植物油和體液等復(fù)雜體系中二階校定量分析應(yīng)用研究

1、植物油和體液等復(fù)雜體系中二階校正法定量分析應(yīng)用研究-畢業(yè)論文 學(xué)校代號(hào) 10532 學(xué) 號(hào) B1011S0022 分 類(lèi) 號(hào) 密 級(jí) 博士學(xué)位論文 植物油和體液等復(fù)雜體系中二階校 正法定量分析應(yīng)用研究 學(xué)位申請(qǐng)人姓名 王建瑤 培 養(yǎng) 單 位 化學(xué)化工學(xué)院 導(dǎo)師姓名及職稱(chēng) 吳海龍 教授 學(xué) 科 專(zhuān) 業(yè) 分析化學(xué) 研 究 方 向 化學(xué)計(jì)量學(xué) 論文提交日期 20 13 年6 月 植物油和體液等復(fù)雜體系中二階校正法定量分析應(yīng)用研究 摘 要 隨著社會(huì)的發(fā)展,儀器技術(shù)的進(jìn)步，越來(lái)越多的分析儀器能夠獲得高維數(shù)據(jù)。 在這種形勢(shì)下，二階校正方法越來(lái)越受到眾多分析領(lǐng)域的關(guān)注。二階校正方法被 廣泛接受的主要原因，是

2、它能夠從許多聯(lián)用方法，如激發(fā)-發(fā)射三維熒光、高效液 相色譜-二極管陣列檢測(cè)器等產(chǎn)生的高維數(shù)據(jù)中提取有效的信息，即使分析體系中 存在未知干擾，也可以完成對(duì)感興趣物質(zhì)的定性、定量分析，這種特殊的優(yōu)勢(shì)被 Booksh 和 Kowalski 命名為“二階優(yōu)勢(shì)” 。二階優(yōu)勢(shì)的存在使得我們能夠從復(fù)雜體 系中直接測(cè)定分析物，這也成為本論文研究的基石。 本論文側(cè)重于二階校正方法在植物油和體液等復(fù)雜體系中的定量分析應(yīng)用研 究。這些體系都包含許多種類(lèi)的化合物，基質(zhì)極為復(fù)雜。分析植物油中的化合物 對(duì)建立植物油的質(zhì)量檢測(cè)非常重要，而體液中藥物和標(biāo)記物含量水平的檢測(cè)則與 人體健康程度密切相關(guān)。目前，用于植物油和體液等復(fù)

3、雜體系分析的最廣泛的方 法是液相色譜法，但是這種方法成本高、耗時(shí)、需要提取等前處理步驟進(jìn)行預(yù)分 離。因此，本論文旨在發(fā)展一種簡(jiǎn)單、快速、有效且無(wú)需提取等預(yù)處理步驟的策 略用于定量分析體液和植物油等復(fù)雜體系中的化合物。基本的研究?jī)?nèi)容概括如下： 第二章：提出了高效液相色譜-二極管陣列檢測(cè)器結(jié)合基于交替懲罰三線(xiàn)性分 解（APTLD ）的二階校正方法，用于快速測(cè)定食用植物油中十種合成酚類(lèi)抗氧化 劑：沒(méi)食子酸丙酯 （PG ）、3-叔丁基-4-羥基茴香醚 （BHA ） 、3,5-二叔丁基-4-羥 基苯酚甲苯 （BHT ）、2,4,5-三羥基苯丁酮 （THBP ）、叔丁基對(duì)苯二酚 （TBHQ ）、 沒(méi)食子酸

