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Development of a novel magnetic molecularly imprinted polymer coating using porous zeolite imidazolate framework-8 coated magnetic iron oxide as carrier for automated solid phase microextraction of estrogens in fish and pork samples Hangzhen Lan a , Ning Gan b,∗ , Daodong Pan a,∗∗ , Futao Hu a , Tianhua Li b , Nengbing Long b , Haoyu Shen c , Yinjie Feng b a
Faculty of Marine Science, Ningbo University, Ningbo 315211, China Faculty of Material Science and Chemical Engineering, Ningbo University, Ningbo 315211, China c Ningbo Institute of Technology, Zhejiang University, Ningbo 315100, China b
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Article history: Received 16 May 2014 Received in revised form 16 July 2014 Accepted 31 August 2014 Available online xxx Keywords: Zeolite imidazolate framework-8 coated magnetic iron oxide Molecularly imprinted polymer coating Automated solid-phase microextraction Estrogens High performance liquid chromatography
a b s t r a c t A high-performance magnetic molecularly imprinted polymer (MIP) coating using zeolite imidazolate framework-8 coated magnetic iron oxide (Fe3 O4 @ZIF-8) as a carrier was developed for simultaneous automated solid phase microextraction of four estrogens in 24 food samples. The coating material, abbreviated as MZMIP, was synthesized through time-efficient layer-by-layer assembling of ZIF-8 and MIP film on Fe3 O4 particles. It was characterized and automatically coated on the surface of SPME fibers by electromagnetic bonding. The extraction performance, reusability, repeatability, and validity of the MZMIP–SPME system was evaluated for high-throughput analysis of estrone (E1), estradiol (E2), estriol (E3), and ethinylestradiol (EE2). Various factors affecting the quality of MZMIP coating were optimized. Compared with traditional magnetic MIP coating based on Fe3 O4 @SiO2 carrier, the MZMIP coating exhibited high extraction capacity and quick adsorption and desorption kinetics to E1, E2, E3, and EE2 owing to the larger amount of imprinting sites in MZMIP. Under optimum conditions, the proposed system requires only 25 min for pretreatment of all 24 samples (62.5 s per sample). The limits of detection and quantitation of the proposed automated system for analysis were found to range from 0.4 to 1.7 and 1.1 to 6.2 ng g−1 , respectively. During analysis of spiked fish and pork, the new coating showed better recovery and selectivity compared with Fe3 O4 @SiO2 @MIP (MMIP) and commercially available SPME. The results indicated that the MZMIP coating could be effectively employed for pretreatment of ultra-trace level of estrogens in food. © 2014 Elsevier B.V. All rights reserved.
1. Introduction Estrogens are a group of potent endocrine-disrupting chemicals, which include naturally occurring estrone (E1), estradiol (E2), estriol (E3), and synthetic ethinylestradiol (EE2). Especially EE2, as a kind of xenoestrogens, has been identified as the most potent estrogenic chemical in all endocrine-disrupting chemicals until date [1]. Estrogens could enter human body through the food chain, disturb the normal endocrine system, and alter the normal reproduction and development of human beings [2,3]. Further, those compounds
∗ Corresponding author. Tel.: +86 574 87608347; fax: +86 574 87608347. ∗∗ Corresponding author. Tel.: +86 574 87609987; fax: +86 574 87609987. E-mail addresses: [email protected] (N. Gan), [email protected] (D. Pan).
have attracted large societal and political attention because of their widespread presence in certain food matrices, such as fish and pork [4–6]. Thus, it is of great significance for food safety supervision to develop sensitive, selective, and simple methodologies to determine estrogens in food. Solid-phase micro-extraction (SPME), pioneered by Arthur and Pawliszyn in 1990s [7], has been widely applied for monitoring estrogens either in food samples, in environment matrices or in biologic samples [8–12]. SPME has many advantages because of its integration of analytes extraction, pre-concentration, and sample clean up in a single step [13–15]. To improve the sample preparation strategies for quick analysis of complex samples and high-throughput analysis, there has been a growing need for an automated SPME method. Recently, Pawliszyn’s research groups have developed many automated SPME devices in multi-well
http://dx.doi.org/10.1016/j.chroma.2014.08.096 0021-9673/© 2014 Elsevier B.V. All rights reserved.
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format using a robotic autosampler to prepare several samples in parallel [16–22]. In our previous work [23], we developed a novel automated SPME device that could simultaneously treat up to 24 samples. In addition, a highly selective and sensitive magnetic molecularly imprinted polymers MMIP-SPME analytical method was established using Fe3 O4 @SiO2 @MIP (MMIP) as a coating of SPME and coupled with high performance liquid chromatography (HPLC) for detection. Fe3 O4 @SiO2 was applied as the matrix of MMIP due to the modifiable silicon hydroxyl on the surface of SiO2 nanoparticle. However, the grafting and molecularly imprinted polymerization procedures on SiO2 were time-consuming. In Ding’s work [24], to modify MIP film using MAA as functional monomer on the surface of Fe3 O4 @SiO2 , the ␥-methacryloxypropyl trimethoxysilane (KH570) was employed to introduce grafting C C group to the surface of Fe3 O4 @SiO2 by reacting in toluene for 24 h at 120 ◦ C. The molecularly imprinted polymerization procedure even reached up to 38 h. In the process, a large volume of organic solvent was needed. To simplify the immobilizing procedure of MIP film on superparamagnetic iron oxide nanoparticles as coating for automated MMIP-SPME, choosing a new carrier instead of Fe3 O4 @SiO2 and polymer monomer instead of MAA was a big challenge. Metal–organic frameworks (MOFs) have recently received considerable attention due to their unusual properties such as large surface area, good thermal stability, and the availability of inpore functionality and outer-surface modification [25]. Owing to the above merits, Kun’s group reported MIP preparation by using MOF-5 as matrix, and the synthesized MOF@MIP showed a homogeneous polymer film, thermal stability, and exhibited a higher specific surface area and a faster transfer-mass speed compared with that of the bulk MIP [26]. Zeolite imidazolate frameworks (ZIFs), a new class of microporous MOFs are synthesized by the crystallization of a transition metal species bound to the nitrogen atom of an imidazolate compound [25]. ZIF-8, which has the formula Zn(MIM)2 (MIM: 2-methylimidazole), has hydrophobic pores [27]. ZIF-8 has been reported to possess good extraction capacity to steroidal hormones due to the presence of a benzene ring by forming a coordinate bond and intermolecular – interactions [28]. Considering the superior properties of ZIF-8 and their potential absorbing capacity for the benzene series and related organic groups, the design and synthesis of magnetic ZIF-8 as a carrier of coating are especially desirable for automated SPME. Some novel monomers that can auto-polymerize without cross-linking or an initiator agent, such as dopamine [29], polydopamine [30,31], or 3-aminobenzeneboronicacid (APBA) [32–34], can be used to quickly prepare the MIP film for the automated SPME method. APBA is an attractive functional monomer that is water-soluble and provides a mild aqueous environment during polymerization [34]. Moreover, the benzene rings of ABPA can be absorbed on ZIF-8, which has imidazole ring, through – stacking interaction. Thus the MIP film using APBA polymer can be easily immobilized on Fe3 O4 @ZIF-8 without any grafting procedure, which can greatly simplify the preparation step of MIP film on the carrier. Furthermore, the MIP film is prepared by an organic solvent-free synthesis. Moreover, the template (E2) can be uniformly and stably distributed on the surface of porous ZIF-8 through – stacking interactions to simply the MIP film immunization procedure. In this work, to develop a new coating with high extraction capacity and stability for the automated SPME system, we prepared a novel MZMIP coating using Fe3 O4 @ZIF-8 as the carrier and ABPA as the functional monomer. The MMIP coating, using Fe3 O4 @SiO2 as the carrier, was also synthesized to compare the extraction performance with the new MZMIP coating. The MZMIPcoated automated SPME system coupled with HPLC was evaluated for high-throughput analysis of estrogens (E1, E2, E3, and EE2) in real fish and pork samples.
