RP-HPLC梯度洗脱法测定桑叶中芦丁和绿原酸的含量

建立了同时测定桑叶中芦丁和绿原酸含量的方法,采用Kromasil C18柱(4.6 mm×250mm,5μm),以甲醇0.5%磷酸的水溶液为流动相,梯度洗脱程序为0 min[V(甲醇) V(0.5%磷酸)=30 70]15 min[V(甲醇) V(0.5%磷酸)=30 70]20 min[V(甲醇) V(0.5%磷酸)=50 50]35 min[V(甲醇) V(0.5%磷酸=50 50];流量0.8 mL.min-1;检测波长350 nm;柱温25℃;光电二极管阵列检测器(PAD)。结果表明,芦丁线性范围为0.218~1.962μg,r=0.999 3,样品的平均加样回收率为101.3%,RSD 1.1%;绿原酸线性范围为0.654~5.886μg,r=0.999 7,样品的平均加样回收率为98.2%,RSD 1.1%;绿原酸线性范围为0.654~5.886μg,r=0.999 7,样品的平均加样回收率为98.2%,RSD1.5%。该法快速简便,专属性强,重复性好,可作为桑叶中芦丁的定量分析方法。     

超高效液相色谱-串联质谱双内标法同时测定复方杏香兔耳风胶囊中的10种有效成分

建立了同时测定复方杏香兔耳风胶囊中原儿茶酸、原儿茶醛、绿原酸、野黄芩苷、异绿原酸C、黄芩苷、木犀草素、芹菜素、白术内酯Ⅲ和白术内酯I等10种有效成分含量的超高效液相色谱-串联质谱（UPLC-MS/MS）双内标分析方法。以咖啡酸和淫羊藿苷为内标（IS）,在ZORBAX RRHD Eclipse Plus C₁₈色谱柱上,以甲醇和含0.3%甲酸的水为流动相进行梯度洗脱分离,流速为0.3 mL/min。在电喷雾电离（ESI）正、负离子切换模式下,采用多重反应监测模式进行检测。结果表明,原儿茶酸、原儿茶醛、绿原酸、野黄芩苷、异绿原酸C、黄芩苷、木犀草素、芹菜素、白术内酯Ⅲ、白术内酯I的线性范围分别为0.00300~24.0 mg/L、0.0170~2.00 mg/L、0.0150~30.0 mg/L、0.00400~30.0 mg/L、0.0105~24.0 mg/L、0.00300~30.0 mg/L、0.00300~5.00 mg/L、0.00600~5.00 mg/L、0.00150~4.00 mg/L、0.000600~0.900 mg/L;检出限分别为1.0、11、5.0、1.5、3.5、1.0、1.0、2.0、0.50、0.20 μg/L。10种成分的加样回收率为92.5%~106%,相对标准偏差均不大于3.2%。该方法快速简便、灵敏度高、重复性好,已成功用于实际样品的分析。     

高效液相色谱用线性梯度洗脱求保留值方程

〕提出了采用线性梯度洗脱求取未知样品中溶质保留值方程的原理及其计算方法,用光二极管阵列扫描检测器解决了梯度洗脱条件下色谱峰的归宿。实验证明：用所得的保留值方程预测等度洗脱下容量因子k'的相对误差不大于3%,而用梯度洗脱下的半峰宽规律可以估算未知样品容量因子k的分布范围,以解决对未知样品是采用等度或梯度洗脱方式。最后,借助计算机仿真方法用于样品的优化,获得了满意的结果。     

高效液相色谱梯度洗脱同时测定化妆品中防腐剂、抗氧化剂和杀菌剂

建立了一种灵敏、快速的高效液相色谱梯度洗脱同时测定化妆品中防腐剂、抗氧化剂和杀菌剂的方法。采用C8柱,以甲醇和水为流动相梯度洗脱。在检测波长273nm用二极管阵列检测器,17min内可将4种防腐剂、2种抗氧化剂和2种杀菌剂分离测定。方法快速准确,用于实际样品的测定。     

高效液相色谱中用于梯度洗脱的有机溶剂的纯度测试

在反相高效液相色谱中,空白梯度基线的漂移和杂质峰,可能由含水溶剂和有机溶剂中杂质所引起的。当含水溶剂通过预柱在线净化后,根据空白梯度色谱图可测试有机溶剂的纯度。按照此法,对国产不同等级的乙腈和甲醇进行了测试,指明这些试剂用于梯度洗脱的可能性。     

高效液相色谱梯度洗脱中弱溶剂的在线净化

利用一个在线净化预柱减少了高压梯度洗脱中由于试剂纯度不够造成的基线噪音。预柱的位置在溶剂泵Ａ和流动相混合器之间,由一个六通阀来控制。在线净化结果良好,有效地除去了空白梯度色谱图中的杂质峰,提高了梯度洗脱的分离重现性。该方法对开展梯度洗脱有实用价值。     

