RETRACTED ARTICLE: The retention behavior of ginsenosides in HPLC and its application to quality assessment of Radix Ginseng

This study systematically investigated the retention behavior of seven neutral ginsenosides Rg₁, Re, Rf, Rb₁, Rb₂, Rc, Rd, and an acidic ginsenoside R₀, the major pharmacologically active components of Radix Ginseng with RP-HPLC. The effects of solvent, pH value, ionic strength of the mobile phase, and column temperature were investigated using an octadecylsiloxanebonded silica gel column. Based on the ginsenosides’ retention characteristics, the concentration of acetonitrile and the gradient of the mobile phase needed to maintain the baseline separation of the major neutral ginsenosides in Radix Ginseng were theoretically predicted. Furthermore, the ionic strength of mobile-phase necessary to achieve good resolution of the neutral ginsenosides and acidic ginsenosides was carefully investigated. According to the results of the quantitative analysis of ginsenosides in eight batches of ginseng samples from different sources, the developed HPLC technique may be a valuable tool for the quality assessment of Radix Ginseng. This article was retracted as it was printed by error in this issue of APR.     

Analysis of Fuzhisan and quantitation of baicalin and ginsenoside Rb<Subscript>1</Subscript> by HPLC-DAD-ELSD

Fuzhisan (FZS) is a traditional Chinese medicine composed of Radix Ginseng Rubra (Araliaceae family), Scutellaria baicalensis Georgi (Labiatae family), Angelica sinensis (Oliv) Diels (Umbelliferae family), Anemone altaica Fisch. Ex C.A. Mey (Araceae family) and Glycyrrhiza uralensis (Leguminosae family). To establish the chemical fingerprint of the components of FZS and quantify the components, baicalin and ginsenoside Rb₁, a high performance liquid chromatography method coupled with diode array and evaporative light scattering detectors (DAD-ELSD) was developed. Separation of 36 components from 12 batches of FZS was performed on a C₁₈ column, with a mobile phase consisting of acetonitrile and 0.1% acetic acid-water, with gradient elution at a column temperature of 30°C. The optimum detection wavelength was set at 335 nm, the drift tube temperature of ELSD was set at 80°C, the carrier gas pressure was 25 psi, and the gain = 10. The similarity among 12 batches of FZS was over 0.95. Five constituents of FZS, namely baicalin, ferulic acid, and ginsenosides Rg₁, Re, and Rb₁, were identified based on their retention times (RT). Calibration curves for baicalin and ginsenoside Rb1 showed good linearity (r ² > 0.9992); recoveries ranged from 95% to 99%. This quantification method is reproducible and simple, and may provide a tool to assess the quality of FZS.     

基于UHPLC-HRMS/MS的鲜人参不同部位中皂苷类成分差异分析

目的 采用UHPLC-HRMS/MS对鲜人参中的三萜皂苷类成分进行分析鉴定,并阐明鲜人参不同部位(主根、侧根、须根和芦头)所含的三萜皂苷类成分的差异性。方法 以水饱和正丁醇为提取溶剂,以水-乙腈为流动相进行梯度洗脱,在ESI负离子模式下建立新鲜人参药材中皂苷类成分的UHPLC-Q-Exactive Orbitrap MS/MS分析表征方法,并采集各样品的高分辨质谱数据。结合色谱保留行为、精确分子量、多级质谱裂解规律及对照品比对等,鉴定各样品中的人参皂苷类成分;依据已鉴定的人参皂苷色谱峰的相对丰度,筛选差异色谱峰,阐明鲜人参各部位差异。结果 从鲜人参不同部位中共检测到68种差异皂苷类成分,其中主根有44种,侧根有47种,须根有52种,芦头有37种。结论 鲜人参不同部位中皂苷类成分差异较大;人参侧根、芦头等非主要药用部位中含有多种人参皂苷,且存在其他部位未检测到的成分,可作为药物研发的重要原料及多种人参皂苷的提取原料。     

