Preconcentration of bisphenols by using calix[4]arene derivatives modified porous polymer monolith coupled with HPLC

Abstract

A functionalized calix[4]arene (alkenyl@C[4]A) was introduced into the poly(butyl methacrylate-ethylene dimethacrylate) monolith inside a capillary to prepare a polymer monolith microextraction (PMME) material (poly(BMA-alkenyl@C[4]A-EDMA)). Various techniques, including scanning electron microscopy, Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, and thermo-gravimetric analysis, were employed to characterize the synthesized poly(BMA-alkenyl@C[4]A-EDMA) monoliths. A new method was developed for the preconcentration and analysis of bisphenols using PMME coupled with high performance liquid chromatography (HPLC). Under the optimized conditions, the monolithic column afforded acceptable linearities, low limits of detection, and good intra-day/inter-day relative standard deviations. The method was applied to the determination of bisphenols in human serum samples with recoveries ranged from 77.6% to 109.7%.

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1. Introduction

Compounds which are able to mimic natural hormones in the endocrine system, commonly called endocrine-disrupting chemicals (EDCs), have become an important issue to investigate in the last few years.^1,2 Among EDCs, bisphenols are a group of chemicals with two hydroxyl phenyl functionalities and include several analogues such as bisphenol A, bisphenol B, bisphenol F, bisphenol AP, bisphenol Z, etc. Bisphenols have been widely used as the base chemical in the manufacturing of polycarbonate plastics and the resin lining of food and beverage cans.^3–7

The extensive use of bisphenols, the hazards associated with them, and increasingly restrictive legal regulations underline the need for efficient and sensitive analytical methods to detect them. Several methods for quantitative analysis of bisphenols have been developed such as HPLC,^8–10 LC/MS,^4,11–14 and GC/MS.^5,15–17 For the determination of complex samples, a sample pretreatment step to separate and/or pre-concentrate the analytes is often required prior to the analysis step. Among the various sample preparation methods, liquid–liquid extraction (LLE),^5,11,16 solid phase extraction (SPE),^8,12 and solid-phase microextraction (SPME)¹⁵ have been applied extensively to the determination of bisphenols.

As one of SPME methods, a novel method termed polymer monolith microextraction (PMME) was introduced by Feng's group in 2006.^18,19 Compared with traditional packed column, porous monolithic column has distinct advantages of high capacity, fast separation speed, low cost, long life, good permeability offered by through pore, excellent biocompatibility, and easy preparation with good control of porosity and diverse surface chemistry.^20,21 Due to the varieties of potential applications of monoliths, it is desirable to explore efficient approaches enabling their functionalization. Conventional functionalization can be classified into two strategies: copolymerization of functional monomers²² and post-polymerization functionalization.^23,24 In both cases, the resulting functionalities depend on the structure and properties of functional monomers. Monolithic columns are usually functionalized by various materials such as graphene,²⁵ nano-oxide,²⁶ metal nano-materials²⁷ to obtain higher selective retention adsorption capacity.

Calix[n]arenes (mainly with n = 4, 6 and 8) are a type of macrocyclic compound containing n-phenol moieties which are connected by methylene bridges to form a hydrophobic cavity, exhibiting superior recognition ability.^28–30 During the last two decades, they have attracted much attention as receptors in supramolecular chemistry.^31–33 This can be largely attributed to the fact that they are attractive host molecules that can be easily functionalized at the narrow (phenolic groups) or at the wide rim (aromatic nuclei) to build suitable binding sites for more target guest species.^34–36

With the extensive and in-depth studies, more and more attention has been paid to calixarenes and their derivatives which maybe processed into materials with potential application perspective in adsorption.^37–40 For example, novel thiacalix[4]arene polymers were investigated for the adsorption of dye.³⁹ The adsorption of the direct black-38 (DB-38) azo dye on potential and newly synthesized p-tert-butylcalix[6]arene based silica resin was also studied.⁴⁰ In Kitano et al.'s work, a selective complexation of guest molecules with calixarene derivatives was realized by employing the optimum cavity size of the latter.⁴¹ However, to the best of our knowledge, there have been no reports on the synthesis and utilization of calixarene derivatives adsorbent materials in combination with PMME for bisphenols.

