基于抗生素残留分离/分析的咪唑型离子液体绿色双水相体系的构建及应用研究
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摘要
抗生素在临床、养殖业和食品加工业中的滥用已经给生态环境和人类健康带来了严重威胁。因此,对环境和食物中抗生素残留的分离/富集方法的建立成为关注的热点。由于环境和食物中抗生素基质成分复杂,干扰多,而且通常为痕量或超痕量存在,必须找到合理的方法对环境和食物样品中的抗生素进行分离/富集,从而达到分离干扰物质,降低检出限和提高测定准确度的目的。
本课题深入研究了咪唑型离子液体-盐双水相体系的液液相平衡行为及其分相能力,为其应用提供了充分的理论依据。在此基础上构建了离子液体双水相萃取和离子液体双水相气浮溶剂浮选体系,将其应用于环境和食物中抗生素残留的分离/富集中。通过优化抗生素萃取、浮选的影响因素,探讨萃取、浮选机理,为离子液体双水相萃取和离子液体双水相气浮溶剂浮选其它生物小分子物质提供可行性研究的可靠依据。主要研究结果如下：
1.咪唑型离子液体-盐双水相体系的构建及液液相平衡行为的研究
(1)通过对一系列咪唑型离子液体-盐双水相体系双节线数据的拟合,筛选得到了适用于咪唑型离子液体-盐双水相体系双节线数据拟合的最佳方程w1=exp(α+bw20.5+Cw2+dw22)。利用滴定法、密度计测定法以及“杠杆规则”计算法得到了离子液体-盐双水相体系的液液相平衡组成,并用Othmer-Tobias方程和Bancroft方程进行关联,结果令人满意。
(2)通过计算不同分相盐在1-丁基-3-甲基咪唑四氟硼酸([C4mim]BF4)水溶液中的有效排除体积,以及绘制以摩尔质量浓度为单位的双节线相图,得出阳离子的盐析能力为Na+>NH4+；阴离子的盐析能力为Po43->C6H5O73->CO32->SO42-≈C4H4O62->C4H4O42->H2PO4->C2H3O2->CI-。同理,根据计算6种分相盐在1-乙基-3-甲基咪哗四氟硼酸([C2mim]BF4)、1-丙基-3-甲基咪唑四氟硼酸([C3mim]BF4)和[C4mim]BF4中的有效排除体积,以及绘制双节线相图,得到离子液体的分相能力顺序为[C4mim]BF4>[C3mim]BF4>[C2mim]BF4。
(3)研究了温度对离子液体-盐双水相体系分相能力的影响。结果表明,随着温度的降低,双节线向左移动,两相区面积扩大,系线斜率增大,离子液体-盐双水相体系的成相能力逐步增强。
2.离子液体-盐双水相体系分离/富集抗生素残留
(1)构建[C4mim]BF4-Na2CO3双水相体系结合分子荧光光谱分离/富集罗红霉素残留。讨论盐的种类、Na2CO3和离子液体用量以及萃取温度的影响。在最佳条件下,该方法能满意地应用于分离/富集环境水样中的罗红霉素残留,检出限为0.03μg/mL,精密度为1.90%,平均萃取率大于90%。离子液体-盐双水相体系的成相机理用水合理论来解释。红外和紫外光谱表征共同证明在萃取过程中离子液体和罗红霉素之间没有发生化学键的变化,离子液体只是起到溶剂的作用。
(2)建立[C4mim]BF4-(NH4)2SO4双水相体系结合高效液相色谱-紫外法萃取磺胺类抗生素残留。实验选取了三种磺胺类抗生素(磺胺甲基嘧啶、磺胺甲恶唑和磺胺甲塞二唑)作为目标物研究和评估实验方法。选用单因素和响应曲面实验设计优化最佳萃取条件。在最佳条件下,将建立的方法用于测定一系列环境和食物样品,加标回收率在83.7～116.5%,方法的检出限为0.15～0.3 ng/mL,检量限为0.5～1.0 ng/mL,低于其它一些预处理技术。
(3)构建[C4mim]BF4-有机盐双水相体系结合高效液相色谱法萃取/分析氯霉素残留。探讨影响氯霉素分配行为的影响因素：有机盐的种类和含量、体系的pH值、[C4mim]BF4的加入体积和萃取温度。热力学研究证明疏水作用是氯霉素萃取的主要驱动力,同时盐析作用也十分关键。在最佳条件下,将该方法成功地用于分离/富集饲养水、牛奶和蜂蜜中的氯霉素残留,加标回收率在90.4～102.7%,方法检出限为0.3 ng/mL,检量限为1.0 ng/mL。此种离子液体-有机盐双水相萃取技术操作简单,并且对环境无污染,在抗生素分离中有着重要的应用价值。
3.离子液体双水相气浮溶剂浮选分离/富集氯霉素及机理研究
(1)建立氯化1-丁基-3-甲基咪唑四氟硼酸([C4mim]Cl)-K2HPO4离子液体双水相气浮溶剂浮选体系结合高效液相色谱法浮选/分离氯霉素残留。讨论了离子液体和盐的种类对离子液体双水相气浮溶剂浮选体系成相行为的影响,详细研究盐种类、K2HPO4浓度、体系pH值、氮气流速、浮选时间和[C4mim]Cl初始加入量对氯霉素浮选效率的影响。在最佳条件下,浮选湖水、饲养水、牛奶和蜂蜜中氯霉素残留,浮选率为97.1～101.9%。该方法的线性范围为0.5～500 ng/mL,方法检出限为0.1 ng/mL,检量限为0.3 ng/mL。该方法优于液液萃取、溶剂浮选以及离子液体双水相萃取,其富集倍数高、减少离子液体用量、对环境友好的优点,在小分子生物物质中的分离/富集具有重要的应用价值。