2. Experimental 2.1. Chemical reagents and materials In this work, estrone (E1, 97%), estradiol (E2, 99%), estriol (E3, 98%), and ethinyloestradiol (EE2, 98%), -naphthol (-nap, 99%) and poly (styrenesulfonate, sodium salt) (PSS, 30 wt%) were obtained from Sigma–Aldrich (Shanghai, China). 3-Amino phenyl boronic acid (APBA) was from J&K Chemical Co. Ltd. Acetonitrile (HPLC grade) was acquired from CNW Technologies (Dusseldorf, Germany). 2-methylimidazole (H-MeIM, 99%) were purchased from Aladdin Chemistry (Shanghai, China). Zinc nitrate hexahydrate (Zn(NO3 )2 ·6H2 O, 99%), iron (III) chloride hexahydrate (FeCl3 ·6H2 O, 99%), sodium acetate (NaAC, 99%) and ethylene glycol ((HOCH2 )2 , 99%) were purchased from Sinopharm Chemical Reagent Co., Ltd., China. All other reagents were of analytical grade. Ultrapure water (Hangzhou Wahaha Foods Co. Ltd., Hangzhou, China) was used throughout this work. A mixed stock solution of the four estrogens (E1, E2, E3, and EE2) was prepared in methanol at a concentration of 100 mg L−1 and stored at 4 ◦ C in darkness. All solutions used for HPLC mobile phase were filtered through a nylon 0.22 m filter prior to use. The commercial SPME fibers with 85 m polyacrylate (PA), 85 m carboxen/polydimethylsiloxane (CAR/PDMS), 65 m polydimethylsiloxane/divinylbenzene (PDMS/DVB), and 50/30 m carboxen/divinylbenzene/polydimethylsiloxane (CAR/DVB/PDMS) coatings, which were purchased from Supelco (Bellefonte, PA, USA) were used for the comparison study. 2.2. Synthesis of MZMIP core–shell particles 2.2.1. Preparation of Fe3 O4 particles The Fe3 O4 particles were synthesized as described in our previous work [23]. 2.2.2. Preparation of Fe3 O4 @ZIF-8 particles The core–shell Fe3 O4 @ZIF-8 particles were prepared according to Zhang et al. [35]. In a typical procedure, 0.05 g Fe3 O4 was added to 150 mL aqueous solution contains of 0.3% poly (styrenesulfonate, sodium salt) (PSS) under ultrasonication for 30 min. The resultant Fe3 O4 @PSS particles were recovered by an external magnetic field and washed with purified water for three times, then redispersed in a mixture containing 150 mL methanol, 1.125 g Zn(NO3 )2 ·6H2 O and 3.11 g 2-methylimidazolate under stirring and the reaction was allowed to proceed at 50 ◦ C for 3 h for ZIF-8 shell growth. Finally, the obtained Fe3 O4 @ ZIF-8 nanoparticles were separated by a magnet and washed with ethanol. 2.2.3. Preparation of MZMIP particles MZMIP was prepared as follows. E2 (0.1 mmol) as the template was first dissolved in 1.25 mL acetonitrile, then mixed with 5 mL phosphate buffer sodium (PBS, pH = 7.2), which contains 0.5 mmol APBA as the functional and cross-linking monomer, and the mixture was then incubated at room temperature for 1 h. After adding Fe3 O4 @ZIF-8 (0.04 g), the solution was incubated for 2 h at room temperature. Prior to use, the Fe3 O4 @ZIF-8 particles were subjected to extensive deionized water and absolute ethanol, and washed thoroughly. Subsequently, a 100 mM aqueous solution of ammonium persulfate (6.5 mL) as initiator was slowly added dropwise to the above solution for about 20 min and the polymerization process was executed at room temperature. After 8 h, Fe3 O4 @ZIF8@MIP (MZMIP) particles were obtained. Finally, MZMIP was collected magnetically, and the template was removed using 20 mL methanol/acetic acid (4/1, v/v) solution (with shaking) until no E2 adsorption was detected by HPLC. The particles were washed with water three times again and vacuum dried at 60 ◦ C.
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The molecularly non-imprinted polymers on the surface of Fe3 O4 @ZIF-8 (MZNIP) were also prepared and processed similarly, except that the template molecule E2 was not added. Fig. 1S shows the procedure used. Further, the molecularly imprinted polymers and molecularly non-imprinted polymers based on Fe3 O4 @SiO2 nanoparticles [23] (MMIP and MNIP, respectively) were also prepared similarly.
2.3. Preparation of MZMIP–SPME fibers Our previous study indicated that our fabricated automated SPME through electromagnetic bonding strategy resulted in high level of physical and chemical stability and reusability [23]. Therefore, we prepare the MZMIP–SPME fibers in this work according to our previous work with slightly modified. It was then dried in the oven at 100 ◦ C for 10 min, this procedure, named as a thermal curing procedure, was useful for the stability of SPME coating [37]. Moreover, MMIP-coated SPME and MNIP-coated SPME fibers were also prepared by a similar way.
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2.7. Sample preparation Pork and fish samples were purchased from retail supermarket in Ningbo, China. These samples were stored at −18 ◦ C before analysis. The pork and fish samples were extracted according to the literature reported [6]. Firstly, the tissue samples were chopped and triturated in a blender. 2 g sample was placed into a 10 mL tube, mixed with 4 mL acetonitrile, and ultrasonicated for 20 min at room temperature. Then, the mixture placed in a high-speed refrigerated centrifuge and centrifuged at 10,000 rpm for 10 min. The extraction solutions were filtrated and concentrated with nitrogen gas, and then were quantified to the volume of 5 mL extraction solvent of n-hexane for the further SPME extraction. For the spiking pork and fish samples, each sample (2 g) was mixed with 4, 10 and 40 L of standard solution (10 mg L−1 ) of four estrogens, and the extraction procedure was the same as described above. The spiking concentrations of fish and pork were obtained with three levels of 20, 50, 200 ng g−1 .
3. Results and discussion 2.4. Automated SPME for determination of four estrogens
3.1. Preparation of MZMIP coating
The current study employed the automated SPME device and robotic autosampler for the sample preparation. The detailed explanation of the automated SPME procedure was the same as that reported in Ref. [23].