反相高效液相色谱法同时测定沙棘叶中绿原酸和芦丁

建立了同时测定沙棘叶中绿原酸和芦丁含量的反相高效液相色谱(RP-HPLC)法。采用Kromasil C18色谱柱(150×4.6mm,5μm),流动相为体积比20∶80的乙腈-1%乙酸水溶液,检测波长358nm。结果表明:绿原酸、芦丁的浓度分别在0.0284～0.1704mg.mL-1(r=0.9995)和0.0566～0.3396mg.mL-1(r=0.9992)范围内与色谱峰面积呈良好线性关系;平均加标回收率分别为100.5%(n=5)和98.5%(n=5)。该方法快速、准确,重现性好,可用于同时测定沙棘叶中绿原酸和芦丁含量。     

梯度洗脱高效液相色谱法测定红花玉兰中4种植物激素

建立了梯度洗脱高效液相色谱法测定红花玉兰中赤霉素(GA3)、生长素(IAA)、脱落酸(ABA)和玉米素(ZT)等4种植物激素的方法。采用Agilent ZORBAX SB-C18柱和紫外检测器,以甲醇和0.1 mol/L乙酸作为流动相进行梯度洗脱,流速1 mL/min,进样量10μL。GA3,IAA和ABA的检测波长为254 nm,柱温35℃;ZT的检测波长为270 nm,柱温40℃。采用外标法进行定量测定,4种植物激素的相关系数均大于0.9990。4种激素的回收率为98.1%~125.2%,相对标准偏差为0.31%~0.92%,日内和日间精密度RSD均<10%。方法可适用于红花玉兰多种组织的植物激素测定,为红花玉兰生长发育特性的研究奠定了基础。     

梯度洗脱高效液相色谱法快速检测厌氧菌代谢物中的有机酸

提出了一种快速分析厌氧细菌代谢物中5种有机酸的高效液相色谱方法。该方法采用乙腈-0.02mol/L磷酸二氢钾缓冲溶液(pH 2.8)作为流动相,流速和流动相中两种组分的比例均采用四元泵程序梯度洗脱方法控制,检测波长为215 nm,柱温30℃时,能够快速、准确地分离和测定细菌培养基中的甲酸、乙酸、乳酸、琥珀酸和丙酸等5种有机酸,总分析时间只需4 m in。方法的相对标准偏差为0.26%~1.26%,回收率95.0%~100.8%,各种有机酸的线性相关系数r≥0.99981,具有较高的精密度和准确度,可以用于细菌代谢物中有机酸的分析。     

Characterization of different RP-HPLC columns by a gradient elution technique

Summary The literature contains no unified testing procedure for evaluation and characterization of reversed-phase (RP) columns of different pore size (surface area) and containing different types of ligand. In our laboratory a testing procedure has been developed using gradient elution under standardized conditions independent of column dimensions and applicable to both narrow-pore and wide-pore columns. Six wide-pore and four narrow-pore columns were investigated and compared. The test solutes were selected to cover a wide range of chemical properties. The evaluation and characterization of the columns was performed on the basis ofk _app retention factors. Principal components analysis (PCA) was performed to reveal similarities and differences among the columns and test substances. The factors obtained characterize the columns according to the extent of various interactions such as hydrophobicity, HB (hydrogen bond)-donor acidity and HB-acceptor basicity. Presented at the 21^st ISC held in Stuttgart, Germany, 15^th–20^th September, 1996     

Solvent gradient elution in micro HPLC

A simple gradient method for capillary column and micro high performance liquid chromatography using either single or multiple mixing chambers is presented. The gradient profile is dependent on the number of the mixing chambers. This system permits solvent gradient elution at flow rates less than 10 μl/min with satisfactory reproducibility.     

梯度洗脱测定植物源调节剂中内源激素方法探讨

建立了梯度洗脱高效液相色谱法同时测定内源激素赤霉素(GA3)、吲哚乙酸(3-IAA)、脱落酸(ABA)、细胞分裂素(6-BA)和α-萘乙酸(α-NAA)等5种植物内源激素的方法,采用Hypersil ODS C18柱和紫外检测器,以甲醇、乙腈和0.6%的乙酸为流动相梯度洗脱,分别在各组分的保留时间段设置其最佳检测波长,在12 min内可将上述5种内源激素的各组分分离完全,峰形良好,该方法的回收率可达到90%～96%,线性相关系数(r)大于0.998,检出限在0.02～0.3 μg/g之间.还探讨了各组分的最佳检测条件和梯度洗脱存在的问题及解决方法.     