We reported the synthesis of alkenyl@C[4]A combined with PMME in this work to set up a simple, economical, and sensitive method. The alkenyl@C[4]A was firstly introduced into poly(butyl methacrylate-ethylene dimethacrylate) monolith as pretreatment materials for the enrichment of bisphenols in human serum. The aim of this study is to explore the application of calixarene derivatives to the microextraction technology as adsorption materials.

2. Experimental section

2.1 Materials and chemicals

Butyl methacrylate (BMA), 3-(trimethoxysilyl) propyl methacrylate, and ethylene dimethacrylate (EDMA) were obtained from Sigma-Aldrich (St. Louis, MO, USA). 1-Ethyl-3-(3-dimethylaminepropyl) carbodiimide (EDC) and N-hydroxysulfosuccinimide (sulfo-NHS) were purchased from Aladdin Reagent (Shanghai, China). n-Propanol, acetone, and 1,4-butanediol were purchased from Tianjin Chemical Plant (Tianjin, China). Azobisisobutyronitrile (AIBN) was obtained from Tianjin Guangfu Fine Chemical Research Institute (Tianjin, China). Methanol (MeOH) and acetonitrile (MeCN) were supplied by Fisher Scientific (Shanghai, China). Sodium hydroxide, and sodium dihydrogen phosphate (NaH₂PO₄) were obtained from Beijing Chemical Works (Beijing, China). Ammonia solution (25%) was from Xilong Chemical Co., Ltd. (Guangdong, China).

2,2-Bis(4-hydroxyphenyl)propane (bisphenol A, BPA), 2,2-bis(4-hydroxyphenyl)butane (bisphenol B, BPB), and 4,4-dihydroxydiphenylmethane (bisphenol F, BPF) were purchased from Aladdin Reagent (Shanghai, China), Dr. EhrenstorferGmbh (Augsburg, Germany), and Alfa Aesar (Shanghai, China). 4,4′-(1-Phenylethylidene)bisphenol (bisphenol AP, BPAP) and 4,4′-cyclo-hexylidenebisphenol (bisphenol Z, BPZ) were obtained from ANPEL Laboratory Technologies Inc. (Shanghai, China).

Carboxylatocalix[4]arenes were synthesized and purified by recrystallization according to a previous literature.^42,43 Ultrapure water was obtained with a Milli-Q SP system (Millipore, Milford, MA, USA). All solvents and solutions for HPLC analysis were filtered through a Millipore filter (0.45 μm). Fused silica capillaries (530 μm i. d.) were purchased from Yongnian Optical Fiber Factory (Handan, China).

2.2 Instrumentation

The morphology and structure of the monolith were observed by an S4800 ESEM Hitachi microscope (SEM, Hitachi, Japan). Fourier-transformed infrared (FTIR) spectra were obtained using a Thermo Nicolet 670 FTIR instrument (Thermo, USA). An X-ray photoelectron spectrometer (Thermo Electron, USA) was used to obtain the XPS data. Thermal gravimetric analysis (TGA) determinations were performed on a Q500 thermal gravimetric analyzer (TA, USA). A KQ-200KDE digital ultrasonic cleaner was obtained from Kunshan Ultrasonic Instruments Co., Ltd. (Kunshan, China). pH measurements of sample solutions were achieved by a pHS-3C digital pH meter (Shanghai Rex Instruments Factory, China). A GenieTouch syringe pump from Kent Scientific (San Francisco, USA) was employed for providing the driving force for extraction.

2.3 Preparation of poly(BMA-alkenyl@C[4]A-EDMA) monolithic column

Firstly, 100 mg carboxylatocalix[4]arene was completely dispersed in 100 mL ultrapure water by ultrasonication for 3 h. Then 0.5 mg EDC and 0.2 mg sulfo-NHS were added and stirred for 30 min, respectively, followed by the addition of 0.5 mL allylamine with stirring all the night. Finally, the mixture were filtered and dried to obtain the alkenyl groups functionalized carboxylatocalix[4]arene (hereafter abbreviated as alkenyl@C[4]A).

Monoliths were synthesized inside the preconditioned fused silica capillary (530 μm i. d. × 20 cm).¹⁹ Briefly, the fused silica capillary was washed subsequently with acetone, 0.1 mol L⁻¹ NaOH, ultrapure water, 0.1 mol L⁻¹ HCl, ultrapure water, and acetone each for 20 min at the flow rate of 1.0 mL min⁻¹. Afterwards the inner wall of the capillary was modified with 3-(trimethoxysilyl) propyl methacrylate (50% in acetone, v/v) to enable covalent attachment of the monolith at 50 °C for 14 h. Then, the residual solution was driven out and the capillary was rinsed thoroughly with acetone. Finally, N₂ was driven to flow through the capillary to dry the inner surface at room temperature prior to use.

