人工神经网络—顺序注射光度分析多组分同时测定技术在环境和药物分析中的应用研究
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摘要
环境样品中的重金属污染物和农药残留物以及药品中的药物成分的检测具有重要的意义。由于环境样品和复方制剂的基体复杂,测定时干扰严重,化学计量学方法是解决此类问题改善分析选择性,提高测定结果准确度的有效方法之一。本文通过构建人工神经网络多元校正模型,对顺序注射分析与分光光度法联用技术在环境和药物分析中的应用进行了系统研究,实现了复杂样品中多种待测组分的同时测定。
化学计量学利用数学和统计学的方法,通过解析测量数据,最大限度地获取关于物质的化学信息,以“数学方法分辨”替代复杂的物理或化学分离,在改善光度分析法选择性的同时显著提高了分析结果的准确度。采用人工神经网络这一化学计量学方法建立的非线性多元校正模型,可以准确地模拟输入信息和响应信号之间的非线性行为,有效地消除或减少待测组分间的交互作用、协同效应和其它未知非线性因素对实验测定结果的影响。然而,在常规的手工操作中存在分析过程费时费力和样品易被污染等弊病。顺序注射分析作为流动注射分析技术的一个分支,有效地克服了上述分析方法的缺陷,它不仅可以精确地控制试剂和试样的体积以及混合时间,极大地降低了试样特别是试剂的消耗,同时显著提高了分析过程的自动化程度和改善了方法的分析性能,可以通过单次测定提供多维分析信息,因而成为流动分析研究中最为活跃的领域之一。本论文的研究目的是将顺序注射光度分析方法与人工神经网络多元校正模型相结合,通过对复杂环境样品和复方制剂的测定,建立一个简便、快速、可靠的多组分流动分析系统。
第一章简述了流动注射分析技术的历史发展过程,对近年来应用于流动注射分析的同时测定技术进行了评述,包括固相萃取法、顺序注射色谱法、多通道管路法、动力学方法、选择性检测器和化学计量学法。
第二章研究发现在碱性介质中,Cu2+、Zn2+、Ni2+、Mn2+与4-(2-吡啶偶氮)-间苯二酚络合产物的吸收光谱严重重叠且体系线性加和性较差,据此将人工神经网络多元校正方法用于这四种金属络合物重叠光谱的分辨解析,提出了停流-顺序注射光度分析技术同时测定四元混合物,即环境水中Cu2+、Zn2+、Ni2+、Mn2+含量的新方法。方法的测定范围为0.1～2.0mg/L,分析速度达到30样/h。对环境标准参考物和实际样品的测定结果与标准参考值相符,与标准测定方法-原子吸收光谱法无显著性差异。
在对实验条件进行优化的同时提出了将逼近度作为优选网络参数的评价标准,综合考虑了校正集和预测集的影响,既保证了网络学习的稳定性,又使校正模型具有较好的预测性。
第三章研究了碱性条件下四种氨基甲酸酯类农药克百威、残杀威、速灭威、仲丁威水解产物与重氮化对硝基苯胺的偶联反应,探讨了该显色体系的反应机理。基于该反应体系在流动分析条件下待测组分随时间变化的浓度梯度信息的差异,提出了顺序注射动力学光度分析联用技术同时测定四种农药含量的新方法。方法的测定范围为0.5～10.0μg/mL,克百威、残杀威、速灭威、仲丁威的检出限分别为0.4,0.3,0.2,0.2μg/mL,分析速度为18样/h。优化了实验条件和网络参数,证实了动力学光谱数据的可靠性(R.S.D.<3.0%,n=7)。
与文献报道的其它同时测定氨基甲酸酯类农药的光度分析方法相比,本法简便、快速、准确度高,用于地表水和水果中克百威、残杀威、速灭威和仲丁威残留量的测定,回收率在84.7～116.0%之间。
第四章实验观察到不同pH下维生素C和扑热息痛的吸收光谱存在明显差异,建立了顺序注射pH梯度技术分光光度法不经分离同时测定维生素C和扑热息痛的新方法。讨论了顺序注射分析系统中pH梯度的建立条件,考察了NaOH溶液浓度及其体积、检测流速等实验参数的影响。在选定的实验条件下,维生素C和扑热息痛的测定范围为1.0～10.0 mg/L,检出限分别为0.3和0.2 mg/L,分析速度为40样/h。将该方法用于维C银翘片中维生素C和扑热息痛含量的同时测定,分析结果与药品标示量相符,加标回收率在92.0～109.1%之间。
本文建立的顺序注射分析方法具有操作简单,分析快速,准确度高,试剂和样品的消耗量少和设备价廉等优点,实现了复杂样品中多种待测组分含量的同时测定,进一步拓宽了顺序注射光度分析法与人工神经网络联用技术在环境和药物分析中的应用。为研究热力学性质,动力学性质或pH效应存在差异而难以实现同时测定的复杂非线性体系提供了新的思路。
The development of new analytical procedures for heavy metal pollutants, pesticide residues and pharmaceutical components in complex environmental and pharmaceutical matrices are highly demanded. Spectrophotometry is one of the most favorable techniques, due to its simplicity, economy, high accuracy and precision for both organic and inorganic components analysis. However, matrix effects and the interaction between analytes are very often problematic for real sample analysis, including environmental samples and Pharmaceuticals. The separation of analytes with the aid of chemometric methods is an ideal approach for eliminating matrix effects and resolving analytical responses of mixtures by mathematical treatment. At this point, sequential injection spectrophotometry coupled with artificial neural networks (ANN) was used to determine multi-components in a single run.
