Preparation and Evaluation of Propranolol-Imprinted Monolithic Stationary Phase by In Situ Technique and Application in Analysis of Propranolol in Biological Samples

Abstract

A propranolol molecule-imprinted monolithic stationary phase (MIMSP) was synthesized by in situ technique. The recognition mechanism of the polymers and the influences of some chromatographic conditions were examined by high-performance liquid chromatography (HPLC). The imprinted polymers showed much higher selectivity for β-blockers than the nonimprinted polymers (NIPs) did, which proves the successful preparation of propranolol-imprinted polymers by using an in situ technique. Then, this technique was used to prepare a molecularly imprinted polymer solid-phase extraction column to concentrate propranolol from biological samples. The results showed that the imprinted solid-phase extraction column could selectively enrich and purify propranolol from biological samples, such as plasma.     

Solid-phase extraction of esculetin from the ash bark of Chinese traditional medicine by using molecularly imprinted polymers

A molecularly imprinted polymer solid-phase extraction method is used to extract esculetin from the ash bark of Chinese traditional medicine. Ratio of ethanol and water as washing solution were investigated. Data of accumulative adsorption on molecularly imprinted polymers from the continuous loading experiment suggests that there are two different kinds of recognition sites in molecularly imprinted polymers. By selecting the washing and eluting solution a scheme was designed to separate esculetin and its analogues including esculin, coumarin, 7-methoxylcoumarin and daphnetin. Finally, by applying the revised scheme esculetin was extracted from the ash bark of Chinese traditional medicine that was purchased from two big drugstores, respectively, with both molecularly imprinted polymers and non-molecularly imprinted polymers.     

Preparation of molecularly imprinted polymers for strychnine by precipitation polymerization and multistep swelling and polymerization and their application for the selective extraction of strychnine from nux‐vomica extract powder

Monodisperse molecularly imprinted polymers for strychnine were prepared by precipitation polymerization and multistep swelling and polymerization, respectively. In precipitation polymerization, methacrylic acid and divinylbenzene were used as a functional monomer and crosslinker, respectively, while in multistep swelling and polymerization, methacrylic acid and ethylene glycol dimethacrylate were used as a functional monomer and crosslinker, respectively. The retention and molecular recognition properties of the molecularly imprinted polymers prepared by both methods for strychnine were evaluated using a mixture of sodium phosphate buffer and acetonitrile as a mobile phase by liquid chromatography. In addition to shape recognition, ionic and hydrophobic interactions could affect the retention of strychnine in low acetonitrile content. Furthermore, molecularly imprinted polymers prepared by both methods could selectively recognize strychnine among solutes tested. The retention factors and imprinting factors of strychnine on the molecularly imprinted polymer prepared by precipitation polymerization were 220 and 58, respectively, using 20 mM sodium phosphate buffer (pH 6.0)/acetonitrile (50:50, v/v) as a mobile phase, and those on the molecularly imprinted polymer prepared by multistep swelling and polymerization were 73 and 4.5. These results indicate that precipitation polymerization is suitable for the preparation of a molecularly imprinted polymer for strychnine. Furthermore, the molecularly imprinted polymer could be successfully applied for selective extraction of strychnine in nux‐vomica extract powder.     

Selective separation of active inhibitors of epidermal growth factor receptor from Caragana jubata by molecularly imprinted solid-phase extraction

A feasibility research was performed to study the possibilities of using a molecularly imprinted polymer as sorbent material in solid-phase extraction for the separation of active inhibitors of epidermal growth factor receptor (EGRF) from Caragana Jubata, a Chinese traditional Tibetan medicine. A molecularly imprinted polymer using quercetin, an active anti-EGFR inhibitor (IC50 = 15 microM), as the template and acrylamide as the functional monomer was prepared. The polymer was evaluated as a selective sorbent in molecularly imprinted solid-phase extraction. The EtOAc extract of Caragana Jubata was loaded on the polymer, and two novel active anti-EGFR inhibitors were found to be selectively retained after washing the polymer with appropriate solvent to disrupt the non-specific interactions occurring between the sample and the polymer matrix, which were identified as (E)-piceatannol (IC50 =4.9 microM) and butein (IC50 = 10 microM). The present work affords us a new potential method for selective separation of bioactive components from herb by using molecularly imprinted polymer as a solid-phase extraction adsorbent.     

