【作者】 张艳；

【导师】 吴海龙； 俞汝勤；

【作者基本信息】 湖南大学 ， 分析化学， 2008， 博士

【摘要】 化学计量学是一门新兴发展的化学学科分支。它运用数学、统计学、计算机科学等方法设计优化化学量测过程,并通过解析化学量测数据最大限度地获取化学及相关信息。分析手段的仪器化和化学体系的复杂化已成为现代分析化学的一个重要特征。化学计量学利用现代分析仪器所产生的庞大的量测信号,为复杂多组分化学体系的定性、定量分析提供了便利的研究手段。化学计量学二阶校正方法具有二阶优势,即使在未知干扰物共存下,依然能对复杂体系中感兴趣组分快速给出定量分析结果,可以解决传统的分离分析手段难以解决的问题,提高分析效率。化学计量学方法与现代分析仪器结合,以“数学分离”代替或者增强“化学分离”,显示出其快速、通用、可靠的特点。它将对各化学分支学科,其中特别是生物化学、食品化学、药物化学等学科带来较大的影响,为直接解决这些分支学科中的实际分析难题提供有力武器。本文作者通过利用LC×LC-DAD、HPLC-DAD及三维荧光光谱等现代分析仪器量测手段,结合化学计量学多维数据阵分析的新方法及新算法,在复杂多组分体系的定量分析方面进行了深入的研究。本论文主要涉及以下几个方面:1.高维量测数据中背景扣除新方法的研究(第二章)针对全二维联用色谱如LC×LC-DAD分析中的背景漂移问题,提出了一种扣除背景的新方法。该方法主要是基于三线性分解原理,对仪器响应数据进行交替三线性分解(ATLD)。在建模时,因样品包含背景漂移信息,如同其他样品组分,将背景漂移看作是一个组分或者因子来考察。该方法首先对原始数据进行三线性分解,然后提取背景组分的信息,并将其从原始数据中减掉,使得分析信号处于一平稳的基线上。同时考察背景漂移和分析组分的信息能明显提高数据质量。本章采用此方法分析了模拟数据以及实验数据中的背景漂移,结果表明该方法能有效地扣除LC×LC-DAD数据中的背景漂移,为进一步进行全二维色谱的定量分析奠定了良好的基础。该方法无需空白样品的分析,并且也无需事先知道样品组分的信息。2.复杂多组分体系的定量分析的研究(第三章-第六章)随着人们对食品质量与安全问题的日益关注,食品分析对检测方法的精确度、时效性要求越来越高,人们在不断地探索研究新技术并应用于食品检测。苏丹红并非食品添加剂,而是一种人工合成的红色染料,常作为一种工业染料,被广泛用于如溶剂、油、蜡、汽油的增色以及鞋、地板等增光方面。不幸的是,苏丹红常被作为增色剂添加入食品如辣椒粉中,严重危害人们的健康。本论文的第三章采用HPLC-DAD分析辣椒粉中苏丹红I和苏丹红Ⅱ,结合二阶校正方法对色谱量测数据进行解析。实验采用甲醇和0.5%(v/v)醋酸水溶液作为色谱条件,样品预处理简便快速。结果表明在辣椒粉中有未知干扰基质存在的情况下,采用二阶校正方法能对该混合物中的苏丹红I和苏丹红Ⅱ给出准确的色谱轮廓、光谱轮廓分辨以及满意的浓度预测。磺胺类(SAs)药物在奶牛业中的应用非常普遍。由于SAs在体内作用和代谢的时间比较长,在动物源性食品中容易残留,危及人体健康。论文的第四章采用HPLC-DAD检测与交替惩罚三线性分解(APTLD)算法相结合对牛奶中的磺胺类兽药残留进行了分析测定,尽管样品中组分分离不完全,并且存在大量干扰组分,但利用化学计量学二阶校正方法的“二阶优势”,仍然能对体系中感兴趣的磺胺组分给出准确的定量分析结果。化学计量学二阶校正方法的应用,为食品成分检测提供了新的检测途径。药物在体液中的分析是现代生物医学领域面临的一个重要问题,传统方法采用色谱分离技术来实现这一目的,通常情况下通过调整色谱柱或者色谱条件分离,但是在某些情况下完全分离是难于实现的,且体液中含有不可预测的基质及其他干扰物。糖皮质激素是由肾上腺皮质分泌的一类甾体激素,为维持生命所必需。超生理量的糖皮质激素具有抗炎、抗过敏和抑制免疫等多种药理作用,临床应用非常广泛。由于氢化可的松和波尼松龙两者结构相似,它们的光谱也很相似,色谱出峰时间接近,因此色谱分析很难达到完全分离。传统色谱分析方法大都需要内标,且其方法仅适合于尿液或者血液中氢化可的松或波尼松龙的测定。本论文的第五章采用甲醇和水作为高效液相色谱的分离条件对血浆和尿液中两种糖皮质激素氢化可的松和波尼松龙进行了同时快速测定,然后结合交替三线性分解(ATLD)算法解析色谱数据,结果满意。该方法无需采用内标,样品预处理简单。美托洛尔为第二代β1受体阻滞剂,临床上用于治疗各种类型高血压、冠心病、心律失常、甲亢等。针对荧光检测具有高灵敏度、高选择性以及检测成本低等特点,在第六章中我们利用美托洛尔的荧光特性,采用激发发射矩阵荧光结合三线性分解算法对美托洛尔的血药浓度进行了定量分析,实现了人血浆中美托洛尔的直接测定。该方法快速简便,无需繁琐的样品预处理,花费成本低廉,定量结果满意。为实现体液中美托洛尔的近实时快速分析打下了基础,为开展临床药理学研究及血药浓度监测提供了一种简便可靠的定量方法,对指导临床安全合理用药具有重要的意义。3.三维数据分析在药物动力学研究中的应用(第七章)研究药物与DNA的相互作用不仅对理解作用机制有很大意义,还可以指导新药的设计。然而,我们对药物小分子与DNA的相互作用机制知之甚少。本文第七章利用平行因子分析算法(PARAFAC)与荧光分析相结合,研究了吡柔比星(THP)、溴化乙锭(EB)与DNA的相互作用。采用PARAFAC处理可以很方便地得到激发发射光谱在不同的反应和平衡混合物中的相对浓度。从PARAFAC分析得到的光谱及浓度图表明,THP能竞争EB与DNA的结合位点,抑制EB和DNA的反应。分析结果表明,THP以嵌入方式与DNA发生相互作用,并且生成了不发荧光的络合物。这些结果对于进一步深入了解THP、EB与DNA作用时的竞争机制很有帮助。化学计量学方法的应用,使得研究药物与DNA的相互作用的直观解析成为可能。即使该混合物中存在着很复杂的化学平衡,也可以很方便地预测感兴趣的组分和DNA的相互作用机制。对有争议的药物小分子与DNA的作用机制的研究提供了有力的佐证,这对于进一步开展新型抗癌药剂的开发和抗癌机理研究具有重要的促进作用。更多还原

【Abstract】 Chemometrics is a developing composite discipline of chemistry. The main focus of chemometrics is on applying the methods of mathematics, statistics and computer sciences to extracting the optimal scheme for chemical measurements and to elucidating the data collected from chemical measurements. Two important characteristics of modern analytical chemistry are that instrumentation of analytical tools and complication of chemical system. Chemometrics uses the enormous signals generated by the modern analytical instruments, and provides a variety of convenient techniques for quantitative analysis of complex multi-component systems. Especially, second-order calibration methodologies have the second-order advantage, allowing analyte concentrations to be estimated even in the presence of uncalibrated interferents. The chemometric methodologies coupled with analytical instruments can improve or replace the“chemical separation”with“mathematical separation”, displaying the characteristic of fast