Fig. 1S shows the simplified scheme of MZMIP coating preparation. The reasons why we choose E2 as template rather than EE2 were as following: (1) according to the literature [6], they also chose E2 as template to prepare the molecularly imprinted polymer in order to analysis of same targets (E1, E2, E3, and EE2) as ours. (2) The structure of E2 was more similar to E1 and E3 than EE2. Furthermore, the structure of E2 is the parent structure of E1, E3, and EE2. Therefore, the choice of E2 as the template molecule could obtain more imprinted cavities for the adsorption of the four target estrogens. Moreover, there is an alkynyl group ( C C) on the 20th site of the parent structure of EE2, showing that it has very different structure with that of E1, E3, and E2 which have carbonyl group or hydroxyl group on the same site. So we chose E2 as template. We choose hydrophilic ABPA as functional monomer because (1) ABPA is a phenylo boric acid compound with benzene ring and could interact with ZIF-8 through electrostatic interaction and – stacking. Even though the pores of ZIF-8 and template E2 are hydrophobic, while APBA is hydrophilic, ZIF-8 and E2 can also react with APBA. (2) According to the literature [38], ZIF-8 could absorb phthalic acid in water efficiently through electrostatic interaction between the positively charged surfaces of ZIF-8 and HPA− or PA2− molecules at high pH. ABPA can also form the anionic compound to react with positively charged surface of ZIF-8. Moreover, the benzene ring of ABPA can also react with the imidazole ring of ZIF-8 and E2 through – stacking. (3) Some literatures have reported that APBA can auto-polymerize without cross-linker or an initiator agent [32–34]. Therefore, we chose hydrophilic APBA as functional monomer. The polymerization conditions (type of polymerization solvent, molar ratio of template to functional monomer, and polymerization time) were optimized to enhance MZMIP coating performance in extraction amount and selectivity. Phosphate buffer saline (PBS, pH = 7.2) was chosen as the polymerization buffer for ABPA, which is a kind of water-soluble functional monomer. However, the template E2 could not dissolve in PBS. Therefore, a certain proportion of organic solvent was needed in the polymerization buffer. 20% of organic solvents (acetonitrile, methanol, acetone, dichloromethane, toluene, and ethanol) were applied. The result shown in Fig. 2S indicated that the MZMIP coating prepared using acetonitrile/PBS (1/4, v/v) exhibited the highest selectivity factors. Further, the MZMIP coating could not recognize the related compound -nap (Fig. 1a), which demonstrated that the MIP prepared
2.5. Comparison of extraction performance Apart from the proposed automated MZMIP–SPME method, the methods were compared using two other analyses which were MMIP-coated SPME extraction analysis (MMIP-SPME) and automated commercial SPME with different coatings such as PA, CAR/PDMS, PDMS/DVB or DVB/CAR/PDMS coating. For MMIP-SPME and commercial SPME fibers, the extraction and desorption procedures were the same as MZMIP-coated SPME. The extraction amounts of E1, E2, E3 and EE2 were calculated with the standard curve method.
2.6. Instruments and HPLC analysis The as-synthesized nanocomposites were characterized with SU-70 scanning electron microscopy (SEM, Hitachi Corporation, Japan), JEOL 2100 transmission electron microscope (TEM), NEXUS 670 infrared Fourier transform spectrometer (Nicolet Thermo, Waltham, MA), Quantum Design Physical Property Measurement System (PPMS-9, Quantum Design, CA) and X-ray diffraction (Bruker, D8 Focus). The chromatographic separation was performed with an Agilent 1260 infinity quaternary liquid chromatography (Hewlett Packard, Wilmington, NC, USA) with a multiple wavelength detector. The chromatographic column was an Agilent Zorbax Eclipse XDB-C18 (150 mm × 4.6 mm I.D., 5 m packing, Agilent Technologies, NC, USA). The analytical column was set at 30 ◦ C. The mobile phase was used at a constant flow rate of 1.0 mL min−1 , and water and acetonitrile as mobile phases A and B respectively. The chromatographic elution and a gradient program with duration of 10 min were optimized to separate the model compounds as follows: liner gradient from 60 to 45% A (0–10 min). Quantitative analyses of compounds were performed using an SPD-detector. Wavelength was set at 280 nm. The data analysis was performed using Chemstation software (Hewlett Packard).
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Fig. 2. The HRTEM image of MZMIP.
Fig. 1. Chromatograms of the standard estrogen (E1, E2, E3, EE2, and -naphthol) solution and after MZMIP-coated SPME treatment (a); effect of MZMIP amount to extraction capacity of MZMIP-coated SPME (b).
using ABPA as the functional monomer and E2 as the template molecule had good selectivity to the four estrogens. Different molar ratios of E2 to ABPA (1:1, 1:2, 1:3, 1:4, 1:5, 1:6) and amounts of MZMIP (2.5, 5, 7.5, 10, 15, 20, 25, and 30 mg, respectively) on the surface of stainless steel fiber were also investigated. According to the results (Fig. 3S and Fig. 1b), the molar ratio of 1:5 and 15 mg MZMIP was chosen for the further study. Furthermore, under optimized condition, the effect of polymerization time was studied, and the results (Fig. 2) indicated that a thin (5 nm), homogeneous and stable MIP film on the surface of Fe3 O4 @ZIF-8 nanoparticles was obtained in 8 h at room temperature, which is much shorter than conventional bulk polymerization (>30 h) [24].