分子印迹技术制备石油有机硫组分固相萃取剂的研究

用分子印迹技术合成了对石油有机硫组分二苯并噻吩(Dibenzothiophene，DBT)具有高效选择性的分子模板聚合物(Molecularly Imprinted Polymer，MIP)，通过静态吸附的方法研究了不同功能单体和致孔剂及其用量对模板聚合物特异性识别能力的影响．实验表明，以4-乙烯基吡啶(4-VP)为功能单体，在甲苯溶剂中聚合得到的固相萃取剂对DBT具有较大的吸附富集能力和识别特性．其饱和吸附容量达到48．3mg／g．     

Magnetic molecularly imprinted polymer based on dynamically established metal-mediated binding sites for the determination of trace captopril in rat plasma

Therapeutic drug monitoring of captopril, which is a commonly used antihypertensive agent in clinical practice, is necessary. However, matrix effect-induced pretreatment is the bottleneck for determination. Metal-mediated molecularly imprinted polymers, an essential branch of molecularly imprinted polymers with better specificity and selectivity, have been used to separate/enrich analytes from complex matrices. In this work, Cu²⁺ was introduced to dynamically establish the binding sites of metal-mediated molecularly imprinted polymer towards captopril. All evidence demonstrated that the metal-mediated molecularly imprinted polymer based on Cu²⁺ coordination obtained a higher adsorption capacity (81.23 mg/g), faster adsorption rate (adsorption equilibrium within 50 min), and better selectivity (with the unrecognized analog). Subsequently, the Cu²⁺-mediated molecularly imprinted polymer was used as dispersive molecularly imprinted solid-phase extraction to successfully establish an analytical platform for the determination of trace captopril in rat plasma. The enrichment factor was up to 20, the detection limit was as low as 0.16 μg/ml, and the average recovery was in the range of 87.51%–98.28% with a relative standard deviation of less than 3.29%. This study provides a promising reference for the preparation of selective adsorbents to improve pretreatment.     

Selective sample treatment using molecularly imprinted polymers

The molecularly imprinted polymers (MIPs) are synthetic polymers possessing specific cavities designed for a target molecule. By a mechanism of molecular recognition, the MIPs are used as selective sorbents for the solid-phase extraction of target analytes from complex matrices. MIPs are often called synthetic antibodies in comparison with immuno-based sorbents; they offer some advantages including easy, cheap and rapid preparation and high thermal and chemical stability. This review describes the use of MIPs in solid-phase extraction with emphasis on their synthesis, the various parameters affecting the selectivity of the extraction, their potential to selectively extract analytes from complex aqueous samples or organic extracts, their on-line coupling with LC and their potential in miniaturized devices.     

An amobarbital molecularly imprinted microsphere for selective solid-phase extraction of phenobarbital from human urine and medicines and their determination by high-performance liquid chromatography

A 40-60 microm amobarbital molecularly imprinted microsphere, used as a solid-phase selective sorbent for extracting phenobarbital from human urine and medicines, was prepared by a suspension polymerization method. A series of binding studies was performed in order to find optimal loading, washing and eluting conditions for solid-phase extraction. Under optimal conditions, good recoveries of phenobarbital in samples were obtained. Normally, molecularly imprinted polymers, prepared in bulk, require laborious work. Significant losses occur during the procedure of grinding and washing. In this work all molecularly imprinted polymers made into microsphere could be utilized, and the cost of the template was reduced too (the price of phenobarbital is twice that of amobarbital). As the phenobarbital to be extracted was different from the template molecule amobarbital, the interference caused by template leaking could be avoided in the assay.     

Molecularly imprinted polymers for solid-phase extraction and solid-phase microextraction: recent developments and future trends

Molecularly imprinted polymers (MIPs) are synthetic polymers having a predetermined selectivity for a given analyte, or group of structurally related compounds, that make them ideal materials to be used in separation processes. In this sense, during past years a huge amount of papers have been published dealing with the use of MIPs as sorbents in solid-phase extraction, namely molecularly imprinted solid-phase extraction (MISPE). Although the majority of these papers were restricted to describe the use of different templates for different applications, several attempts proposing new alternatives to minimize the inherent drawbacks of the preparation and use of MIPs (i.e. template bleeding, tedious synthesis procedure, etc.) have been reported. Thus, this paper does not pretend to be a collection of MISPE-related papers but to give an overview on the significant attempts carried out during recent years to improve the performance of MIPs in solid-phase extraction. In addition, the use of MIPs packed in high performance liquid chromatography (HPLC) columns for the direct injection of crude sample extracts and the preparation of imprinted fibres for solid-phase microextraction will be also discussed.     