HPLC法测定石柱参药材及其片剂中5种人参皂苷的含量

目的建立测定石柱参药材及其片剂中5种人参皂苷含量的方法,为石柱参的质量控制提供科学依据。方法色谱柱为Kromasil C18柱;流动相为乙腈-水,梯度洗脱;柱温为25℃;流速为1.0 mL.min-1;检测波长为203 nm。结果5种人参皂苷的色谱峰与相邻色谱峰分离良好。人参皂苷Rg1质量浓度在19.8~198 mg.L-1内、人参皂苷Re质量浓度在20.6~206 mg.L-1内、人参皂苷Rb1质量浓度在33.0~330 mg.L-1内、人参皂苷Rc质量浓度在18.0~180 mg.L-1内、人参皂苷Rb2质量浓度在13.0~130 mg.L-1内与峰面积呈良好的线性关系。石柱参药材中人参皂苷Rg1、Re、Rb1、Rc、Rb2的平均回收率分别为99.8%、98.3%、99.2%、95.4%、96.8%,RSD分别为3.0%、3.0%、2.6%、2.2%、2.8%(n=6);石柱参片剂中人参皂苷Rg1、Re、Rb1、Rc、Rb2的平均回收率分别为100.2%、99.0%、99.7%、96.4%、98.4%,RSD分别为2.2%、3.1%、3.6%、2.6%、2.8%(n=6)。结论本实验中所建立的方法可用于石柱参药材及其片剂的质量控制。     

人参的抗癌作用及其机制研究进展

《中草药》杂志是由中国药学会和天津药物研究院共同主办的国家级期刊,月刊,国内外公开发行。本刊创始于1970年1月,1992年荣获首届全国优秀科技期刊评比一等奖;2001年荣获中国期刊方阵"双奖期刊";2003年荣获第二届     

基于近红外光谱分析技术的注射用益气复脉(冻干)红参醇提过程在线监测

目的 利用近红外光谱分析技术建立注射用益气复脉（冻干）主要原料红参醇提过程中3种单体皂苷——人参皂苷Rg₁、Re和Rb₁的定量模型,实现提取过程中关键指标的快速检测。方法 在线采集红参醇提过程的近红外光谱,以超高效液相色谱（UPLC）法测定提取过程药液中人参皂苷Rg₁、Re和Rb₁的量为参考值,采用偏最小二乘法建立光谱与测定值之间的定量校正模型,进而对提取过程进行在线分析。结果 人参皂苷Rg₁和Re的建模波段均为9 403.7～7 498.3 cm^-1和6 102～5 446.3 cm^-1组合波段;人参皂苷Rb₁的建模波段为5 774.1～5 446.3 cm^-1。人参皂苷Rg₁、Re、Rb₁定量模型的交叉验证决定系数（R²）分别为99.40、99.44、99.41,交叉验证均方根误差分别为5.18、2.77、11.00。结论 所建立的3种单体皂苷定量模型预测性能良好,能够有效测定红参醇提过程中人参皂苷Rg₁、Re和Rb₁的量。     

人参皂苷水溶液热稳定性研究

目的 研究加热时间对人参皂苷水溶液的影响及受热后人参皂苷含量的变化情况。方法 红参须浓缩液加热不同时间,采用高效液相色谱法测定其人参皂苷Rg1、Re、Rb1、Rc和Rd的含量。结果 5种人参皂苷在加热6 h内,发生不同程度的降解反应,二醇类人参皂苷Rb1、Rc和Rd在加热2-3 h时,含量呈明显下降趋势,人参皂苷Rd降解速率最慢。三醇类人参皂苷Rg1和Re在加热3 h内含量快速下降,3 h后趋于平缓。结论 在常压受热条件下,人参皂苷水溶液主要成分人参皂苷Rg1、Re、Rb1、Rc和Rd的含量,随加热时间延长而不断下降,3 h后下降速率减缓。三醇类人参皂苷较二醇类对热更为敏感。     

Biomass and content of ginsenosides and polyacetylenes in American ginseng roots can be increased without affecting the profile of bioactive compounds

