基于UPLC/Q-TOF-MS分析附子半夏配伍相反的物质基础

利用超高效液相色谱-飞行时间质谱联用技术(UPLC/Q-TOF-MS)分析附子半夏药对配伍相反的物质基础,从化学成分层次阐释其配伍相反机制.基于UPLC/Q-TOF-MS建立附子半夏药对配伍后生物碱类成分的化学指纹图谱,通过主成分分析法和正交偏最小二乘判别法分析药对配伍在合煎过程中的生物碱类成分的含量变化,找出差异变化显著的化学成分.结果表明正离子模式时附子半夏药对合煎液中次乌头碱,中乌头碱,乌头碱,去氧乌头碱,10-OH-中乌头碱,10-OH-乌头碱等的含量明显增高,而中乌头原碱,去乙酸中乌头原碱,去乙酸次乌头原碱,苯甲酰乌头原碱,苯甲酰次乌头原碱,10-OH-苯甲酰中乌头原碱等含量降低.附子半夏药对配伍应用时双酯型二萜生物碱的含量明显增高,而单酯型二萜生物碱的含量明显降低,这可能是附子半夏药对配伍相反作用的物质基础.     

RP-HPLC法测定黑顺片中6种酯型生物碱的含量

建立黑顺片中6种酯型生物碱含量的高效液相测定方法。采用岛津C18色谱柱(250 mm×4.6 mm,5μm),以乙腈–四氢呋喃(体积比为25∶15)为流动相A,以0.1 mol/L乙酸铵溶液(每1 000 m L加冰乙酸0.5 m L)为流动相B梯度洗脱,流量为1.0 ml/min,检测波长为235 nm,柱温为35℃。苯甲酰新乌头原碱、苯甲酰乌头原碱、苯甲酰次乌头原碱、新乌头碱、次乌头碱、乌头碱进样量分别在0.222~4.430,0.060~1.119,0.144~2.878,0.015~0.297,0.025~0.496,0.011~0.229μg范围内与色谱峰面积线性呈良好的线性关系,相关系数r均大于0.999 5,检出限分别为2.10,2.70,2.20,2.60,1.60,3.10 ng,样品在12 h内稳定。苯甲酰新乌头原碱、苯甲酰乌头原碱、苯甲酰次乌头原碱、新乌头碱、次乌头碱、乌头碱的加标回收率分别为98.7%~102.2%,98.6%~102.3%,99.0%~101.9%,102.0%~97.3%,102.1%~98.3%,95.7%~103.5%,测定结果的相对标准偏差分别为2.5%,2.5%,2.2%,2.3%,2.2%,1.7%(n=6)。该方法简便,测定结果准确、可靠,可为黑顺片的质量评价和质量控制提供参考。     

复方中药四逆汤中乌头碱类二萜生物碱的电喷雾串联质谱研究

利用电喷雾串联质谱方法不经柱分离而直接分析复方中药四逆汤中二萜生物碱的组成,乌头碱类二萜生物碱在质谱条件下形成质子化分子,一种生物碱对应一个分子离子峰.检测到了苯甲酰单酯型、双酯型和脂类生物碱等18种二萜生物碱,其中苯甲酰中乌头原碱等单酯型生物碱是四逆汤中的主要生物碱成分.     

大孔树脂对麻黄细辛附子汤生物碱成分静态吸附及解吸附性能研究

采用高效液相色谱法(HPLC)测定麻黄细辛附子(MXF)汤复方水提液中盐酸麻黄碱、盐酸伪麻黄碱、苯甲酰新乌头原碱、苯甲酰乌头原碱、苯甲酰次乌头原碱含量,考察了4种大孔树脂(HPD–100,HPD–722,HPD–400,HPD–600型)对5种生物碱的静态吸附、解吸附性能,筛选出分离纯化麻黄细辛附子汤中生物碱的最佳树脂。结果显示,HPD–722型树脂对5种生物碱的吸附过程稳定且吸附量大,具有较高的解吸附率(87.26%),优于其它3种树脂。HPD–722型大孔树脂适用于富集麻黄细辛附子汤中的生物碱类化合物。     

