高效液相色谱法测定皱皮木瓜饮片中齐墩果酸和熊果酸

建立高效液相色谱法测定皱皮木瓜饮片中齐墩果酸、熊果酸含量的方法.选用Kromasil C18色谱柱(250 mm×4.6 mm,5μm)为分离柱,流动相为甲醇–0.3％磷酸溶液(体积比为87:13),流量为1.0 mL/min,柱温为25℃,检测波长为210 nm.齐墩果酸、熊果酸的质量浓度分别在65.76～205.5...     

固相萃取-高效液相色谱法测定枣中熊果酸和齐墩果酸

采用正交设计方法优化枣中熊果酸和齐墩果酸的提取工艺,并建立熊果酸和齐墩果酸的固相萃取-高效液相色谱测定方法.样品经乙醇提取,减压浓缩,C18固相萃取小柱净化后,采用高效液相色谱方法分离,以甲醇-磷酸盐溶液作流动相,等度洗脱,紫外检测器检测.齐墩果酸和熊果酸在0.01～1.0mg/mL浓度范围内与峰面积线性良好,线性相关系数分别为0.9998和0.9999.样品的平均加标回收率为95.1%～98.8%.方法的检出限为1.0～3.7μg/mL(S/N=3),同一样品测定结果的相对标准偏差小于2.0%(n=7).方法具有简单、快速,灵敏、可靠的特点,适用于枣及枣类产品中熊果酸和齐墩果酸含量的测定.     

圆萼刺参中的芦丁,熊果酸和齐墩果酸的定性定量分析

采用薄层色谱法(TLC)和反相高效液相色谱法(RP-HPLC)对藏药材圆萼刺参中的芦丁,熊果酸和齐墩果酸进行定性和定量的分析。定性分析:TLC法检定芦丁和熊果酸和齐墩果酸成分,薄层色谱条件是以V(乙酸乙酯)∶V(丁酮)∶V(甲酸)∶V(水)=10∶6∶1∶2为展开剂,喷以10 g/L NaNO2的1%甲醇溶液在105℃检定芦丁,以V(CHCl3)∶V(乙酸乙酯)=1∶1为展开剂,喷以V(H2SO4)∶V(甲醇)=1∶2溶液在105℃检定熊果酸和齐墩果酸。定量分析:RP-HPLC法测定圆萼刺参中芦丁,流动相:V(甲醇)∶V(0.4%H3PO4溶液)=38∶62;检测波长340 nm;RP-HPLC法测定熊果酸和齐墩果酸,流动相:V(甲醇)∶V(0.2%H3PO4)溶液=85∶15;检测波长215 nm;圆萼刺参中的齐墩果酸,在本种植物中首次发现。     

强阳离子交换色谱分离多肽混合物的条件优化

以酵母全蛋白提取液的胰蛋白酶切产物为研究对象,对多维色谱分离中常用的强阳离子交换色谱的分离条件,包括上样量、盐的种类、调节缓冲液pH值的酸的种类及有机溶剂的比例进行了考察和比较。结果表明,在强阳离子交换色谱分离线性上样量范围内,在氯化铵溶液作为流动相,用磷酸调节流动相缓冲液pH值至2.7,且流动相中乙腈的体积分数为30%条件下进行梯度洗脱时,可获得最佳的分离结果。本结果可为采用二维色谱-质谱策略分析多肽混合物时的第一维强阳离子交换色谱分离条件的选择提供有益的参考。     

UPLC-MS/MS法测定中药材中齐墩果酸与熊果酸的含量

建立了一种同时检测中药材中齐墩果酸(OA)和熊果酸(UA)含量的超高效液相色谱串联质谱方法。采用超高效液相色谱-三级四极杆质谱(UPLC-TQMS)法对样品进行测定,Waters Acquity UPLC BEH C_(18)色谱柱(50 mm×2.1 mm,1.7μm)进行分离,以乙腈-5 mmol/L乙酸铵水溶液(氨水调至pH 9.24)为流动相,梯度洗脱;负离子模式下检测。结果表明,齐墩果酸和熊果酸在0.5~50.0 ng/mL浓度范围内线性关系良好,相关系数(r~2)分别为0.999 8和0.999 7;检出限(S/N=3)分别为0.006 6,0.012 8 ng/mL,定量下限(S/N=10)分别为0.002 0,0.003 8 ng/mL;对OA和UA进行加标回收实验,平均回收率分别为101.1%和100.8%,相对标准偏差(RSD,n=9)分别为1.8%和0.04%。对10种不同中药材中齐墩果酸和熊果酸含量进行检测,结果表明该方法快速简便、准确度高、重现性好,可用于含有齐墩果酸和熊果酸的中药材含量测定。     