3.2. Characterization of synthesized particles Morphological features of Fe3 O4 @ZIF-8 and MZMIP were characterized by SEM and TEM. The diameter of the Fe3 O4 @ZIF-8 particle was about 400 nm, and Fig. 4S(a) indicated that the Fe3 O4 particles were fully coated by ZIF-8, which was consistent with published literature [36]. After the thin imprinted film further covered the Fe3 O4 particles, the surface of the nanoparticles became smooth, suggesting the successful formation of imprinted polymer layer (Fig. 4S(b)). The coated ZIF-8 shell had an average thickness of 20 nm and the imprinted polymer layer exhibited the average
thickness of 5 nm, as calculated from the TEM and HRTEM images (Fig. 4S (c–d) and Figs. 2 and 3c and d). Vibrating sample magnetometry (VSM) was employed to study the magnetic properties of Fe3 O4 , Fe3 O4 @ZIF-8 and MZMIP. Fig. 3a shows the magnetic hysteresis loops of the dried samples at room temperature. The saturation magnetization values obtained at room temperature were 76.94, 50.33, and 35.71 emu g−1 for Fe3 O4 , Fe3 O4 @ZIF-8, and MZMIP, respectively. The MZMIP particles with the magnetization value could immobilize on the surface of stainless steel fiber in the presence of a strong magnetic field (ECS). They could also be removed rapidly after the magnetic field is removed for renewing the SPME coating. The X-ray power diffraction (XRD) patterns for the synthesized pure ZIF-8, Fe3 O4 , Fe3 O4 @ZIF-8 and MZMIP are shown in Fig. 3b. In the 2 range of 5◦ –80◦ , the relative intensity and peak positions of the ZIF-8 were consistent with previous publications [25]. Six characteristic peaks for Fe3 O4 (2 = 29.74◦ , 34.96◦ , 42.54◦ , 52.85◦ , 56.64◦ , 62.38◦ ) were observed in Fe3 O4 , and the peak positions at the corresponding 2 value were indexed as (2 2 0), (3 1 1), (4 0 0), (4 2 2), (5 1 1) and (4 4 0), respectively. As shown in the Fe3 O4 @ZIF8 curve, the characteristic peaks of both Fe3 O4 and ZIF-8 were detected in the obtained Fe3 O4 @ZIF-8 core–shell particles. The XRD pattern of MZMIPs particles was very similar to that of Fe3 O4 @ZIF-8, which indicated successfully imprinted layer coating. The products of Fe3 O4 , ZIF-8, Fe3 O4 @ZIF-8 and Fe3 O4 @ZIF8@MIP were investigated by FT-IR spectroscopy (Fig. 3c). The bond at 574 cm−1 is attributed to the stretch of Fe O. Compared with the infrared data of Fe3 O4 , the characteristic peaks of Z–N group at about 416 cm−1 , imidazole ring group at about 500–1500 cm−1 , C H stretch at 2930 cm−1 and 3145 cm−1 indicate the formation of ZIF-8 coating on the surface of Fe3 O4 . As can be seen from the curve of Fe3 O4 @ZIF-8@MIP, the peaks of C-N group at 1364 cm−1 and N H group at 1569 cm−1 indicated that the MIP film on the surface of Fe3 O4 @ZIF-8 was successfully obtained [38]. 3.3. Extraction capacity The extraction capacity studies of MZMIP- and Fe3 O4 @ZIF8@NIP (MZNIP)-coated fiber were performed with a serious of E1, E2, E3, EE2 mixed standard solutions of 0.1–10 mg L−1 in nhexane. Fig. 4a shows the extraction amount curves of the MZMIPand MZNIP-coated fibers to each estrogen. It was obvious that
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Fig. 3. The hysteresis loops of Fe3 O4 , Fe3 O4 @ ZIF-8, and MZMIP. The insert shows the separation and redispersion process of a solution of MZMIP in the absence (left) and presence (right) of an external magnetic field (a); XRD patterns of ZIF-8, Fe3 O4 , Fe3 O4 @ ZIF-8, and MZMIP (b); FT-IR spectra of Fe3 O4 , ZIF-8, Fe3 O4 @ZIF-8, and MZMIP (c).
the extraction amounts of four estrogens increased along with the increase of concentration in range of 0.1–2 mg L−1 for both MZMIPand MZNIP-coated fibers. For the MZMIP-coated fibers, the extraction reached equilibrium when the concentration was 4 mg L−1 , and the extraction capacities for E1, E2, E3, and EE2 were about 538, 753, 575, and 435 ng, respectively. The extraction capacities of MZMIPcoated fibers for E1, E2, E3, and EE2 were about 2.69, 3.35, 2.75, and 2.32 times as much as that with the MZNIP-coated fibers. It was indicated that MZMIP-coated fiber exhibited high extraction capabilities and good selectivities to the four estrogens because of its tailor-made specific recognition mechanism and large amount of MZMIP. The extraction efficiency of the MIP film is also crucially depended on the surface properties of carrier. Usually, Fe3 O4 @SiO2 was widely used as carrier for preparation of MIP for extraction of estrogens. Therefore, for comparison purpose, the adsorption isotherm of estrogens on MMIP-coating was also investigated. Fig. 4b shows that the amount of adsorbed E1, E2, E3, and EE2 on MMIP was about 309, 385, 328, and 216 ng, respectively, which were lower than those of the MZMIP coating for each estrogen and will affect the sensitivity of analytical method. At the same time, the selectivity of MMIP coating (2.10, 2.18, 1.95 and 1.69 for E1, E2, E3 and EE2, respectively) was also weaker than MZMIP coating. Fe3 O4 @ZIF-8 as the carrier material of imprinted polymer plays a key role. The reasons maybe as follows: (1) there are many surface-open Zn ion sites and imidazole ligands on the surface of Fe3 O4 @ZIF-8particles, which could strongly adsorb E2
through a coordinate bond between Zn2+ and hydroxyl group of the template molecular, and intermolecular – interactions between imidazole ligand and naphthalene ring of E2 [27]. Moreover, the pores of ZIF-8 are hydrophobic, which could also help for bonding hydrophobic E2. All these interactions can facilitate uniform and large distribution of the template molecule on the surface of porous Fe3 O4 @ZIF-8 for further creating the imprinted film. (2) The imidazole ring of ZIF-8 can react with APBA as monomer through – stacking interaction, which facilitate the immobilization of a stable MIP film on the surface of Fe3O4@ZIF-8particles. After the templates were eluted, more imprinted sites could be obtained due to the largely bonded template molecules. However, the hydrophilic Fe3 O4 @SiO2 has weak absorption capacity with hydrophobic E2 for creating imprinted sites and preparing MIP film. (3) The rugged and porous surface of Fe3 O4 @ZIF-8 as carrier can result in a larger specific surface area of MIP film than the relatively smooth Fe3 O4 @SiO2 (see Fig. 4S(b)). The nitrogen adsorption desorption measurements were employed to evaluate the surface properties between MMIP and MZMIP. The BET surface area, porous value, and pore size of MZMIP were 184.59 m2 g−1 , 0.21 cm3 g−1 , and 9.18 nm, respectively. But for MMIP, the parameters were 87.1 m2 g−1 , 0.085 cm3 g−1 , and 5.24 nm, respectively. Therefore, all these results may be attributed to the fact that MZMIP could have higher extraction capacity than MMIP. Moreover, in order to investigate the reproducibility of extraction efficiencies, the intraday, interday, and fiber-to-fiber repeatability of extraction by MZMIP-coated fiber were studied
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Fig. 4. Extraction amount curves of MZMIP (solid line) and MZMIP (dotted line) fibers to E1 (), E2 (), E3 (), and EE2 (䊉) mixed solution in n-hexane of 0.1–10 mg L−1 (a); extraction amount curves of Fe3 O4 @SiO2 @MIP (solid line) and Fe3 O4 @SiO2 @NIP (dotted line) fibers to E1 (), E2 (), E3 (), and EE2 (䊉) mixed solution in n-hexane of 0.1–5 mg L−1 (b).