4、辛酯（OG ）、沒(méi)食子酸月桂酯（DG ）、去甲二氫愈創(chuàng)木酚（NDGA ）、 乙氧喹啉（EQ ）和 2,6-二叔丁基-4-羥甲基苯酚（Ionox- 100 。交替懲罰三線(xiàn)性分 解算法（APTLD ）能夠正確的解析包含重疊峰以及基線(xiàn)漂移的二階數(shù)據(jù)，從而實(shí) 現(xiàn)植物油中抗氧化劑的快速測(cè)定。該方法無(wú)需提取步驟，避免了分析物的流失， 并能在較短的時(shí)間內(nèi)完成十種物質(zhì)的洗脫，減少了有機(jī)溶劑對(duì)環(huán)境的污染。我們 計(jì)算了預(yù)測(cè)均方差、線(xiàn)性相關(guān)系數(shù)、檢測(cè)下線(xiàn)以及加標(biāo)回收率等參數(shù)用以驗(yàn)證新 方法的有效性。預(yù)測(cè)結(jié)果令人滿(mǎn)意，檢測(cè)下限能夠滿(mǎn)足大多數(shù)國(guó)家測(cè)定植物油中 酚類(lèi)抗氧化劑的需求。 第三章：提出了高效液相色譜-熒光檢測(cè)器

5、結(jié)合基于交替三線(xiàn)性分解（ATLD ） 的二階校正方法，用于同時(shí)測(cè)定六種植物油中的七種合成酚類(lèi)抗氧化劑。此方法 無(wú)需提取步驟，樣品稀釋后即可直接進(jìn)樣，在實(shí)驗(yàn)過(guò)程中也無(wú)需清洗色譜柱。所 有物質(zhì)的洗脫可在短時(shí)間內(nèi)完成。在多種植物油所含未知組分對(duì)分析物形成干擾 的情況下，依然可以完成準(zhǔn)確的定量分析。此方法的檢測(cè)下限遠(yuǎn)遠(yuǎn)低于抗氧化劑 II 植物油和體液等復(fù)雜體系中二階校正法定量分析應(yīng)用研究 的常規(guī)限量水平，并且比第二章方法的檢測(cè)下限低一個(gè)數(shù)量級(jí)。 第四章：發(fā)展了一種新的算法，自加權(quán)整體最小二乘三線(xiàn)性分解算法（SWTT ）， 用于解析三維熒光模擬數(shù)據(jù)以及實(shí)驗(yàn)數(shù)據(jù)。我們?cè)O(shè)計(jì)了三組模擬數(shù)據(jù)，分別從噪 聲、共線(xiàn)

6、性以及背景干擾強(qiáng)度三個(gè)方面對(duì)自加權(quán)交替三線(xiàn)性分解（SWATLD ），平 行因子分析交替最小二乘（PARAFAC-ALS ）和新算法SWTT 做了比較，并從預(yù)測(cè) 能力、出錯(cuò)次數(shù)以及與真實(shí)光譜的擬合程度等幾個(gè)方面對(duì)各個(gè)算法做了評(píng)價(jià)。當(dāng) 共線(xiàn)性、噪聲以及干擾強(qiáng)度水平較低時(shí)，三個(gè)算法都能得到準(zhǔn)確穩(wěn)定的結(jié)果，但 是當(dāng)其中之一水平較高時(shí)，SWATLD 算法就無(wú)法得到正確的分辨，而 PARAFAC-ALS 算法則更容易陷入沼澤，運(yùn)行出錯(cuò)。SWTT 算法通過(guò)利用整體最小 二乘和權(quán)重因子，提高了穩(wěn)定性，從一定程度上改善了雙因子退化問(wèn)題，并能得 到更加準(zhǔn)確的結(jié)果。此外，我們還將三種算法應(yīng)用于化妝品中雌酮和雌二醇的