Fifty selected roots from a 7-year-old American ginseng (Panax quinquefolium L.) plant population grown in Denmark, with root weights varying from 191 to 490 g fresh weight (FW), were investigated for bioactive ginsenosides and polyacetylenes (PAs) in order to determine the correlation between the content of ginsenosides and PAs and root FW. PAs (falcarinol, panaxydol) and ginsenosides (Rb₁, Rb₂, Rb₃, Rc, Rd, Re, Rg₁) were extracted from roots by sequential extraction with ethyl acetate and 80% methanol, respectively, and quantified in extracts by reverse-phase high-performance liquid chromatography (HPLC) using photodiode array detection. Total concentrations of PAs and ginsenosides varied between 150 and 780 mg/kg FW and 5,920 and 15,660 mg/kg FW, respectively. No correlation existed between the content of ginsenosides and PAs and root FW or between the total concentration of ginsenosides and PAs. Strong significant correlation was found between total content of ginsenosides and ginsenoside Rb₁ (r = 0.8190, P < 0.0001) and between total content of PAs and falcarinol (r = 0.9904, P < 0.0001). Based on the results of this study, it was concluded that it is possible to select large American ginseng roots for increased biomass production and concentration of bioactive ginsenosides and PAs without affecting the profile of bioactive compounds. Ginsenoside Rb₁ and falcarinol were found to be important selection parameters for identifying superior genotypes with the highest content of bioactive compounds.     

Comparative analysis of ginsenosides in different root parts and cultured cells from Chinese ginseng

HPLC analysis of extracts of different parts of two six-year-old Panax ginseng (Ginseng & Tobacco Company, South Korea) plantation roots demonstrated the presence of seven detectable ginsenosides (Rb₁, Rb₂, Rc, Rd, Rf, Rg₁, and Re). The total contents were 1.16 and 0.64% for main roots, 4.00 and 4.60% for rhizomes, 6.79, 4.38% for side roots (d = 0.4–0.5 cm), and 13.26 and 11.37 for fine roots (d = 0.1–0.15 cm). Four new ginseng callus lines were originated from side and fine roots. HPLC analysis of the qualitative and quantitative composition of the biomass after the first year of cultivation showed that all lines contained the full spectrum of ginsenosides (Rb₁, Rb₂, Rc, Rd, Rf, Rg₁, and Re), the total content of which varied from 0.2 to 0.4%. HPLC monitoring of these callus lines over five years of cultivation demonstrated gradual disappearance of glycosides Rb₂, Rc, Rd, and Rf from the ginsenoside spectrum. Loose calluses growing in medium containing 0.5 and 0.2 mg-liter 6-benzylaminopurine were used to start suspension cultures L-1 and L-2, respectively. HPLC analysis of the qualitative and quantitative composition of glycosides in suspension cultures showed that both lines contained only ginsenosides Rb₁, Rg₁, and Re, with a predominance of R_e ginsenoside. __________ Translated from Khimiko-Farmatsevticheskii Zhurnal, Vol. 42, No. 1, pp. 33–38, January, 2008.     

一测多评法同步测定人参和三七药材中多种人参皂苷的含量

通过建立人参皂苷Rb₁与其他8种皂苷间的紫外相对校正因子(RCF)，实现只用一个对照品测定人参和三七药材中多个人参皂苷类成分的含量，以解决人参等药材质量控制中，对照品供应不足问题。结果表明，在一定的线性范围内，人参皂苷Rb₁与Rg₁、 Re、 Rf、 Rh₁、 Rc、 Rb₂、 Rb₃、 Rd间的RCF值分别为1.400， 1.215， 1.517， 1.801， 0.944， 1.012， 1.143， 1.135，且在不同实验条件下重现性良好(RSD=0.30%～3.9%)。本方法只需测定人参和三七药材中Rb₁的含量，其余人参皂苷含量由其RCF值计算得到，实现一测多评；并与常规外标法比较，两种药材中一测多评法与外标法所得结果均无显著性差异；所建立的校正因子可同时用于人参及三七药材及其相关产品的定量分析及质量评价。     