A combination of spectrophotometry and appropriate chemometric methods was widely applied in the multicomponent analysis, due to simple operation, low cost instrument, reasonable accuracy and precision and simultaneous determination of multiple species without carrying out any complex pretreatments. Among these approaches, an excellent alternative is the use of ANN algorithms by their high efficiency as predictors for non-linear systems. After proper training, ANN can accurately model the non-linear behavior between input variables and responses and eliminate or reduce the effects of the interaction between analytes, the synergistic effect (non-additivity of reaction rates), the multi-step process and any other unknown non-linearity. However, manual procedures are tedious and time-consuming, and are sometimes complicated. Such disadvantages in manual procedures can be easily overcome by flow injection analysis (FIA), since FIA technique can precisely and reproducibly control the timing of the measurement of the chemical reaction and the mixing of the solutions of reagent and sample with rapid and low consumption of sample throughput. It is also highly versatile, in terms of the types of data it can provide. Sequential injection analysis (SIA), as a branch of sampling means of FIA, the advantages of SIA over FIA are the simpler flow manifold, the lower consumption of reagents, the easier and more convenient PC-control of the instrumental parameters and the greater potential for fluidic handling. All these superiorities have made SIA technique coupled with spectrophotometry and ANN calibration model a powerful analytical tool in multicomponent analysis. In this thesis, three simple, rapid and reliable procedures for the simultaneous determination of multiple species in complex environmental and pharmaceutical matrices were proposed.
Chapter 1 of the thesis briefly reviewed the development of FIA method, and the strategies used in flow systems to achieve multicomponent analysis include solid phase extractions, sequential injection chromatography, multichannel manifolds, kinetics, selective detectors and chemometrics.
In Chapter 2 of the thesis, a stopped-flow sequential injection spectrophotometry for the simultaneous determination of Cu(Ⅱ), Zn(Ⅱ), Ni(Ⅱ) and Mn(Ⅱ) using an ANN algorithm for multivariate calibration was developed. The procedure was based on the reaction of these metal ions with 4-(2-pyridilazo) resorcinol in an alkaline medium to form colored complexes. The degree of approximation, a new evaluation criterion of the networks parameters was employed to prove the accuracy of the predicted results. The experimental variables were studied and determination ranges were obtained in the concentration range of 0.1-2.0 mg L'1 for each analyte, at a sampling frequency of 30 h-1, respectively. The proposed method was validated and the determination results for environmental water samples were statistically similar to those from the atomic absorption spectrometry. The results obtained for analysis of reference samples of environmental water were in good agreement with the certified values.
