Particle Column Research Articles

Diffusion and Dispersion in Tubes in Supercritical Fluid Chromatography Using Sub-2 µm Packings

Detailed plots of diffusion coefficients, Dm, versus % methanol in CO2 and pressure, P, are presented. Using sub-2 µm particle columns with small ID connector tubing make for large pressure drops, ∆P, across both the system and the column. This creates low average Dm, and large gradients in Dm, slowing the chromatography. The ∆P with 75 µm ID tubing varies non-linearly with flow creating U shaped plots of reduced plate height versus flow rate. Plumbing does but should never change h versus F plots. Using larger ID tubes to estimate the extra-column variance $$\sigma_{e - c}^{2}$$ of smaller tubes, by keeping the aspect ratio (L/r) constant, assumes the same constant Dm in both tubes, which is not true in SFC. Contrary to prediction, calculated tube plate count, N, indicates connector tubing is more dispersive in SFC compared to high performance liquid chromatography (HPLC). Current supercritical fluid chromatographs (SFC) have up to 20 × higher variance than appropriate for use with 2.1 and 3 mm columns ≤ 100 mm long, packed with sub-2 µm particles. Lower volume UV detector flow cells are needed for use with sub-2 µm particles. The modular nature of instruments makes it difficult to further minimize tube lengths while retaining column temperature control. Very small ID tubes, like 75 µm should probably be avoided. It remains difficult to characterize and minimize $$\sigma_{e - c}^{2}$$ .

Simultaneous HPLC-DAD determination of seven intense sweeteners in foodstuffs and pharmaceuticals using a core-shell particle column

As diabetes and obesity have become an epidemic across the world, sweetener compounds are commonly used due to medical need and aesthetic worries. Because of legal limitations and synergistic effects, sweeteners are used together; as a result, their simultaneous determination has become a current topic. A novel HPLC method for simultaneous determination of acesulfame-K, saccharin, aspartame, alitame, neohesperidin dihydrochalcone, rebaudioside-A and neotame in various matrices was described. The proposed method was applied using a core-shell (superficially porous) silica particle column as a stationary phase, and a mobile phase consisting of acetonitrile, phosphate buffer and water. The method provided limits of detection in the range of 0.03–1.12 mg L−1, with recoveries between 95.69–103.99 % and precision better than 1.90 %. The developed method was successfully applied to 65 different commercial foodstuffs and pharmaceutical products by using easy to perform sample preparation applications, including steps such as simple dissolution, dilution and filtration. It was found that none of the analyzed food samples was above the legally imposed limits and contained no alitame, that was not allowed to be used. It was also observed that the results were in accordance with the product label information and did not exceed the limit values for foods.

Analysis of magneto rheological elastomers for energy harvesting systems

Magneto Rheological Elastomers (MREs) are composite materials based on an elastomer filled by magnetic particles. Anisotropic MRE can be easily manufactured by curing the material under homogeneous magnetic field which creates column of particles. The magnetic and elastic properties are actually coupled making these MREs suitable for energy conversion. From these remarkable properties, an energy harvesting device is considered through the application of a DC bias magnetic induction on two MREs as a metal piece is applying an AC shear strain on them. Such strain therefore changes the permeabilities of the elastomers, hence generating an AC magnetic induction which can be converted into AC electrical signal with the help of a coil. The device is simulated with a Finite Element Method software to examine the effect of the MRE parameters, the DC bias magnetic induction and applied shear strain (amplitude and frequency) on the resulting electrical signal.