(2)以氯霉素为目标物,构建离子液体-K2HPO4双水相气浮溶剂浮选体系。利用四因素三水平响应曲面实验设计法优化氯霉素的影响因素,筛选出的最佳条件为：离子液体选择[C6mim]Cl, K2HPO4浓度为0.74 g/mL、浮选时间为50 min、浮选流速为50 mL/min,此时氯霉素的分配系数为405.71,浮选率为93.16%。三维响应曲面图可以得到影响氯霉素分配系数和浮选率的影响因素强度依次为浮选时间>浮选流速>IL的种类> K2HPO4的浓度。离子液体双水相气浮溶剂浮选机理是由离子液体双水相体系的成相机理和溶剂浮选机理两部分构成。
The abuse of antibiotics in clinical medicine, aquaculture and food processing industry has been posed a serious threat to the ecological environment and human health. Therefore, the establishment of the method for separation and enrichment of antibiotic residues in the environment and food becomes the focal point of attention. Due to the composition of antibiotics matrix is complicated and usually trace or ultra-trace exists, we must find a reasonable method for separation and enrichment of antibiotics in the environment and food. It will achieve the separation of interfering substances, lower detection limits and improve the measurement accuracy of the purpose.
This thesis deeply researches into the behavior of liquid-liquid equilibrium (LLE) of iminazole-ionic liquid aqueous two-phase system (ILATPS) and its phase-separation abilities, so this provides sufficient theoretical foundation for its application. On the basis of which, an ionic liquid aqueous two-phase extraction (ILATPE) system and an ionic liquid aqueous two-phase flotation (ILATPF) system were established, they can be applied to separate and concentrate antibiotic residues in the environment and food. Various factors influencing the extraction and flotation of antibiotics were optimized; the mechanisms of extraction and flotation were discussed. These studies provide a feasibility evidence on separation and concentration of other biological small molecules by ILATPE and ILATPF. The main results are as follows:
1. Construction of ILATPS and the research of LLE
(1) Through the fitting of a series of iminazole ILATPS of binodal data, the optimum equation was selected, and it was w,= exp(a+bw20.5+cw2+dw22). Based on the titration method, the densimeter method and "lever rule" calculation, the LLE compositions were obtained. The equations of Othmer-Tobias and Bancroft were used to fitting the LLE data, and the result was satisfactory.