In Chapter 3 of the thesis, a new procedure was developed for the simultaneous kinetic spectrophotometric determination of a quaternary carbamate pesticide mixture consisting of carbofuran, propoxur, metolcarb and fenobucarb using SIA. The procedure was based upon the different kinetic properties between the analytes reacted with reagent in flow system in the non-stopped-flow mode, in which their hydrolysis products coupled with diazotized p-nitroaniline in an alkaline medium to form the corresponding colored complexes. The absorbance data from SIA peak time profile were recorded at 510 nm and resolved by the use of ANN for multivariate quantitative analysis. The experimental variables and main networks parameters were optimized and each of the pesticides could be determined in the concentration range of 0.5-10.0μg mL-1, at a sampling frequency of 18 h-1. The proposed method was compared to other spectrophotometric methods for simultaneous determination of mixtures of carbamate pesticides, and it was proved to be adequately reliable and was successfully applied to the simultaneous determination of the four pesticide residues in water and fruit samples, obtaining the satisfactory results based on recovery studies (84.7-116.0%).
In Chapter 4 of the thesis, a sequential injection pH gradient spectrophotometry was proposed for the simultaneous determination of vitamin C and paracetamol in pharmaceuticals. The method was based on the significant differences between absorption spectra of vitamin C and paracetamol at different pH, and a pH gradient in the range of 9.2-10.9 was produced by the dispersion of NaOH solution in a flow system using SI technique. ANN algorithms were applied to resolving overlapping spectra of the two pharmaceutical components and to the simultaneous determination of their concentrations. Some experimental variables including NaOH concentration and its volume, sample volume, flow rate were studied, respectively. Under the selected experimental conditions, the determination ranges were obtained in the concentration range of 1.0-10.0 mg L-1 for vitamin C and paracetamol, with the limits of detection of 0.3 and 0.2 mg L-1 at a sampling frequency of 40 h-1, respectively. The proposed method was applied to the simultaneous determination of vitamin C and paracetamol in vitamin C yinqiao tables, and the determination results were not statistically different to the claimed values and the recoveries for all components were in the range of 92.0-109.1%.
In conclusion, this thesis demonstrated a multicomponent analytical method by sequential injection spectrophotometry with artificial neural networks multivariate calibration model. These procedures are simple, rapid, accurate and low reagents and samples consumption, and they were successfully applied for the determination several analytes in environmental and pharmaceutical samples in a single run. Thus, it infers that the proposed method provides a very interesting way for the simultaneous quantification of multiple species of the different thermodynamic, kinetic or pH properties in non-linear systems.
化学计量学利用数学和统计学的方法,通过解析测量数据,最大限度地获取关于物质的化学信息,以“数学方法分辨”替代复杂的物理或化学分离,在改善光度分析法选择性的同时显著提高了分析结果的准确度。采用人工神经网络这一化学计量学方法建立的非线性多元校正模型,可以准确地模拟输入信息和响应信号之间的非线性行为,有效地消除或减少待测组分间的交互作用、协同效应和其它未知非线性因素对实验测定结果的影响。然而,在常规的手工操作中存在分析过程费时费力和样品易被污染等弊病。顺序注射分析作为流动注射分析技术的一个分支,有效地克服了上述分析方法的缺陷,它不仅可以精确地控制试剂和试样的体积以及混合时间,极大地降低了试样特别是试剂的消耗,同时显著提高了分析过程的自动化程度和改善了方法的分析性能,可以通过单次测定提供多维分析信息,因而成为流动分析研究中最为活跃的领域之一。本论文的研究目的是将顺序注射光度分析方法与人工神经网络多元校正模型相结合,通过对复杂环境样品和复方制剂的测定,建立一个简便、快速、可靠的多组分流动分析系统。
第一章简述了流动注射分析技术的历史发展过程,对近年来应用于流动注射分析的同时测定技术进行了评述,包括固相萃取法、顺序注射色谱法、多通道管路法、动力学方法、选择性检测器和化学计量学法。