柱切换液相色谱法快速定量检测参附注射液中乌头碱类生物碱和人参皂苷

利用柱切换液相色谱,建立了参附注射液中苯甲酰新乌头原碱、苯甲酰乌头原碱、苯甲酰次乌头原碱、新乌头碱、次乌头碱和乌头碱6种乌头碱类生物碱,以及Rg1、Re、Rf、Rb1、Rc、Ro、Rb2、Rb3、Rd 9种人参皂苷的分析方法。首先利用强阳离子交换的在线固相小柱选择性富集和净化样品中生物碱类成分,优化了色谱条件;并采用EC－C18柱作为人参皂苷的分析柱,通过优化实验条件,结合柱切换方式,去除了样品中辅料等大极性基质成分对色谱柱的污染,实现了生物碱分析和人参皂苷分析的自动切换。结果显示,样品中的生物碱和人参皂苷分离良好,线性相关系数（r2）均大于0.999,连续进样精密度的相对标准偏差（RSD） < 2.0%,重复性的RSD < 2.0%;其中6种生物碱的平均回收率为95.1%～98.6%,检出限为4.0～8.2 ng/mL;9种人参皂苷的平均回收率为91.7%～104%。所构建的基于柱切换液相色谱技术的在线固相萃取方法能够有效去除样品中的基质干扰,快速完成参附注射液中3种单酯型生物碱和9种人参皂苷的快速定量,同时也可对3种双酯型生物碱进行限量检测,可应用于药物的质量评价。     

离子液体均匀提取-高效液相色谱法测定参附注射液中4种单酯型乌头碱的含量

摘要 采用离子液体均匀提取-高效液相色谱法测定了参附注射液中苯甲酰乌头碱（BA）、苯甲酰新乌头碱（BM）、苯甲酰次乌头碱（BH）和苯甲酰脱氧乌头碱（BD）的含量,通过优化离子液体的用量,样品溶液的pH值,提取时间和盐浓度等参数,确定最佳提取条件为：pH为中性,盐浓度为5%,采用0.058g的[C6Mim][ BF4]和0.26g的 NH4PF6作为提取剂,提取1min,可有效地将目标物从5mL注射液中提取出来。在此实验条件下测得四种单酯型乌头碱的检测限分别为45.5（BM）,19.8(BA),26.3(BH)和12.2(BD)μg L-1,对参附注射液样品进行分析,加样回收率范围在93.74-107.91%,RSD值均小于4.34%。     

离心超滤质谱筛选川乌中与人血清白蛋白相互作用的乌头类生物碱

采用离心超滤质谱技术从川乌总生物碱提取物中筛选与人血清白蛋白相互作用的乌头类生物碱成分,并用LC-MSn技术对筛选出的活性成分进行了分离鉴定.结果表明,从川乌总生物碱中筛选并鉴定出9种与人血清白蛋白存在相互作用的乌头类生物碱:苯甲酰新乌头原碱、苯甲酰乌头原碱、10-OH-中乌头碱、中乌头碱、10-OH-乌头碱、乌头碱、次乌头碱、脱氧乌头碱和3,13-脱氧乌头碱.     