(2) The salting-out abilities of salt ions were obtained by calculating the effective excluded volume (EEV) of different species of phase-separation salts in aqueous l-butyl-3-methylimidazolium tetrafluoroborate ([C4mimJBF4) solutions, and drawing binodal curves in molar mass concentration. The phase-separation abilities of cation are in the order Na+>NH4+, and the phase-separation abilities of anion are PO43->C6H5O73->CO32->SO42-≈C4H4O62->C4H4O42->H2PO4->C2H3O2->Cl- Similarly, the phase-separation abilities of ionic liquids were obtained by calculating the EEV of six species of phase-separation salts in aqueous 1-ethyl-3-methylimidazolium tetrafluoroborate ([C2mim]BF4), 1-propyl-3-methylimi-dazolium tetrafluoroborate ([C3mim]BF4) or [C4mim]BF4, and drawing binodal curves in molar mass concentration. The phase-separation abilities of ionic liquids are in the order [C4mim]BF4> [C3mim]BF4> [C2mim]BF4.
(3) The effect of temperature on ILATPSs was investigated. When the temperature decreased, the binodal curves shifted to the left, then the area of two-phase region and the slope of tie-line simultaneously increased, so the ability of forming ILATPSs was increased gradually.
2. Separation and concentration of antibiotic residues using ILATPS
(1) An ILATPS consisting of [C4mim]BF4 and Na2CO3, which was a sample pretreatment technique coupled with molecular fluorescence spectrophotomery, was developed for simultaneous separation and enrichment of roxithromycin. The extraction yield of roxithromycin in [C4mim]BF4-Na2CO3 ILATPS was influenced by the types of salts, the concentrations of Na2CO3 and [C4mim]BF4, as well as the extracting temperature. Under the optimum conditions, this method was practical when applied to the analysis of roxithromycin in real water samples with the detection limit of 0.03μ.g/mL, relative standard deviation (RSD) of 1.9%, and the average extraction efficiency was up to 90.7%. The mechanism of ILATPS formation was discussed by hydration theory, and the extraction mechanism of the [C4mim]BF4-salt ILATPS was investigated by FT-IR spectroscopy and UV-vis spectroscopy. The results demonstrated that no chemical (bonding) interactions were observed between ionic liquid and roxithromycin, while the nature properties of the roxithromycin are not altered, the ionic liquid just as the role of the solvent.
(2) [C4mim]BF4-(NH4)2SO4 ILATPS coupled with high-performance liquid chromatography with ultraviolet detection (HPLC-UV) was developed for the extraction of sulfonamides. Three sulfonamides, sulfamerazine, sulfamethoxazole and sulfamethizole were selected as model compounds for developing and evaluating the method. The effects of various experimental parameters in extraction step were studied using two optimisation methods, one variable at a time and the response surface methodology (RSM) approach. Under the optimum conditions, this method was practical when applied to the analysis of sulfonamides in the environment and food. The detection limits (S/N=3) and quantification limits (S/N=10) of the proposed method for the target compounds were achieved within the range of 0.15-0.3 ng/mL and 0.5-1.0 ng/mL from spiked samples, respectively, and the average recovery rate was in the range of 83.7～116.5%, which was lower than or comparable with other reported approaches applied to the determination of the same compounds.
(3) A sample pretreatment procedure coupled with HPLC was developed for the analysis of chloramphenicol that exploits an aqueous two-phase system based on [C4mim]BF4 and organic salt using a liquid-liquid extraction technique. The influence factors on partition behaviors of chloramphenicol were studied, including the type and amount of salts, the pH value, the volume of [C4mim]BF4, and the extraction temperature. Thermodynamic studies indicated that hydrophobic interactions were the main driving force, and salting-out effects was also important for the transfer of the chloramphenicol. Under the optimal conditions, this method was practical when applied to the analysis of chloramphenicol in feed water, milk, and honey samples with the limit of detection of 0.3 ng/mL and the limit of quantification of 1.0 ng/mL. The recovery of CAP was 90.4-102.7%. This novel process is much simpler and more environmentally friendly and is suggested to have important applications for the separation of antibiotics.