毛细管胶束电动色谱法测定夏枯草中两种有效成分

建立了毛细管胶束电动色谱法（ＭＥＫＣ）同时分离和测定夏枯草中齐墩果酸和熊果酸异构体含量的新方法。以１５ｍｍｏｌ／Ｌ磷酸氢二钠（Ｎａ２ＨＰＯ４）、１５ｍｍｏｌ／Ｌ硼砂（Ｎａ２Ｂ４Ｏ７）、１０ｍｍｏｌ／Ｌ十二烷基硫酸钠（ＳＤＳ）、５％（Ｖ／Ｖ）乙醇的缓冲溶液作为运行电解质溶液,在２０ｍｉｎ实现了齐墩果酸和熊果酸的基线分离。同时也研究了ＳＤＳ浓度和有机溶剂（乙醇）浓度对待测物质分离的影响。研究了不同提取     

酸性药物在微乳液相色谱体系中的保留行为

以酸性药物萘普生、布洛芬、双氯芬酸、苯丙氨酸和2,4-二氯苯氧基乙酸为研究对象,探讨了酸性药物在微乳液相色谱体系中保留机理,建立了酸性药物的容量因子(k′)与表面活性剂浓度、助表面活性剂浓度、亲脂性有机溶剂浓度以及流动相pH值之间的相关模型。结果表明,表面活性剂、助表面活性剂和亲脂性有机溶剂的浓度以及流动相pH值的变化对酸性药物的容量因子的影响完全与理论模型相吻合。所建立的保留模型能较好地反映在微乳液相色谱体系中微乳液组成和pH值对酸性药物保留行为的影响。     

中性溶质在反相毛细管电色谱中的保留行为

分别在以甲醇、乙腈、异丙醇和四氢呋喃为有机改性剂的４种二元流动相体系中对中性溶质在反相毛细管电色谱中的保留行为进行了研究。不仅考察了有机改性剂的种类和浓度对电渗淌度的影响,而且建立了溶质容量因子与有机改性剂在流动相中体积分数间的定量关系,此外还对样品在反相毛细管电色谱和反相毛细管高效液相色谱中的保留行为进行了比较,发现中性溶质在这两种分离模式中的容量因子基本相同。     

在18—冠—6硅胶键合固定相上有机酸及其异构体的高效液相色谱研究

本文报导了一种新型18—冠—6硅胶键合固定相(P18—C—6SB)在有机酸分析中的应用。流动相组成为Cu~(2+)离子、甲醇和HAC—NaAc缓冲溶液。研究了流动相中甲醇体积比、Cu~(2+)离子浓度、缓冲溶液浓度及pH值等因素对容量因子(K’)的影响。在优化条件下,成功地分离了包括氨基取代苯甲酸异构体在内的多种芳香酸。     

羟丙基-β-环糊精修饰毛细管胶束电动色谱法测定夏枯草中的齐墩果酸和熊果酸

建立了羟丙基-β-环糊精修饰毛细管胶束电动色谱同时分离测定夏枯草中的齐墩果酸和熊果酸的方法。以pH 9.0的10 mmol/L Na2B4O7、10 mmol/LNaH2PO4、50 mmol/L十二烷基硫酸钠（SDS）、15 mmol/L羟丙基-β-环糊精（HP-β-CD）、体积分数6%甲醇作为背景电解质溶液,15 min内实现了两种待测组分的基线分离。齐墩果酸和熊果酸分别在22.5-360 mg/L、48.8-780 mg/L范围内线性关系良好,检出限分别为3.2 mg/L和3.8 mg/L。两者的峰面积和迁移时间的相对标准偏差分别为2.2%、0.5%和2.6%、0.6%,回收率分别在93.8%-102.1%和96.7%-105.2%之间。     

