Synthesis and application of hypercrosslinked polymers with weak cation-exchange character for the selective extraction of basic pharmaceuticals from complex environmental water samples

The synthesis of high specific surface area sorbents (HXLPP-WCX) in the form of hypercrosslinked polymer microspheres with narrow particle size distributions, average particle diameters around 6 μm, and weak cation-exchange (WCX) character, is described. The WCX character arises from carboxylic acid moieties in the polymers, derived from the comonomer methacrylic acid. A novel HXLPP-WCX sorbent with an attractive set of chemical and physical properties was then used in an off-line solid-phase extraction (SPE) protocol for the selective extraction of a group of basic compounds from complex environmental samples, a priority being the clean separation of the basic compounds of interest from acidic compounds and interferences. The separation power of the new sorbent for basic pharmaceuticals was compared to two commercially available, mixed-mode sorbents, namely Oasis WCX and Strata-X-CW. Under identical experimental conditions, HXLPP-WCX was found to deliver both higher capacity and better selectivity in SPE than either of the two commercially available materials. In an optimised SPE protocol, the HXLPP-WCX sorbent gave rise to quantitative and selective extractions of low μg l⁻¹ levels of basic pharmaceuticals present in 500 ml of river water and 250 ml of effluent waste water.     

Mechanistic study of the sorption properties of OASIS® HLB and its use in solid-phase extraction

Summary The solvation parameter model is used to characterize the sorption properties of the porous polymer Oasis^? HLB for solid-phase extraction with water and water-methanol mixtures as a sample solvent. Increasing solute size and electron lone pair interactions favor retention from water. Oasis^? HLB is not competitive with water for dipole-type and hydrogen-bond interactions, which result in lower analyte retention. The selectivity of Oasis^? HLB is different to porous graphic carbon (Hypercarb^?), a conventional poly (styrene-divinylbenzene) porous polymer sorbent (PLRP-S 100) and two silica-based, octadecylsiloxane-bonded sorbents with a high and a low carbon loading, respectively. Because of selectivity differences no single sorbent is ideal for the extraction of analytes possessing a wide range of polar interactions. Oasis^? HLB is preferred for the extraction of low molecular mass and polar compounds, PLRP-S 100 for the extraction of higher molecular mass compounds of moderate polarity, and the silica-based octadecylsiloxane sorbent with a high carbon loading is the best compromise for the extraction of compounds that cover a wide polarity range. For methanol-water mixtures as a sample solvent PLRP-S 100 is the best general choice with Oasis^? HLB preferred for the extraction of strong hydrogen-bond acids. Hypercarb^? is shown to have favorable retention properties for solid-phase extraction with the except for its low surface area.     

Online capillary weak cation exchange enrichment hyphenated to nanospray mass spectrometry for quantitation of a basic pegvisomant derived peptide

A multidimensional LC–MS/MS configuration is described for targeted quantitation of a highly basic peptide chemically derived from pegvisomant as a surrogate for the intact protein. The method developed for an immunogenicity assay employed orthogonal separation of the target peptide commencing with flowing microgram quantities of total digests from a protein G extraction of human serum through a weak cation exchange (WCX) monolithic trap column. The basic peptide of interest was retained on the WCX column. Following a washing procedure, peptides were eluted with acetic acid and retained on a down-stream reverse phase trap compatible with online nanoflow LC–MS/MS. Such a LC configuration including the use of other sorbents such as strong cation exchange media is proposed as an enabling tool in assay development for quantitative protein bioanalysis by LC–MS/MS.     

