草乌花及其煎煮液中二萜生物碱的电喷雾串联质谱研究

目的对草乌花及其煎煮液中的二萜生物碱进行定性分析,说明煎煮前后化学成分发生的变化. 方法用注射泵自动进样,电喷雾离子阱串联质谱直接分析草乌花及其煎煮液中生物碱混合物.结果在生草乌花中发现3个新生物碱,草乌花煎煮后其中的双酯型生物碱和三酯型生物碱都发生水解,前者水解为苯甲酰乌头原碱和乌头原碱类生物碱,后者水解为3-乙酰-乌头原碱类生物碱.结论该法简便、快速、灵敏、特异性强,为乌头属植物煎煮液中的生物碱分析提供了新途径.     

附子配伍原则的电喷雾质谱研究

目的从化学角度探讨附子配伍的原则。方法利用电喷雾质谱分析和比较生附子及其复方中药煎煮液中二萜类生物碱的变化规律；测定附子及其他中药煎煮液的pH值并用电喷雾质谱考察酸性对药材中乌头碱水解的影响。结果生附子单煎煮及与甘草共煎后双酯型乌头生物碱大部分发生水解，生附子与半夏或五味子共煎生物碱水解受抑制，生附子和甘草单煎液的酸性远低于半夏或五味子单煎液，中乌头碱在酸性条件下不水解。结论酸性是影响乌头碱类双酯型生物碱水解的重要因素，强酸条件下水解受抑制，附子煎煮液毒性增加。     

附子炮制品及其煎煮液6种生物碱的含量测定

目的从附子炮制品来源、煎煮、配伍等方面评价其的安全性和有效性,为临床用药的提供一定参考依据。方法采用高效液相色谱法同时测定不同来源附子炮制品、单方煎煮和复方煎煮液中6种生物碱含量,综合评价附子炮制品的质量及煎煮、配伍对其有效成分和毒性成分的影响。结果不同来源黑顺片中6种生物碱含量差异较大,单酯型生物碱在0.303 5~1.394 8 mg/g之间,双酯型生物碱的含量在0~0.094 0 mg/g之间;炮附片生物碱含量偏低,单酯型生物碱在0.099 0~0.213 5 mg/g之间;两种炮制品经煎煮后,双酯型生物碱基本消失,单酯型生物碱含量也有所降低,其中黑顺片单煎液的溶出率为58.92%,炮附片单煎液的溶出率为87.87%,其复方煎煮液中单酯型生物碱含量均比单煎液偏高。结论 12批黑顺片中6种生物碱含量均符合药典要求,炮附片中生物碱含量偏低,不合格现象明显,煎煮液中双酯型生物碱含量基本消失,毒性更小,与甘草和干姜配伍后,有增效作用。     

草乌花总生物碱的纯化工艺研究

目的:优选草乌花总生物碱的纯化工艺。方法:采用酸碱滴定法测定草乌花总生物碱的含量。以树脂型号、上样药质量浓度、交换速度为考察因素,以最大吸附量、解吸率和总生物碱质量分数为指标,优化离子树脂纯化草乌花总生物碱的工艺并进行验证。结果:选择树脂型号为732型阳离子交换树脂、上样液质量浓度为0.32 g/L、交换速度为7倍柱体积(BV)/h为最优纯化工艺。验证试验中纯化后总生物碱质量分数平均值为86.88%(RSD=0.52%,n=3),解吸率平均值为92.81%(RSD=0.40%,n=3);3批样品草乌花总碱提取转移率平均值为81.76%,纯化转移率平均值为89.47%。结论:建立的纯化工艺稳定、可行,且转移效率较高。     

附子不同配伍药对中生物碱成分的电喷雾质谱分析

以传统中医用药附子的配伍药对为研究对象，考察附子与不同中药配伍对附子中生物碱的影响规律，揭示配伍减毒的科学内涵。利用电喷雾质谱技术和内标法，分别对生附子，生附子加炙甘草、白芍、干姜、大黄共煎液和药渣中乌头类生物碱进行系统考察。与生附子相比，双酯型生物碱在附子加炙甘草、大黄、白芍、干姜共煎液中的含量降低；附子加炙甘草、白芍、干姜共煎液中的脂型生物碱含量增高。附子与炙甘草、白芍、干姜配伍的解毒机制是使毒性较大的双酯型生物碱转化为毒性小的脂型生物碱；与大黄配伍的解毒机制是药物所含成分与附子中的双酯型生物碱络合生成难溶于水的复合物，使双酯型生物碱的含量降低。本法对深入研究乌头属植物的配伍作用机制具有借鉴作用。     