Determination and pharmacokinetic study of chlorogenic acid in rat dosed with Yin-Huang granules by RP-HPLC

A reversed-phase high-performance liquid chromatographic (RP-HPLC) method was described for the determination of chlorogenic acid (CGA) in rat plasma using protocatechuic acid as internal standard (IS). CGA in plasma was extracted with acetonitrile, which also acted as deproteinization agent. Chromatographic separation was performed on a Kromasil C18 column with methanol-0.2 m acetic acid (pH 3.0, 25:75, v/v) as mobile phase at a flow-rate of 1.0 mL/min with an operating temperature of 30 degrees C and UV detection at 300 nm. The standard curve was found to be linear over the concentration ranges of 0.4-2.5 microg/mL and 2.5-40 microg/mL, and the limit of quantification (LOQ) was 0.4 microg/mL. The analytical precision and accuracy were validated by relative standard deviation (RSD) and relative error, which were in ranges 3.14-10.78% and -2.20-5.00%, respectively. The average recovery of CGA was 87.59%. The method was successfully applied to the pharmacokinetic study of CGA in Yin-Huang granules.     

Optimization of stepwise gradient elution in columns chromatography

Summary A general equation for the final retention of a solute chromatographed under conditions of stepwise gradient elution has been derived. The elution process and the distances travelled by solutes as a function of eluent volume were simulated by computer for the optimization of stepwise gradient prorams from isocratic HPLC data. The validity of the equations was experimentally veritied.     

A new approach to the prediction of RP-HPLC retention times in gradient elution from isocratic data

Summary A general chromatographic model has been set up starting from a set of equations based on the concept of the velocity of a solute along the column. The composition of the mobile phase is taken into account solely as a numerical factor entering into suitable equations and totally independent of the chemical-properties of the constituents. A few isocratic experimental runs are necessary as input data, and subsequently a small amount of computational effort is sufficient to make predictions of retention times under gradient elution conditions for solutes of whatever chemical structure. The prediction errors are dependent on the steepness of the linear gradient chosen but are, in any case, acceptably low.     

Determination of inorganic ions in mineral water by gradient elution moving boundary electrophoresis

A sensitive method was developed for the determination of the major inorganic ions in commercial mineral waters using gradient elution moving boundary electrophoresis with capacitively coupled contactless conductivity detection. This application was the first to demonstrate the separation of cations and anions simultaneously using gradient elution moving boundary electrophoresis. Seven ionic analytes (calcium, chloride, magnesium, nitrate, potassium, sodium, and sulfate) were separated in less than 7 min with detection values in the low μmol/L to sub-μmol/L range. Calculated values of the major ions in three commercial mineral waters were compared to reported values with good correlation. In another application, phosphate and arsenate were separated in less than 2 min with limits of detection of 300 and 140 nmol/L, respectively. For all standard analyses, the RSD for migration times and peak areas were under 3%.     

反相高效液相色谱法测定豆制品中两种异黄酮甙元的含量

建立了反相高效液相色谱测定异黄酮甙元的方法。采用Hypersil BDS C18色谱柱(250×4.6 mm i.d.,5μm),流动相为甲醇(A)、0.5%乙酸水溶液(B),流速为1.0mL/min,柱温为35℃,检测波长为260 nm,进样量为10μL。结果表明:大豆黄素的线性范围为0.0450~0.3150μg,加标回收率为99.05%,相对标准偏差(RSD)为1.31%;染料木黄酮的线性范围为0.0448~0.3136μg,加标回收率为98.82%,RSD为0.50%。方法可用于豆豉及其它大豆制品中异黄酮甙元的测定。     

Isocratic and gradient elution in micellar liquid chromatography with Brij‐35

Polyoxyethylene(23)lauryl ether (known as Brij‐35) is a nonionic surfactant, which has been considered as an alternative to the extensively used in micellar liquid chromatography anionic surfactant sodium lauryl (dodecyl) sulfate, for the analysis of drugs and other types of compounds. Brij‐35 is the most suitable nonionic surfactant for micellar liquid chromatography, owing to its commercial availability, low cost, low toxicity, high cloud temperature, and low background absorbance. However, it has had minor use. In this work, we gather and discuss some results obtained in our laboratory with several β‐blockers, sulfonamides, and flavonoids, concerning the use of Brij‐35 as mobile phase modifier in the isocratic and gradient modes. The chromatographic performance for purely micellar eluents (with only surfactant) and hybrid eluents (with surfactant and acetonitrile) is compared. Brij‐35 increases the polarity of the alkyl‐bonded stationary phase and its polyoxyethylene chain with the hydroxyl end group allows hydrogen‐bond interactions, especially for phenolic compounds. This offers the possibility of using aqueous solutions of Brij‐35 as mobile phases with sufficiently short retention times. The use of gradients of acetonitrile to keep the concentration of Brij‐35 constant is another interesting strategy that yields a significant reduction in the peak widths, which guarantee high resolution.