Investigation of elutions from a surfactant immobilized solid phase extraction sorbent based upon the hydrophobicity of the trapped species

A unique solid phase extraction (SPE) sorbent having a removable “stationary phase” is presented. This removable phase consists of alkyltrimethylammonium surfactant, which is initially immobilized onto hydrophilic strong cation exchange resin. The surfactant chain through hydrophobic interactions extracts hydrophobic analytes in the same manner as conventional bonded alkyl moieties on silica-based non-polar sorbents. For the extraction of very hydrophobic species with conventional sorbents, solvents such methylene chloride and benzene are needed to break strong hydrophobic interactions for efficient elutions. These solvents however are toxic to the analyst and present a significant environmental concern. Using a removable “stationary phase”, hydrophobic interactions need not be broken between the analyte and the sorbent. In the presented approach, the surfactant (“stationary phase”) is removed via ion exchange with exchange ions in very mild aqueous-based and instrument compatible solutions. The analyte, being associated with the surfactant, is also removed in the process. Very efficient elutions of analytes, regardless of hydrophobicity, under mild and more favorable environmental conditions are a direct benefit of having a removable “stationary phase”. Rinse solution parameters explored include exchange cation type and concentration, and alcohol type and concentration. The extraction of three test molecules of varying hydrophobicity, naphthalene, pyrene and benzo(ghi)perylene, is investigated using this sorbent material.     

Determination of pharmaceuticals in wastewaters using solid-phase extraction-liquid chromatography-tandem mass spectrometry

Four different commercial sorbents for solid-phase extraction have been evaluated for the extraction of a group of acidic pharmaceuticals in terms of selectivity and capacity: Oasis hydrophilic-lipophilic balance (HLB), Oasis MAX (strong anion exchange), Oasis WAX (weak anion exchange) and a commercial available molecularly imprinted polymer specific for non-steroidal anti-inflammatory drugs. Among the sorbents studied, molecularly imprinted polymer proved to be very effective in the reduction of matrix interferences and the selective extraction of acidic pharmaceuticals, such as salicylic acid, ibuprofen, fenoprofen, diclofenac and naproxen, among others, from effluent wastewater samples. Moreover, molecularly imprinted solid-phase extraction protocol was applied to liquid chromatography coupled to tandem mass spectrometry (MS/MS) with the purpose of evaluating the clean-up effect on ion suppression/enhancement when the complexity of the samples increases and a reduction of this effect was observed. Molecularly imprinted solid-phase extraction followed by liquid chromatography coupled to ultraviolet detection and liquid chromatography coupled to tandem mass spectrometry validation methodologies with effluent wastewaters were developed, obtaining recoveries between 70 and 85% and limits of detection at low levels of μg/L (0.15-1 μg/L) and ng/L (0.5-2 ng/L), respectively. The final application of molecularly imprinted solid-phase extraction and liquid chromatography coupled to MS/MS detection showed the presence of acidic pharmaceuticals studied in this work in effluent wastewaters (相似文献   

单分散大孔亲水弱阳离子交换填料的制备及其对鱼精蛋白的分离纯化

以自制的5.0 μm单分散大孔亲水交联聚甲基丙烯酸环氧丙酯(PGMA/EDMA)微球为基质,对其表面进行化学改性,合成弱阳离子交换色谱填料(WCX)。详细考察了该填料对标准蛋白质的分离性能、表面亲水性能、稳定性和重现性以及流速对蛋白保留的影响。实验结果表明,该色谱填料对蛋白的分离性能、重现性及稳定性良好;在流速为3 mL/min时,采用线性梯度洗脱,6 min内可分离4种标准碱性蛋白质,以溶菌酶测定的该填料的动力学吸附容量为29.86 mg/g。将其用于鱼精蛋白的分离纯化,经反相高效液相色谱测定纯化后鱼精蛋白的纯度为99.2%;与商品Shodex IEC SP-825强阳离子交换色谱柱比较,纯化结果几乎一样。     

Preparation of weak cation exchange packings based on monodisperse poly(glycidyl methacrylate-<Emphasis Type="Italic">co</Emphasis>-ethylene dimethacrylate) beads and their chromatographic properties

The monodisperse, macroporous poly(glycidyl methacrylate- co-ethylene dimethacrylate) beads were synthesized by a single-step swelling and polymerization method. Based on this media, a weak cation exchange (WCX) stationary phase for HPLC was synthesized by a new chemically modified method. The prepared resin has advantages for biopolymer separation, high column efficiency, low column backpressure, high protein mass recovery, and good resolution for proteins. The measured bioactivity recovery for lysozyme was 98+/-5%. The dynamic protein loading capacity of the WCX packings was 17.3 mg g(-1). The experimental results show that the synthesized WCX resin has very weak hydrophobicity.     