基于最小二乘支持向量机的油页岩含油率近红外光谱分析

为了提高油页岩含油率近红外光谱分析建模的预测精度和稳定性,开展了基于最小二乘支持向量机(LS-SVM)建模方法的对比研究.采用主成分-马氏距离(PCA-MD)和基于蒙特卡洛采样(MCS)2种方法进行了奇异样本的检测,采用径向基核函数的LS-SVM、偏最小二乘(PLS)和反向传播神经网络(BPANN)3种方法进行建模方法对比.结果表明,对于64个油页岩岩芯样本,与PCA-MD方法相比,采用MCS方法剔除奇异样本后所建PLS模型的预测精度提高了28%.对于MCS方法剔除奇异样本后的58个样品,采用KennardStone法划分了44个样品的校正集和14个样品的预测集,采用2阶导数和标准化预处理方法,建立了100个LS-SVM的校正模型,模型的预测决定系数R2平均值达到0.90以上,高于PLS和BPANN模型的对应值;且R2的变化量(0.02)小于BPANN模型的对应值(0.32).因此,MCS奇异样本检测结合LS-SVM方法可提高油页岩含油率样本建模的精度和稳定性.     

乌头碱水解产物的研究

乌头碱为C_(19)二萜类双酯型生物碱,是乌头类中药的主要毒性成分和活性成分,乌头碱具有很强的抗炎和镇痛活性~([1]),在临床上被用于抗肿瘤药物,由于其治疗剂量与中毒剂量接近,必须经炮制后使用.乌头碱水解产物已有报道,但主要集中在苯甲酰乌头原碱和乌头原碱上,焦乌头碱很少提及,本研究利用硅胶柱色谱法对乌头碱的水解产物进行了分离和纯化,利用质谱和核磁共振谱确定了3种主要产物的结构,为研究乌头碱类生物碱的水解产物提供了新的数据.     

近红外光谱快速测定红花逆流提取过程中羟基红花黄色素A的含量

采用近红外光谱(NIRS)透射法对红花罐组式逆流提取过程中羟基红花黄色素A(Hydroxysafflor yellow A,HSYA)的含量进行快速无损的测定.在红花逆流提取过程中,以高效液相色谱法(HPLC)为对照分析方法,测定提取液中羟基红花黄色素A的含量,运用偏最小二乘(PLS)法建立NIR光谱与羟基红花黄色素A的HPLC分析值之间多元校正模型,并对逆流提取过程的未知样本进行含量预测.校正模型相关系数达到0.982,预测相关系数达到0.965,RMSEC和RMSEP分别为0.053和0.075,RSEC和RSEP分别为3.96%和5.25%.结果表明,NIRS可以作为一种准确、快速、无损的检测方法用于检测中药逆流提取过程有效成分含量变化规律.     

液相色谱-质谱法研究附子生物碱的煎煮减毒机理

建立了基于液相色谱-质谱分析的附子中生物碱指纹分析方法,并将其应用于附子煎煮减毒的机理研究。基于质谱特征鉴定了生物碱标准品在加热回流过程中产生的6种降解产物,确定了双酯型生物碱的煎煮条件降解规律。在此基础上研究了附子饮片煎煮不同时间条件下生物碱的指纹变化,结果表明,长时间煎煮可明显降低双酯型生物碱和生物脂碱的浓度,为附子的安全用药提供了科学依据。     

The comparative research on constituents of Radix Aconiti and its processing by HPLC quadrupole TOF-MS

Based upon the regulations stipulated by the State Food and Drug Administration of China, only the processed, detoxified tubers and roots of Aconitum are allowed to be administered orally, used in clinical decoctions and adopted as raw materials for pharmaceutical manufacturing, so the processing principle of preparation of Radix Aconiti is important for ensuring the Radix Aconiti praeparata quality. A simple approach was described for HPLC‐Q‐TOF‐MS screening and identification of many of the aconitine alkaloids present in unprocessed Radix Aconiti and Radix Aconiti praeparata. To compare their fingerprints, the processing principle of preparation of Radix Aconiti was developed. Twenty‐nine compounds and 26 compounds were assigned to aconitine alkaloids and tentatively identified by comparing accurate mass and fragments information with that of the authentic standards or by mass spectrometry analysis and retrieving the reference literature. The nonester alkaloids were almost the same. The diester diterpene alkaloids were decreased, the monoester‐diterpene alkaloids were increased and lipo‐alkaloids decreased obviously in the processing of the preparation. These transformed components could be regarded as potential chemical markers that can be used to distinguish between raw and processed herbs. Copyright © 2012 John Wiley & Sons, Ltd.     