一些酸性药物在正相硅胶/反相洗脱色谱中保留机理研究

用液相色谱法中“溶质计量置换保留模型”(SDM-R)的两个线性公式,分析了正相硅胶/反相洗脱色谱体系中酸性药物分子与固定相和流动相分子间的相互作用机理,研究了流动相pH值、离子强度、温度和强溶剂浓度改变对溶质保留的影响。从而阐明了酸性药物的保留机理为一种以反相色谱为主的与正相色谱共存的混合保留机理。此方法可以作为药物制剂和中药材中酸性药物成分的分离测定方法。     

人参的分析——Ⅳ.人参皂甙的高效液相色谱测定

本文报道了人参中六种主要皂甙(Rb₁,Rb₂,Rc,Rd,Re,Rg)的HPLC测定法,用氨基键合相柱,示差折光检测,流动相为甲醇中乙腈—乙二醇—醋酸铵(0.14 mol/L)(30:70:5:10.6),同时还改进了纯化方法,此法快速、重现性好,与薄层比色法比较结果基本一致。     

基于近红外光谱技术的红参多指标成分快速分析

目的 利用近红外光谱（near infrared spectroscopy,NIR）技术对红参药材中的水分、人参皂苷和糖类成分进行快速分析,实现红参药材质量的快速评价。方法 用62批红参样品作为校正集,分别建立水分、人参皂苷和糖类成分的定量模型,对模型进行系统的优化,并用另外38批样品作为验证集对建立的模型进行验证和评价。结果 所建的模型对红参水分含量、人参皂苷Rg₁和Re总含量、人参皂苷Rb₁含量及麦芽糖和蔗糖总含量均有较好的预测能力,预测相对偏差均<15%,所建NIR快速检测方法的精密度和重复性结果均较好。结论 该方法可以对红参药材的关键质控指标进行快速检测,结果可靠,是对传统分析方法的有效补充,可以有效地指导后道工序的生产,从而避免因药材质量不合格而造成不必要的浪费。     

Importance of injection solution composition for LC-MS-MS methods

For the first time, the influence of the injection solution composition on the quality of LC–MS–MS methods, in terms of column efficiency and peak shape, was systematically investigated. Various types of compounds, including polar ionic acidic, polar ionic basic and non-polar neutral compounds, were prepared in different solutions ranging from 100% water to 100% acetonitrile. Different volumes of these solutions were injected onto either C₁₈ or silica columns connected to tandem mass spectrometry. The mobile phases consisted of acetonitrile, water, and small amounts of volatile acid or buffer. On silica columns, the influence of injection solution on the peak shape and column efficiency was straightforward. The sharpest peaks and the highest column efficiency were obtained with 100% acetonitrile as the injection solvent. On C₁₈ columns, this type of influence was less clear due to the dual retention mechanism of the bonded phase and of the residual silanol groups. On C₁₈ column, retention due to residual silanol groups was significant even with a mobile phase containing less than 50% acetonitrile. Poor peak shape was observed when the injection solution had a stronger eluting strength than mobile phase, particularly for early eluting peaks.     

不同参类中人参皂苷Rg_1、Re、Rb_1的质量测定

目的:建立超高效液相色谱法(UPLC)同时测定人参、红参、西洋参中人参皂苷Rg1、Re、Rb1质量方法。方法:采用AcquityUPLCBEHC18色谱柱(2.1mm×100mm,1.7μm),以乙腈-水为流动相进行梯度洗脱,流速为0.5mL/min,检测波长为203nm,柱温为40℃。结果:人参、红参、西洋参中人参皂苷Rg1、Re、Rb1在14min内得到完全分离并进行了质量测定。结论:应用UPLC法能够同时测定人参、红参、西洋参中人参皂苷Rg1、Re、Rb1质量,大大提高了人参皂苷类成分的分析速度,减少了溶剂消耗,为人参皂苷类成分质量测定提供了参考。     

Change of the content of chemical constituents and anti-oxidative action of the decoction of radix ginseng combined with Flos Lonicerae, radix Polygoni Multiflori and Radix Astragali