Molecularly imprinted solid-phase extraction of cocaine metabolites from aqueous samples

Cocaine is a well-known drug of abuse which, when ingested nasally or by smoking, undergoes a number of biotransformation and degradation reactions. In the present work, a synthetic analogue of the cocaine metabolite benzoylecgonine was prepared and used as a template molecule in the preparation of a series of molecularly imprinted polymers (MIPs). Molecularly imprinted solid-phase extraction (MISPE) conditions were established under which benzoylecgonine in aqueous samples could be selectively extracted and quantified at clinically relevant concentrations (μg/ml). Under optimised MISPE conditions, recoveries of analyte were high (>70%) and excellent discrimination between imprinted and non-imprinted materials observed.     

Synthesis and characterization of photo‐responsive magnetic molecularly imprinted microspheres for the detection of sulfonamides in aqueous solution

A dual responsive molecularly imprinted polymer sensitive to both photonic and magnetic stimuli was successfully prepared for the detection of four sulfonamides in aqueous media. The photoresponsive magnetic molecularly imprinted polymer was prepared by surface imprinting polymerization using superparamagnetic Fe₃O₄ nanoparticles functionalized with a silica layer as a support, water‐soluble 4‐[(4‐methacryloyloxy)phenylazo]benzenesulfonic acid as the functional monomer, and sulfadiazine as the template. The magnetic molecularly imprinted polymers showed specific affinity to sulfadiazine and its structural analogs in aqueous media. Upon alternate irradiation at 365 and 440 nm, the quantitative bind and release of the four sulfonamides by magnetic molecularly imprinted polymers occurred. Furthermore, the prepared magnetic molecularly imprinted polymers were used as solid‐phase extraction material selectively extracted the four sulfonamides from water samples with good recoveries. Thus, a simple, convenient, and reliable detection method for sulfonamides in the environment based on responsive magnetic molecularly imprinted polymers was successfully established.     

On-line solid-phase extraction with molecularly imprinted polymers to selectively extract substituted 4-chlorophenols and 4-nitrophenol from water

Three polymers have been synthesised using 4-chlorophenol (4-CP) as the template, following different protocols (non-covalent and semi-covalent) and using different functional co-monomers, 4-vinylpyridine (4-VP) and methacrylic acid (MAA). The polymers were evaluated to check their selectivity as molecularly imprinted polymers (MIPs) in solid-phase extraction (SPE) coupled on-line to liquid chromatography. The solid-phase extraction procedure using MIPs (MISPE), including the clean-up step to remove any interferences, was optimised. The 4-VP non-covalent polymer was the only one which showed a clear imprint effect. This MIP also showed cross-reactivity for the 4-chloro-substituted phenols and for 4-nitrophenol (4-NP) from a mixture containing the 11 priority EPA (Environmental Protection Agency) phenolic compounds and 4-chlorophenol. The MIP was applied to selectively extract the 4-chloro-substituted compounds and 4-NP from river water samples.     

Selective Preconcentration of Mezlocillin from Eggs by Molecularly Imprinted Polymers on Silica

Surface molecularly imprinted polymers for mezlocillin were used for the selective solid-phase extraction of mezlocillin from eggs. The molecularly imprinted polymers were prepared using mezlocillin as the template, methacrylic acid as the functional monomer, ethylene glycol dimethacrylate as the cross-linker, silica particle as the support and were characterized by infrared spectroscopy, scanning electron microscopy, and thermogravimetric analysis. The adsorption properties of polymers were investigated in detail. The obtained polymers provided high imprinting efficiency to mezlocillin with an imprinting factor of 3.72 and were used as selective sorbents for mezlocillin. Molecularly imprinted solid-phase extraction followed by high-performance liquid chromatography was used for the determination of mezlocillin in eggs and the conditions were evaluated. The average recovery of mezlocillin in fortified eggs was between 69.8 and 82.3% with a relative standard deviation less than 6.9%. The results demonstrate the application of molecularly imprinted polymers for the extraction of mezlocillin from biological samples.     

分子印迹技术在样品前处理中的应用

分子印迹聚合物具有选择性高、稳定性好及制备简单的特点,可用于生物、医药、环境样品等复杂基体中痕量分析物的高选择性分离与富集,因此在样品前处理中的应用特别引人关注.本文介绍了分子印迹技术的基本原理,综述了分子印迹技术在样品前处理中应用的研究进展.     