Charge variant analysis of protein-based biopharmaceuticals using two-dimensional liquid chromatography hyphenated to mass spectrometry

The profile of charge variants represents an important critical quality attribute of protein-based biopharmaceuticals, in particular of monoclonal antibodies, and must therefore becontrolled. In this work, 2D-LC methods for charge variant analysis were developed using a strong cation-exchange chromatography (SCX) as first dimension (1D) separation. Non-porous SCX (3 µm) particle columns and different mobile phases were evaluated using a test mixture of some standard proteins of different size and pI (comprising myoglobin, bovine serum albumin, cytochrome c, lysozyme and β-lactoglobulin) and two monoclonal IgG1 antibodies (NIST mAb and Secukinumab). The most promising 1D eluent for SCX was a salt-mediated pH-gradient system using a ternary mobile phase system with 2-(N-morpholino)ethanesulfonic acid, 1,3-diamino-2-propanol and sodium chloride. For the second dimension (2D), a desalting reversed-phase liquid chromatography (RP-LC) was chosen to enable the hyphenation of the charge variant separation with mass spectrometric (MS) detection. While for intact mAbs the 2D just served for desalting without additional selectivity, the 2D contributed some orthogonal selectivity for the mAb fragment separation. Various core-shell and monolithic columns were tested and variables such as gradient time and flow rate systematically optimized. Unexpectedly, a C4 400 Å column (3.4 µm diameter with 0.2 µm porous shell) provided higher peak capacities compared to the same 1000 Å column (2.7 µm diameter with 0.5 µm porous shell). A thinner shell appeared to be more advantageous than wider pores under high flow regime. An ultra-fast RP-LC method with a run time of one minute was developed using trifluoroacetic acid which was later replaced by formic acid as additive for better MS compatibility. The successful hyphenation of the two orthogonal separation modes, SCX and RP-LC, could be demonstrated in the multiple heart-cutting and the full comprehensive mode. MS analysis using a high-resolution quadrupole time-of-flight instrument enabled to identify different glycoforms and some major charge variants of the antibody at the intact protein level as well as on the subunit level (Fc/2, Lc, Fd’) in a middle-up approach by 2D-LC-ESI-MS analysis.

Role of porogenic solvent type on the performance of a monolithic imprinted column

Due to many advantages, the use of imprinted monolith columns has become the choice to replace conventional particle columns. An imprinted monolith column has high permeability with minimal backpressure. Several factors can affect the performance of a column, such as the condition of synthesis, back pressure and the nature of the stationary phase. The latter feature relies on its constituent components, one of which lies in the pore structure. Another factor that influences the structure and morphology of pores is the porogenic solvent, where the selection of a porogen must be based on the solubility of all the polymer-forming components in it and must provide a sufficient pore size to facilitate the flow of the mobile phase. The use of porogenic solvents with different polarity levels and mixtures of more than one type of porogen can affect the pore structure of the formed polymers and, thus, also affect the performance of the resulting column. In this review, we will further discuss the role of porogenic solvent—polar or non-polar, single or mixed—on the performance of monolithic polymer columns as well as its applications.

Rapid and Economic Determination of 13 Steviol Glycosides in Market-Available Food, Dietary Supplements, and Ingredients: Single-Laboratory Validation of an HPLC Method.

Steviol glycosides, obtained from leaves of Stevia rebaudiana Bertoni (stevia) or produced via bioconversion and biosynthesis, are diterpenes used by the food/dietary supplement industry as zero-calorie sweeteners derived from natural sources. JECFA 2017 is the most updated international standardized method but it runs for 80 min per sample with suboptimal separations on several critical pairs for its high-performance liquid chromatography-ultraviolet (HPLC-UV) determination. We developed and validated a rapid and economic HPLC-UV method using the superficially porous particle column to determine 13 steviol glycosides (stevioside, dulcoside A, rubusoside, steviobioside, and rebaudioside A-F, I, M, and N). Baseline separation with a minimum resolution of 1.5 for 13 steviol glycosides was achieved within only 14 min of separation time. The hydrocarbon stationary phase with additional steric interactions from the isobutyl side chains on the C18 ligand was shown to be an important contributor to chromatographic selectivity of several critical pairs of steviol glycosides. The method was proven to perform suitably on columns from three different manufacturers and two HPLC instruments. The method was further used to perform a single-lab validation on eight food and supplement products with multiple matrices. The results ranged from 0.05% w/w rebaudioside A for a hard-candy finished product to 100.8% w/w purity for a rebaudioside M raw ingredient. The validation test results showed that the method was linear, suitable, specific, accurate, and precise. The method is therefore suitable to be considered as a new industrial standard for quality control analysis for stevia products.