第二章研究发现在碱性介质中,Cu2+、Zn2+、Ni2+、Mn2+与4-(2-吡啶偶氮)-间苯二酚络合产物的吸收光谱严重重叠且体系线性加和性较差,据此将人工神经网络多元校正方法用于这四种金属络合物重叠光谱的分辨解析,提出了停流-顺序注射光度分析技术同时测定四元混合物,即环境水中Cu2+、Zn2+、Ni2+、Mn2+含量的新方法。方法的测定范围为0.1～2.0mg/L,分析速度达到30样/h。对环境标准参考物和实际样品的测定结果与标准参考值相符,与标准测定方法-原子吸收光谱法无显著性差异。
在对实验条件进行优化的同时提出了将逼近度作为优选网络参数的评价标准,综合考虑了校正集和预测集的影响,既保证了网络学习的稳定性,又使校正模型具有较好的预测性。
第三章研究了碱性条件下四种氨基甲酸酯类农药克百威、残杀威、速灭威、仲丁威水解产物与重氮化对硝基苯胺的偶联反应,探讨了该显色体系的反应机理。基于该反应体系在流动分析条件下待测组分随时间变化的浓度梯度信息的差异,提出了顺序注射动力学光度分析联用技术同时测定四种农药含量的新方法。方法的测定范围为0.5～10.0μg/mL,克百威、残杀威、速灭威、仲丁威的检出限分别为0.4,0.3,0.2,0.2μg/mL,分析速度为18样/h。优化了实验条件和网络参数,证实了动力学光谱数据的可靠性(R.S.D.<3.0%,n=7)。
与文献报道的其它同时测定氨基甲酸酯类农药的光度分析方法相比,本法简便、快速、准确度高,用于地表水和水果中克百威、残杀威、速灭威和仲丁威残留量的测定,回收率在84.7～116.0%之间。
第四章实验观察到不同pH下维生素C和扑热息痛的吸收光谱存在明显差异,建立了顺序注射pH梯度技术分光光度法不经分离同时测定维生素C和扑热息痛的新方法。讨论了顺序注射分析系统中pH梯度的建立条件,考察了NaOH溶液浓度及其体积、检测流速等实验参数的影响。在选定的实验条件下,维生素C和扑热息痛的测定范围为1.0～10.0 mg/L,检出限分别为0.3和0.2 mg/L,分析速度为40样/h。将该方法用于维C银翘片中维生素C和扑热息痛含量的同时测定,分析结果与药品标示量相符,加标回收率在92.0～109.1%之间。
本文建立的顺序注射分析方法具有操作简单,分析快速,准确度高,试剂和样品的消耗量少和设备价廉等优点,实现了复杂样品中多种待测组分含量的同时测定,进一步拓宽了顺序注射光度分析法与人工神经网络联用技术在环境和药物分析中的应用。为研究热力学性质,动力学性质或pH效应存在差异而难以实现同时测定的复杂非线性体系提供了新的思路。
The development of new analytical procedures for heavy metal pollutants, pesticide residues and pharmaceutical components in complex environmental and pharmaceutical matrices are highly demanded. Spectrophotometry is one of the most favorable techniques, due to its simplicity, economy, high accuracy and precision for both organic and inorganic components analysis. However, matrix effects and the interaction between analytes are very often problematic for real sample analysis, including environmental samples and Pharmaceuticals. The separation of analytes with the aid of chemometric methods is an ideal approach for eliminating matrix effects and resolving analytical responses of mixtures by mathematical treatment. At this point, sequential injection spectrophotometry coupled with artificial neural networks (ANN) was used to determine multi-components in a single run.
A combination of spectrophotometry and appropriate chemometric methods was widely applied in the multicomponent analysis, due to simple operation, low cost instrument, reasonable accuracy and precision and simultaneous determination of multiple species without carrying out any complex pretreatments. Among these approaches, an excellent alternative is the use of ANN algorithms by their high efficiency as predictors for non-linear systems. After proper training, ANN can accurately model the non-linear behavior between input variables and responses and eliminate or reduce the effects of the interaction between analytes, the synergistic effect (non-additivity of reaction rates), the multi-step process and any other unknown non-linearity. However, manual procedures are tedious and time-consuming, and are sometimes complicated. Such disadvantages in manual procedures can be easily overcome by flow injection analysis (FIA), since FIA technique can precisely and reproducibly control the timing of the measurement of the chemical reaction and the mixing of the solutions of reagent and sample with rapid and low consumption of sample throughput. It is also highly versatile, in terms of the types of data it can provide. Sequential injection analysis (SIA), as a branch of sampling means of FIA, the advantages of SIA over FIA are the simpler flow manifold, the lower consumption of reagents, the easier and more convenient PC-control of the instrumental parameters and the greater potential for fluidic handling. All these superiorities have made SIA technique coupled with spectrophotometry and ANN calibration model a powerful analytical tool in multicomponent analysis. In this thesis, three simple, rapid and reliable procedures for the simultaneous determination of multiple species in complex environmental and pharmaceutical matrices were proposed.