3. Separation and enrichment of chloramphenicol using ILATPF and mechanism research
(1) ILATPF which combined ILATPS based on [C4mim]Cl and K2HPO4 with solvent sublation was developed for the analysis of chloramphenicol coupled with HPLC. In ILATPF systems, phase behaviors of the ILATPF were studied for different types of ionic liquids and salts. The sublation efficiency of chloramphenicol in [C4mim]C1-K2HPCO4 ILATPF was influenced by the types of salts, the concentration of K2HPO4 in aqueous solution, solution pH, nitrogen flow rate, sublation time and the amount of [C4mim]Cl. Under the optimum conditions, this method was practical when applied to the analysis of chloramphenicol in lake water, feed water, milk, and honey samples with the linear range of 0.5～500 ng/mL. The method yielded limit of detection of 0.1 ng/mL and limit of quantification of 0.3 ng/mL. The recovery of CAP was 97.1-101.9% from aqueous samples of environmental and food samples by the proposed method. Compared with liquid-liquid extraction, solvent sublation and ionic liquid aqueous two-phase extraction, ILATPF can not only separate and concentrate chloramphenicol with high sublation efficiency, but also efficiently reduce the wastage of IL. This novel technique is much simpler and more environmentally friendly and is suggested to have important applications for the concentration and separation of other small biomolecules.
(2) An IL-K2HPO4 ILATPF was established to separate chloramphenicol. The response surface experimental design method of four factors and three levels was used to optimize the impact factors of flotation of chloramphenicol, and pick up the best conditions:the ionic liquid was [C6mim]Cl, the concentration of K2HPO4 was 0.74 g/mL, the flotation time was 50 min, and the flow rate was 50 mL/min. Under the best conditions, the partition coefficient of chloramphenicol was 405.71, and the flotation rate was 93.16%. According to the three-dimensional response surface chart, we concluded that the order of influencing factors on the partition coefficient and the flotation rate of chloramphenicol was the time of flotation> flow rate of flotation> the type of IL> the concentration of K2HPO4. The mechanism of ILATPF contained two principal processes. One was the mechanism of IL-salt ILATPS formation, the other was solvent sublation.
本课题深入研究了咪唑型离子液体-盐双水相体系的液液相平衡行为及其分相能力,为其应用提供了充分的理论依据。在此基础上构建了离子液体双水相萃取和离子液体双水相气浮溶剂浮选体系,将其应用于环境和食物中抗生素残留的分离/富集中。通过优化抗生素萃取、浮选的影响因素,探讨萃取、浮选机理,为离子液体双水相萃取和离子液体双水相气浮溶剂浮选其它生物小分子物质提供可行性研究的可靠依据。主要研究结果如下：
1.咪唑型离子液体-盐双水相体系的构建及液液相平衡行为的研究