Stability-indicating LC-MS/MS and LC-DAD methods for robust determination of tasimelteon and high resolution mass spectrometric identification of a novel degradation product

Novel stability-indicating HPLC methods have been presented for the analysis of Tasimelteon (TSM) besides its main degradation products in bulk and pseudo tablet formulations in this study. For the LC-DAD method; quantitation of TSM and its separation with other degradation products were achieved on an Ascentis ExpressTM pentafluorophenylpropyl (F5)-bonded fused-core silica particle column (2.7 μm particle size 100 × 4.6 mm, Supelco) using the mobile phase consisted of acetonitrile: acetate buffer (0.025 M, pH 4.5): water (40:10:50, v/v/v); the elution was performed at 0.8 mL min−1 flow rate, detecting the compounds at 281 nm. In addition to regular in- house validation studies, the developed method was tested on another UPLC instrument to assess its ruggedness for possible method transfer demands. For the LC–MS/MS method, analyte quantitation was achieved on a second-generation monolithic silica column (Chromolith™ High-Resolution RP-18e, 100 × 4.6 mm from Merck KGaA, Germany); the mobile phase was a mixture with 0.1 % (v/v) formic acid in water and 0.1 % (v/v) formic acid in acetonitrile (60: 40 (v/v), pH = 2.5). The instrumental and analytical performances of all three instrumental systems were compared with each other in terms of the working range, LOD, and LOQ. In addition to the above, a new degradation product was identified using LCMS-IT-TOF system. It can be concluded that among the similar ones in the literature, this study is the most comprehensive method optimization and validation study to date about TSM, in which a novel degradation product was also reported.

A Novel HPLC Method for Simultaneous Determination of Methyl, Ethyl, n-propyl, Isopropyl, n-butyl, Isobutyl and Benzyl Paraben in Pharmaceuticals and Cosmetics.

The alkyl esters of p-hydroxybenzoic acid at the C-4 position, "the parabens," including methyl, ethyl, propyl, and butyl, are widely used as antimicrobial preservatives in foods, cosmetics, and pharmaceuticals. Official regulations on the use of these compounds make their analysis essential for the estimation of their exposure. On this basis, the presented study was realized to develop a simple, selective and cheap high-performance liquid chromatographic method for the quantitative determination of methylparaben, ethylparaben (EP), n-propyl paraben (NPP), isopropyl paraben (IPP), n-butyl paraben (NBP), isobutyl paraben (IBP) and benzyl paraben (BP) in pharmaceuticals and cosmetic products. The chromatographic separation of the analytes was achieved under flow rate gradient elution conditions using a C18-bonded core-shell silica particle column (2.6 μm particle size, 150 × 3.0 mm from Phenomenex Co.). The samples were injected into the system as aliquots of 1.0 μL, and the compounds were detected by using a photodiode array detector set at 254 nm wavelength. With this technique, seven paraben derivatives can be determined in the concentration range of 250-2000 ng/mL. The recovery of the method is in the range of 99.95-13.84%, and the RSD is at a maximum value of 3.95%. The proposed method was fully validated and successfully applied to different pharmaceutical and cosmetic samples (n=16), including syrups, suspensions, oral sprays, gels, etc. At least one paraben derivative was detected in six samples and was determined quantitatively. The maximum amount of a paraben derivative found in the analyzed samples was 321.7 ng/mL, which was MP. To the best of our knowledge, this is the first LC method, which is applicable both on pharmaceutical and cosmetic samples.