and universal, dependable. It gives active impact on these disciplines of chemistry, especially biochemistry and food chemistry and pharmaceutical chemistry, and also provides a variety of powerful techniques to resolve difficult problems of complex chemical systems. In this paper, the author utilized the instruments such as LCxLC-DAD and HPLC-DAD, EEMs coupled with multi-way data analysis methodologies, and did some work in the research of complex multi-component systems.1. The removal of background drift in LCxLC-DAD measurements (Chapter 2)A novel technique for removal of three-dimensional background drift in comprehensive two-dimensional (2D) liquid chromatography coupled with diode array detection (LCxLC-DAD) data was proposed. The basic idea is to perform trilinear decomposition on the instrumental response data, which is based on the alternating trilinear decomposition (ATLD) algorithm. In model construction, the background drift is modeled as one component or factor as well as the analytes of interest, hence, the drift is explicitly included into the calibration. The method involves performing trilinear decomposition on the raw data, then extracting the background component and subtracting this background data from the raw data, leaving the analytes’signal on a flat baseline. Simultaneous evaluation of three-dimensional background drift and true signals may improve the quality of the data. This method has been applied to the determination and removal of three-dimensional background drifts in simulated multidimensional data as well as experimental comprehensive two-dimensional liquid chromatographic data. It was shown that this technique yields a good removal of background drift, without the need to perform a blank chromatographic run, and requires no prior knowledge about the sample composition.2. The quantitative analysis of complex multi-component systems (Chapter 3 to 6)With the increasing attention on the quality and security of food, analytical measurements with high accuracy and efficiency are required urgently. New analytical techniques are investigated and applied in food analysis. Sudan dyes are synthetic colorants and are widely used in household products such as waxes, floor and shoe polishes. Unfortunately, Sudan dyes are used as synthetic colorants in food illegally. In Chapter 3, the Sudan dyes in hot chilli samples were analyzed with HPLC-DAD. With second-order calibration methodologies such as the self-weight alternating trilinear decomposition (SWATLD), the satisfactory prediction results has be obtained even in the presence of uncalibrated intertering components. The sample preparation was based on solvent extraction, and internal standard was not required. Quantification was carried out with simple mobile phase.Sulfonamides constitute a group of drugs which are widely used in veterinary. Their metabolizing process are slow in animal body, thus, residues of these drugs may remain in food of animal origin. In Chapter 4, a new method based on the use of HPLC-DAD coupled with the alternating penalty trilinear decomposition (APTLD) was applied to the rapid determination of sulfonamides in milk samples. Though the chromatographic and spectral peaks of the components were highly overlapped, the good quantitative results have been obtained owing to the second-order advantage. The application of second-order calibration methodologies provides a new technique to food analysis.Drug analysis in body fluids is an important issue in biomedical field. Chromatographic separation techniques are usually used in drug analysis. In