(1)通过对一系列咪唑型离子液体-盐双水相体系双节线数据的拟合,筛选得到了适用于咪唑型离子液体-盐双水相体系双节线数据拟合的最佳方程w1=exp(α+bw20.5+Cw2+dw22)。利用滴定法、密度计测定法以及“杠杆规则”计算法得到了离子液体-盐双水相体系的液液相平衡组成,并用Othmer-Tobias方程和Bancroft方程进行关联,结果令人满意。
(2)通过计算不同分相盐在1-丁基-3-甲基咪唑四氟硼酸([C4mim]BF4)水溶液中的有效排除体积,以及绘制以摩尔质量浓度为单位的双节线相图,得出阳离子的盐析能力为Na+>NH4+；阴离子的盐析能力为Po43->C6H5O73->CO32->SO42-≈C4H4O62->C4H4O42->H2PO4->C2H3O2->CI-。同理,根据计算6种分相盐在1-乙基-3-甲基咪哗四氟硼酸([C2mim]BF4)、1-丙基-3-甲基咪唑四氟硼酸([C3mim]BF4)和[C4mim]BF4中的有效排除体积,以及绘制双节线相图,得到离子液体的分相能力顺序为[C4mim]BF4>[C3mim]BF4>[C2mim]BF4。
(3)研究了温度对离子液体-盐双水相体系分相能力的影响。结果表明,随着温度的降低,双节线向左移动,两相区面积扩大,系线斜率增大,离子液体-盐双水相体系的成相能力逐步增强。
2.离子液体-盐双水相体系分离/富集抗生素残留
(1)构建[C4mim]BF4-Na2CO3双水相体系结合分子荧光光谱分离/富集罗红霉素残留。讨论盐的种类、Na2CO3和离子液体用量以及萃取温度的影响。在最佳条件下,该方法能满意地应用于分离/富集环境水样中的罗红霉素残留,检出限为0.03μg/mL,精密度为1.90%,平均萃取率大于90%。离子液体-盐双水相体系的成相机理用水合理论来解释。红外和紫外光谱表征共同证明在萃取过程中离子液体和罗红霉素之间没有发生化学键的变化,离子液体只是起到溶剂的作用。
(2)建立[C4mim]BF4-(NH4)2SO4双水相体系结合高效液相色谱-紫外法萃取磺胺类抗生素残留。实验选取了三种磺胺类抗生素(磺胺甲基嘧啶、磺胺甲恶唑和磺胺甲塞二唑)作为目标物研究和评估实验方法。选用单因素和响应曲面实验设计优化最佳萃取条件。在最佳条件下,将建立的方法用于测定一系列环境和食物样品,加标回收率在83.7～116.5%,方法的检出限为0.15～0.3 ng/mL,检量限为0.5～1.0 ng/mL,低于其它一些预处理技术。
(3)构建[C4mim]BF4-有机盐双水相体系结合高效液相色谱法萃取/分析氯霉素残留。探讨影响氯霉素分配行为的影响因素：有机盐的种类和含量、体系的pH值、[C4mim]BF4的加入体积和萃取温度。热力学研究证明疏水作用是氯霉素萃取的主要驱动力,同时盐析作用也十分关键。在最佳条件下,将该方法成功地用于分离/富集饲养水、牛奶和蜂蜜中的氯霉素残留,加标回收率在90.4～102.7%,方法检出限为0.3 ng/mL,检量限为1.0 ng/mL。此种离子液体-有机盐双水相萃取技术操作简单,并且对环境无污染,在抗生素分离中有着重要的应用价值。
3.离子液体双水相气浮溶剂浮选分离/富集氯霉素及机理研究
(1)建立氯化1-丁基-3-甲基咪唑四氟硼酸([C4mim]Cl)-K2HPO4离子液体双水相气浮溶剂浮选体系结合高效液相色谱法浮选/分离氯霉素残留。讨论了离子液体和盐的种类对离子液体双水相气浮溶剂浮选体系成相行为的影响,详细研究盐种类、K2HPO4浓度、体系pH值、氮气流速、浮选时间和[C4mim]Cl初始加入量对氯霉素浮选效率的影响。在最佳条件下,浮选湖水、饲养水、牛奶和蜂蜜中氯霉素残留,浮选率为97.1～101.9%。该方法的线性范围为0.5～500 ng/mL,方法检出限为0.1 ng/mL,检量限为0.3 ng/mL。该方法优于液液萃取、溶剂浮选以及离子液体双水相萃取,其富集倍数高、减少离子液体用量、对环境友好的优点,在小分子生物物质中的分离/富集具有重要的应用价值。
(2)以氯霉素为目标物,构建离子液体-K2HPO4双水相气浮溶剂浮选体系。利用四因素三水平响应曲面实验设计法优化氯霉素的影响因素,筛选出的最佳条件为：离子液体选择[C6mim]Cl, K2HPO4浓度为0.74 g/mL、浮选时间为50 min、浮选流速为50 mL/min,此时氯霉素的分配系数为405.71,浮选率为93.16%。三维响应曲面图可以得到影响氯霉素分配系数和浮选率的影响因素强度依次为浮选时间>浮选流速>IL的种类> K2HPO4的浓度。离子液体双水相气浮溶剂浮选机理是由离子液体双水相体系的成相机理和溶剂浮选机理两部分构成。
The abuse of antibiotics in clinical medicine, aquaculture and food processing industry has been posed a serious threat to the ecological environment and human health. Therefore, the establishment of the method for separation and enrichment of antibiotic residues in the environment and food becomes the focal point of attention. Due to the composition of antibiotics matrix is complicated and usually trace or ultra-trace exists, we must find a reasonable method for separation and enrichment of antibiotics in the environment and food. It will achieve the separation of interfering substances, lower detection limits and improve the measurement accuracy of the purpose.