with 200 g L−1 E1, E2, E3, and EE2 mixed standard solutions, and RSDs (n = 6) of 1.7–2.6%, 2.9–4.5%, and 2.3–3.8% for extraction amounts of four estrogens were obtained, respectively. 3.4. The reusability, stability and preservability of MZMIP coating To investigate the robustness and reusability of the developed coating, MZMIP-coated stainless steel fiber was applied for 250 repetitious SPME operations. Performance of the MZMIP coating was evaluated through extraction recovery and physical stability. The coating demonstrated good reproducibility for at least 250 times with recovery over 80% and related standard deviation (RSD)
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Journal of Chromatography A journal homepage: www.elsevier.com/locate/chroma
Development of a novel magnetic molecularly imprinted polymer coating using porous zeolite imidazolate framework-8 coated magnetic iron oxide as carrier for automated solid phase microextraction of estrogens in fish and pork samples Hangzhen Lan a , Ning Gan b,∗ , Daodong Pan a,∗∗ , Futao Hu a , Tianhua Li b , Nengbing Long b , Haoyu Shen c , Yinjie Feng b a
Faculty of Marine Science, Ningbo University, Ningbo 315211, China Faculty of Material Science and Chemical Engineering, Ningbo University, Ningbo 315211, China c Ningbo Institute of Technology, Zhejiang University, Ningbo 315100, China b
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Article history: Received 16 May 2014 Received in revised form 16 July 2014 Accepted 31 August 2014 Available online xxx Keywords: Zeolite imidazolate framework-8 coated magnetic iron oxide Molecularly imprinted polymer coating Automated solid-phase microextraction Estrogens High performance liquid chromatography
a b s t r a c t A high-performance magnetic molecularly imprinted polymer (MIP) coating using zeolite imidazolate framework-8 coated magnetic iron oxide (Fe3 O4 @ZIF-8) as a carrier was developed for simultaneous automated solid phase microextraction of four estrogens in 24 food samples. The coating material, abbreviated as MZMIP, was synthesized through time-efficient layer-by-layer assembling of ZIF-8 and MIP film on Fe3 O4 particles. It was characterized and automatically coated on the surface of SPME fibers by electromagnetic bonding. The extraction performance, reusability, repeatability, and validity of the MZMIP–SPME system was evaluated for high-throughput analysis of estrone (E1), estradiol (E2), estriol (E3), and ethinylestradiol (EE2). Various factors affecting the quality of MZMIP coating were optimized. Compared with traditional magnetic MIP coating based on Fe3 O4 @SiO2 carrier, the MZMIP coating exhibited high extraction capacity and quick adsorption and desorption kinetics to E1, E2, E3, and EE2 owing to the larger amount of imprinting sites in MZMIP. Under optimum conditions, the proposed system requires only 25 min for pretreatment of all 24 samples (62.5 s per sample). The limits of detection and quantitation of the proposed automated system for analysis were found to range from 0.4 to 1.7 and 1.1 to 6.2 ng g−1 , respectively. During analysis of spiked fish and pork, the new coating showed better recovery and selectivity compared with Fe3 O4 @SiO2 @MIP (MMIP) and commercially available SPME. The results indicated that the MZMIP coating could be effectively employed for pretreatment of ultra-trace level of estrogens in food. © 2014 Elsevier B.V. All rights reserved.
1. Introduction Estrogens are a group of potent endocrine-disrupting chemicals, which include naturally occurring estrone (E1), estradiol (E2), estriol (E3), and synthetic ethinylestradiol (EE2). Especially EE2, as a kind of xenoestrogens, has been identified as the most potent estrogenic chemical in all endocrine-disrupting chemicals until date [1]. Estrogens could enter human body through the food chain, disturb the normal endocrine system, and alter the normal reproduction and development of human beings [2,3]. Further, those compounds
∗ Corresponding author. Tel.: +86 574 87608347; fax: +86 574 87608347. ∗∗ Corresponding author. Tel.: +86 574 87609987; fax: +86 574 87609987. E-mail addresses: [email protected] (N. Gan), [email protected] (D. Pan).
have attracted large societal and political attention because of their widespread presence in certain food matrices, such as fish and pork [4–6]. Thus, it is of great significance for food safety supervision to develop sensitive, selective, and simple methodologies to determine estrogens in food. Solid-phase micro-extraction (SPME), pioneered by Arthur and Pawliszyn in 1990s [7], has been widely applied for monitoring estrogens either in food samples, in environment matrices or in biologic samples [8–12]. SPME has many advantages because of its integration of analytes extraction, pre-concentration, and sample clean up in a single step [13–15]. To improve the sample preparation strategies for quick analysis of complex samples and high-throughput analysis, there has been a growing need for an automated SPME method. Recently, Pawliszyn’s research groups have developed many automated SPME devices in multi-well
http://dx.doi.org/10.1016/j.chroma.2014.08.096 0021-9673/© 2014 Elsevier B.V. All rights reserved.
Please cite this article in press as: H. Lan, et al., Development of a novel magnetic molecularly imprinted polymer coating using porous zeolite imidazolate framework-8 coated magnetic iron oxide as carrier for automated solid phase microextraction of estrogens in fish and pork samples, J. Chromatogr. A (2014), http://dx.doi.org/10.1016/j.chroma.2014.08.096
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format using a robotic autosampler to prepare several samples in parallel [16–22]. In our previous work [23], we developed a novel automated SPME device that could simultaneously treat up to 24 samples. In addition, a highly selective and sensitive magnetic molecularly imprinted polymers MMIP-SPME analytical method was established using Fe3 O4 @SiO2 @MIP (MMIP) as a coating of SPME and coupled with high performance liquid chromatography (HPLC) for detection. Fe3 O4 @SiO2 was applied as the matrix of MMIP due to the modifiable silicon hydroxyl on the surface of SiO2 nanoparticle. However, the grafting and molecularly imprinted polymerization procedures on SiO2 were time-consuming. In Ding’s work [24], to modify MIP film using MAA as functional monomer on the surface of Fe3 O4 @SiO2 , the ␥-methacryloxypropyl trimethoxysilane (KH570) was employed to introduce grafting C C group to the surface of Fe3 O4 @SiO2 by reacting in toluene for 24 h at 120 ◦ C. The molecularly imprinted polymerization procedure even reached up to 38 h. In the process, a large volume of organic solvent was needed. To simplify the immobilizing procedure of MIP film on superparamagnetic iron oxide nanoparticles as coating for automated MMIP-SPME, choosing a new carrier instead of Fe3 O4 @SiO2 and polymer monomer instead of MAA was a big challenge. Metal–organic frameworks (MOFs) have recently received considerable attention due to their unusual properties such as large surface area, good thermal stability, and the availability of inpore functionality and outer-surface modification [25]. Owing to the above merits, Kun’s group reported MIP preparation by using MOF-5 as matrix, and the synthesized MOF@MIP showed a homogeneous polymer film, thermal stability, and exhibited a higher specific surface area and a faster transfer-mass speed compared with that of the bulk MIP [26]. Zeolite imidazolate frameworks (ZIFs), a new class of microporous MOFs are synthesized by the crystallization of a transition metal species bound to the nitrogen atom of an imidazolate compound [25]. ZIF-8, which has the formula Zn(MIM)2 (MIM: 2-methylimidazole), has hydrophobic pores [27]. ZIF-8 has been reported to possess good extraction capacity to steroidal hormones due to the presence of a benzene ring by forming a coordinate bond and intermolecular – interactions [28]. Considering the superior properties of ZIF-8 and their potential absorbing capacity for the benzene series and related organic groups, the design and synthesis of magnetic ZIF-8 as a carrier of coating are especially desirable for automated SPME. Some novel monomers that can auto-polymerize without cross-linking or an initiator agent, such as dopamine [29], polydopamine [30,31], or 3-aminobenzeneboronicacid (APBA) [32–34], can be used to quickly prepare the MIP film for the automated SPME method. APBA is an attractive functional monomer that is water-soluble and provides a mild aqueous environment during polymerization [34]. Moreover, the benzene rings of ABPA can be absorbed on ZIF-8, which has imidazole ring, through – stacking interaction. Thus the MIP film using APBA polymer can be easily immobilized on Fe3 O4 @ZIF-8 without any grafting procedure, which can greatly simplify the preparation step of MIP film on the carrier. Furthermore, the MIP film is prepared by an organic solvent-free synthesis. Moreover, the template (E2) can be uniformly and stably distributed on the surface of porous ZIF-8 through – stacking interactions to simply the MIP film immunization procedure. In this work, to develop a new coating with high extraction capacity and stability for the automated SPME system, we prepared a novel MZMIP coating using Fe3 O4 @ZIF-8 as the carrier and ABPA as the functional monomer. The MMIP coating, using Fe3 O4 @SiO2 as the carrier, was also synthesized to compare the extraction performance with the new MZMIP coating. The MZMIPcoated automated SPME system coupled with HPLC was evaluated for high-throughput analysis of estrogens (E1, E2, E3, and EE2) in real fish and pork samples.