Ginsenosides in the decoction of Radix Ginseng, Radix Ginseng with Flos Lonicerae, Radix Polygoni Multiflori or Radix Astragali have been investigated by high performance liquid chromatography (HPLC) and electrospray ionization mass spectrometric method (ESI-MS). Change of the content of ginsenosides was nonlinear in diverse combinative proportion of Radix Ginseng with Flos Lonicerae, while the stripping of ginsenosides was promoted by a small amount of Radix Polygoni Multiflori. In the combinative decoction of Radix Ginseng with Radix Astragali, ginsenosides contents were increased compared to single decoction of Radix Ginseng. Besides, ferric reducing antioxidant power (FRAP) method was developed for determination of the total antioxidative activity of n-butanol and water-soluble extracts from the decoction. The experimental results showed that antioxidative activity was better in the combinative decoction than that in single decoction, and the FRAP values of n-butanol extract were also greater compared with that of water extract.     

Drug-biomacromolecule interaction V

The binding properties of three ginsenosides, Rb₁, Rc and Re, to bovine and human serum albumins have been examined by fluorescence probe technique. 1-anilinonaphthalene-8-sulfonate (ANS) was used as the fluorescence probe. Protopanaxatriol glycoside, Re, did not quench the fluorescence of ANS to the bovine serum albumin. Competitive bindings between protopanaxadiol glycosides, Rb₁ and Rc, and ANS were observed. The numbers of binding sites of bovine serum albumin for Rb₁ and Rc are both 3.3. The binding constants for Rb₁ and Rc with bovine serum albumin were 1.91×10⁴M^?1 and 1.04×10⁴M^?1, respectively. The ginsenosides, Rb₁, Rc and Re did not quench the fluorescence of ANS bound to human serum albumin.     

Effects of the Ginsenosides Rg1 and Rb1 on Morphine-induced Hyperactivity and Reinforcement in Mice

Recent studies have demonstrated that ginseng saponin inhibits the hyperactivity and conditioned place-preference response induced by psychostimulants and opiates. This seems to occur by direct or indirect modulation of dopaminergic activity. However, it is not known which components of ginseng saponin are active. These experiments were conducted to determine the effects of the ginsenosides Rb₁ and Rg₁, major components of the protopanaxadiol and protopanaxatriol fractions of ginseng saponin, on morphine-induced hyperactivity and conditioned place-preference. Morphine-induced hyperactivity, but not apomorphine-induced climbing behaviour, was inhibited by both Rb₁ and Rg₁. These findings confirm the hypothesis that ginsenosides modulate catecholaminergic activity preferentially at pre-synaptic sites. Morphine-induced conditioned place-preference was inhibited by Rg₁, but not by Rb₁. It has previously been shown that at low doses Rb₁ and Rg₁ are equally effective at inhibition of catecholamine secretion at the pre-synaptic site, but that at high doses Rg₁ is a more effective inhibitor. This observation might explain our finding that morphine-induced conditioned place-preference was inhibited by Rg₁ only. Our findings suggest that Rg₁, a component of ginseng saponin with appropriate activity, might be a useful agent for prevention and treatment of the adverse effects of morphine.     

UPLC法评价硫磺熏蒸对西洋参皂苷类成分的影响

目的：评价硫磺熏蒸对西洋参皂苷类成分的影响。方法：建立超高效液相色谱法,测定并比较分析硫磺熏蒸前后西洋参中人参皂苷Rg₁、Re和Rb₁的含量。结果：硫磺熏蒸后西洋参中人参皂苷Rg₁、Re和Rb₁总量仍符合《中国药典》规定;但当硫磺熏蒸致二氧化硫残留量大于400 mg·kg^-1时,人参皂苷Re、Rb₁的含量及人参皂苷Rg₁、Re和Rb₁三者的总量显著降低。当二氧化硫残留量不大于150 mg·kg^-1时,人参皂苷Rg₁、Re和Rb₁的含量及其总量基本不受影响。结论：《中国药典》规定的西洋参二氧化硫残留量不得大于150 mg·kg^-1有其科学合理性,硫磺过度熏蒸西洋参（二氧化硫残留量大于400mg·kg^-1）对西洋参中皂苷类成分的含量有显著影响。