川乌不同煎煮时间总生物碱与酯型生物碱含量变化趋势对比

目的:以川乌为研究对象,比较其在不同煎煮时间的煎剂中总生物碱、酯型生物碱类成分的变化趋势。方法:采用滴定法及高效液相色谱法,对川乌在不同煎煮时间煎剂中的总生物碱及酯型生物碱进行测定。并以此为指标,考察其在不同煎煮时间的煎剂中有效成分及主要毒性成分变化趋势。结果:在煎煮0～4h的川乌煎剂中,随煎煮时间延长,其有效成分总生物碱含量呈现上升趋势,而主要毒性成分则呈现下降趋势。结论:为了使毒性药物川乌的使用符合"低毒高效",完善药典相关内容,应进一步明确双酯型生物碱与单酯型苯甲酰类乌头碱在川乌及其制剂中的含量及二者的比例。     

草乌与贝母配伍禁忌的化学研究

目的:探讨草乌与贝母在煎煮过程及体外模拟消化环境中生物碱类成分的变化规律,初步阐明草乌配伍贝母毒性作用变化原因。方法:采用酸性染料比色法、改良异羟肟酸铁分光光度法和HPLC法,测定草乌与贝母不同配比合煎液及人工胃、肠液处理后溶液中总生物碱和酯型生物碱、乌头碱类成分的含量,并通过主成分分析(PCA)和聚类分析(HCA)综合考察各配比和处理方法对毒性作用变化情况。结果:通过PCA和HCA的综合考察,两者得到的结果相似,均显示11个样品被分为3组,其中样品1为Ⅰ组,样品2、5、9、10、11为Ⅱ组,样品3、4、6、7、8为Ⅲ组,样品的分类结果可能与草乌生物碱和贝母生物碱含量有关;草乌与贝母在一定的比例下毒性增加,尤其是其合煎剂经胃液处理后毒性进一步增大,而经过肠液处理后毒性有一定的降低。结论:"草乌反贝母"是在一定条件下的配伍禁忌,可能与配伍比例和药物体内释放部位有一定的关系。     

不同煎煮时间对熟附子生物碱的影响

目的 通过测定熟附子不同煎煮时间的生物碱含量,确定最佳的煎煮时间,为规范化的煎煮工艺提供参考.方法 采用HPLC和UV方法分别测定熟附子煎煮液中6种生物碱和总生物碱的含量,并以此含量为指标综合评价其煎煮工艺.结果 熟附子主要含有单酯型生物碱和微量的双酯型生物碱,煎煮0.5h后,3种双酯型生物碱完全消失,而煎煮1h...     

HPLC测定草乌与贝母配伍后乌头碱类成分含量的变化

目的探讨生草乌与贝母不同配比在煎煮过程及体外模拟消化环境中乌头碱类成分的变化规律。方法采用HPLC法,测定生草乌与浙贝母不同配比水煎液及其经人工胃、肠液解离后的溶液中乌头碱及其水解产物焦乌头碱、苯甲酰乌头碱和苯甲酸的含量。结果除草乌贝母(2∶1)合煎液外,草乌贝母不同配比合煎或单煎后混合溶液中乌头碱的含量均较草乌单煎液明显提高。经人工胃肠液处理后各配伍液中乌头碱类成分的含量明显降低,但水解产物焦乌头碱、苯甲酰乌头碱与乌头碱的构成比较处理前明显提高。结论草乌与贝母一定比例下的配伍,毒性成分明显增大,证实了"草乌反贝母"的科学性。     

不同煎煮时间对熟附子生物碱的影响

目的通过测定熟附子不同煎煮时间的生物碱含量,确定最佳的煎煮时间,为规范化的煎煮工艺提供参考。方法采用HPLC和UV方法分别测定熟附子煎煮液中6种生物碱和总生物碱的含量,并以此含量为指标综合评价其煎煮工艺。结果熟附子主要含有单酯型生物碱和微量的双酯型生物碱,煎煮0.5 h后,3种双酯型生物碱完全消失,而煎煮1 h后,总单酯型生物碱和总生物碱均达到了峰值,分别为:2.900 1 mg/g和4.832 0 mg/g,随着煎煮时间的延长,单酯型生物碱和总生物碱的含量均逐渐下降。结论熟附子的最佳煎煮时间为1 h。     

不同品种附子生物碱和多糖含量的比较

目的:比较不同品种附子生物碱和多糖的含量。方法:采用酸性染料比色法测定附子总生物碱含量,反相高效液相色谱法测定双酯型生物碱含量,蒽酮-硫酸法测定多糖含量。结果:不同品种附子总生物碱和双酯型生物碱含量差异较大,附子多糖含量差异不显著。附子炮制后,黑附片中总生物碱、双酯型生物碱含量显著下降,附子多糖含量稍有增加。结论:本试验结果可为附子品种选育提供一定的科学依据。     

Diterpenoid alkaloids from the roots of Aconitum brachypodum Diels

A new diterpenoid alkaloid, named bullatine H (1), along with 10 known diterpenoid alkaloids were isolated from the roots of Aconitum brachypodum Diels (Ranunculaceae). The structure of 1 was elucidated by analysis of its spectroscopic data. It should be noted that compound 1 is the first example with 11, 13-dioxygenated denudatine-type diterpenoid alkaloid isolated from Aconitum brachypodum.     