In a typical process, the polymerization mixture for preparing the poly(BMA-alkenyl@C[4]A-EDMA) monolith containing 180 mg BMA, 120 mg EDMA, 396 mg n-propanol, 304 mg 1,4-butanediol, 4 mg AIBN, and 15 mg alkenyl@C[4]A were mixed thoroughly by ultrasonication and purged with N₂ for 10 min to remove O₂. After the capillary was filled with the polymerization mixture, the two ends were immediately closed with a silicone rubber. The reaction was kept at 60 °C for 22 h.

2.4 Polymer monolith microextraction procedure

The synthesized poly(BMA-alkenyl@C[4]A-EDMA) monolithic column was employed for the extraction of bisphenols. A typical procedure was as follows. For the adsorption step, the sample solution (1 mL) was pumped through the capillary at the flow rate of 0.05 mL min⁻¹, and then 0.5 mL phosphate buffer (0.02 mol L⁻¹, pH = 3) was driven at the same flow rate to get rid of the residual matrix in order to avoid the interference for separation. Then the residual phosphate buffer solution was expelled from the monolithic capillary. For the desorption step, the eluent of 1% ammonia solution/MeOH (1 : 1, v/v) was injected in the monolithic capillary at 0.05 mL min⁻¹ and the eluate was collected for the HPLC analysis.

2.5 HPLC analytical conditions

Chromatographic analysis was performed on an Agilent 1100 liquid chromatography system equipped with a quaternary pump and degasser, a thermostated autosampler and column compartment, a multiple wavelength detector, and Chem-Station software. A reversed phase Agilent Zorbax Eclipse XDB-C18 column (4.6 mm × 250 mm, 5 μm) was employed for the chromatographic separation. A phenomenex C18 security guard column (4.0 mm × 3.0 mm) from Phenomenex, Torrance, CA was used to protect the column. The mobile phase consisted of A and B, where A represented water and B represented MeOH. A gradient program was used for elution: 0 min, 60% B; 0–25.0 min, 60–80% B; 25.0–25.5 min, 80–60% B. The flow rate was set at 0.8 mL min⁻¹ while the column temperature was maintained at room temperature. The injection volume was 10 μL and the detection wavelength was set at 225 nm.

2.6 Sample preparation

Human serum samples were collected from a local hospital. All blood samples were allowed to clot at room temperature for 15–30 min and then refrigerated until centrifugation at 4 °C for 15 min. The serum was transferred into 10 mL centrifuge tubes and then frozen at −20 °C. Before use, the serum was thawed at room temperature, and the samples were five times diluted with phosphate buffer (0.02 mol L⁻¹, pH = 3). Then the sample solution was degassed and homogenized with an ultrasonic bath and filtered through a 0.45 μm membrane filter prior to analysis.

3. Results and discussion

3.1 Preparation and characterization of the alkenyl@C[4]A

The composite of alkenyl@C[4]A was prepared as mentioned above. Allylamine and carboxylatocalix[4]arene were connected in the presence of EDC/NHS which aided in the formation of amide bond. The alkenyl@C[4]A was represented by FTIR and XPS (see Fig. S1†). When allylamine bonded with carboxylatocalix[4]arene, the peak at 1610 cm⁻¹ in Fig. S1A† was obviously stronger, which might be because of the generation of amide bond and increasing numbers of C=C bond. The primary characteristic N 1s spectrum peak appeared at 399.9 eV but moved to 401.9 eV (Fig. S1B†) due to the formation of –NHCOCH₂–, which further confirmed the presence of amide bond.

3.2 Preparation and characterization of poly(BMA-alkenyl@C[4]A-EDMA) monolithic column

The poly(BMA-alkenyl@C[4]A-EDMA) monolith was synthesized by one-step method and the preparation scheme was illustrated in Fig. 1. In the synthesis process, effects of the monomer/porogen weight ratios were optimized to achieve the best combination for each component. Results were listed in Table S1,† showing that the monolith named C-3 presented the optimal perviousness. Subsequently, the amount of alkenyl@C[4]A was further investigated by polymer reaction liquid property, permeability, extraction efficiency and morphology. The effects of different additive amount of alkenyl@C[4]A on polymer properties and perviousness were shown in Table S2 and Fig. S2.† We chose the column with 10–40 mg alkenyl@C[4]A to investigate the extraction efficiency and found that the enrichment capacity of poly(BMA-alkenyl@C[4]A-EDMA) monolith added with 15 mg alkenyl@C[4]A was the best one. Finally, 15 mg alkenyl@C[4] (1.5 wt%) was added into the monolith in subsequent experiments.