A Simple Methodology for Preparing Synthetic Multiple-Interaction Chiral Stationary Phase Column for Chiral Drug Analysis

A simple methodology for preparing synthetic multiple-interaction (SMI) chiral stationary phase (CSP) column is described in this report, The column was synthesized by an in situ derivatizing procedure and can be easily prepared with a standard analytical HPLC set-up. The column is a no frills, low cost one, and can be tailored to various packing particle sizes and column lengths. The efficiency of the column is high; over 5,000 plates per 10 cm Applications on resolving some racemic drugs in plasma and formulation are illustrated.

Artificial neural network model development for prediction of nonlinear flow in porous media

To explore non-linear flow behaviour in porous media, e.g., packed bed of particles, this study focuses on the prediction of pressure drop/gradient by establishing empirical relations for Forchheimer coefficients based on artificial neural network (ANN) development. To facilitate this process, a large set of experimental data for pressure drop of water flow in various porous media is collected: part from the present experiment and other from those available in the literature. The newly generated ANN model is developed by considering the key influencing factors such as particle density, diameter, and shape, and column porosity. The new model shows excellent performance on the prediction of flow behaviour over previous empirical formula, which is attributed to the ANN's advantages on considering the inter-factor connections and their influences on the results. This proposed approach can provide a new means for analysing the nonlinear flow regime through porous media based on limited dataset.

Two-dimensional separation by sequential injection chromatography.

Sequential injection chromatography (SIC) is an alternative for fast chromatographic separations with low consumption of organic solvents. However, its separation capacity is restricted by the use of short chromatographic columns and the limitations for gradient elution. The present work aimed to expand the analytical capacity of SIC by exploiting a multidimensional approach with two chromatographic columns, different in their separation mechanisms, which increased the selectivity and peak resolution. The viability of the proposal was demonstrated by separation of aromatic biogenic amines (histamine, tyramine, phenylethylamine, and tryptamine), whose unidimensional separation was not achieved either by using cyanopropyl or C18 chromatographic columns. In the two-dimensional approach, the fraction of the eluate unresolved in the first dimension (containing tyramine and phenylethylamine) was collected in a sampling loop and, subsequently, inserted in the second chromatographic dimension (heart-cutting mode). Under the optimized conditions, the first chromatographic dimension was based on a cyanopropyl monolithic column and an aqueous mobile phase composed of phosphoric acid solution, pH 2.5, while the second dimension employed a C18 superficially porous particle column and a mobile phase composed of acetonitrile and phosphoric acid aqueous solution, pH 2.5 (7:93, v/v). The total analysis time was 8 min, and a resolution of 1.72 was achieved between the nearest peaks (tyramine and phenylethylamine). Linear responses were obtained within 10 and 50 mg L-1 (r > 0.997), with detection limits estimated at 2.7, 7.7, 1.9, and 0.3 mg L-1, for histamine, tyramine, phenylethylamine, and tryptamine, respectively, and a coefficient of variation of 3.0% (n=12).

Development of a rapid reversed-phase liquid chromatographic method for total free thiol quantitation in protein therapeutics

Free thiols, or unpaired cysteines, are important product quality attributes in the therapeutic proteins due to their potential impact on the protein structure, bioactivity and stability. While many free thiol quantitation methods were developed for specific therapeutic formats, an unmet need still exists for a multiproduct, high-throughput method for free thiol quantitation. In this study, a workflow was established that combines N-cyclohexylmaleimide (NcHM) derivatization and high-throughput reversed-phase ultra-high performance liquid chromatography (RP-UHPLC) separation with superficially porous particle (SPP) column for quantitating total free thiols in monoclonal antibodies (mAbs), fragment antigen-binding (Fab), and bispecific antibodies (BsAbs). The NcHM derivatization increases the hydrophobicity of the free thiol variants and allows the further separation and quantitation with RP-UHPLC. A thorough evaluation of sample preparation, column selection, chromatographic condition and LC–MS peak identification was performed to optimize and characterize the method outputs. Method optimization resulted in a 42-minute total analysis time. Method qualification demonstrated suitable accuracy, precision, linearity, specificity and robustness. This high-throughput method is not only used for quantitation of total free thiols for both in-process testing and drug substance/drug product batch testing, but also for provide the positional distribution of the free thiols in the protein.