2. Experimental 2.1. Chemical reagents and materials In this work, estrone (E1, 97%), estradiol (E2, 99%), estriol (E3, 98%), and ethinyloestradiol (EE2, 98%), -naphthol (-nap, 99%) and poly (styrenesulfonate, sodium salt) (PSS, 30 wt%) were obtained from Sigma–Aldrich (Shanghai, China). 3-Amino phenyl boronic acid (APBA) was from J&K Chemical Co. Ltd. Acetonitrile (HPLC grade) was acquired from CNW Technologies (Dusseldorf, Germany). 2-methylimidazole (H-MeIM, 99%) were purchased from Aladdin Chemistry (Shanghai, China). Zinc nitrate hexahydrate (Zn(NO3 )2 ·6H2 O, 99%), iron (III) chloride hexahydrate (FeCl3 ·6H2 O, 99%), sodium acetate (NaAC, 99%) and ethylene glycol ((HOCH2 )2 , 99%) were purchased from Sinopharm Chemical Reagent Co., Ltd., China. All other reagents were of analytical grade. Ultrapure water (Hangzhou Wahaha Foods Co. Ltd., Hangzhou, China) was used throughout this work. A mixed stock solution of the four estrogens (E1, E2, E3, and EE2) was prepared in methanol at a concentration of 100 mg L−1 and stored at 4 ◦ C in darkness. All solutions used for HPLC mobile phase were filtered through a nylon 0.22 m filter prior to use. The commercial SPME fibers with 85 m polyacrylate (PA), 85 m carboxen/polydimethylsiloxane (CAR/PDMS), 65 m polydimethylsiloxane/divinylbenzene (PDMS/DVB), and 50/30 m carboxen/divinylbenzene/polydimethylsiloxane (CAR/DVB/PDMS) coatings, which were purchased from Supelco (Bellefonte, PA, USA) were used for the comparison study. 2.2. Synthesis of MZMIP core–shell particles 2.2.1. Preparation of Fe3 O4 particles The Fe3 O4 particles were synthesized as described in our previous work [23]. 2.2.2. Preparation of Fe3 O4 @ZIF-8 particles The core–shell Fe3 O4 @ZIF-8 particles were prepared according to Zhang et al. [35]. In a typical procedure, 0.05 g Fe3 O4 was added to 150 mL aqueous solution contains of 0.3% poly (styrenesulfonate, sodium salt) (PSS) under ultrasonication for 30 min. The resultant Fe3 O4 @PSS particles were recovered by an external magnetic field and washed with purified water for three times, then redispersed in a mixture containing 150 mL methanol, 1.125 g Zn(NO3 )2 ·6H2 O and 3.11 g 2-methylimidazolate under stirring and the reaction was allowed to proceed at 50 ◦ C for 3 h for ZIF-8 shell growth. Finally, the obtained Fe3 O4 @ ZIF-8 nanoparticles were separated by a magnet and washed with ethanol. 2.2.3. Preparation of MZMIP particles MZMIP was prepared as follows. E2 (0.1 mmol) as the template was first dissolved in 1.25 mL acetonitrile, then mixed with 5 mL phosphate buffer sodium (PBS, pH = 7.2), which contains 0.5 mmol APBA as the functional and cross-linking monomer, and the mixture was then incubated at room temperature for 1 h. After adding Fe3 O4 @ZIF-8 (0.04 g), the solution was incubated for 2 h at room temperature. Prior to use, the Fe3 O4 @ZIF-8 particles were subjected to extensive deionized water and absolute ethanol, and washed thoroughly. Subsequently, a 100 mM aqueous solution of ammonium persulfate (6.5 mL) as initiator was slowly added dropwise to the above solution for about 20 min and the polymerization process was executed at room temperature. After 8 h, Fe3 O4 @ZIF8@MIP (MZMIP) particles were obtained. Finally, MZMIP was collected magnetically, and the template was removed using 20 mL methanol/acetic acid (4/1, v/v) solution (with shaking) until no E2 adsorption was detected by HPLC. The particles were washed with water three times again and vacuum dried at 60 ◦ C.
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The molecularly non-imprinted polymers on the surface of Fe3 O4 @ZIF-8 (MZNIP) were also prepared and processed similarly, except that the template molecule E2 was not added. Fig. 1S shows the procedure used. Further, the molecularly imprinted polymers and molecularly non-imprinted polymers based on Fe3 O4 @SiO2 nanoparticles [23] (MMIP and MNIP, respectively) were also prepared similarly.
2.3. Preparation of MZMIP–SPME fibers Our previous study indicated that our fabricated automated SPME through electromagnetic bonding strategy resulted in high level of physical and chemical stability and reusability [23]. Therefore, we prepare the MZMIP–SPME fibers in this work according to our previous work with slightly modified. It was then dried in the oven at 100 ◦ C for 10 min, this procedure, named as a thermal curing procedure, was useful for the stability of SPME coating [37]. Moreover, MMIP-coated SPME and MNIP-coated SPME fibers were also prepared by a similar way.
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2.7. Sample preparation Pork and fish samples were purchased from retail supermarket in Ningbo, China. These samples were stored at −18 ◦ C before analysis. The pork and fish samples were extracted according to the literature reported [6]. Firstly, the tissue samples were chopped and triturated in a blender. 2 g sample was placed into a 10 mL tube, mixed with 4 mL acetonitrile, and ultrasonicated for 20 min at room temperature. Then, the mixture placed in a high-speed refrigerated centrifuge and centrifuged at 10,000 rpm for 10 min. The extraction solutions were filtrated and concentrated with nitrogen gas, and then were quantified to the volume of 5 mL extraction solvent of n-hexane for the further SPME extraction. For the spiking pork and fish samples, each sample (2 g) was mixed with 4, 10 and 40 L of standard solution (10 mg L−1 ) of four estrogens, and the extraction procedure was the same as described above. The spiking concentrations of fish and pork were obtained with three levels of 20, 50, 200 ng g−1 .
