Analysis of the Nucleoside Content of Cordyceps sinensis Using the Stepwise Gradient Elution Technique of Thin-Layer Chromatography

Nucleoside is the main class of active components in Cordyceps sinensis. Thin-layer chromatography (TLC) is one of the most commonly used methods in pharmacopoeias for analyzing chemical components of herbal medicine. Since the isocratic elution method cannot be applied successfully in TLC analysis for separating all the nucleoside components, the stepwise gradient elution has been developed in this work to separate eight nucleoside standards with success. In this way, quantitative analyses of the samples of Cordyceps sinensis were achieved via the pro-posed TLC procedure coupled with the scanning densitometric techniques of CAMAG and TLCQA methods for qualitative and quantitative analysis.

染发剂中2-氨基-4-硝基苯酚和2-氨基-5-硝基苯酚的液相色谱-质谱法测定方法研究

A new method of 2-amino-4-nitrophenol and 2-amino-5-nitrophenol in hair dye determined by liquid chromatography-mass spectrometry.Chromatographic separation was carried out on a Atlantis C18 (2.1 mm i.d.×150 mm,5 μm)column with gradient elution with methanol-acetonitrile at a flow rate 0.2 mL/min.The mass spectrometry was operated in the positive electrospray ionization mode.Quantity was performed using multiple reaction monitoring of the product ion transitions m/z153.0>m/z106(17eV) for 2-amino-4-nitrophenol and m/z153.0>m/z122(15eV) for 2-amino-5-nitrophenol.Linearity of peak was obtained over a range of concentration is from 0.2 to 100 mg/L.The limit is 0.2 mg/L of the method.The proposed method was accurate and rapid with wide linear range.The repeatability and the reproducibility of the method were excellent.It has been applied to detect the real with satisfactory results.

梯度洗脱层析法

层析法已广泛应用于分离纯化合成或天然的产物。一般的柱层析虽可用于分离各种结构类型的化合物,但对于在结构上近似、极性相差不大的混合物分离效率较差。应用梯度洗脱的柱层析法可提高分离效率,有时会取得满意的结果。

Protein retention in dextran-grafted cation exchange chromatography:The influence of pHs,counterions and polymer structure

Polymer-grafted ion exchange adsorbents were of great interest for the development of high-performance protein chromatography in biopharmaceutical and related fields.In this work,protein retention was systematically investigated in ion exchange chromatography packed respectively with dextran-grafted cation exchange adsorbents containing sulphopropyl(SP)ligand,SP Sepharose XL and Capto S,and non-grafted cation exchange adsorbent,SP Sepharose FF,using five proteins.With an increase of buffer p Hs,retention factors of proteins decreased among all the adsorbents,demonstrating the dominant role of electrostatic interaction for protein binding on cation exchange adsorbents.The evidences further revealed that the scattered positive charges on the surface of protein molecules,rather than net charge of protein molecule,determined protein retention on cation exchange adsorbent.Likely,counterions including NH4^+,K^+,Na^+and Mg^2+exhibited distinct influence on protein retention.It was well ascribed to solvent-mediated indirect ion-macromolecule interactions and direct ion-macromolecule interactions.Compared with SP Sepharose FF,polymer structure in dextran-grafted cation exchange adsorbents ultimately brought about different ligand distributions and smaller pore sizes,thereby regulating protein retention in cation exchange chromatography.By comparing the retention of myoglobin andβ-lactoglobulin B in SP Sepharose XL and Capto S,we reasonably speculated that the enhancement of nonelectrostatic interaction caused by reducing the space arm length was a major reason for an increasing retention factor of myoglobin in Capto S.The results in this research help us understand adsorption mechanism of protein in polymer-grafted adsorbents and give scientific guidance for the development of chromatographic materials.

Novel computer-assisted approach to quick prediction and optimization of gradient separation for online enrichment-reversed phase liquid chromatography tandem system

An algorithm capable of predicting and optimizing the gradient separation of LC×LC system was developed in this paper.Two groups of structural analogues,five ginsenosides as well as eight bisphenols,which were difficult to discriminate in routine analysis,were used to verify the effectiveness of the proposed algorithm in fast separation optimization.Average errors of retention times below 1%were found in the retention prediction for all types of gradient programs,implying that the theory could lead to high quality in prediction of the retention times under gradients elution.Meanwhile,84%of relative average deviations(RADs)between the predicted peak width and the measured ones were less than 20%.The larger deviation occurred at the time when the peak appeared while the gradient of the mobile phase changed,which led the deviations increased to 20%–42%.In all,method development and optimization for LC×LC tandem system was realized by the homemade user-friendly software.The present protocol may turn on great opportunities for the convenient method development in analysis of trace structural analogues in environmental,food and biological samples.

RP-HPLC梯度洗脱法测定吡嘧司特钾滴眼液的有关物质

目的建立RP-HPLC梯度洗脱法测定吡嘧司特钾滴眼液的有关物质。方法采用WATERS Sunfire C 18分析柱(250 mm×4.6 mm,5μm),流动相A为甲醇-三氟乙酸(1→1000)(1∶4),流动相B为甲醇-三氟乙酸(1→1000)(3∶2),检测波长为260 nm,柱温为40℃。结果在选定的色谱条件下,吡嘧司特钾与有关物质完全分离;滴眼液中基质无干扰。结论本法具有灵敏高、分离效果佳、检查结果准确等优势,可用于吡嘧司特钾滴眼液的有关物质检查。