This thesis deeply researches into the behavior of liquid-liquid equilibrium (LLE) of iminazole-ionic liquid aqueous two-phase system (ILATPS) and its phase-separation abilities, so this provides sufficient theoretical foundation for its application. On the basis of which, an ionic liquid aqueous two-phase extraction (ILATPE) system and an ionic liquid aqueous two-phase flotation (ILATPF) system were established, they can be applied to separate and concentrate antibiotic residues in the environment and food. Various factors influencing the extraction and flotation of antibiotics were optimized; the mechanisms of extraction and flotation were discussed. These studies provide a feasibility evidence on separation and concentration of other biological small molecules by ILATPE and ILATPF. The main results are as follows:
1. Construction of ILATPS and the research of LLE
(1) Through the fitting of a series of iminazole ILATPS of binodal data, the optimum equation was selected, and it was w,= exp(a+bw20.5+cw2+dw22). Based on the titration method, the densimeter method and "lever rule" calculation, the LLE compositions were obtained. The equations of Othmer-Tobias and Bancroft were used to fitting the LLE data, and the result was satisfactory.
(2) The salting-out abilities of salt ions were obtained by calculating the effective excluded volume (EEV) of different species of phase-separation salts in aqueous l-butyl-3-methylimidazolium tetrafluoroborate ([C4mimJBF4) solutions, and drawing binodal curves in molar mass concentration. The phase-separation abilities of cation are in the order Na+>NH4+, and the phase-separation abilities of anion are PO43->C6H5O73->CO32->SO42-≈C4H4O62->C4H4O42->H2PO4->C2H3O2->Cl- Similarly, the phase-separation abilities of ionic liquids were obtained by calculating the EEV of six species of phase-separation salts in aqueous 1-ethyl-3-methylimidazolium tetrafluoroborate ([C2mim]BF4), 1-propyl-3-methylimi-dazolium tetrafluoroborate ([C3mim]BF4) or [C4mim]BF4, and drawing binodal curves in molar mass concentration. The phase-separation abilities of ionic liquids are in the order [C4mim]BF4> [C3mim]BF4> [C2mim]BF4.
(3) The effect of temperature on ILATPSs was investigated. When the temperature decreased, the binodal curves shifted to the left, then the area of two-phase region and the slope of tie-line simultaneously increased, so the ability of forming ILATPSs was increased gradually.
2. Separation and concentration of antibiotic residues using ILATPS
(1) An ILATPS consisting of [C4mim]BF4 and Na2CO3, which was a sample pretreatment technique coupled with molecular fluorescence spectrophotomery, was developed for simultaneous separation and enrichment of roxithromycin. The extraction yield of roxithromycin in [C4mim]BF4-Na2CO3 ILATPS was influenced by the types of salts, the concentrations of Na2CO3 and [C4mim]BF4, as well as the extracting temperature. Under the optimum conditions, this method was practical when applied to the analysis of roxithromycin in real water samples with the detection limit of 0.03μ.g/mL, relative standard deviation (RSD) of 1.9%, and the average extraction efficiency was up to 90.7%. The mechanism of ILATPS formation was discussed by hydration theory, and the extraction mechanism of the [C4mim]BF4-salt ILATPS was investigated by FT-IR spectroscopy and UV-vis spectroscopy. The results demonstrated that no chemical (bonding) interactions were observed between ionic liquid and roxithromycin, while the nature properties of the roxithromycin are not altered, the ionic liquid just as the role of the solvent.
