Study On Application Of Rapid Detection Technique In The Enviromental Medicine Based On The Novel Molecularly Imprinted Composite Materials

With the development of life science and enviromental science, health effects of enviromental pollution are investigated deeply based on the personal exposure, individual effect, population health effect. The personal exposure is obtained from the determination of pollutant in complex enviromental and foodstuff samples. The individual effect is investigated based on the distribution and metabolism of enviromental pollutant in the body. The population health effect is investigated based on the detection of corresponding biomarker in a large number of human samples. Therefore, study on rapid determination of trace pollutant in enviromental and foodstuff samples and then high-throughput analysis of corresponding biomarker in biological samples are beneficial to discuss the comprehensive evaluation for health effects of enviromental pollution, which plays an important role in enviromental medicine. Nowadays, with the development of modern analytical technique, highly sensitive, selective and stability instruments are on the state of tending to mature developing. Therefore, the sample pretreatment is the main step of rapid and high-throughout detection technique. The development of automatic, simple and rapid, highly effective and sensitive sample pretreatment is favourable to shorten analytical time, improve the reproducibility, accuracy and precision of the results, avoid secondary pollution and reduce the running cost. However, because of the complex sample matrix and low selectivity of traditional technique, several matrix components are simultaneously extracted, which can interfere with the determination of target compounds. Therefore, it is imperative for us to develop rapid and high-throughout detection technique based on new high selective materials.Molecularly imprinted polymers (MIPs) are tailor-made materials that can exhibit high affinity and selectivity towards a given analyte, or a group of structurally related compounds. Furthermore, these materials have excellent mechanical strength, long lifetime and strong durability to heat, acid and base conditions. Recently, successful applications of MIPs have been described in the literatures. Current trends in sample pretreatment are clearly towards developing mutifunctional imprinted composite materials, which display dual character by combining the specific properties of functional nanoparticles with the recognition capabilities of MIPs. Because of the complementary advantage, these composites have been recognized as promising and innovative materials and received increased interest in the practical application. The whole study is composed of the following three parts:Part I:Automated on-line analysis of trace tetracycline antibiotics in foodstuff samples based on mixed-templates molecularly imprinted polymersWe developed in the present study a set of molecularly-imprinted polymers (MIPs) by precipitation polymerization using four members of tetracycline antibiotics (TCs) as templates (Oxytetracycline, Tetracycline, Chlortetracycline and Doxycycline) and studied the recognition mechanism of these polymers for TCs. Based on the imprinting effect of different MIPs (TCs), Oxytetracycline and Chlortetracycline were chosen as mixed-templates to synthesize high selective MIPs for recognition of TCs groups. It was shown that the optimal imprinting factors of TCs on mixed-templates MIPs were more than 6.0 and the maximum binding amount of TCs were about 27μmol g-1 (Oxytetracycline),35 μmol g-1 (Tetracycline),35μmol g-1 (Chlortetracycline) and 39μmol g-1 (Doxycycline), respectively. Furthermore, an automated system has been developed for determination of trace tetracycline antibiotics (TCs) in foodstuff samples, based on on-line molecularly imprinted solid-phase extraction (MISPE) coupling to high-performance liquid chromatography (HPLC). Under the optimal on-line MISPE-HPLC condition,10 milliliter foodstuff samples were injected to the MISPE column and then the matrix was washed out. By rotating the switching valve, TCs were transferred to the analytical column and then separated by HPLC. Because the sample pretreatment and chromatographic separation were earried out simultaneously, the whole analytical time (17 min) was significantly shortened compared with conventional off-line sample pretreatment techniques. The detection limits ranged from 0,8 to 1.3 ng g-1. The enhancement factors were in the range of 159-410. The spiked recoveries of TCs in real foodstuff samples ranged from 91.6% to 107.6% and the relative standard deviations was not higher than 4.0%. It was shown that this method could be used to determine trace TCs in complex foodstuff samples, which laid the foundation for the evaluation of foodstuff pollution and investigation of health effect.PartⅡ:Rapid electrochemical determination of trace 2,4-dinitrophenol in surface water samples based on hydrophilic molecularly imprinted polymers/Ni fiber compositeHydrophilic molecular imprinted polymers (MIPs) were synthesized by two-step polymerization, using 2,4-dinitrophenol (2,4-DNP) as the template, acrylamide as the monomer and glycidilmethacrylate (GMA) as the pro-hydrophilic co-monomer. In this study, preparation of hydrophilic MIPs was optimized and the rebinding capacity, selectivity and rebinding kinetic behavior of hydrophilic MIPs were investigated in water media and acetonitrile media. It was shown that GMA could induce a hydrophilic behavior to the imprinted particles and had low ability to interfere the pre-polymerization of template/monomer. Therefore, hydrophilic MIPs could greatly reduce the non-specific hydrophobic interaction between the template and MIPs, and enhance the accessibility of 2,4-DNP in water medium to the imprinted cavities. Compared with traditional MIPs, hydrophilic MIPs exhibited excellent selectivity and rebinding capacity for nitrophenol compounds in water media. Furthermore, a rapid, sensitive and selective electrochemical method was proposed for the determination of 2,4-dinitrophenol (2,4-DNP) in surface water samples, using hydrophilic MIPs as the recognition element and nickel (Ni) fiber as the catalytic element. The parameters affecting the analytical performance were investigated. Under the optimized conditions, the linear range was 0.7-30μL-1 and the detection limit was 0.1μg L-1. Finally, the proposed method was applied to measure 2,4-DNP in surface water samples. The spiked recoveries were changed from 91.3% to 102.6% and the RSD was not higher than 5.1%. There was no statistically significant difference between the results obtained by the proposed method and the traditional chromatographic method. It was shown that this method could be used to determine trace 2,4-DNP in enviromental samples, which laid the foundation for the evaluation of environmental pollution and investigation of health effect. PartⅢ:Rapid and high-throughput determination of urinary lysozyme based on novel magnetic molecularly imprinted polymersIn this work, we presented a method for preparing multifunctional lysozyme imprinted nanoparticles (magnetic susceptibility, molecular recognition and environmental response). The magnetic susceptibility was imparted through the successful encapsulation of Fe3O4 nanoparticles. Selective lysozyme recognition depended on molecularly imprinted film. Moreover, it was also a hydrophilic stimuli-responsive polymer, which could undergo a reversible change of imprinted cavity in response to a small change in the environmental conditions. Thus, magnetic molecularly imprinted nanoparticles had high adsorption capacity (0.11 mg mg-1), controlled selectivity and direct magnetic separation (22.1 emu g-1) in crude samples. Furthermore, a rapid, low cost and selective chemiluminescence method coupled with magnetic molecularly imprinted polymers extraction was developed to detect lysozyme in human urine samples. Compared with traditional solid-phase extraction, this method could achieve selective extraction of lysozyme, avoid the time consuming elution from a column or centrifugation steps, and then showed great potential in the high-throughput screening of clinical samples. Under the optimal conditions, the whole analytical procedure was completed within 12 min and spiked recovery ranged from 90.1% to 103.7%(R.S.D≤6.7%). The limit of quantitation was 5 ng mL-1. Furthermore, the results obtained by the proposed method were linearly correlated to those by commercial lysozyme detection kit (r=0.9595, n=26). Finally, the validated method was used to measure the urinary lysozyme of renal disease patients and healthy controls. The results confirmed the reliability and practicality of the protocol and revealed a good perspective of this method for estimating the early damage of renal tubules caused by enviromental pollution.