Influence of mobile phase acid-base equilibria on the chromatographic behaviour of protolytic compounds

A review about the influence of mobile phase acid-base equilibria on the liquid chromatography retention of protolytic analytes with acid-base properties is presented. The general equations that relate retention to mobile phase pH are derived and the different procedures to measure the pH of the mobile phase are explained. These procedures lead to different pH scales and the relationships between these scales are presented. IUPAC rules for nomenclature of the different pH are also presented. Proposed literature buffers for pH standardization in chromatographic mobile phases are reviewed too. Since relationships between analyte retention and mobile phase pH depends also on the pKa value of the analyte, the solute pKa data in water-organic solvent mixtures more commonly used as chromatographic mobile phase are also reviewed. The solvent properties that produce variation of the pKa values with solvent composition are discussed. Chromatographic examples of the results obtained with the different procedures for pH measurement are presented too. Application to the determination of aqueous pKa values from chromatographic retention data is also critically discussed.     

Rapid analysis of triterpenic acids by liquid chromatography using porous graphitic carbon and evaporative light scattering detection

An original system which uses Porous Graphitic Carbon as support and a mixture of organic solvents as mobile phase is proposed for the analysis of triterpenic acids by liquid chromatography. The separation of betulinic acid, ursolic acid, oleanolic acid, and 18alpha- and 18beta-glycyrrhetinic acids was carried out within a short time and monitored by evaporative light scattering detection as universal detection method. Molecular modelling studies show that the main contribution to the selectivity comes from the electrostatic interaction characterised by the dipole moment of the products.     

Retention behavior of o-phthalic, 3-nitrophthalic, and 4-nitrophthalic acids in ion-suppression reversed-phase high performance liquid chromatography using acids instead of buffers as ion-suppressors

In reversed-phase high performance liquid chromatography, the logarithm of the retention factor, log k, is usually correlated with the logarithm of the octanol-water partition coefficient, log Kow. The k and Kow of an ionizable analyte are greatly influenced by the mobile phase pH. In this paper, log kw of diprotic o-phthalic, 3-nitrophthalic, and 4-nitrophthalic acids, are obtained by extrapolation to pure aqueous fraction of mobile phase in ion-suppression reversed-phase high performance liquid chromatography with acetic acid and perchloric acid as the ion-suppressors. The Kow values of the three analytes are calibrated according to the apparent octanol-water partition coefficient, Kow, under different pH conditions, and the log K"ow values show a much better correlation with log kw than do log Kow. The influences of two ion-suppressors, acetic and perchloric acids, on the retention behavior of these diprotic acids at different pH are contrasted. An abnormal trend is found in the k vs. pHw plot of the acetic acid system when the methanol content is low. A possible reason is that acetic acid is an even stronger organic modifier than methanol, besides being an ion-suppressor. The results make the selection of mobile phase for the separation of acidic compounds by ion-suppression reversed-phase high performance liquid chromatography direct, accurate, and practical.     

Determination of oleanolic and ursolic acid in Chinese herbs using HPLC and γ-CD as mobile phase modifier

To separate and determine oleanolic acid and ursolic acid, a rapid and accurate HPLC using γ‐CD as the mobile phase additive was developed. The effect of CD nature and concentration, and the acidity of the mobile phase on the chromatographic behavior of two bioactive triterpenes were systematically studied. Two bioactive triterpenes were completely separated (R = 3.11) on a Kromasil^® C₁₈ column (150×4.6 mm id, 5 μm) with the mobile phase consisting of acetonitrile/0.1% phosphoric acid with 2 mM γ‐CD as the mobile phase modifier (60:40, v/v). The flow rate was set at 1.0 mL/min and the eluent was detected at 210 nm for two bioactive triterpenes. The linearity of the method was excellent (r=0.9999) over the studied range of 6–300 μg/mL for oleanolic acid, and 12–600 μg/mL for ursolic acid. The LOD and LOQ were 1.5 and 5.0, 1.0 and 3.0 μg/mL for oleanolic acid and ursolic acid, respectively. The optimized method was successfully applied to separate and determine two bioactive triterpenes in five Chinese herbs. It is concluded that this method could be used for rapid and accurate qualitative and quantitative analysis of the two bioactive triterpenes in Chinese herbs.     