On-line solid-phase extraction with a monolithic weak cation-exchange column and simultaneous screening of alpha1-adrenergic receptor antagonists in human plasma

An on-line SPE-HPLC method, using a monolithic poly(glycidyl methacrylate-co-ethylene glycol dimethacrylate) (poly(GMA-EDMA)) based weak cation-exchange (WCX) column, was developed for simultaneous determination of alpha1-adrenergic receptor antagonists in human plasma. The monolithic WCX column was prepared by an in-situ polymerization protocol and modified stepwise with ethylenediamine and chloroacetic acid. On connecting this column to an injection valve, an on-line SPE protocol could be established for removal of matrices (mainly proteins and lipids) and preconcentration of four alpha1-adrenergic receptor antagonists in human plasma. This method was validated and then used for determination of terazosin, alfuzosin, prazosin, and doxazosin in clinical plasma samples. For all analytes, each calibration curve was found to be linear over a range of 0.005-5 microg/mL (R>0.997), and the limit of detection for each analyte was 0.5 ng/mL. Recovery (>80%) and precision (RSD<15%) for inter- and intra-day assay were tested at three concentration levels of each analyte and showed acceptable results for quantitative assay. Real samples from hypertensive patients were monitored and results were in agreement with those of the conventional liquid-liquid extraction-HPLC method. Furthermore, due to its good permeability and biocompatibility, the monolithic WCX sorbent could be reused more than 300 times. The proposed method was especially appropriate for multi-analyte monitoring in plasma samples.     

Sample preparation on polymeric solid phase extraction sorbents for liquid chromatographic-tandem mass spectrometric analysis of human whole blood--a study on a number of beta-agonists and beta-antagonists

Alternative strategies for sample preparation of human blood samples were evaluated including protein precipitation (PP) and solid phase extraction (SPE) on Waters Oasis polymeric columns. Gradient chromatography within 15 min was performed on a Hypersil Polar-RP column combined with a Sciex API 2000 triple quadrupol instrument equipped with an electro-spray interface. Beta-agonists and beta-antagonists available on the Swedish market were included in the study. A combination of zinc sulphate and ethanol was found effective for PP. A clear supernatant was achieved that either could be injected directly on the LC-MS-MS system for analysis or transferred to a SPE column for further extraction and analyte concentration. Retention on the hydrophilic-lipophilic balanced sorbent HLB as well as the mixed mode cationic MCX and anionic MAX sorbents were investigated. On HBL the relative lipophilicity of the target analytes was investigated. At a high pH when the amino alcohols are deprotonised the more non-polar analytes (e.g., carvediol, betaxolol, bisoprolol and propranolol) were well retained on the sorbent and for the majority methanol content higher than 50% in water (v/v) was needed for elution. Some analytes though, with additional weak acidic functionalities (fenoterol, salbutamol, sotalol, and terbutaline) were poorly retained. On MAX the retention of these weak acids was improved when loaded under basic conditions but under neutral conditions analyte recoveries was comparable with HLB. On MCX all the analytes were well retained allowing a wash step of 100% methanol at neutral and low pH. By applying the supernatant from PP in combination with an additional portion of aqueous formic acid (2%) the analytes could be loaded and retained. High extraction recoveries were found for most analytes but for a few, significant losses were seen during PP (e.g., formoterol) and/or evaporation (e.g., fenoterol, formoterol, labetalol and terbutaline). The effectiveness of the sample preparation was evaluated by ESI ion-suppression studies by post column infusion of the target analyte. An ethanol zinc sulphate aq mixture was found to be more effective than acetonitrile, methanol or ethanol for PP of human whole blood samples. Beside suppression by salts in the front peak, only limited suppression from other artefacts such as more lipophilic compounds was found late in the chromatograms. Some tendency though to concentrate more lipophilic artefacts on the Oasis sorbents was seen. These findings show that the Oasis MCX sorbent is well suited for sample preparation of beta-agonists and beta-antagonists from human whole blood if the objective is to cover a great number of the analytes in the same assay.     