some complex multi-component systems, it is difficult to get completely separation of the components, and inaccuracy results could be obtained. However, the application of second-order calibration methodologies can improve or replace the“chemical separation”with“mathematical separation”, displaying the characteristic of fast and universal, dependable.Glucocorticoids are important in predicting against shock, stress, inflammation, etc. It plays an important role in human physiology, and is being a useful marker for the diagnosis under pathologic conditions. Due to the structural similarity between cortisol and prednisolone, it is not surprising that the two analytes have similar spectral profiles and close retention time. In Chapter 5, second-order calibration based on ATLD is employed to determine the concentration of cortisol and prednisolone in body fluids from HPLC-DAD data. In this work, solvent extraction was used in sample preparation, since it is simple and convenient. A simple mobile phase consisting of methanol and water was used. Though heavily overlapped chromatographic peaks of the analytes and interferences were obtained and the spectra of these species were also overlapped, the powerful ATLD algorithm can resolve the overlapped peaks into corresponding chromatographic, spectral and concentration profiles even in the presence of unknown coeluting interferences.Metoprolol, aβ1-selectiveβ-adrenergic receptor-blocking agent, is clinically used in the treatment of arrhythmia, angina pectoris and hypertension. As direct spectrofluorimetric analysis has the characteristic of high sensitivity and selectivity, as well as its relative low cost, in Chapter 6, a new spectrofluorimetric method for the direct quantitive analysis of metoprolol in plasma is presented. It is based on the use of EEMs and second-order calibration algorithms. This methodology enables accurate and reliable measurement of the analyte concentration, even in the presence of unknown and uncalibrated fluorescent components. Simple sample pretreatment step was required.3. Three-way data analysis in the research of pharmaceutical dynamics process (Chapter 7)The study of interactions between drugs and DNA are significant not only in understanding the mechanism of interaction, but also in giving a good guidance for designing new drugs. However, the mechanisms of interactions between molecules and DNA were still little known. In this study, the competitive interactions of THP and EB with DNA were investigated with fluorescent spectrophotometry. Spectroscopic studies, including fluorescent spectra analysis and parallel factor analysis (PARAFAC) of excitation-emission three-way data array were carried out on the competitive interactions of the complex. With PARAFAC, the excitation and emission spectra as well as the relative concentrations of co-existing species in different reaction and equilibrium mixtures can be directly and conveniently obtained. The obtained results showed that THP competed against EB to bind to DNA. The results also indicated that the mode of binding of the complex to DNA was intercalation action. It is valuable for providing a deeper insight into the interaction mechanism of THP and EB with DNA. The PARAFAC algorithm proved a convincing method for studying the competitive interactions of complex with DNA. The application of chemometric methods can provide more intuitionistic information for the mechanism study of drugs and DNA. It also provides a helpful verification for the controversial interaction mechanism between the drugs and DNA.更多还原