Experimental investigation of the retention factor dependency of eddy dispersion in packed bed columns and relation to knox's empirical model parameters.

We report on a systematic and comprehensive (0.7≤k'' ≤ 122) experimental study of the effect of the zone retention factor k'' on eddy dispersion (heddy) in packed bed columns for liquid chromatography. The values for heddy are obtained by subtracting rigorously estimated contributions to the total plate height from longitudinal diffusion (hB) and the mobile (hCm) and stationary zone (hCs) mass transfer resistances. For the first time, hCm-values are calculated using an expression for the Sherwood number (Sh) that has been established and validated in the relevant velocity range. Experiments were carried out on both a fully-porous and a core-shell particle column. In both cases, the eddy dispersion systematically decreased with increasing retention factor k'', dropping 0.5 to 0.8 reduced plate height units when going from the lowest to the highest k''. To establish a simple empirical fitting equation that can represent the observed effects, the widely used power law-based Knox model has been extended to express the dependence of its A- and n-parameters on the retention factor.

Determinations of Labile Nutrients (Methylcobalamin, 5-Methyltetrahydrofolate, and Trans Menaquinone-7) in Oil- and Beadlet-Filled, Gastric-Resistant Capsule

Abstract Objectives The dietary supplement industry has grown substantially, and advances in technology have played important roles in the development of new forms of dietary supplements and delivery systems. While it is recommended to use compendia methods for testing dietary supplements, the analytical methods in place are often developed for single ingredients or simple matrices and are not always fit to be used for solving modern matrix challenges. Oil- and beadlet-filled, gastric acid-resistant capsule (OBGRC) is designed to improve nutrient delivery and absorption. However, it is challenging to accurately analyze labile nutrients in OBGRCs because nutrients within different physical locations are difficult to separate, and lengthy sample preparations lead to nutrient degradation. We developed and validated analytical methods to determine methylcobalamin (MeB12), 5-methyltetrahydrofolate (5-MTHF), and trans menaquinone-7 (MK-7) in OBGRCs. Methods MeB12 and 5-MTHF were extracted by completely dissolving the OBGRC shells using aqueous buffers and then applying hexane-facilitated liquid-liquid extraction to remove the oil phase and determined by RP-HPLC using superficially porous particles columns. For MK-7 analysis, the whole OBGRC was cut to open and the oil phase was collected to dissolve in a mixture of dimethyl sulfoxide/tetrahydrofuran/ethanol before analyzed by RP-HPLC with fluorescence detection after on-line, post-column zinc reduction. The methods were subjected to single-laboratory validations according to the United States Pharmacopeia (USP) guidelines for linearity, suitability, detection limits, specificity, accuracy, and precision. Results The methods achieved chromatographic resolution of target analytes without the expensive requirement of ultra-high pressure liquid chromatography and/or mass spectrometry. The methods have wide analytical ranges (from at least 50% to 250% of the input concentrations), high precision (repeatability relative standard deviations ranging from 1.04% to 4.90%), and high accuracy (spike recovery rate ranging from 89.2% to 108%). Conclusions All three methods passed USP method validation criteria and have the potential to be widely adopted to analyze MeB12, 5-MTHF, and MK-7 in similar matrices for quality control purposes. Funding Sources The research funding was provided by Ritual and Eurofins.