Synthesis and physicochemical properties of chelating sorbents containing functional groups of N-aryl-3-aminopropionic acids

Two types of chelating sorbents with different types of addition of iminodipropionate groups to a polymeric matrix were synthesized: carboxyethylated aminopolystyrene (sorbent 1) based on linear polystyrene and carboxyethylaminomethylpolystyrene (sorbent 2) based on the copolymer of styrene and divinylbenzene. The ionization constants and concentrations of functional groups of the sorbents (exchange capacity for hydrogen ions) were determined. The sorbents exhibit high selectivity for copper(II) ions with the maximum of sorption from ammonia—acetate buffer solutions lying in a range of pH 5.0–7.5. The time needed for a solution of copper(II)—sorbent system with continuous stirring to reach exchange equilibrium is 3.5 and 2 h for sorbents 1 and 2, respectively. The exchange capacity for copper(II) ions is 2.54 and 0.10 mmol g⁻¹, respectively. Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 5, pp. 800–806, May, 2006.     

“Click chemistry” preparation of WCX packings for protein separation

Click chemistry was applied to immobilize three kinds of alkyne-carboxylic acids onto azide-modified silica gel to prepare three novel stationary phases for weak cation exchange chromatography(WCX).The developed protocol combines the benefits of operational simplicity,exceptionally mild conditions and high surface loadings.Six kinds of standard proteins were separated completely on the novel packings.Compared with commercial WCX columns,the three kinds of novel WCX packings prepared by click chemistry approach have better resolution and selectivity.Lysozyme was purified successfully from egg white with the novel WCX column by one step.The purity was more than 97%and a high specific activity was achieved to be 81,435 U/mg.The results illustrate the potential of click chemistry for preparation of stationary phase for IEC.     

Fast purification process optimization using mixed-mode chromatography sorbents in pre-packed mini-columns

Pre-packed MediaScout MiniChrom columns of 2.5, 5 and 10 mL were investigated for screening three mixed-mode chromatography sorbents (HEA, PPA and MEP HyperCel). Packing performance was of good quality and the three sorbents displayed higher capacity than traditional HIC sorbents in physiological-like conditions. Each sorbent offered a unique selectivity. Bovine beta-lactoglobulin was partially purified after loading milk whey directly on HEA HyperCel sorbent. The combination of small pre-packed columns and SELDI-MS appeared to be a valuable strategy for high-throughput screening of chromatography sorbents and for enabling rapid process development and optimization.     

Comparison of mixed-mode anion-exchange performance of N-vinylimidazole-divinylbenzene sorbent

A newly synthesized copolymer based on N-vinylimidazole-divinylbenzene (NVIm-DVB) was evaluated as a mixed-mode anion-exchange sorbent for SPE, since the NVIm monomer apart from its hydrophilic properties can be protonated at a certain pH, and then performs as an anion-exchanger. To investigate the behavior of the NVIm-based sorbent, the SPE performance was evaluated under reversed-phase (RP), weak anion-exchange, and strong anion-exchange conditions. The results for the NVIm-DVB sorbent were also compared to commercial reference sorbents from each group: Oasis HLB, Oasis WAX, and Oasis MAX, respectively. SPE results from this evaluation showed that NVIm-DVB can be used as an RP material, compared to Oasis HLB. It also has the potential to act as a strong anion-exchange sorbent, compared to Oasis MAX, since under the proper conditions it was able to fraction and quantitatively recover a group of selected solutes.     

