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.     

Quantitative determination of diterpenoid alkaloids in four species of Aconitum by HPLC

A high performance liquid chromatography (HPLC) method has been developed for the determination of five principal alkaloids (benzoylmesaconine, mesaconitine, aconitine, hypaconitine and deoxyaconitine) found in four species of genus Aconitum. The five alkaloids were analyzed simultaneously with an XTerraRP18 column by gradient elution using 0.03 M ammonium hydrogen carbonate-acetonitrile as mobile phase. The recovery of the method was 94.6-101.9%, and all the alkaloids showed good linearity (gamma = 0.9999) in a relatively wide concentration range. The results indicated that contents of alkaloids in Aconitum varied significantly from species to species; hence quality control of Aconitum drugs is very necessary.     

Norditerpenoid alkaloids from Aconitum manshuricum

A new norditerpenoid alkaloid, manshuritine (1) has been isolated from Aconitum manshuricum, together with seven known alkaloids; beiwudine (2), beiwutine (3), 16-epi-pyromesaconitine (4), mesaconitine (5), aconitine (6), hypaconitine (7) and 14-benzoylmesaconine (8). The structure of the new compound was elucidated on the basis of spectral data and chemical correlations.     

对中国药典附子中乌头硷限量检查的商榷

用高效液相色谱法测定生附子。炮制附片中的乌头硷、中乌头硷和次乌头硷含量，以及３种生物硷的水解速度，发现炮制附片中主要含次乌头硷，而药典只限定乌头硷含量，显得不够完善。故在此基础上探讨了药典中增加次乌头硷含量限定内容的可能性。     

Chemical constituents and analgesic activity of Aconitum kusnezoffii Reichb.

In the present study,we aimed to investigate the chemical constituents and analgesic activity of Aconitum kusnezoffii Reichb. The isolation and purification of components were achieved by a series of chromatography, including silica gel, Sephadex LH-20 and HPLC. By using spectroscopic analysis, their structures were identified. Using PDE-4A as analgesic target, moleculardocking was conducted between isolated compounds by using Schrodinger software. Neoline is a typical non-ester diterpene alkaloid. It was studied by using the mouse torsion body method and hot plate method. A total of 12 diterpene alkaloids were obtainedand identified as Mesaconitine(1), Bewutine (2), Bewudine (3), Songoramine (4), Songorine (5), Neoline (6), Talasamine (7), isotalatizidine (8), Hokbusine A (9), Mesaconine (10), 8-OEt-14-benzoylmesaconine (11), 8-Methoxy-14-benzoyl-beiwutinine (12).Compounds 9 and 12 were isolated from Aconitum kusnezoffii Reichb. for the first time. Twelve diterpenealkaloids could act on the analgesic target. Neoline is a typical non-ester diterpene alkaloid. It had significant analgesic effect. Diterpene alkaloids were the main components of Aconitum kusnezoffiiReichb., and they had good analgesic activity.     

Separation and identification of Aconitum alkaloids and their metabolites in human urine.

To study the safety of Aconitum medicinal herbs in clinic and identify Aconitum alkaloids poisoning in forensic medicine, Aconitum alkaloids and their metabolites were separated and identified in human urine by liquid chromatography-electrospray ionization-multi-stage mass spectrometry (LC-ESI-MS(n)) and chemical pathway of metabolism was investigated. The alkaloids and their metabolites in the urine sample were extracted with solid-phase cartridges and separated by HPLC with acetonitrile-water-formic acid (40:60:0.5) mobile phase. Structures of five metabolites and three parent Aconitum alkaloids were identified with multi-stage mass spectrometry data through comparison with authentic substances as aconitine (M(1)), mesaconitine (M(2)), hypaconitine (M(3)), benzoylaconine (M(4)), benzoylmesaconine (M(5)), benzoylhypaconine (M(6)), 16-O-demethylaconitine (M(7)) and 16-O-demethylhypaconitine (M(8)), respectively. Among them, M(8) was identified and reported for the first time. Metabolic pathways of Aconitum alkaloids in human body were proposed.     

全固态乌头碱电化学检测器的研制及其在流动注射分析中的应用

报道一种新型结构的全固态乌头碱电化学检测器的研制及其在流动注射分析中的应用。采用流动注射分析法对川乌、草乌及其中成药(小活络丸)中剧毒成分(双酯型生物碱)进行了测试,方法简便快速,结果同光度法接近。本文还提出了电化学法研究乌头碱水解动力学原理。在pH 6.5,温度98℃的条件下测得乌头碱水解成苯甲酰乌头碱的速度常数为1.36×10^-2min^-1。     

一测多评法测定乌头类药材中生物碱

目的 建立一测多评法同时定量测定乌头类药材中6种乌头类生物碱.方法 采用高效液相色谱法同时测定乌头类药材中6种生物碱,以乌头碱为内参物,测定其与次乌头碱、新乌头碱、苯甲酰乌头原碱、苯甲酰次乌头原碱、苯甲酰新乌头原碱的相对校正因子,利用乌头碱和相对校正因子对其他5种生物碱成分进行测定,同时利用外标法测定这6种成分,比较两种测定方法的差异,验证一测多评法的可行性和准确性.结果 在一定线性范围内,乌头碱与次乌头碱、新乌头碱、苯甲酰乌头原碱、苯甲酰次乌头原碱和苯甲酰新乌头原碱的相对校正因子分别为0.780、1.008、0.836、0.907、0.987,在不同实验条件下相对校正因子重现性良好,不同乌头类药材中6种生物碱成分含量计算值与实测值间无显著性差异.结论 一测多评法在乌头类药材及制剂质量控制中应用是可行的、准确的.     