Diterpenoid alkaloid toxicosis in cattle in the Swiss Alps

Between 1995 and 1999, several cattle of a group of 80 heifers died acutely on a pasture in the Swiss Alps. The animals were Found dead between July 9th and 15th eachyear. Only 1 animal was examined on post-mortem, and no significant lesions were found. Aconitum vulpera, A napellus, and Delphinium elatum were identified in the pasture. The presence of diterpenoid alkaloid-containing plants in the pasture, the rapid death of the animals, and the lack of pathologic lesions suggested diterpenoid alkaloid toxicosis as a cause of death. A multiresidue alkaloid screen using gas chromatography with a mass spectrometric detector was employed on rumen, abomasal, small intestine, and cecal contents from the I heifer. Deltaline, deltamine, and lycoctonine were identified. Aconitine was found in all gastrointestinal samples using a sensitive and highly specific liquid chromatography/mass spectrometry methodology for aconitine analysis. The findings ofditerpenoid alkaloids in the gastrointestinal contents confirmed exposure to Delphinium and Aconitum spp, possibly resulting in sudden death.     

Diterpenoid alkaloids from the roots of Aconitum cochleare

From the roots of Aconitum cochleare Woroschin, collected in Turkey, a new diterpenoid alkaloid named acochlearine has been isolated along with the known diterpenoid alkaloids talatisamine, 14-O-acetyltalatisamine, senbusine C and condelphine. The structure for acochlearine was established on the basis of 1H, 13C, DEPT, homonuclear 1H COSY, NOESY, HSQC and HMBC NMR studies.     

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.     

Distribution of Aconitum alkaloids in body fluids and tissues in a suicidal case of aconite ingestion

A case involving a suicidal ingestion of Aconitum tubers is presented. A 40-year-old woman in Hokkaido, Japan ingested ground aconite and died of aconite intoxication about 4 h after ingestion. The Aconitum alkaloids were quantitated using gas chromatography-selected ion monitoring from extracts of the body fluids and organs. The blood and urine concentrations of jesaconitine, the main alkaloid of the aconite in this case, were 69.1 ng/mL and 237.8 ng/mL, respectively. Higher values of the alkaloid were demonstrated in the kidneys, the liver, and in the bile rather than other organs or serum, suggesting the alkaloids were eliminated by the liver and kidneys. In the gastrointestinal tract, the highest value of jesaconitine (471.3 ng/g) was in the ileal contents. These findings show that Aconitum alkaloids were found in the liver and kidneys in much higher concentrations than in serum and suggest that they were eliminated not only via urine but also in feces. Feces may be useful to detect Aconitum alkaloid if other biological samples are not available.     

Diterpenoid alkaloids from Aconitum kirinense

A new C₁₈-diterpenoid alkaloid, kirinenine A (1), was isolated from the root of Aconitum kirinense, along with eight known diterpenoid alkaloids. The structures of all compounds were characterized on the basis of extensive NMR and MS analyses and by comparison with literature values, and the new one was further confirmed by X-ray crystallographic diffraction. All the compounds were isolated from the title plant for the first time.     

Studies on structural elucidation of Aconitum diterpenoid alkaloid by LC-APCI-MS and effects of Aconitum diterpenoid alkaloid on cutaneous blood flow

The chemical constituents of Aconitum yesoense var. macroyesoense and Aconitum japonicum were examined using high-resolution spectral analysis. Twelve novel alkaloids were isolated from A. yesoense var. macroyesoense together with 20 known alkaloids. Eight novel alkaloids were isolated from A. japonicum together with 15 known alkaloids. An HPLC-atmospheric pressure chemical ionization-mass spectrometry (HPLC-APCI-MS) method was useful for the simultaneous determination of 21 Aconitum alkaloids found in A. yesoense var. macroyesoense and A. japonicum. These compounds were fairly stable under the conditions used, and the protonated molecules or fragment ions characteristic of the molecule appeared as base peaks in the mass spectra and were used for selected ion monitoring. HPLC-APCI-MS is a very promising approach for structural investigations of positional isomers and stereoisomers. This method was applied successfully to stereoisomeric Aconitum alkaloids differing in configuration at C-1, -6, or -12. Comparison of the APCI spectra showed that the abundance of fragment ions was significantly higher for the C-1, -6, or -12 beta-form alkaloid than for C-1, -6, or -12 alpha-form alkaloid. The main alkaloid constituents in the root of A. yesoense var. macroyesoense, Aconitum alkaloids of the C20-diterpenoid type, kobusine and pseudokobusine, and their acyl derivatives were examined for their peripheral vasoactivities by measuring laser-flowmetrically the cutaneous blood flow in the hind foot of mice after intravenous administration. It is thought that the hydroxyl groups of alkaloids, especially a free OH group of pseudokobusine at C-6, were important for action on the peripheral vasculature leading to dilatation, and the results indicated that esterification of the hydroxyl group at C-15 with either anisoate, veratroate, or p-nitroben-zoate may contribute to enhancement of the activity of the parent alkaloids.