7、同 時(shí)測(cè)定，SWATLD 無(wú)法正確分辨，SWTT 和 PARAFAC-ALS 則能夠給出合理的結(jié) 果，并且兩者的結(jié)果極為相似。 第五章：一些研究表明，人體體液中異黃蝶呤的增加和癌癥之間有一定的聯(lián) 系，可將其含量作為醫(yī)學(xué)中判斷某些癌癥病患的重要指標(biāo)之一。因此，本文通過(guò) 結(jié)合二階校正方法和三維熒光技術(shù)發(fā)展了一種簡(jiǎn)單、快速并具有高靈敏度和高選 擇性的的方法用于測(cè)定人體尿液中異黃蝶呤的含量。雖然尿液中其它蝶呤類(lèi)化合 物的激發(fā)、發(fā)射光譜與異黃蝶呤的光譜嚴(yán)重重疊，但是由于我們方法具有二階優(yōu) 勢(shì)，在無(wú)需任何預(yù)處理進(jìn)行化學(xué)或物理分離的情況下就可對(duì)異黃蝶呤的含量進(jìn)行 直接測(cè)定。我們還采用常規(guī)的高效液相色譜-熒光

8、檢測(cè)器（HPLC-FLD ）方法對(duì)相 同的尿液樣本進(jìn)行了測(cè)定，兩種方法得到的異黃蝶呤的含量基本一致，證明了我 們方法的可靠性。 第六章：采用交替三線(xiàn)性分解（ATLD ）和交替歸一加權(quán)殘差（ANWE ）法與 激發(fā)-發(fā)射熒光矩陣相結(jié)合，測(cè)定細(xì)胞培養(yǎng)基中的阿霉素。預(yù)測(cè)結(jié)果令人滿(mǎn)意。此 外，還將細(xì)胞中的四種內(nèi)源熒光物加入到細(xì)胞培養(yǎng)基，用以模擬細(xì)胞內(nèi)熒光干擾 環(huán)境，在此環(huán)境下，我們提出的方法依然能成功測(cè)定了阿霉素的含量。本方法最 大的優(yōu)勢(shì)在于無(wú)需對(duì)細(xì)胞培養(yǎng)基做任何預(yù)處理，即可直接取樣測(cè)定，操作簡(jiǎn)單， 測(cè)定快速，對(duì)樣本也無(wú)破壞性。結(jié)果表明二階校正方法結(jié)合激發(fā)發(fā)射熒光矩陣 （EEM ）適用于培養(yǎng)基中藥物的測(cè)

9、定，為細(xì)胞的體外藥物研究提供了一種綠色的 新方法。 第七章：本章通過(guò)結(jié)合具有高靈敏度的三維熒光技術(shù)和具有高選擇性的基于 自加權(quán)交替歸一殘差擬合（SWANRF ）算法的二階校正方法，在背景干擾較強(qiáng)的 化妝品樣本中同時(shí)測(cè)定了丁基羥基茴香醚（BHA ）和沒(méi)食子酸丙酯（PG ）。我們 采用的這種策略，符合綠色化學(xué)的原則，使用極少量的有機(jī)溶劑就可完成兩種抗 氧化劑的測(cè)定。此外，我們的方法無(wú)需對(duì)樣本進(jìn)行濃縮或洗脫分離的等預(yù)處理步 III 植物油和體液等復(fù)雜體系中二階校正法定量分析應(yīng)用研究 驟，從而減少了分析時(shí)間，降低了實(shí)驗(yàn)成本。與 HPLC （AOAC 983.15 ，1994）方 法相比，EEM-SWA

10、NRF 方法更加的靈敏、簡(jiǎn)單、快速、有效且無(wú)污染。 關(guān)鍵詞：二階校正方法；高效液相色譜-二極管陣列檢測(cè)器；高效液相色譜-熒 光檢測(cè)器；激發(fā)-發(fā)射熒光矩陣；植物油；合成酚類(lèi)抗氧化劑；體液； 定量分析 IV 植物油和體液等復(fù)雜體系中二階校正法定量分析應(yīng)用研究 Abstract As society develops, a wide range of analytical instrumentation is available that enables high dimensionality data to be obtained. In this situation, second-order

11、calibration methods has gained widespread acceptance by the analytical community, because it exploit the intrinsic data structure of hyphenated analytical methods, such as excitation-emission fluorescence matrix spectroscopy and chromatographic separation coupled to diode array detection, to enable