A novel sorbent for the determination of clenbuterol in bovine liver.

The use of three C18 sorbents in matrix solid phase dispersion (MSPD) for the determination of clenbuterol in bovine liver fortified at 5 ng g-1 is described. MSPD grade C18 sorbents give rise to more efficient blending and packing of the material for subsequent washing and analyte elution in comparison with a non-MSPD grade C18 sorbent. Following enzymatic deconjugation of the liver extracts, radioimmunoassay is used as the method of determination. The mean recovery of clenbuterol with all sorbents is comparable and within the range 86-96% in two intra-assay studies (n = 3). The liver extracts in each case are highly coloured. The variation in recovery is observed to be lowest with MSPD grade C18 (end-capped). This sorbent was used in further studies to evaluate the use of solid phase extraction (SPE), post MSPD, with normal phase aminopropyl or mixed mode cation exchange columns for extract purification. The mean recovery of clenbuterol (n = 4, inter-assay study) following MSPD and normal phase SPE clean-up was 95 +/- 15% and 89 +/- 9% at fortification levels of 1 and 2.5 ng g-1, respectively.     

Development of imprinted materials for the selective extraction of nitroaromatic explosives

Molecularly imprinted sorbents were synthesized and used as selective extraction sorbents for the analysis of nitroaromatic explosives. Their synthesis by radical polymerization using organic monomers and by sol–gel approach using organosilanes was considered to develop a selective sorbent. The sol–gel approach with phenyltrimethoxysilane (PTMS) as monomer and 2,4-dinitrotoluene (2,4-DNT) as template gave the most promising results. An optimized procedure adapted to the selective treatment of aqueous samples was then developed and applied to various target explosives. For the first time four nitroaromatic compounds were retained on the molecularly imprinted silica (MIS) with extraction recoveries between 29% and 81%, while only low recoveries were obtained on the non-imprinted sorbent, thus highlighting the high degree of selectivity. The MIS was then used for the clean-up of a sample containing motor oil spiked with 2,4-DNT and 2,4,6-trinitrotoluene (2,4,6-TNT). The results were compared with those obtained using a conventional sorbent (Oasis HLB). The cleanest chromatogram obtained using the MIS emphasized the high potential of the MIS as selective sorbent.     

Synthesis of Monodisperse Poly（glycidylmethacrylate-co-ethylene dimethacrylate） Beads and Their Application in Separation of Biopolymers

Introduction Since 19541 the polymeric separation media has attracted much attention due to their chemical stability over the entire pH range. The rigid, highly cross-linked styrene copolymers were first used for chromatography by Moore.2 The macroporous copolymers currently available are not only chemically stable but also more resistant to mechanical forces prevailing in a column and therefore are comparable to the traditional packings based on silica gel. Most polymer separation media are …     

Preparation and characterization of a novel dual‐retention mechanism mixed‐mode stationary phase with PEG 400 and succinic anhydride as ligand for protein separation in WCX and HIC modes

A novel dual‐retention mechanism mixed‐mode stationary phase based on silica gel functionalized with PEG 400 and succinic anhydride as the ligand was prepared and characterized by infrared spectra and elemental analysis. Because of the ligand containing PEG 400 and carboxyl function groups, it displayed hydrophobic interaction chromatography (HIC) characteristic in a high‐salt‐concentration mobile phase, and weak cation exchange chromatography (WCX) characteristic in a low‐salt‐concentration mobile phase. As a result, it can be employed to separate proteins with both WCX and HIC modes. The resolution and selectivity of the stationary phase was evaluated under both HIC and WCX modes with protein standards, and its performance was comparable to that of conventional ion‐exchange chromatography and HIC columns. The results indicated that the novel dual‐retention mechanism column, in many cases, could replace two individual WCX and HIC columns as a ‘2D column’. In addition, the mixed retention mechanism of proteins on this ‘2D column’ was investigated with stoichiometric displacement theory for retention of solute in liquid chromatography in detail in order to understand why the dual‐retention mechanism column has high resolution and selectivity for protein separation under WCX and HIC modes, respectively. Based on this ‘2D column’, a new 2DLC technology with a single column was developed. It is very important in proteome research and recombinant protein drug production to save column expense and simplify the processes in biotechnology. Copyright © 2013 John Wiley & Sons, Ltd.     

