of these herbal soups and meals, including two reports of large clusters of cases (n = 19–45) and two reports of cases (n = 15–156) managed by two hospitals over a period of 4.5 to 5 years. The herbal formulae used did not adhere to the suggested guidelines, with regarding to the doses (50–500 g instead of 3–30 g per person) and types (raw instead of processed) of aconite roots used. The quantities of Aconitum alkaloids involved were huge, taking into consideration the doses of aconite roots used to prepare herbal soups/meals and the amounts of aconite roots and herbal soups/meals consumed. In a large cluster of cases, despite simmering raw “caowu” (the root tuber of A. kusnezoffii) in pork broth for 24 h, all 19 family members who consumed this soup and boiled “caowu” developed poisoning. Severe or even fatal aconite poisoning can occur after consumption of herbal soups and foods prepared from aconite roots. Even prolonged boiling may not be protective if raw preparations and large quantities of aconite roots are used. The public should be warned of the risk of severe poisoning related to the culinary and traditional medicinal uses of aconite roots.     

附子新型炮制品中乌头类生物碱测定及其强心作用研究

目的 比较附子新型炮制品与传统炮制品中乌头碱类水平及强心作用,为附子炮制工艺改进提供药理学依据。方法 采用高效液相(HPLC)色谱法测定生附子及黑顺片、高温片、高压片、微波片4种附子炮制品及其煎煮液中6种乌头类生物碱水平;应用Langendorff大鼠离体心脏灌流法,测定附子4种炮制品对心率(HR)、左心室收缩压(LVSP)、舒张末压(LVEDP)、左心室内压变化速率(±dp/dt_max)的影响,分析其强心作用。结果 与生附子比较,附子经炮制后,毒性成分双酯型生物碱减低,单酯型生物碱逐渐增加;与黑顺片比较,高压片中生物碱得到最大限度保留,单酯型生物碱水平高于黑顺片,双酯型生物碱水平较低,说明附子高压炮制是一种“低毒高效”的工艺;而高温片和微波片中单、双酯型生物碱水平均有所降低,其成分有一定的流失;4种炮制品煎煮液中6种生物碱溶出率与炮制品中生物碱水平呈正相关。与对照组比较,4种炮制品均能够显著升高离体大鼠心脏的HR、LVSP、±dp/dt_max,降低LVEDP,且呈一定剂量相关性;与黑顺片组比较,3种新型炮制品HR、LVSP、LVEDP、±dp/dt_max均差异不显著;在相同剂量下,高压片中HR、LVSP、±dp/dt_max值最高,LVEDP值较低。结论 附子高压炮制工艺是一种“高效低毒”炮制工艺,且强心作用效果较好。     

乌头安全有效性研究进展及其临床应用前景

目的 分析文献中乌头疗效与安全应用的研究进展、用药特点与注意事项,促进该药材及其复方制剂的临床合理与广泛的应用。方法 通过检索秦汉时期至今的主要书籍和文献,从乌头的药理作用、药效物质、炮制方法、药代动力学、民间应用、临床疗效等多方面进行系统的总结和归纳,并提出乌头安全有效的应用前景和现代制剂研制的基本要求。结果 乌头有抑瘤、镇痛、提高免疫力、抗炎等作用,其药效物质为乌头类生物碱,其毒性大小为乌头碱、新乌头碱、次乌头碱>苯甲酰乌头碱、苯甲酰新乌头碱、苯甲酰次乌头碱>乌头原碱、新乌头原碱、次乌头原碱;体内研究显示乌头在大鼠体内吸收迅速,代谢较快,但不同文献中同一成分生物半衰期数据差异较大,变化幅度分别为：乌头碱0.936~20.130 h,新乌头碱1.490~20.930 h,次乌头碱1.930~17.300 h,导致体内消除速率和蓄积时间明显不同;临床使用中出现的毒副作用主要与合理炮制、恰当配伍、正确服用、控制剂量,避免误服等操作的失当以及个体差异相关。结论 乌头应用的民间基础扎实,镇痛等疗效确切,炮制工艺、药理毒理与临床应用等数据翔实,建议加强体内研究数据的确证,开展临床安全的再评价,进一步阐明乌头的作用机制和量效关系。     

The relationship between growth of the aerial part and alkaloid content variation in cultivated <Emphasis Type="Italic">Aconitum carmichaeli</Emphasis> Debeaux