Highly selective fenuron-imprinted polymer with a homogeneous binding site distribution prepared by precipitation polymerisation and its application to the clean-up of fenuron in plant samples

In the present study, two different imprinted polymers were synthesised by precipitation polymerisation using methacrylic acid (MAA) or 4-vinylpyridine (VP) as monomer and fenuron (FEN, a phenylurea herbicide) as template. After template removal, their ability to recognise fenuron was evaluated and the optimum loading, washing and elution conditions were established. From this study, it was concluded that imprinted binding sites were not formed in the vinylpyridine-based polymer. However, methacrylic acid-based polymer was able to recognise fenuron with high affinity and to selectively retain it from a mixture of several phenylurea herbicides. In addition, different rebinding experiments were carried out and the experimental binding isotherms were fitted to the Langmuir-Freundlich (LF) isotherm in order to assess the binding site distribution. It was concluded that the methacrylic acid-based polymer possesses a homogeneous binding site distribution and permits to achieve quantitative recoveries in a wide concentration range in molecularly imprinted solid-phase extraction (MISPE) processes. The developed MISPE procedure using methacrylic acid-based polymeric micro-spheres was evaluated for the trace-enrichment and clean-up of fenuron from plant sample extracts.     

The application of pseudo template molecularly imprinted polymer to the solid-phase extraction of cyromazine and its metabolic melamine from egg and milk

The combination of molecularly imprinted polymer with high performance liquid chromatography has been developed to determine cyromazine and its metabolic melamine in some samples. However, the potential risk of template leakage used in molecularly imprinted polymer is a major disadvantage. To solve this problem, 2-(4,6-diamino-1,3,5-triazin-2-ylamino) ethanethiol disulfide, a molecule that shares the similar imprinting sites with cyromazine and melamine, was selected as pseudo template to prepare molecularly imprinted polymer. Methacrylic acid, ethylene glycol dimethyl acrylate and toluene were selected as functional monomer, crosslinker and porogen, respectively. The molecular recognition property and binding capability of cyromazine and melamine were evaluated by adsorption test and Scatchard analysis. The results showed that the molecularly imprinted polymer based on pseudo template had more excellent affinity and selectivity for cyromazine and melamine. The resulting molecularly imprinted polymer was used as a solid-phase extraction material to enrich cyromazine and melamine in egg and milk samples for high performance liquid chromatography analysis. The solid-phase extraction process was carefully optimized. It was found that when different concentration of cyromazine and melamine standards were spiked into samples, satisfactory recovery rate of cyromazine and melamine were obtained as 85.6-98.8% with relative standard deviation <5.5%.     

替代模板分子印迹技术在样品前处理中的应用

分子印迹聚合物具有抗恶劣环境、选择性高、稳定性好等特点,广泛应用于复杂样品的前处理。采用结构类似物作为替代模板分子,可以解决分子印迹聚合物制备时目标物溶解性差的问题,替代模板分子印迹聚合物不仅对目标分析物具有选择性识别能力,还可以避免模板泄露对痕量分析造成的影响。本文综述了替代模板分子印迹技术在样品前处理中的应用进展,包括替代模板分子印迹技术在固相萃取、固相微萃取、色谱固定相、基质固相分散萃取中的应用,最后对替代模板分子印迹技术在未来的样品前处理中的研究进行了展望。     

分子印迹固相萃取技术在天然产物有效成分分离分析中的应用进展

天然产物体系复杂,尤其是一些活性成分含量较低,采用一般的方法对其进行分离富集难度较大。分子印迹聚合物具有良好的亲和性和专一的选择性,将分子印迹固相萃取技术应用于天然药物资源样品前处理过程,能够选择性地分离富集复杂基质中的目标成分。本文对近几年分子印迹固相萃取技术在天然产物有效成分分离分析中的应用进行了总结,分析物包括黄酮类、多元酚类、生物碱类、有机酸类、苯丙素类、萜类以及其他一些类型的生物活性成分。     

Synthesis and properties of molecular imprints of darifenacin: The potential of molecular imprinting for bioanalysis

Summary A molecularly imprinted polymer has been developed which subsequently demonstrated an ability to selectively retain darifenacin (UK-88, 525-S) from aqueous acetonitrile when used as a stationary phase in HPLC columns and as a packing in solid-phase extraction cartridges. The imprinted polymer is applicable to a wide range of analytical methods including extraction from plasma, purification of radiolabelled UK-88,525, chiral separations and separation of metabolites and structural analogues. The polymer is able to extract darifenacin directly from a protein-precipitated human plasma/acetonitrile (1:1 v/v) mixture with 100% recovery. The imprinted polymer can also effect a repurification of¹⁴C-labelled darifenacin. The drawbacks of molecular imprints for ultra-trace bioanalysis (in the sub-nanogram/mL range) are discussed. These centre on the difficulty of removing all the template from the polymer and the consequent effects of template bleed on assay precision and accuracy when used as solid-phase extraction cartridges. Possible solutions to this problem are considered. Presented at: Affinity Chromatography Conference, Cambridge, UK, July 1–3, 1997.