微波辅助提取/HPLC分析石蒜中的生物碱

石蒜科石蒜属(amaryllidaceae)植物,含有石蒜碱、力可拉敏和加兰他敏(图1)等生物碱,具有镇静、镇痛、解热、降压、抗炎、催吐、抗病毒、抗癌等作用[1]。目前石蒜生物碱的提取主要采用溶剂回流法(SRE),但SRE提取效率低、耗时、耗溶剂,而且有效成分易被破坏[2-3]。微波辅助提取法     

HPLC法同时测定石蒜中加兰他敏和石蒜碱

建立了高效液相色谱同时测定石蒜中加兰他敏和石蒜碱含量的方法。 采用phenomenex-C18色谱柱,以乙腈(A)-0.1%TFA水溶液(B)为流动相梯度洗脱,检测波长289 nm。 结果表明,加兰他敏和石蒜碱均在0.5～10 mg/L(r=0.9999)呈现良好的线性关系,最低检测限(S/N=3)分别为0.09和0.15 mg/L,平均加样回收率分别为99.53%和96.82%。 该方法简单、快速、准确,适合于石蒜中加兰他敏和石蒜碱的测定。     

微波辅助提取石蒜和虎杖中有效成分的动力学模型

基于微波辅助提取(MAE)中药材中化学成分的非稳态扩散过程, 根据Fick第二定律建立了石蒜中石蒜碱、力可拉敏和加兰他敏以及虎杖中白藜芦醇和大黄素微波辅助提取过程的动力学模型. 研究了提取时间、提取温度及药材粒度等因素对石蒜中石蒜碱、力可拉敏和加兰他敏以及虎杖中大黄素和白藜芦醇的提取率的影响, 采用Matlab软件编程对动力学模型进行回归分析, 拟合的动力学模型与实验结果吻合. 根据模型计算了石蒜中石蒜碱、力可拉敏和加兰他敏以及虎杖中大黄素和白藜芦醇在MAE提取过程中的扩散系数D, 与溶剂加热回流法(SRE)比较, 引入增强因子γ, 表征了微波对溶质分子扩散传质的影响及其对不同基质药材作用的差异.     

离子液体微波辅助萃取石蒜中生物碱的研究

以离子液体溶液为溶剂,研究了离子液体微波辅助萃取石蒜中石蒜碱、力克拉敏和加兰他敏生物碱。考察了离子液体的种类和浓度对萃取石蒜生物碱的影响,并与传统有机溶剂和传统萃取技术进行了对比研究。结果表明,1.0mol/L[bmim]Cl溶液为溶剂,液固比(mL∶g)15∶1,80℃微波辅助萃取10min,石蒜碱、力可拉敏和加兰他敏生物碱的萃取率分别为2.730、0.857和0.179mg/g。与传统的萃取方法比较,本方法快速高效、环境友好。     

超高效液相色谱-四极杆线性离子阱质谱法同时检测石蒜属3种生物碱

基于超高效液相色谱-串联四极杆/线性离子阱质谱(UPLC-QTRAP-MS/MS),建立了一种同时检测石蒜属植物中加兰他敏(Gal)、力可拉敏(Lycm)和石蒜碱(Lyc)含量的方法。提取液超声波辅助提取离心后,经Waters ACQUITY UPLC BEH C18(150 mm×2.1 mm,1.7μm)色谱柱,以水溶液(0.1%甲酸)-乙腈为流动相梯度洗脱,流速0.2 m L/min,在正离子扫描下以MRM模式进行分析。结果表明,Gal、Lycm和Lyc在0.5~500μg/L质量浓度范围内线性关系良好,相关系数达0.999 9~1.000 0,可在6.0 min内得到检测结果,Gal、Lycm和Lyc的保留时间分别为2.86、2.31、1.95 min。3种物质的加标回收率为97.2%~98.6%,相对标准偏差为0.8%~4.3%。应用此方法测得石蒜属12个种的Gal、Lycm和Lyc含量分别在21.98~496.77、0.17~467.21、9.34~4 510.18μg/g干重之间,3种物质含量在种间差异均较大。     

石蒜科生物碱的研究Ⅴ.臭水仙中的生物碱及新生物碱臭水仙碱的结构

从臭水仙(Narcissus papyraceus Ker-Gawl)的鳞茎中分离和鉴定了六个已知生物碱:石蒜碱(1)、多花水仙碱(2)、加兰他敏(3)、力可拉敏(4)、假石蒜碱(5)和maritidine(6).此外还分到一种新生物碱臭水仙碱(Papyramine),根据化学和光谱(UV,IR,ORD,NMR)数据证明其化学结构为7.     