Fig. 1 Synthesis process of poly(BMA-alkenyl@C[4]A-EDMA) monolithic material.

SEM images reflected the diverse morphology feature of the monolithic materials. Fig. 2 showed SEM images of the monolithic capillary columns containing varying quantities of alkenyl@C[4]A (0, 15, 40 mg). There are homogeneous porous features in the network skeleton of the polymer monolith, which guarantees fast dynamic transport and high efficient enrichment in applications.²⁴ Compared with the image in Fig. 2A, the morphology features were much rougher in Fig. 2B and C, which were caused by embedding alkenyl@C[4]A. In Fig. 2C, increasing alkenyl@C[4]A amount resulted in the agglomeration phenomenon and the pore size and porosity of the prepared monolith gradually reduced. The throughout pores and porosity affect the permeability of the monolithic column and further influence the substantial contributions to the adsorption efficiency.⁴⁴ So, 15 mg alkenyl@C[4] (1.5 wt%) was chosen in the experiments.

Fig. 2 SEM images of poly(BMA-alkenyl@C[4]A-EDMA) monolithic columns containing varying quantities of alkenyl@C[4]A: (A) 0 mg, (B) 15 mg, and (C) 40 mg.

FT-IR spectra of poly(BMA-EDMA) and poly(BMA-alkenyl@C[4]A-EDMA) monoliths between 4000 and 400 cm⁻¹ were shown in Fig. 3A. In Fig. 3A(b), the infrared absorption peak at 1626 cm⁻¹ was the characteristic peak of amide bond. The band at 3417 cm⁻¹ was attributed to –OH and N–H bonds stretching vibrations, and the band at 2962 cm⁻¹ was C–H stretching vibration. The successful introduction of alkenyl@C[4]A could be proved from these absorption peaks.

Fig. 3 (A), (a) IR spectra of poly(BMA-EDMA) and (b) poly(BMA-alkenyl@C[4]A-EDMA); (B) XPS spectra of poly(BMA-alkenyl@C[4]A-EDMA); (C) TGA curves of poly(BMA-EDMA) and poly(BMA-alkenyl@C[4]A-EDMA).

As is known, X-ray photoelectron spectroscopy (XPS) can provide an indication of the chemical environment of atoms. Therefore, the element composition loading on the poly(BMA-alkenyl@C[4]A-EDMA) column was detected by XPS. In Fig. 3B, XPS analysis was performed to identify the elemental composition with the spectra peaks of C 1s, O 1s, and N 1s. The spectra indicated the materials were successfully connected with alkenyl@C[4]A.

To examine the thermal properties and stability of poly(BMA-EDMA) and poly(BMA-alkenyl@C[4]A-EDMA) monoliths, TGA analysis was carried out under N₂ atmosphere at a heating rate of 10 °C min⁻¹ over the temperature range of 30–800 °C. The mass loss which takes place below 200 °C is probably due to the physical desorption of water and the evaporation of methanol. The tiptop pyrolysis temperatures of poly(BMA-EDMA) were 172 and 279 °C, while that of poly(BMA-alkenyl@C[4]A-EDMA) was about 340 °C in Fig. 3C. For poly(BMA-alkenyl@C[4]A-EDMA) materials, when the temperature raised from 276 to 470 °C, the weight loss might be attributed to the decomposition of alkenyl@C[4]A. The pyrolysis behavior of poly(BMA-EDMA) and poly(BMA-alkenyl@C[4]A-EDMA) materials ended in 400 °C and 470 °C, respectively. In conclusion, the thermal stability of poly(BMA-alkenyl@C[4]A-EDMA) materials were relatively strong.