3. Results and discussion 2.4. Automated SPME for determination of four estrogens
3.1. Preparation of MZMIP coating
The current study employed the automated SPME device and robotic autosampler for the sample preparation. The detailed explanation of the automated SPME procedure was the same as that reported in Ref. [23].
Fig. 1S shows the simplified scheme of MZMIP coating preparation. The reasons why we choose E2 as template rather than EE2 were as following: (1) according to the literature [6], they also chose E2 as template to prepare the molecularly imprinted polymer in order to analysis of same targets (E1, E2, E3, and EE2) as ours. (2) The structure of E2 was more similar to E1 and E3 than EE2. Furthermore, the structure of E2 is the parent structure of E1, E3, and EE2. Therefore, the choice of E2 as the template molecule could obtain more imprinted cavities for the adsorption of the four target estrogens. Moreover, there is an alkynyl group ( C C) on the 20th site of the parent structure of EE2, showing that it has very different structure with that of E1, E3, and E2 which have carbonyl group or hydroxyl group on the same site. So we chose E2 as template. We choose hydrophilic ABPA as functional monomer because (1) ABPA is a phenylo boric acid compound with benzene ring and could interact with ZIF-8 through electrostatic interaction and – stacking. Even though the pores of ZIF-8 and template E2 are hydrophobic, while APBA is hydrophilic, ZIF-8 and E2 can also react with APBA. (2) According to the literature [38], ZIF-8 could absorb phthalic acid in water efficiently through electrostatic interaction between the positively charged surfaces of ZIF-8 and HPA− or PA2− molecules at high pH. ABPA can also form the anionic compound to react with positively charged surface of ZIF-8. Moreover, the benzene ring of ABPA can also react with the imidazole ring of ZIF-8 and E2 through – stacking. (3) Some literatures have reported that APBA can auto-polymerize without cross-linker or an initiator agent [32–34]. Therefore, we chose hydrophilic APBA as functional monomer. The polymerization conditions (type of polymerization solvent, molar ratio of template to functional monomer, and polymerization time) were optimized to enhance MZMIP coating performance in extraction amount and selectivity. Phosphate buffer saline (PBS, pH = 7.2) was chosen as the polymerization buffer for ABPA, which is a kind of water-soluble functional monomer. However, the template E2 could not dissolve in PBS. Therefore, a certain proportion of organic solvent was needed in the polymerization buffer. 20% of organic solvents (acetonitrile, methanol, acetone, dichloromethane, toluene, and ethanol) were applied. The result shown in Fig. 2S indicated that the MZMIP coating prepared using acetonitrile/PBS (1/4, v/v) exhibited the highest selectivity factors. Further, the MZMIP coating could not recognize the related compound -nap (Fig. 1a), which demonstrated that the MIP prepared
2.5. Comparison of extraction performance Apart from the proposed automated MZMIP–SPME method, the methods were compared using two other analyses which were MMIP-coated SPME extraction analysis (MMIP-SPME) and automated commercial SPME with different coatings such as PA, CAR/PDMS, PDMS/DVB or DVB/CAR/PDMS coating. For MMIP-SPME and commercial SPME fibers, the extraction and desorption procedures were the same as MZMIP-coated SPME. The extraction amounts of E1, E2, E3 and EE2 were calculated with the standard curve method.
2.6. Instruments and HPLC analysis The as-synthesized nanocomposites were characterized with SU-70 scanning electron microscopy (SEM, Hitachi Corporation, Japan), JEOL 2100 transmission electron microscope (TEM), NEXUS 670 infrared Fourier transform spectrometer (Nicolet Thermo, Waltham, MA), Quantum Design Physical Property Measurement System (PPMS-9, Quantum Design, CA) and X-ray diffraction (Bruker, D8 Focus). The chromatographic separation was performed with an Agilent 1260 infinity quaternary liquid chromatography (Hewlett Packard, Wilmington, NC, USA) with a multiple wavelength detector. The chromatographic column was an Agilent Zorbax Eclipse XDB-C18 (150 mm × 4.6 mm I.D., 5 m packing, Agilent Technologies, NC, USA). The analytical column was set at 30 ◦ C. The mobile phase was used at a constant flow rate of 1.0 mL min−1 , and water and acetonitrile as mobile phases A and B respectively. The chromatographic elution and a gradient program with duration of 10 min were optimized to separate the model compounds as follows: liner gradient from 60 to 45% A (0–10 min). Quantitative analyses of compounds were performed using an SPD-detector. Wavelength was set at 280 nm. The data analysis was performed using Chemstation software (Hewlett Packard).
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Fig. 2. The HRTEM image of MZMIP.
Fig. 1. Chromatograms of the standard estrogen (E1, E2, E3, EE2, and -naphthol) solution and after MZMIP-coated SPME treatment (a); effect of MZMIP amount to extraction capacity of MZMIP-coated SPME (b).
using ABPA as the functional monomer and E2 as the template molecule had good selectivity to the four estrogens. Different molar ratios of E2 to ABPA (1:1, 1:2, 1:3, 1:4, 1:5, 1:6) and amounts of MZMIP (2.5, 5, 7.5, 10, 15, 20, 25, and 30 mg, respectively) on the surface of stainless steel fiber were also investigated. According to the results (Fig. 3S and Fig. 1b), the molar ratio of 1:5 and 15 mg MZMIP was chosen for the further study. Furthermore, under optimized condition, the effect of polymerization time was studied, and the results (Fig. 2) indicated that a thin (5 nm), homogeneous and stable MIP film on the surface of Fe3 O4 @ZIF-8 nanoparticles was obtained in 8 h at room temperature, which is much shorter than conventional bulk polymerization (>30 h) [24].