Chapter 1 of the thesis briefly reviewed the development of FIA method, and the strategies used in flow systems to achieve multicomponent analysis include solid phase extractions, sequential injection chromatography, multichannel manifolds, kinetics, selective detectors and chemometrics.
In Chapter 2 of the thesis, a stopped-flow sequential injection spectrophotometry for the simultaneous determination of Cu(Ⅱ), Zn(Ⅱ), Ni(Ⅱ) and Mn(Ⅱ) using an ANN algorithm for multivariate calibration was developed. The procedure was based on the reaction of these metal ions with 4-(2-pyridilazo) resorcinol in an alkaline medium to form colored complexes. The degree of approximation, a new evaluation criterion of the networks parameters was employed to prove the accuracy of the predicted results. The experimental variables were studied and determination ranges were obtained in the concentration range of 0.1-2.0 mg L'1 for each analyte, at a sampling frequency of 30 h-1, respectively. The proposed method was validated and the determination results for environmental water samples were statistically similar to those from the atomic absorption spectrometry. The results obtained for analysis of reference samples of environmental water were in good agreement with the certified values.
In Chapter 3 of the thesis, a new procedure was developed for the simultaneous kinetic spectrophotometric determination of a quaternary carbamate pesticide mixture consisting of carbofuran, propoxur, metolcarb and fenobucarb using SIA. The procedure was based upon the different kinetic properties between the analytes reacted with reagent in flow system in the non-stopped-flow mode, in which their hydrolysis products coupled with diazotized p-nitroaniline in an alkaline medium to form the corresponding colored complexes. The absorbance data from SIA peak time profile were recorded at 510 nm and resolved by the use of ANN for multivariate quantitative analysis. The experimental variables and main networks parameters were optimized and each of the pesticides could be determined in the concentration range of 0.5-10.0μg mL-1, at a sampling frequency of 18 h-1. The proposed method was compared to other spectrophotometric methods for simultaneous determination of mixtures of carbamate pesticides, and it was proved to be adequately reliable and was successfully applied to the simultaneous determination of the four pesticide residues in water and fruit samples, obtaining the satisfactory results based on recovery studies (84.7-116.0%).
In Chapter 4 of the thesis, a sequential injection pH gradient spectrophotometry was proposed for the simultaneous determination of vitamin C and paracetamol in pharmaceuticals. The method was based on the significant differences between absorption spectra of vitamin C and paracetamol at different pH, and a pH gradient in the range of 9.2-10.9 was produced by the dispersion of NaOH solution in a flow system using SI technique. ANN algorithms were applied to resolving overlapping spectra of the two pharmaceutical components and to the simultaneous determination of their concentrations. Some experimental variables including NaOH concentration and its volume, sample volume, flow rate were studied, respectively. Under the selected experimental conditions, the determination ranges were obtained in the concentration range of 1.0-10.0 mg L-1 for vitamin C and paracetamol, with the limits of detection of 0.3 and 0.2 mg L-1 at a sampling frequency of 40 h-1, respectively. The proposed method was applied to the simultaneous determination of vitamin C and paracetamol in vitamin C yinqiao tables, and the determination results were not statistically different to the claimed values and the recoveries for all components were in the range of 92.0-109.1%.
In conclusion, this thesis demonstrated a multicomponent analytical method by sequential injection spectrophotometry with artificial neural networks multivariate calibration model. These procedures are simple, rapid, accurate and low reagents and samples consumption, and they were successfully applied for the determination several analytes in environmental and pharmaceutical samples in a single run. Thus, it infers that the proposed method provides a very interesting way for the simultaneous quantification of multiple species of the different thermodynamic, kinetic or pH properties in non-linear systems.
引文
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