一种同时检测石蒜属植物中3种生物碱含量的方法

正申请公布号:CN107561186A申请公布日:2018.01.09申请人:上海市农业科学院摘要本发明公开一种同时检测石蒜属植物中3种生物碱含量的方法,能够对石蒜属植物中的生物碱类物质——加兰他敏、立可拉敏和石蒜碱进行快速提取并定性定量分析。     

微波辅助提取-气相色谱-质谱法测定石蒜中加兰他敏和力可拉敏

采用乙醇(95+5)溶液于70℃提取石蒜中加兰他敏和力可拉敏,所得提取液经过滤后旋转蒸发至干。残渣溶于盐酸(2+98)溶液中,用氯仿萃取3次以除去杂质,用氨水调节水相酸度至pH 10,再用氯仿萃取3次,合并萃取液并蒸发至干。用甲醇溶解残渣,所得溶液用DB-5MS毛细管柱分离,进样量为1μL。质谱分析中采用电子轰击离子源及在(m/z)50～550范围内全扫描方式检测。检索图谱库NBS 75K.L及NIST 147对样品作定性分析,并根据[M~+]286及[M~+]288的丰度分别对加兰他敏和力可拉敏作定量分析。对方法做回收及精密度试验,测得回收率在93.0%～106.0%之间,相对标准偏差(n=9)为5.5%(加兰他敏)和2.4%(力可拉敏)。     

微波辅助提取石蒜和虎杖中有效成分的热力学机理研究

采用微波辅助提取(MAE)技术研究了石蒜和虎杖两种不同植物中石蒜碱、力可拉敏和加兰他敏以及白藜芦醇和大黄素提取过程的热力学机理. 以溶剂回流提取方法(SRE)作为对比, 采用一种简单的测定提取分配系数的方法, 计算了这些组分在两种提取过程中的热力学函数ΔH0, ΔS0和ΔG0, 对其化学结构与极性以及在MAE过程中的热力学行为进行了讨论, 并用扫描电镜法观察了MAE和SRE提取后样品的细胞结构. 结果表明, 石蒜和虎杖的提取是一个吸热熵增的过程, 微波的作用导致石蒜和虎杖细胞结构发生显著变化, 使MAE热力学函数变化较大, 其提取过程的热力学行为特征与SRE明显不同, 但提取效率提高.     

微波辅助提取石蒜和虎杖中有效成分的动力学机理

以鳞茎药材石蒜(Lycoris radiata)和根茎药材虎杖(Rhizma Polygoni Cuspidati)作为研究对象,采用微波辅助提取(MAE)石蒜中的石蒜碱、力可拉敏和加兰他敏以及虎杖中的大黄素和白藜芦醇,研究了提取过程的动力学机理。分别对这5种组分在提取温度、提取时间、搅拌速度和颗粒度等因素影响下MAE过程的动力学行为和特征进行了探讨,并与常规的溶剂回流提取法(SRE)进行对比研究,采用扫描电镜观察了MAE和SRE提取后样品的表面细胞结构。结果表明,石蒜和虎杖的MAE提取过程具有明显不同的动力学特征,其机理分别基于内部扩散传质控制和细胞破壁引起的界面反应控制。与SRE相比,MAE能引起石蒜和虎杖的细胞结构发生显著变化,降低了表观活化能Ea,组分的表观速率常数明显增大。     

Simultaneous determination of galanthamine and lycorine in Lycoris radiata by a capillary electrophoresis with an electrochemiluminescence method

A novel capillary electrophoresis with electrochemiluminescence determination method was developed for the determination of two alkaloids based on the electrochemiluminescence signal enhancement effect of the tertiary amine group on tris(2,2′‐bipyridyl)ruthenium(II). A linear relationship between the electrochemiluminescence peak area and concentrations of galanthamine and lycorine in the range of 0.07 ～ 17 μg/mL and 0.07 ～ 18 μg/mL was obtained and the detection limit was 0.008 and 0.002 μg/mL, respectively. The method is selective, simple, and convenient. It had been successfully applied to the analysis of galanthamine and lycorine in Lycoris radiata samples purchased from a local market.     