In order to examine the permeability of the monolithic column, the backpressure was investigated and the values of permeability (K) were estimated using the following equation: K = FηL/SΔP,⁴⁵ where K is the permeability, F is the flow rate of the pump, η is the solvent viscosity, L is the column length, S is the innercross-sectional area of the column, and ΔP is the back pressure. The permeability value of the poly(BMA-alkenyl@C[4]A-EDMA) monolith was calculated to be 2.91 × 10⁻¹³ m² when methanol was selected as the mobile phase at 20 °C with the flow rate was set at 0.5 mL min⁻¹. This result also implied that collapse could be avoided when the monolith was subjected to flow through liquids.

3.3 Optimization of polymer monolith PMME conditions

In order to achieve the best extraction efficiency for the poly(BMA-alkenyl@C[4]A-EDMA) monolith, various parameters including sample pH, sample volume, sample flow rate, eluent type, and eluent flow rate, were investigated when the concentration of bisphenols was set at 1.0 μg mL⁻¹.

To obtain high extraction efficiency for desorption, eluent is crucial in PMME procedures. Eluents with different alkaline and polar types were investigated including MeOH, 1% ammonia solution/MeOH (1 : 1, v/v), ACN, and 1% ammonia solution/ACN (1 : 1, v/v). As shown in Fig. 4A, 1% ammonia solution/MeOH (1 : 1, v/v) manifested better and more balanced elution ability for all the targets than the other three eluents. A possible explanation may be as follows. The interactions between the polymer monolith and targets are hydrogen bonds formed by amide groups at the surface of the porous polymer monolithic column and phenolic hydroxyl groups of bisphenols and π–π interactions between alkenyl@C[4]A and bisphenols. Therefore, strongly polar eluent may contribute to the desorption of the analytes from the polar adsorbent. In addition, the high pH caused by the addition of ammonia solution goes against the formation of hydrogen bonds, which facilitates desorption. Therefore, 1% ammonia solution/MeOH (1 : 1, v/v) was employed as the eluent in subsequent experiments.

Fig. 4 Effects of (A) type of eluent, (B) sample pH, and (C) sample volume on the extraction efficiency.

As a significant parameter, sample pH dependencies were investigated in the range of 2.0–7.0 to obtain the maximum adsorption capacity (Fig. 4B). Results demonstrated that the highest extraction efficiency achieved at around pH 3.0, which might be explained by the hydrophobic interaction, hydrogen bonding interaction, and electrostatic repulsion between the analytes and the monolithic material.¹⁹ When pH was too high, the hydrogen bonding interaction between the monolith and phenolic groups of bisphenols decreased. When pH was too low, the electrostatic repulsion increased, resulting in the decrease of extraction efficiency. Thus, pH 3.0 was selected in the following studies. The influence of sample volume was evaluated in the range of 0.6–1.4 mL. Results were demonstrated in Fig. 4C, illustrating that a significant increasing tendency of the peak areas was observed when the sample volume increased from 0.6 to 1.4 mL and the extraction equilibrium did not reach a maximum. It could be expected that the peak areas would stop increasing after the sample volume reached a certain value. Taking the time of experiment into consideration, 1.0 mL sample volume was selected for further experiments.

Sample flow and eluent flow rates are also important parameters, which not only affect the adsorption and desorption, but also control analysis time. In consideration of the perviousness of the monolith and the effect of extraction, flow rate values were investigated in the range of 0.02–0.1 mL min⁻¹ and the optimal values were obtained at the flow rate of 0.05 mL min⁻¹. Too low flow rate is not favorable for the mass transfer during the extraction. In addition, low sample rate leads to long operation time. However, too high flow rate is not conducive to sufficient adsorption and desorption.^24,25 Therefore, 0.05 mL min⁻¹ was selected as sample and eluent flow rates in this work.

3.4 Method validation

Under the optimal experimental conditions, analytical features of the developed method including linearity, limits of detection (LOD), limits of quantitation (LOQ), relative standard deviation (RSD), and enrichment factor (EF) were summarized in Table 1. The calibration curves of the analytes were constructed with the linear range (LR) of 0.005–5 μg mL⁻¹. The analytes showed good linearity with R² values greater than 99%. According to IUPAC recommendations, LOD and LOQ, based on the signal-to-noise (S/N) ratio of 3 and 10, were calculated to be in the range of 0.0012–0.0024 μg mL⁻¹ and 0.0038–0.0080 μg mL⁻¹, respectively. The reproducibility of the method was evaluated by RSD values. The intra-day and inter-day RSDs, determined in 5 replicates for all the target analytes, were in the ranges of 4.4–6.8% and 5.7–9.1%, respectively. The EF values of 5 analytes were 11.4–40.6, calculated as the ratios between the slopes of the linear calibration curves with and without employing the PMME method. The reproducibility of the preparation and practicability of the monolith are very significant factors for the performance evaluation. The intra-batch and inter-batch RSDs were also determined to be lower than 7.6 and 8.7%. The results demonstrated good reproducibility of the monolithic capillary preparation. Furthermore, poly(BMA-alkenyl@C[4]A-EDMA) monolith showed high stability in use (200 times within two months) without significant changes in the extraction efficiency.