(2) [C4mim]BF4-(NH4)2SO4 ILATPS coupled with high-performance liquid chromatography with ultraviolet detection (HPLC-UV) was developed for the extraction of sulfonamides. Three sulfonamides, sulfamerazine, sulfamethoxazole and sulfamethizole were selected as model compounds for developing and evaluating the method. The effects of various experimental parameters in extraction step were studied using two optimisation methods, one variable at a time and the response surface methodology (RSM) approach. Under the optimum conditions, this method was practical when applied to the analysis of sulfonamides in the environment and food. The detection limits (S/N=3) and quantification limits (S/N=10) of the proposed method for the target compounds were achieved within the range of 0.15-0.3 ng/mL and 0.5-1.0 ng/mL from spiked samples, respectively, and the average recovery rate was in the range of 83.7～116.5%, which was lower than or comparable with other reported approaches applied to the determination of the same compounds.
(3) A sample pretreatment procedure coupled with HPLC was developed for the analysis of chloramphenicol that exploits an aqueous two-phase system based on [C4mim]BF4 and organic salt using a liquid-liquid extraction technique. The influence factors on partition behaviors of chloramphenicol were studied, including the type and amount of salts, the pH value, the volume of [C4mim]BF4, and the extraction temperature. Thermodynamic studies indicated that hydrophobic interactions were the main driving force, and salting-out effects was also important for the transfer of the chloramphenicol. Under the optimal conditions, this method was practical when applied to the analysis of chloramphenicol in feed water, milk, and honey samples with the limit of detection of 0.3 ng/mL and the limit of quantification of 1.0 ng/mL. The recovery of CAP was 90.4-102.7%. This novel process is much simpler and more environmentally friendly and is suggested to have important applications for the separation of antibiotics.
3. Separation and enrichment of chloramphenicol using ILATPF and mechanism research
(1) ILATPF which combined ILATPS based on [C4mim]Cl and K2HPO4 with solvent sublation was developed for the analysis of chloramphenicol coupled with HPLC. In ILATPF systems, phase behaviors of the ILATPF were studied for different types of ionic liquids and salts. The sublation efficiency of chloramphenicol in [C4mim]C1-K2HPCO4 ILATPF was influenced by the types of salts, the concentration of K2HPO4 in aqueous solution, solution pH, nitrogen flow rate, sublation time and the amount of [C4mim]Cl. Under the optimum conditions, this method was practical when applied to the analysis of chloramphenicol in lake water, feed water, milk, and honey samples with the linear range of 0.5～500 ng/mL. The method yielded limit of detection of 0.1 ng/mL and limit of quantification of 0.3 ng/mL. The recovery of CAP was 97.1-101.9% from aqueous samples of environmental and food samples by the proposed method. Compared with liquid-liquid extraction, solvent sublation and ionic liquid aqueous two-phase extraction, ILATPF can not only separate and concentrate chloramphenicol with high sublation efficiency, but also efficiently reduce the wastage of IL. This novel technique is much simpler and more environmentally friendly and is suggested to have important applications for the concentration and separation of other small biomolecules.
(2) An IL-K2HPO4 ILATPF was established to separate chloramphenicol. The response surface experimental design method of four factors and three levels was used to optimize the impact factors of flotation of chloramphenicol, and pick up the best conditions:the ionic liquid was [C6mim]Cl, the concentration of K2HPO4 was 0.74 g/mL, the flotation time was 50 min, and the flow rate was 50 mL/min. Under the best conditions, the partition coefficient of chloramphenicol was 405.71, and the flotation rate was 93.16%. According to the three-dimensional response surface chart, we concluded that the order of influencing factors on the partition coefficient and the flotation rate of chloramphenicol was the time of flotation> flow rate of flotation> the type of IL> the concentration of K2HPO4. The mechanism of ILATPF contained two principal processes. One was the mechanism of IL-salt ILATPS formation, the other was solvent sublation.
引文
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