高效液相色谱法测定麻黄及其制剂中的麻黄类生物碱和川芎嗪

 用RP HPLC分离测定了麻黄及其制剂中的麻黄类生物碱及川芎嗪。采用色谱柱Nova Pak C18(15 0mm×3.9mmi.d .) ,二极管阵列检测器 (DAD)。测定麻黄类生物碱时 ,以甲醇 0 .0 2mol/LKH2 PO4 乙酸 三乙胺 (体积比为 4∶96∶0 .2∶0 .0 1)为流动相 ,在 2 10nm波长下检测 ;测定川芎嗪时 ,以甲醇 水 乙酸 (体积比为 35∶6 5∶0 .5 )为流动相 ,在 2 90nm波长下检测。分别测定了麻黄原药材、中成药 (小儿清肺丸、鹭鸶咳丸 )中麻黄碱、伪麻黄碱、去甲基麻黄碱、去甲基伪麻黄碱和川芎嗪的含量。     

Comparison of different HPLC systems for analysis of galantamine and lycorine in various species of Amaryllidaceae family

Retention parameters of galantamine and lycorine standards were determined on different columns, i.e., octadecyl silica, SM C18, and strong cation-exchange (SCX) columns with different aqueous mobile phases. Retention of alkaloids was investigated on C18, SM C18 columns with mobile phase containing 5% MeCN, 20% acetate buffer at pH 3.5, and 0.025 ML^?1 diethylamine (DEA), and on SCX column with mobile phase containing 8% MeCN and phosphate buffer at pH 2.5. Better results were also obtained in ion-exchange chromatographic system. On the basis of results obtained in different chromatographic systems, simple, rapid, and sensitive high-performance liquid chromatography methods were developed for determining lycorine and galantamine in plant extracts from various species belonging to Amaryllidaceae family. Extracts were prepared from various parts of plants collected at different times of the growing season.     

高效液相色谱法测定淀粉及淀粉制品中的顺丁烯二酸与顺丁烯二酸酐总含量

建立了基于高效液相色谱(HPLC)测定淀粉及其制品中顺丁烯二酸和顺丁烯二酸酐总含量的方法。通过优化得到最佳样品前处理条件为乙醇体积分数5%,超声时间10 min。色谱分离检测的最佳分析条件为:流动相:甲醇-1‰磷酸(2∶98),色谱柱:Plastisil ODS C18(250 mm×4.6 mm,5μm),检测波长214 nm,流速1.0 mL/min,柱温30℃。该方法对顺丁烯二酸的定量下限为5.0 mg/kg,线性范围为0.25～100 mg/L,相关系数为0.999 7,平均加标回收率为88%～89%,相对标准偏差(n=5)小于2%,能够满足实际检测需要。     

高效液相色谱法同时测定阿达帕林凝胶剂中的主药阿达帕林和两种防腐剂

建立了同时测定阿达帕林凝胶剂中主药阿达帕林和防腐剂(苯氧乙醇、对羟基苯甲酸甲酯)含量的反相高效液相色谱法(HPLC)。采用Tigerkin C18柱(150 mm×4.6 mm,5 μm),以pH 3.0的0.02 mol/L醋酸盐缓冲液-四氢呋喃-乙腈为流动相进行梯度洗脱,在270 nm波长条件下检测。线性范围:苯氧乙醇,10～100 mg/L(r=0.9999);对羟基苯甲酸甲酯,4～40 mg/L(r=0.9999);阿达帕林,4～40 mg/L(r=0.9999)。3种组分的平均回收率为98.0%～98.6%。该方法简便、可靠,可用于阿达帕林凝胶剂中阿达帕林、苯氧乙醇和对羟基苯甲酸甲酯的同时测定。     