Processed root of aconite, Aconitum carmichaeli Debeaux—known as bushi in Japan—is indispensable for treating diseases among elderly persons in Japanese and Chinese traditional medicine. Its active component is bushi diester alkaloid (BDA), which consists of aconitine (ACO), mesaconitine (MES), hypaconitine (HYP), and jesaconitine (JES). Since an overdose of BDA results in severe side effects, the BDA content should be within safe limits. However, the BDA content of raw aconite root, even that produced by standard cultivation procedures, varies greatly. In this study, to clarify the cause of BDA variation, we examined the weight and BDA content of each part of cultivated A. carmichaeli: the aerial part, the mother tuberous root (MT), the daughter tuberous root (DT), and the rootlet (RL). We found the following positive relationships: between aerial part weight and DT weight, aerial part weight and BDA content in stem of apex, and BDA content in stem of apex and total BDA of DT attached to the plant. Furthermore, DT belonging to a higher weight group showed less BDA content variation. In addition, BDA of DT and those of MT and RL differ in both content and composition. In conclusion, it was suggested that the weight or the size of the aerial part was a good marker for monitoring BDA content and its variation in the tuberous root, and it was found to be desirable to prevent mixing MT and RL at harvest.     

The effect of planting depth,disbudding, and root pruning on root weight and aconite alkaloid content of the tuberous root of aconite

The tuberous root of aconite, Aconiti tuber, contains tonic but poisonous aconite alkaloids, such as aconitine, hipaconitine and mesaconitine. The standardization of the cultivation protocols must be established, since the quality of the crude drug, especially the content of aconite alkaloids, is influenced by cultivation conditions. Here, we report the effects of planting depth, disbudding and root pruning on the content of the aconite alkaloids as well as on the weight of the root tuber of aconite. The daughter root tubers of shallow-planted plants showed a significantly higher content of aconitine, hipaconitine and sum of aconite alkaloids per unit weight than those of deeply planted root tubers (control group), while the average weight per root tuber was lower than that of the control group. The total weight of the daughter root tubers per plant of the disbudding group was significantly higher than that of other groups, including the root-pruning group whose average weight per tuber was the highest. In conclusion, disbudding is effective in achieving both higher content of aconite alkaloids and heavier total weight of the root tuber.     

生川乌瓜蒌不同配伍中乌头类生物碱在大鼠体内的药动学研究

目的考察生川乌瓜蒌不同配伍比例醇提液中乌头类生物碱在大鼠体内的药动学研究,比较瓜蒌对其药动学行为的影响。方法建立灵敏、专属、快速的测定大鼠血浆中6种乌头类生物碱的液相色谱-串联质谱方法。大鼠随机分为3组,分别ig生川乌与瓜蒌1∶0、6∶1、1∶6配伍醇提液,在不同的时间点采血分析,经DAS 2.0软件计算主要药动学参数。结果乌头碱、次乌头碱、新乌头碱的药动学参数tmax和t1/2发生了显著变化,且变化趋势基本一致,在生川乌与瓜蒌6∶1配伍组tmax显著减小(P0.05),生川乌与瓜蒌1∶6配伍组tmax显著增加(P0.01、0.001),且t_(1/2)显著降低(P0.05、0.01),提示生川乌与瓜蒌不同配伍比例影响了3种生物碱的吸收和消除过程。结论生川乌与瓜蒌配伍影响了乌头类生物碱的药动学过程,配伍比例是其配伍禁忌的重要条件。     

Development and validation of a highly sensitive UPLC-MS/MS method for simultaneous determination of aconitine,mesaconitine, hypaconitine,and five of their metabolites in rat blood and its application to a pharmacokinetics study of aconitine,mesaconitine, and hypaconitine

1. A rapid, specific and sensitive method was developed for the simultaneous determination of eight Aconitum alkaloids: aconitine (AC), mesaconitine (MA), hypaconitine (HA), benzoylaconine (BAC), benzoylmesaconine (BMA), benzoylhypaconine (BHA), aconine and mesaconine in rat blood by ultra-performance liquid chromatography coupled tandem mass spectrometry (UPLC-MS/MS).

2. The UPLC-MS/MS system coupled with an electrospray ionization (ESI) source was operated in a positive mode via multiple-reaction monitoring (MRM). Samples were treated with methanol to remove protein prior to analysis by UPLC-MS/MS. The analytes were separated with a Waters C18 column (1.7 µm, 50?×?2.1?mm) and a gradient elution using acetonitrile and 0.1% formic acid-water as the mobile phases.

3. The linear response range was from 0.125 to 1000 nmol/L for these eight alkaloids and the correlation coefficients (r² values) were all higher than 0.997. The method was validated with respect to precision, accuracy, recovery, matrix effect, carryover effect and sample stability, and found to be within the acceptable limits.

4. The developed and validated method was successfully applied to simultaneously determine the eight Aconitum alkaloids in rats blood after intravenous administration of a mixture of AC, MA and HA.