Comparison of several solid-phase extraction sorbents for continuous determination of amines in water by gas chromatography-mass spectrometry

A semiautomatic method has been proposed for the determination of different types of amines in water samples including anilines, chloroanilines, N-nitrosamines and aliphatic amines. The analytes were retained on a solid-phase extraction sorbent column and after elution, 1 μL of the extract was analysed by gas chromatography coupled with electron impact ionization mass spectrometry. A systematic overview is given of the advantages and disadvantages of several sorbents (LiChrolut EN, Oasis HLB, RP-C₁₈, graphitized carbon black, fullerenes and nanotubes) in the retention of amine compounds and based on sensitivity, selectivity and reliability. The retention efficiency for the studied amines was higher (ca. 100%) with LiChrolut EN and Oasis HLB than it was with RP-C₁₈ and fullerenes (53 and 62%, respectively, on average). Detection limits of 0.5-16 ng L⁻¹ for the 27 amines studied were obtained when using a sorbent column containing 75 mg of LiChrolut EN for 100 mL of sample, the RSD being lower than 6.5%. The method was applied with good accuracy and precision in the determination of amines in various types of water including river, pond, tap, well, drinking, swimming pool and waste.     

Comparison of Hydrophilic Polymeric Sorbents for On-Line Solid-Phase Extraction of Polar Compounds from Aqueous Samples

Two 4-vinylimidazole-divinylbenzene (4VIm-DVB) polymers were synthesized and applied as sorbents for on-line solid-phase extraction (SPE) followed by liquid chromatography for analyzing polar compounds in aqueous samples. The new sorbents (4VIm-DVB) were compared to another sorbent that had been previously synthesized by our group (N-vinylimidazole-divinylbenzene (NVIm-DVB)) and to the commercial OASIS^® HLB and Strata^TM X. All the sorbents enabled 100 mL of sample to be on-line concentrated with good recoveries for the studied polar compounds. Real water samples were analyzed using NVIm-DVB and OASIS^® HLB as SPE sorbent, for which the best results were obtained.     

Determination of nicardipine and amlodipine in human plasma using on-line solid-phase extraction with a monolithic weak cation-exchange column

An on-line solid-phase extraction (SPE)-HPLC method was developed for simultaneous screening of nicardipine and amlodipine in human plasma. A short monolithic poly(glycidyl methacrylate-co-ethylene glycol dimethacrylate) [p(GMA-EDMA)]-based weak cation-exchange (WCX) column was prepared and employed as the selective extraction sorbent, which exhibited good permeability and biocompatibility. During the on-line SPE protocol, high-abundance proteins (human serum albumin, immunoglobulin G, immunoglobulin A and transferrin) and most matrixes in plasma were fast removed while nicardipine and amlodipine were effectively trapped on this monolithic column. Furthermore, the monolithic WCX sorbent could be continuously reused more than 300 times without obvious changes in analytes extraction and proteins cleanup. The proposed method was linear over a range of 0.5-50.0 ng mL⁻¹ for both analytes with a linear regression coefficient greater than 0.998, and the limit of detection (LOD) for each analyte was 0.2 ng mL⁻¹. Validation assays also demonstrated acceptable precision and adequate recovery for simultaneous quantitative screening of nicardipine and amlodipine in human plasma. Real plasma samples from hypertensive patients receiving a dosing of 5 mg antagonists were examined by using the proposed method. Results indicated that the on-line SPE-HPLC method could be applied for simultaneously monitoring of nicardipine and amlodipine in clinical plasma samples.