Rapid analysis of 14 ultraviolet absorbents in plastic food contact materials by supercritical fluid chromatography on Sub-2-micron particles

A rapid and reliable method based on supercritical fluid chromatography (SFC) technology was established for the analysis of 14 ultraviolet (UV) absorbents in plastic food contact materials. The separation was optimized by employing different chromatographic conditions including stationary phases, organic modifiers, column temperatures and back pressures to achieve appropriate selectivity and resolution. The 14 UV absorbents were analyzed within 4.5 min on the sub-2-micron particles column (ACQUTY UPC2 HSS C18 SB) via gradient elution with methanol as the organic modifier. The calibrations were carried out in the concentration range of 0.1–50.0 mg/L (except for UV-329 and UV-320 were 0.2–50.0 mg/L, UV-360 was 0.3–50.0 mg/L), and excellent linearity was observed with correlation coefficients (R2) greater than 0.99. The limits of quantification (LOQ) were in the range of 0.10–0.30 µg/g. The recoveries for accuracy were ranged from 90.4 to 114.1%, and the relative standard deviation (RSD) for repeatability was between 0.67 and 7.23%, respectively. The established method was successfully applied for the analysis of UV absorbents in five plastic food contact materials samples. The SFC method developed in this study can provide an alternative route for routine analysis of UV absorbents in the future.

Investigation of mass transfer properties and kinetic performance of high-efficiency columns packed with C18 sub-2μm fully and superficially porous particles.

Three columns packed with 2.0μm superficially porous particles, 1.7μm fully porous particles, and monodisperse 1.9μm fully porous particles with narrow particle size distribution have been deeply characterized from a kinetic point of view. The 1.9μm column showed excellent kinetic performance, comparable to that of the superficially porous one. These two columns also exhibit flatter c-branches of the van Deemter curve compared to the 1.7μm fully porous particles column, resulting in smaller loss of efficiency when they are operated at higher flow rates than the optimal ones. The independent evaluation of each contribution to band broadening has revealed that the difference in kinetic performance comes from the very small eddy dispersion contribution on the 1.9μm column, surprisingly even lower than that of the superficially porous one. This finding suggests a very good packing of the monodisperse 1.9μm column. On the other hand, the potential of 1.7μm fully porous particles is completely broken down by the strong frictional heating effect already arising at relatively low flow rates.

Tracking particles in sands based on particle shape parameters

The acquisition of particle kinematics of deforming sands generally requires tracking the particles within the specimen. For this purpose, one particle tracking method based on particle polar radius functions (PR-Track) and a second method based on the SH spectrum of particle spherical harmonics (SH-Track) are presented. Both methods are applied to the acquisition of particle kinematics of a Leighton Buzzard sand sample undergoing shearing in a miniature loading apparatus using X-ray micro-tomography. The results are compared to those from a particle volume-based particle tracking method (PV-Track). It is found that PR-Track and SH-Track return particle tracking results with a high precision which is comparable to PV-Track. For the tested sample, PR-Track is about 0.4 times faster than SH-Track. Furthermore, PR-Track and SH-Track have a much lower computational efficiency than PV-Track. However, PR-Track and SH-Track do not require particles to be displaced in a similar manner to their neighbours, which is assumed by PV-Track. Particle tracking results from PR-Track and SH-Track are not sensitive to search region sizes. These features imply that PR-Track and SH-Track are capable of tracking particles of granular media undergoing a large and complicated deformation. This capability is verified in a simple case study by applying the methods to track particles undergoing particle column collapse.

Rattler wedging and force chain buckling: metastable attractor dynamics of local grain rearrangements underlie globally bistable shear banding regime