应用遗传算法和PLS的近红外光谱预测玉米中淀粉含量的研究

以普通玉米籽粒为试验材料,在应用遗传算法结合偏最小二乘回归法对近红外光谱数据进行特征波长选择的基础上,应用偏最小二乘回归法建立了特征波长测定玉米籽粒中淀粉含量的校正模型．试验结果表明,基于11个特征波长所建立的校正模型,其校正误差（RMSEC）、交叉检验误差（RMSECV）和预测误差（RMSEP）分别为0.30％、0.35％和0.27％,校正数据集和独立的检验数据集的预测值与实际测定值之间的相关系数分别达到0.9279和0.9390,与全光谱数据所建立的预测模型相比,在预测精度上均有所改善,表明应用遗传算法和PLS进行光谱特征选择,能获得更简单和更好的模型,为玉米籽粒中淀粉含量的近红外测定和红外光谱数据的处理提供了新的方法与途径．     

Rapid determination of yunaconitine and related alkaloids in aconites and aconite‐containing drugs by ultra high‐performance liquid chromatography–tandem mass spectrometry

Yunaconitine (YAC) is a toxic aconite alkaloid that is considered to be a hidden aconite poison since it is frequently found in body fluids from aconite poisoning patients, but has not been well studied in commonly used herbal drugs. In this paper, a rapid and sensitive ultra high‐performance liquid chromatography–tandem mass spectrometry (UHPLC‐MS/MS) detection combined with microwave‐assisted extraction (MAE) was developed for high throughput simultaneous determination of YAC and six other toxic aconite alkaloids in 31 samples of crude, processed aconites and aconite‐containing drugs. The optimized method showed excellent linearity, precision, accuracy and recovery for all target compounds with short run time. YAC was detected in some samples with contents from 0.015 to 10.41 mg/g. This is the first report on the determination of YAC in Radix Aconiti, Radix Aconiti Kusnezoffii and aconite‐containing drugs. This newly developed method facilitates the rapid screening of YAC and related toxic aconite alkaloids and allows YAC to be used as a chemical marker for the quality control of aconites and aconite‐containing drugs. Copyright © 2012 John Wiley & Sons, Ltd.     

Determination of polyphenols in oats by near-infrared spectroscopy (NIRS) and two-dimensional correlation spectroscopy

Two-dimensional correlation spectroscopy (2DCOS) and near-infrared spectroscopy (NIRS) were used to determine the polyphenol content in oat grain. A partial least squares (PLS) algorithm was used to perform the calibration. A total of 116 representative oat samples from four locations in China were prepared and the corresponding near-infrared spectra were measured. Two-dimensional correlation spectroscopy was employed to select wavelength bands for the PLS regression model for the polyphenol determination. The number of PLS components and intervals was optimized according to the coefficients of determination (R²) and root mean square error of cross validation (RMSECV) in the calibration set. The performance of the final model was evaluated using the correlation coefficient (R) and the root mean square error of validation (RMSEV) in the prediction set. The results showed the band corresponding to the optimal calibration model was between 1350 and 1848?nm and the optimal spectral preprocessing combination was second derivative with second smoothing. The optimal regression model was obtained with an R² of 0.8954 and an RMSECV of 0.06651 in the calibration set and R of 0.9614 and RMSEV of 0.04573 in the prediction set. These measurements reveal the calibration model had qualified predictive accuracy. The results demonstrated that the 2DCOS with PLS was a simple and rapid method for the quantitative determination of polyphenols in oats.     