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。     

不同产地哈萨克药材白喉乌头中总生物碱含量的分析

目的测定不同产地白喉乌头所含总生物碱的含量。方法采用索式提取法对药材进行提取,结合酸溶碱沉和萃取的方法从药材中提取白喉乌头总生物碱;利用紫外光谱对经过酸性染料络合的生物碱进行分析,检测其中总生物碱的含量。结果新源县、巩留县和昭苏县所产白喉乌头原药材中含有总生物碱分别为2.246,4.682和3.597 mg.g 1;回归方程为Y=11.5X+0.121,r=0.999 6,乌头碱浓度在6～36μg.mL 1内呈良好的线性关系。结论采用紫外-可见分光光度法测定不同产地白喉乌头中总生物碱的含量,具有操作简便、结果准确的特点;其结果可为该药材的活性和毒性物质基础研究提供依据。     

双向发酵对川乌指纹图谱及乌头碱类成分含量的影响

目的:观察川乌被19种药用真菌发酵后,乌头碱、新乌头碱、次乌头碱含量的变化及高效液相色谱(HPLC)指纹图谱的差异。方法:将川乌作为"药性基质",在一定的条件控制下,应用双向发酵技术,使其被槐耳、灵芝、红栓等19种药用真菌发酵,产生药材的"药性菌质",并采用反相高效液相色谱法对"药性菌质"和生药材中所含的乌头碱、新乌头碱和次乌头碱进行含量测定,比较其含量的变化及指纹图谱的差异。结果:在一定的时间范围内,大多数的菌株在川乌基质上的适应性良好,菌丝体生长旺盛,如代号A、C、D、E、F、M、Q、V、SW、い等真菌。发酵之后的川乌中乌头碱、新乌头碱及次乌头碱含量与生药材比较多数有明显的降低。结论:双向发酵后多数"药性菌质"中乌头碱、新乌头碱和次乌头碱含量明显降低了,提示双向发酵是生物技术在中医药领域中应用的有益尝试。     

Determination of aconitine-type alkaloids as markers in fuzi (Aconitum carmichaeli) by LC/(+)ESI/MS(3)

LC/(+)ESI/MS(3) was used to determine aconitine, mesaconitine, and hypaconitine as target markers in crude methanol extracts of (i) the raw lateral roots of Aconitum carmichaeli, (ii) roots treated by three different refining processes, and (iii) eight generally available traditional Chinese medicine (TCM) preparations containing fuzi (treated lateral roots of A. carmichaeli). The optimal ionization behavior resulted when using electrospray ionization (ESI) in positive-ion mode with 0.005% TFA as an additive in the mobile phase. The consecutive reaction monitoring (CRM) mode provided additional improvements in selectivity, which was exploited to minimize the noise and interference problems. Employing this approach, aconitine and mesaconitine were found to decompose readily during the refining processes, but hypaconitine remains present at the same content, presumably because of its characteristic chemical structure. Thus, treated and untreated fuzi samples can be distinguished by monitoring the ratio of aconitine and mesaconitine to hypaconitine. The limits of detection (LODs) for these three markers were 0.05, 0.08, and 0.03 ng/ml. The linearity range for the three marker compounds was 0.1-1,000 ng/ml. The analysis time was 12 min per sample.     

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.     

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.     

川乌氯仿部位的化学成分研究

目的研究川乌氯仿部位的化学成分。方法通过硅胶、碱性氧化铝柱色谱进行分离纯化,并根据理化性质和波谱数据鉴定化合物的结构。结果分离得到9个化合物,分别鉴定为次乌头碱(1)、金色酰胺醇酯(2)、乌头碱(3)、松胞素B2(4)、宋果灵(5)、新乌头碱(6)、多根乌头碱(7)、北乌碱(8)、附子灵(9)。结论化合物2为首次从乌头属植物中分离得到,化合物4为首次从植物界中分离得到。     

Neuropharmacological effects of Aconiti Lateralis Radix Praeparata

Aconiti Lateralis Radix Praeparata (Fuzi in Chinese), which are the lateral roots of Aconitum Carmichaelii Debx, is widely used in China to treat many neurological diseases. Fuzi, in its various forms, has many neuropharmacological effects. It can act as an analgesic and help with depression, epilepsy, and dementia. However, the neuropharmacological effects of Aconiti Lateralis Radix Praeparata are seldom comprehensively reviewed. In this review, the neuropharmacological activities of some components contained in Aconiti Lateralis Radix Praeparata are considered. These include aconitine, mesaconitine, hypaconitine, total alkaloid, polysaccharide-1, benzoylmesaconine, fuziline, songorine, and napelline. We also specifically discuss the antidepressant effects of total alkaloids and polysaccharide-1. This review may provide a theoretical basis for further utilization of Aconiti Lateralis Radix Praeparata for diseases that affect the central nervous system.     

草乌中有毒生物碱的提取分离

针对药材中乌头碱,中乌头碱、次乌头碱的提取分离方法进行了研究,本方法提取回收率与碱性苯溶液提取方法相似.方法中所用试剂成本低、毒性小,适用于乌头碱、中乌头碱、次乌头碱的提取分离以及对照品的制备.     

草乌花及其煎煮液中二萜生物碱的电喷雾串联质谱研究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。