12、one to quantify the analyte, even in the presence of interferences not modelled in the calibration stage. This property of second-order calibration methods has been termed the second -order advantage by Booksh and Kowalski. This advantage makes it possible to directly quantify the interesting analyt

13、es from complex system and provides the main motivation for the present parper . In this paper, second-order calibration methods are employed for the quantitative analysis in complex systems mainly focused on vegetable oil and body fluid . These complex systems are made of complex mixtures of compou

14、nds of a wide range of chemical classes. Characterization of compounds in vegetable oils is of great importance in establishing the oil quality, and it is important to determine the levels of medicines or markers in the body fluid for understanding how healthy people are. The most commonly used anal

15、ytical techniques for analysis of these compounds are liqu id chromatography. These techniques are however expensive, time -consuming and require a pre-separation step before the analysis. Therefore, the aim of this paper is to couple the second-order calibration methods with high-performance liquid

16、 chromatography-diode array detection HPLC-DAD , high-performance liquid chromatography-fluorescence detection HPLC-FLD or excitation -emission fluorescence matrix EEM for the development and optimization of a simple, rapid, efficient, fast and no-extraction methodology for quantitative analysis in

17、complex systems, such as vegetable oil and body fluid . The contents of our studies can be summarized as: Chapter 2:A new chromatographic methodology is presented for fast analysis of ten synthetic phenolic antioxidants in five kinds of oil samples: propyl gallate PG , 2, 4, 5-trihydroxybutyrophenon

18、e THBP , tert-butylhydroquinone TBHQ , nordihydroguaiaretic acid NDGA , ethoxyquin EQ , 3 -tert-butyl -4- hydroxyanisole V 植物油和體液等復(fù)雜體系中二階校正法定量分析應(yīng)用研究 BHA , octyl gallate OG , 2, 6-di-tert-butyl -4-hydroxymethyphenol Ionox - 100 , dodecyl gallate DG , 3, 5-di-tert-butyl -4-hydroxytoluene BHT . The alt

19、ernating penalty trilinear decomposition APTLD algorithm has shown to be an excellent tool for modelling the second-order data, where overlapping peak and baseline drift were existed, making the fast determination and resolution of the phenolic antioxidants in oils possible. The extraction is unnece

20、ssary and the ten antioxidants can be eluted within 6 mins. For the validation of the method, linearity, root-mean-square error of prediction RMSEP and limit of detection LOD have been performed. The results of overall predictions are desirable and the LOD of our method is enough to detect SPAs in e

21、dible vegetable oils in most cases . Chapter 3: A liquid chromatographic method has been developed, in combination with the second-order calibration method based on the alternating trilinear decomposition ATLD algorithm with fluorescence detection FLD , for the simultaneous determination of seven ph

22、enolic antioxidants in six kinds of oil samples. The extraction is unnecessary and a column washing is also not required. After a simple dilution step, oil samples can be directly injected into the detecting system and the elution is accomplished in a short time 6 min . Overl apped peaks have been s

23、uccessfully resolved and the quantitative results of the analytes of interest have been accurately estimated. The limits of this method are much lower than the imum residue level established for the antioxidants. It is about one -tenth of the value previously proposed by Chapter 2. Chapter 4: A nove

24、l algorithm named as self -weighted total least -squares alternating trilinear decomposition SWTT has been proposed for quantitative analysis of EEM fluorescence data. Based on different input parameters including collinearity, interference intensity and noise level in three simulated data arrays, t

25、he performances of the self -weighted alternating trilinear decomposition SWATLD , the parallel factor analysis -alternating least squares PARAFAC-ALS and the SWTT algorithms were evaluated in terms of predicting ability, error times and consistency of resolved and real profiles. When the collineari

26、ty, interference intensity and noise level of data are low, the results obtained by the three algorithms are nearly the same. However, the SWATLD can not get satisfactory results and the PARAFAC -ALS easily trap into swamps many times when the leve is high. By utilizing the alternating total least-s