基于偏最小二乘与广义回归神经网络的近红外光谱测定土豆中3种营养成分的研究

偏最小二乘(partial least squares,PLS)与广义回归神经网络(generalized regression neural networks,GRNN)联用对土豆样品建立起粗纤维、淀粉、蛋白质含量的预测校正模型,用PLS法将原始数据压缩为主成份,取前3个主成份的12个特征吸收峰输入GRNN网络,网络光滑因子σi为0.1.PLS-GRNN模型对样品3个组分含量的预测决定系数(R2)分别为: 0.945、 0.992、 0.938.结果表明,近红外光谱技术可以快速、准确地同时测定土豆中的粗纤维、淀粉、蛋白质,该方法可应用于果蔬产业的品质管理与控制.     

利用液质联用技术分析干姜对乌头类生物碱在大鼠肠内吸收的影响

为从吸收的角度考察干姜对乌头类双酯型生物碱的解毒机理, 采用外翻肠囊法展开实验. 利用超高液相与三重四极杆质谱联用技术定量检测双酯型生物碱成分, 采用标准曲线法计算乌头碱、中乌头碱、次乌头碱在肠囊内吸收的绝对含量, 采用质谱峰面积直接分析其它双酯型生物碱的相对变化, 结果加入干姜提取液后, 乌头碱、中乌头碱、次乌头碱的单位肠管面积累计吸收量均降低, 10-羟基中乌头碱的的累积峰面积降低; 加入维拉帕米后, 双酯型生物碱的单位肠管面积累计吸收量及累积峰面积均增加; 向含有地高辛的肠营养液中加入干姜提取液后, 地高辛在各实验时间点的单位肠管面积累计吸收量均降低, 根据以上结果推测干姜抑制乌头类双酯型生物碱在大鼠肠囊内吸收的可能机制是通过诱导肠内P-葡糖蛋白, 从而抑制作为P-葡糖蛋白底物的双酯型生物碱的吸收, 最终起到减毒作用.     

偏最小二乘近红外光谱法测定瘦肉脂肪酸组成的研究

利用偏最小二乘将瘦肉的近红外光谱数据分别与其棕榈酸、棕榈油酸、硬脂酸、油酸、亚油酸含量建立校正模型,并用交互校验和外部检验来考查模型的可靠性.各脂肪酸模型的校正相关系数分别为0.9998、0.9844、0.9963、0.9754、0.9969,均方估计残差(RMSEC)分别为0.0231、0.0485、0.111、0.373、0.311,交互校验均方残差(RMSECV)分别为0.509、0.115、0.225、0.848、0.649.应用所建立的各脂肪酸近红外模型对瘦肉脂肪酸组成进行预测,并对各脂肪酸的预测值与气相色谱法测定值进行配对t-检验,结果表明两者差异均不显著(p>0.05).     

Averaged and weighted average partial least squares

Two alternative partial least squares (PLS) methods, averaged PLS and weighted average PLS, are proposed and compared with the classical PLS in terms of root mean square error of prediction (RMSEP) for three real data sets. These methods compute the (weighted) average of PLS models with different complexity. The prediction abilities of the alternative methods are comparable to that of the classical PLS but they do not require to determine how many components should be included in the model. They are also more robust in the sense that the quality of prediction depends less on a good choice of the number of components to be included. In addition, weighted average PLS is also compared with the weighted average part of LOCAL, a published method that also applies weighted average PLS, with however an entirely different weighting scheme.     

短波近红外光谱法对蛇床子SFE萃取产物的定量分析

利用中药蛇床子CO2超临界萃取(SFE)的萃取物的短波近红外漫反射光谱(800～1100 nm),以HPLC分析值作参比值,采用化学计量学中的偏最小二乘法(PLS)建立短波近红外漫反射光谱与蛇床子SFE萃取物中主要成分蛇床子素和欧前胡素间定量分析数学模型.实现了快速、无损的测定双组分中药的有效成分.讨论了光谱的预处理方法和主成分数对PLS定量预测蛇床子萃取物中蛇床子素和欧前胡素含量能力的影响,并对预测集样品进行预测.