pH dependence of the hydrophobicity of beta-blocker amine compounds measured by counter-current chromatography

The octanol-water partition coefficients (Poct) of 17 antiadrenergic beta-blocker compounds were determined by counter-current chromatography (CCC). Since CCC uses a biphasic liquid system, the octanol-water liquid system was used with essentially an octanol stationary phase and aqueous buffer mobile phase. The Poct coefficients were obtained directly without any extrapolation. The measured Poct values were in the 0.0015-4070 range (-2.8 < log Poct < 3.6). Since the beta-blocking agents are ionizable compounds, the Poct values obtained were strongly dependent on the aqueous-phase pH. The apparent Poct coefficients of the beta-blockers were determined at three different pH values (approximately 3, 7 and 11) using 0.01 M ammonium phosphate buffers saturated with octanol. A model allowed us to obtain the molecular and ionic Poct value using the solute pKa with these three experimental octanol-water coefficients. Often, the Poct coefficients of the molecular forms obtained with the CCC method differ significantly from computed literature values and/or experimental values obtained by extrapolation. Relationships between biological properties and hydrophobicity were also examined.     

Effect of mobile phase composition, pH and buffer type on the retention of ionizable compounds in reversed-phase liquid chromatography: application to method development

Optimizing separation of ionizable compounds in order to find robust conditions has become an important part of method development in liquid chromatography. This work is an attempt to explain the observed variations of retention of acid and basic compounds with the organic modifier content in the mobile phase, according to various factors: the type of modifier, the type of buffer, the temperature and of course the type of solute. This is done by considering the variation of the so-called chromatographic pKa which refers to the pH measured in the aqueous medium and is determined from retention data. A procedure is described that accurately relates, from nine experiments, retention to solvent composition and pH. The limits of such a procedure are evaluated and two examples of optimized separations of basic compounds are given.     

Determination of oleanolic acid and ursolic acid in cornel by cyclodextrin-modified micellar electrokinetic chromatography

A cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC) method was established for the determination of oleanolic acid and ursolic acid in cornel. The two components were separated in the running buffer of 40 mmol/L sodium borate containing 5% methanol, 25 mmol/L SDS and 15 mmol/L hydroxypropyl-beta-cyclodextrin (HP-beta-CD). The applied voltage was 24 kV. The wavelength of detection was 200 nm. The temperature was kept at 25 C. Cinnamic acid was used as the internal standard. The analytical performance of the method was tested with respect to linearity, precision and recovery. The calibration curves were linear in the range of 10.15-243.6 microg/mL, r=0.9993 (oleanolic acid) and 10.07-241.7 microg/mL, r=0.9994 (ursolic acid); the intra-day precision (RSD) was less than 3.7% (oleanolic acid) and 4.1% (ursolic acid); the inter-day precision (RSD) was less than 4.2% (oleanolic acid) and 4.9% (ursolic acid). The limits of detection were 1.6 microg/mL for both components. The method proved to be sensitive, rapid, accurate and suitable for the determination of oleanolic acid and ursolic acid in cornel.     

齐墩果酸与熊果酸结构修饰物的药理活性和构效关系研究进展

齐墩果酸(OA)和熊果酸(UA)均属于五环三萜类化合物,广泛存在于自然界中,具有多种显著的生物活性。本文综述了近年来齐墩果酸及熊果酸结构修饰物的药理活性和构效关系的研究进展。     

High-Speed Separation of Oleanolic acid and Ursolic acid from the Fruit of Ligustrum Lucidum ait and Crataegus by Cyclodextrin-Modified Micellar Electrokinetic Chromatography

ABSTRACT

A cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC) technique for determining oleanolic acid and ursolic acid in the fruit of Ligustrum lucidum Ait and Crataegus was developed. The two bio-active components were successfully separated within 4 minutes using a pH 8.5 sodium borate buffer containing 60mM SDS and 2mM γ-CD. The correlation coefficients of the linear calibration graphs for the analyses exceeded 0.9980. The effects of pH, SDS, γ-CD, sodium borate and temperature on separation were also investigated.