27、quares principle and weight factor, the SWTT algorithm can improve the two-factor degeneracy problem, which is difficult to handle for the PARAFAC -ALS algorithm, and obtain more stable and more precise results. In addition, the SWTT and VI 植物油和體液等復(fù)雜體系中二階校正法定量分析應(yīng)用研究 PARAFAC-ALS method was successful

28、ly applied to the simultaneous analysis f or estriol and estrone in liquid cosmetic samples. Chapter 5: Several studies have shown that an increase in amounts of isoxanthopterin can be associated with the presence of cancer, thus its level is importance parameter in clinical diagnosis. A simple, rap

29、id, sensitive and selective method for determining isoxanthopterin content in human urine samples has been developed using secon-order calibration mehtod coupled with EEM. With the application of a second Corder advantage, the proposed strategy could be utilized for a direct concentration determinat

30、ion of isoxanthopterin without any pretreatment step, even the spectra of isoxanthopterin is highly overlapping with other pterin compounds neopterin, biopterin, pterin and pterin-6-carboxylic acid . The values of the isoxanthopterin from healthy volunteers, which is botained by our method, were in

31、good agreement with those obtained by HPLC -FLD method. Chapter 6: A simple and rapid method is proposed for quantitative analysis of adriamycin in cell culture media without any pretreatment procedure. It combined second-order calibration methods based on both an alternating trilinear decomposition

32、 ATLD and an alternating normalization-weighter error ANWE algorithms combined with excitation-emission fluorescent matrix spectra. The satisfactory results showed that the second-order calibration method can be applied to directly quantify adriamycin in cell culture media. Moreover, in the analysis

33、 of cell culture media including four interferences which are the fluoresc ent materials in cell, the adriamycin also can be accurately determinated . It is found that second-order calibration method is appropriate for quantitative analysis of drug content in cell culture media even in the presence

34、of natural fluorescent interfe rences of cell. Chapter 7: This work presents a novel approach for simultaneous determination of butylated hydroxyanisole BHA and propyl gallate PG in a very interfering environment by combining the sensitivity of molecular fluorescence and the selectivi ty of second-o

35、rder calibration method based on the self -weighted alternating normalized residue fitting SWANRF algorithm . This method uses very small amounts of organic reagents and the quantification of PG and BHA were accomplished using green-chemistry principles. In addition, the proposed method avoids preco

36、ncentration and elution procedures, so it considerably decreases the analytical time and the experimental expenses . Because the instrument involved in the measurement is nonsophisticated, the experiments could be carried out in routine laboratories. Compared to the HPLC method the AOAC official met

37、hod 983.15 , 1994 , the VII 植物油和體液等復(fù)雜體系中二階校正法定量分析應(yīng)用研究 EEM-SWANRF is a more sensitive, simpler, faster, cleaner and more powerful method. Keywords: Seconder-order method; HPLC-DAD; HPLC-FLD ; Excitation-emission fluorescent matrix; Vegetable oils ; Synthetic phenolic antioxidants; Body fluid ; Quantitative analysis VIII 植物油和體液等復(fù)雜體系中二階校正法定量分析應(yīng)用研究 目 錄 學(xué)位論文原創(chuàng)性聲明. I 學(xué)位論文版權(quán)使用授權(quán)書(shū). I 摘 要.II Abstract .V 第 1 章 緒 論. 1 1.1 分析化學(xué)概論. 1 1.2 化學(xué)計(jì)量學(xué)概論. 2 1.3 張量校正研究進(jìn)展. 3 1.4 高效液相色譜結(jié)合二階校正方法在復(fù)雜體系分析中的應(yīng)用. 7 1.4.1 高效液相色譜方法. 7 1.4.2 高效液相色譜-二極管陣列檢測(cè)器（HPLC-DAD ）結(jié)合二階校正方法 . 8 1.4.3 高效液相色譜-快速掃描熒光檢測(cè)器 （