Table 1 Analytical performance of the target bisphenols with the PMME-HPLC method under optimized conditions (n = 5)

Analytes	Linear range (μg mL^−1)	R^2	LOD (μg mL^−1)	LOQ (μg mL^−1)	EF	RSD (%)
						Intra-day	Inter-day
BPF	0.005–5	0.9989	0.0024	0.0080	18.2	6.8	7.5
BPA	0.005–5	0.9979	0.0012	0.0040	11.4	5.0	8.6
BPB	0.005–5	0.9949	0.0019	0.0064	17.8	5.6	6.3
BPAP	0.005–5	0.9969	0.0012	0.0038	40.6	4.4	5.7
BPZ	0.005–5	0.9999	0.0012	0.0042	33.8	6.2	9.1

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The comparison study between direct HPLC analysis and PMME using poly(BMA-EDMA) and poly(BMA-alkenyl@C[4]A-EDMA) monoliths was performed under the optimized conditions (sample pH of 3.0, sample volume of 1.0 mL, sample and eluent flow rates of 0.05 mL min⁻¹). For poly(BMA-EDMA) monolith, there are hydrophobic groups on the surface of monolithic column, resulting in the extraction capability for bisphenols. When alkenyl@C[4]A was introduced into the polymerization mixture, poly(BMA-alkenyl@C[4]A-EDMA) monolith demonstrated the preferable extraction ability. This may be ascribed to several aspects: first, amide groups at the surface of the porous polymer monolithic column, which can form hydrogen bonds with the phenolic hydroxyl groups of bisphenols, so it is expected to present good extraction capability; second, the role of alkenyl@C[4]A employed in the present study was not only to enhance hydrophobic interaction with bisphenols, but also to π–π interactions between alkenyl@C[4]A and bisphenols. Therefore, as obviously shown in Fig. 5, a significant enhancement of peak area was observed after extracted by poly(BMA-alkenyl@C[4]A-EDMA) monolith, indicating the remarkable preconcentration ability of the column for all the targets.

Fig. 5 Comparison of analytical performance of direct HPLC analysis with that of poly(BMA-EDMA) monolithic column and poly(BMA-alkenyl@C[4]A-EDMA) monolithic column.

3.5 Application to real samples

The performance of the present PMME-HPLC method was tested to determine the bisphenols in human serum samples from local hospital. All the real samples were spiked with 5 standard solutions of bisphenols at two concentration levels to assess the matrix effects, Level 1 (0.02 μg mL⁻¹) and Level 2 (0.05 μg mL⁻¹), respectively. As shown in Table 2, the recovery values demonstrated that a sufficient recovery could be achieved in the range of 77.6–109.7% with RSD values between 0.5% and 7.9% (n = 3). The consequences in Table 2 also demonstrated that the content of BPA and BPZ is relatively larger in human serum sample than other bisphenols. The results suggest that the present PMME-HPLC method is effective when applied to the determination of bisphenols in human serum sample.

Table 2 Recoveries (%) of the five bisphenols in samples (n = 3)