3.2. Characterization of synthesized particles Morphological features of Fe3 O4 @ZIF-8 and MZMIP were characterized by SEM and TEM. The diameter of the Fe3 O4 @ZIF-8 particle was about 400 nm, and Fig. 4S(a) indicated that the Fe3 O4 particles were fully coated by ZIF-8, which was consistent with published literature [36]. After the thin imprinted film further covered the Fe3 O4 particles, the surface of the nanoparticles became smooth, suggesting the successful formation of imprinted polymer layer (Fig. 4S(b)). The coated ZIF-8 shell had an average thickness of 20 nm and the imprinted polymer layer exhibited the average
thickness of 5 nm, as calculated from the TEM and HRTEM images (Fig. 4S (c–d) and Figs. 2 and 3c and d). Vibrating sample magnetometry (VSM) was employed to study the magnetic properties of Fe3 O4 , Fe3 O4 @ZIF-8 and MZMIP. Fig. 3a shows the magnetic hysteresis loops of the dried samples at room temperature. The saturation magnetization values obtained at room temperature were 76.94, 50.33, and 35.71 emu g−1 for Fe3 O4 , Fe3 O4 @ZIF-8, and MZMIP, respectively. The MZMIP particles with the magnetization value could immobilize on the surface of stainless steel fiber in the presence of a strong magnetic field (ECS). They could also be removed rapidly after the magnetic field is removed for renewing the SPME coating. The X-ray power diffraction (XRD) patterns for the synthesized pure ZIF-8, Fe3 O4 , Fe3 O4 @ZIF-8 and MZMIP are shown in Fig. 3b. In the 2 range of 5◦ –80◦ , the relative intensity and peak positions of the ZIF-8 were consistent with previous publications [25]. Six characteristic peaks for Fe3 O4 (2 = 29.74◦ , 34.96◦ , 42.54◦ , 52.85◦ , 56.64◦ , 62.38◦ ) were observed in Fe3 O4 , and the peak positions at the corresponding 2 value were indexed as (2 2 0), (3 1 1), (4 0 0), (4 2 2), (5 1 1) and (4 4 0), respectively. As shown in the Fe3 O4 @ZIF8 curve, the characteristic peaks of both Fe3 O4 and ZIF-8 were detected in the obtained Fe3 O4 @ZIF-8 core–shell particles. The XRD pattern of MZMIPs particles was very similar to that of Fe3 O4 @ZIF-8, which indicated successfully imprinted layer coating. The products of Fe3 O4 , ZIF-8, Fe3 O4 @ZIF-8 and Fe3 O4 @ZIF8@MIP were investigated by FT-IR spectroscopy (Fig. 3c). The bond at 574 cm−1 is attributed to the stretch of Fe O. Compared with the infrared data of Fe3 O4 , the characteristic peaks of Z–N group at about 416 cm−1 , imidazole ring group at about 500–1500 cm−1 , C H stretch at 2930 cm−1 and 3145 cm−1 indicate the formation of ZIF-8 coating on the surface of Fe3 O4 . As can be seen from the curve of Fe3 O4 @ZIF-8@MIP, the peaks of C-N group at 1364 cm−1 and N H group at 1569 cm−1 indicated that the MIP film on the surface of Fe3 O4 @ZIF-8 was successfully obtained [38]. 3.3. Extraction capacity The extraction capacity studies of MZMIP- and Fe3 O4 @ZIF8@NIP (MZNIP)-coated fiber were performed with a serious of E1, E2, E3, EE2 mixed standard solutions of 0.1–10 mg L−1 in nhexane. Fig. 4a shows the extraction amount curves of the MZMIPand MZNIP-coated fibers to each estrogen. It was obvious that
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Fig. 3. The hysteresis loops of Fe3 O4 , Fe3 O4 @ ZIF-8, and MZMIP. The insert shows the separation and redispersion process of a solution of MZMIP in the absence (left) and presence (right) of an external magnetic field (a); XRD patterns of ZIF-8, Fe3 O4 , Fe3 O4 @ ZIF-8, and MZMIP (b); FT-IR spectra of Fe3 O4 , ZIF-8, Fe3 O4 @ZIF-8, and MZMIP (c).
the extraction amounts of four estrogens increased along with the increase of concentration in range of 0.1–2 mg L−1 for both MZMIPand MZNIP-coated fibers. For the MZMIP-coated fibers, the extraction reached equilibrium when the concentration was 4 mg L−1 , and the extraction capacities for E1, E2, E3, and EE2 were about 538, 753, 575, and 435 ng, respectively. The extraction capacities of MZMIPcoated fibers for E1, E2, E3, and EE2 were about 2.69, 3.35, 2.75, and 2.32 times as much as that with the MZNIP-coated fibers. It was indicated that MZMIP-coated fiber exhibited high extraction capabilities and good selectivities to the four estrogens because of its tailor-made specific recognition mechanism and large amount of MZMIP. The extraction efficiency of the MIP film is also crucially depended on the surface properties of carrier. Usually, Fe3 O4 @SiO2 was widely used as carrier for preparation of MIP for extraction of estrogens. Therefore, for comparison purpose, the adsorption isotherm of estrogens on MMIP-coating was also investigated. Fig. 4b shows that the amount of adsorbed E1, E2, E3, and EE2 on MMIP was about 309, 385, 328, and 216 ng, respectively, which were lower than those of the MZMIP coating for each estrogen and will affect the sensitivity of analytical method. At the same time, the selectivity of MMIP coating (2.10, 2.18, 1.95 and 1.69 for E1, E2, E3 and EE2, respectively) was also weaker than MZMIP coating. Fe3 O4 @ZIF-8 as the carrier material of imprinted polymer plays a key role. The reasons maybe as follows: (1) there are many surface-open Zn ion sites and imidazole ligands on the surface of Fe3 O4 @ZIF-8particles, which could strongly adsorb E2
through a coordinate bond between Zn2+ and hydroxyl group of the template molecular, and intermolecular – interactions between imidazole ligand and naphthalene ring of E2 [27]. Moreover, the pores of ZIF-8 are hydrophobic, which could also help for bonding hydrophobic E2. All these interactions can facilitate uniform and large distribution of the template molecule on the surface of porous Fe3 O4 @ZIF-8 for further creating the imprinted film. (2) The imidazole ring of ZIF-8 can react with APBA as monomer through – stacking interaction, which facilitate the immobilization of a stable MIP film on the surface of Fe3O4@ZIF-8particles. After the templates were eluted, more imprinted sites could be obtained due to the largely bonded template molecules. However, the hydrophilic Fe3 O4 @SiO2 has weak absorption capacity with hydrophobic E2 for creating imprinted sites and preparing MIP film. (3) The rugged and porous surface of Fe3 O4 @ZIF-8 as carrier can result in a larger specific surface area of MIP film than the relatively smooth Fe3 O4 @SiO2 (see Fig. 4S(b)). The nitrogen adsorption desorption measurements were employed to evaluate the surface properties between MMIP and MZMIP. The BET surface area, porous value, and pore size of MZMIP were 184.59 m2 g−1 , 0.21 cm3 g−1 , and 9.18 nm, respectively. But for MMIP, the parameters were 87.1 m2 g−1 , 0.085 cm3 g−1 , and 5.24 nm, respectively. Therefore, all these results may be attributed to the fact that MZMIP could have higher extraction capacity than MMIP. Moreover, in order to investigate the reproducibility of extraction efficiencies, the intraday, interday, and fiber-to-fiber repeatability of extraction by MZMIP-coated fiber were studied
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Fig. 4. Extraction amount curves of MZMIP (solid line) and MZMIP (dotted line) fibers to E1 (), E2 (), E3 (), and EE2 (䊉) mixed solution in n-hexane of 0.1–10 mg L−1 (a); extraction amount curves of Fe3 O4 @SiO2 @MIP (solid line) and Fe3 O4 @SiO2 @NIP (dotted line) fibers to E1 (), E2 (), E3 (), and EE2 (䊉) mixed solution in n-hexane of 0.1–5 mg L−1 (b).
with 200 g L−1 E1, E2, E3, and EE2 mixed standard solutions, and RSDs (n = 6) of 1.7–2.6%, 2.9–4.5%, and 2.3–3.8% for extraction amounts of four estrogens were obtained, respectively. 3.4. The reusability, stability and preservability of MZMIP coating To investigate the robustness and reusability of the developed coating, MZMIP-coated stainless steel fiber was applied for 250 repetitious SPME operations. Performance of the MZMIP coating was evaluated through extraction recovery and physical stability. The coating demonstrated good reproducibility for at least 250 times with recovery over 80% and related standard deviation (RSD)