Pressurized liquid extraction and anticholinesterase activity-based thin-layer chromatography with bioautography of Amaryllidaceae alkaloids

Modern extraction technique-pressurized liquid extraction (PLE) was optimised for extraction of lycorine and galanthamine (Amaryllidaceae alkaloids) from Narcissus jonquilla ‘Pipit’. Crude extracts were purified on Oasis MCX cartridges, and the alkaloids eluted with 80-100% recoveries using methanol-10% ammonia solution (3:1, v/v). Quantitative results were obtained by both HPTLC-densitometry on silica gel plates and RP-HPLC with diode array (DAD) on XTerra C₁₈ stationary phase. Both methods were fully validated in terms of specificity, precision (including intra- and inter-day measurements), LOD and LOQ values, correlation of UV spectra and linearity of calibration curves. The methods were also well correlated each other with correlation coefficients (r) 0.98823 and 0.99081, respectively, for the mean values of galanthamine and lycorine.Among the investigated solvents methanol and 1% tartaric acid methanolic solution at default conditions (120 °C, p = 60 bar, time: 10 min, one static cycle) permit the highest yields of the total sum of the alkaloids, whereas for toluene the lowest amounts were measured. Lycorine to galanthamine mean ratios were dependant on the type of solvent used, and in toluene galanthamine and related alkaloids were preferably extracted.In temperature experiments for galanthamine, the levels of this compound increased from the temperature of 20 till 150 °C in the investigated solvent systems, then decreased with slight increase from the temperature of 175 to 200 °C in 1% tartaric acid methanolic solution. When lycorine was analysed, similar trends were observed, however the maximum of the concentration was measured at a temperature about 125 °C. The ratios of the mean values of these two compounds differed in temperature-dependant experiments in both solvent systems.Further more, two TLC with bioautography approaches were used in screening for anticholinesterese properties of the extracts. No qualitative differences were found among the different solvent extracts, and AChE inhibition was correlated with galanthamine and related compounds.In conclusion, optimised PLE was much more effective than previously applied hot-solvent extraction, microwave-assisted extraction (MAE) or ultrasound-assisted extraction (USAE).     

Acetylcholinesterase-inhibiting alkaloids from Zephyranthes concolor

The bulbs and aerial parts of Zephyranthes concolor (Lindl.) Benth. & Hook. f. (Amaryllidaceae), an endemic species to Mexico, were found to contain the alkaloids chlidanthine, galanthamine, galanthamine N-oxide, lycorine, galwesine, and epinorgalanthamine. Since currently only partial and low resolution (1)H-NMR data for chlidanthine acetate are available, and none for chlidanthine, its 1D and 2D high resolution (1)H- and (13)C-NMR spectra were recorded. Unambiguous assignations were achieved with HMBC, and HSQC experiments, and its structure was corroborated by X-ray diffraction. Minimum energy conformation for structures of chlidanthine, and its positional isomer galanthamine, were calculated by molecular modelling. Galanthamine is a well known acetylcholinesterase inhibitor; therefore, the isolated alkaloids were tested for this activity. Chlidanthine and galanthamine N-oxide inhibited electric eel acetylcholinesterase (2.4 and 2.6 × 10(-5) M, respectively), indicating they are about five times less potent than galanthamine, while galwesine was inactive at 10(-3) M. Inhibitory activity of HIV-1 replication, and cytotoxicity of the isolated alkaloids were evaluated in human MT-4 cells; however, the alkaloids showed poor activity as compared with standard anti-HIV drugs, but most of them were not cytotoxic.     

高效液相色谱法测定新雪颗粒中栀子苷的含量

建立了新雪颗粒中栀子苷含量的高效液相色谱测定方法。色谱柱为Diamonsil C18(200 mm×4.6 mm i.d., 5 μm), 流动相为乙腈-水(体积比为15∶85),流速1.0 mL/min,检测波长238 nm,进样量20 μL。栀子苷在25~400 mg/L时其浓度与峰面积呈良好的线性关系,方法平均回收率为101.2%,相对标准偏差(RSD)为1.6％。