Sample			BPF	BPA	BPB	BPAP	BPZ
Sample 1	Measured (ng mL^−1)		<LOD	130.0	<LOD	<LOD	140.0
	Recovery ± RSD (%)	Level 1	77.6 ± 3.6	83.4 ± 0.8	105.4 ± 1.6	99.2 ± 7.0	90.9 ± 4.9
		Level 2	90.0 ± 1.7	103.8 ± 4.9	91.3 ± 5.0	99.7 ± 4.8	92.9 ± 6.6
Sample 2	Measured (ng mL^−1)		<LOD	63.7	<LOD	31.2	26.2
	Recovery ± RSD (%)	Level 1	89.5 ± 4.8	85.6 ± 4.7	95.2 ± 6.0	89.7 ± 7.9	79.6 ± 2.9
		Level 2	92.0 ± 2.3	99.7 ± 4.2	90.1 ± 2.6	100.0 ± 1.1	87.2 ± 7.7
Sample 3	Measured (ng mL^−1)		<LOD	830.0	<LOD	<LOD	37.2
	Recovery ± RSD (%)	Level 1	85.1 ± 0.9	93.9 ± 5.4	79.0 ± 4.8	88.6 ± 1.3	100.8 ± 5.9
		Level 2	86.6 ± 6.6	85.6 ± 1.7	87.4 ± 5.8	88.8 ± 7.9	90.3 ± 1.5
Sample 4	Measured (ng mL^−1)		<LOD	<LOD	<LOD	<LOD	<LOD
	Recovery ± RSD (%)	Level 1	88.0 ± 6.6	101.8 ± 1.7	87.2 ± 4.5	96.2 ± 5.9	100.1 ± 4.9
		Level 2	88.7 ± 4.6	95.2 ± 0.5	93.7 ± 2.5	83.0 ± 1.7	85.0 ± 5.7
Sample 5	Measured (ng mL^−1)		<LOD	<LOD	<LOD	<LOD	<LOD
	Recovery ± RSD (%)	Level 1	102.2 ± 5.7	83.9 ± 5.7	100.4 ± 5.2	89.7 ± 0.6	84.2 ± 2.7
		Level 2	97.1 ± 4.4	101.9 ± 7.8	107.5 ± 5.7	92.5 ± 2.9	92.6 ± 6.1
Sample 6	Measured (ng mL^−1)		<LOD	<LOD	<LOD	23	3.9
	Recovery ± RSD (%)	Level 1	94.4 ± 0.9	87.3 ± 5.4	94.4 ± 4.8	92.5 ± 1.3	93.1 ± 5.9
		Level 2	95.4 ± 6.6	107.1 ± 1.7	102.7 ± 5.8	87.6 ± 7.9	100.7 ± 1.5
Sample 7	Measured (ng mL^−1)		<LOD	54.4	<LOD	<LOD	29.1
	Recovery ± RSD (%)	Level 1	88.4 ± 6.6	109.7 ± 1.7	85.4 ± 4.5	78.0 ± 5.9	89.4 ± 4.9
		Level 2	97.1 ± 4.6	92.3 ± 5.0	104.6 ± 2.5	103.3 ± 1.7	80.7 ± 5.7
Sample 8	Measured (ng mL^−1)		<LOD	<LOD	<LOD	<LOD	119.5
	Recovery ± RSD (%)	Level 1	86.5 ± 5.7	93.8 ± 5.7	109.1 ± 5.2	83.5 ± 0.6	93.2 ± 2.7
		Level 2	79.0 ± 4.4	79.0 ± 7.8	92.2 ± 5.7	93.2 ± 2.9	96.8 ± 6.1

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4. Conclusions

In summary, we designed and synthesized a new type of poly(BMA-alkenyl@C[4]A-EDMA) monolithic materials via a simple procedure. Alkenyl@C[4]A was successfully modified into the polymer monolith through the in situ polymerization method. The obtained material was characterized by SEM, FT-IR, XPS, and TGA, demonstrated the connection of alkenyl@C[4]A and poly(BMA-EDMA) monolith. Based on the novel adsorbent material, a PMME-HPLC method for the simultaneous extraction and detection of bisphenols, BPA, BPB, BPF, BPAP and BPZ, were set up. The adsorption procedure is mainly controlled by various interactions involving hydrogen bond, π–π complexation, and hydrophobic interactions. Under the optimal extractive conditions, the preconcentration efficiency for bisphenols of poly(BMA-alkenyl@C[4]A-EDMA) was obviously higher than that of pure methacrylate-based monoliths. The method was applied to the determination of bisphenols in human serum samples with recoveries ranged from 77.6% to 109.7% with relative standard deviations less than 7.9%. It can be expected that the prepared macrocyclic compound or supramolecular complex decorated polymer monolith as a PMME method will be an excellent chromatographic separation media.

Acknowledgements

The project was supported by National Natural Science Foundation of China (No. 21575049) and Jilin Provincial Science & Technology Department (No. 20140101112JC).

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Footnote

† Electronic supplementary information (ESI) available. See DOI: 10.1039/c6ra03352k

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