Grain rearrangements govern the mechanical response of granular materials under load. A comprehensive time series analysis of macroscopic stress, previously undertaken for the persistent shear banding regime, uncovered a bistable dynamical system with two interconnected attractor basins—one tied to jamming, the other to unjamming. Here we examine the state space of local fabric of a 3D granular material to uncover the underlying grain rearrangements which induce the system to switch basins and manifest macroscopic quasistationary dynamics. The fabric state space comprises metastable attractors organized according to structural determinacy: hypostatic, isostatic and hyperstatic. Although trajectories overwhelmingly favor identity transitions which preserve local fabric, rare nonidentity transitions hold the key to basin switching. Jamming basin dynamics is governed by force chain transitions in the hyperstatic region: here redundant constraints enable force reconfigurations with little to no change in the local fabric. But as force chains become overloaded, they buckle. Buckling and attendant dilatancy push grains to the isostatic region. This in turn triggers unjamming and ensuant transitions to the hypostatic region where rattlers, though few, dominate dynamics. Rattlers provide wedge-like supports to unstable misaligned particle columns, promoting force chain formation and return to the hyperstatic region. The lateral reinforcement of an isostatically constrained grain in a trimer by the addition of a contact and/or a 3-cycle is the most probable nonidentity transition during nascent force chain evolution.

High-performance liquid chromatography (HPLC)-fluorescence method for determination of bisphenol A diglycidyl ether (BADGE) and its derivatives in canned foods

Bisphenol A diglycidyl ether (BADGE) is used as a raw material for the production of epoxy resins and PVC organosols, which are commonly applied as inner coatings for food cans. BADGE and its derivatives can migrate from coatings to foodstuffs during processing and storage thereby creating adverse health issues. In this work, a method based on high-performance liquid chromatography (HPLC)-fluorescence detection (FLD) method was developed for the rapid determination of BADGE and its five derivatives in canned foods. Modeling software DryLab® was applied for the optimization of separation conditions. An adequate separation was achieved in 5 min including equilibration time, using a core–shell particle column; such application has not been reported so far. Also, the results showed that LOD varied from 0.01 to 0.20 ng/g, while LOQ varied from 0.03 to 0.66 ng/g, and RSD was found to be <8.64%. The analytical recoveries ranged from 70.46 to 103.44%. Excellent validation data revealed that this method is suitable for the investigation of can coating-to-food migration of BADGE and its derivatives. The HPLC-FLD method is rapid, inexpensive and highly efficient, which could be applicable for safety inspection of food contact materials involving BADGE and its derivatives.

Isolation and purification of four phenolic glycoside isomers from the leaves of Idesia polycarpa Maxim. by high-speed counter-current chromatography and preparative high-performance liquid chromatography.

We established an efficient method using high-speed counter-current chromatography (HSCCC) combined with preparative high-performance liquid chromatography (prep-HPLC) for isolating and purifying phenolic glycoside isomers. The method involves a rapid and sensitive ultra-performance liquid chromatography-under voltage(UPLC-UV) technique using a sub-2μm core-shell particle column for qualitative and quantitative analysis of four phenolic glycoside isomers from Idesia polycarpa Maxim. leaves. The partially purified samples from ethyl acetate extraction of ethanol extracts of I. polycarpa Maxim. leaves were obtained by HSCCC with a two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water solution (3:5:3:5, v/v/v/v) to obtain fractions I and II, which contain two phenolic glycoside isomers. The two fractions were further isolated by prep-HPLC to yield compounds 1, 2, 3, and 4 with purities of 98.29%, 98.71%, 98.49% and 98.52%, and total recoveries of 93.5%, 72.2%, 75.5% and 88.3%, respectively. Compound 1 was first isolated from I. polycarpa Maxim., while compound 2 was reported to be a new phenolic glycoside, which is 1-[(6'-O-(Z)-p-coumaroyl)-β-D-glucopyranosyl]-oxy-2-phenol. The chemical structures of the four phenolic glycoside isomers were analyzed and confirmed by UPLC, UV, electrospray ionization mass spectrometry (ESI-MS), fourier transform infra-red (FT-IR), 1H-nuclear magnetic resonance (1H NMR), 13C-nuclear magnetic resonance (13C NMR) and 2D nuclear magnetic resonance (2D NMR) spectra. This study opens prospects for broad industrial applications of HSCCC/prep-HPLC for the isolation and purification of isomers.