摘要
化学发光分析是根据化学反应产生的光辐射(化学发光)确定物质含量
的一种痕量分析方法。化学发光分析具有灵敏度高、线性范围宽、分析速度
快以及仪器设备相对简单便宜等优点,并已经成为现代痕量分析中一个十分
活跃的研究和应用领域。目前,这种方法在痕量金属离子、无机阴离子、生
物分子、致癌物质、药物化学、环境科学及临床医学等方面得到了广泛的应
用。
本论文包括两部分。第一部分是综述,这部分结合各种化学发光反应体
系,评述了1985年以来,特别是最近5年来国内外药物化学发光分析方面
的应用研究进展。
第二部分是研究报告。详细研究了铁氰化钾-鲁米诺体系和铁氰化钾-鲁
米诺-亚铁氰化钾体系的化学发光动力学性质,对影响体系化学发光强度及
信噪比的各种因素进行了研究。利用维生素K_3对铁氰化钾-鲁米诺体系化学
发光的抑制作用、异烟肼对K_3Fe(CN)_6-luminol体系化学发光的增强作用以
及尿酸对铁氰化钾-鲁米诺-亚铁氰化钾体系化学发光的增强作用,通过对实
验条件的优化,建立了简单、灵敏、快速测定VK_3、异烟肼和尿酸的流动注
射化学发光新方法,并探讨了化学发光反应的机理。
建立了做透析-流动注射-化学发光检测联用系统,并用来研究了盐酸四
环素与牛血清白蛋白的相互作用。首先利用K_3Fe(CN)_6-luminol-K_4Fe(CN)_6化
学发光体系,通过对氧化剂浓度、背景抑制剂浓度和样品流速等实验条件的
优化,结合流动注射技术,建立了盐酸四环素的FIA-CL分析方法,然后对
微透析探针进行了校正,测定了不同灌流速率下探针的相对回收率。在选定
的实验条件下,对盐酸四环素-牛血清白蛋白之间的相互作用进行了体外研
究,结果表明简单、快速的微透析-流动注射-化学发光联用技术运用于药物
与蛋白的相互作用研究。
(1)铁氰化钾溶液与鲁米诺碱性溶液混合时产生化学发光,维生素K_3
的水溶液抑制此反应的化学发光强度,基于此,建立了测定VK_3的流动注射
化学发光新方法,该法简单、快速。化学发光强度与维生素K_3的浓度在0-
1μg/mL范围内成正比,检测限(3σ)为0.01μg/mL,相对标准偏差为0.16%
(n=11,0.4μg/mL MSB)。用于VK_3片剂及注射液中MSB的测定,结果满
意。
摘 要
门)铁氰化钾与碱性鲁米诺溶液混合产生化学发光,异烟胁增强其化
学发光强度,基于此,建立了一种测定异烟胁的流动注射化学发光分析新方
法。在0—lug/mL的异烟姘浓度范围内,化学发光强度与异烟胁浓度呈良
好的线性关系,检测限* d为 0刀3 ng/mL,对 0.lug/mL的异烟姘进行 11次
电 平行测定,相对标准偏差(RSD)为l·2%。用于异烟盼药片及针剂的测定,结
果令人满意。
(3)在碱性条件下,铁氰化钾氧化鲁米诺,产生化学发光,尿酸对该
体系的化学发光有显著的增强作用(亚铁氰化钾存在时)。基于此,建立了
一种直接测定尿酸的流动注射化学发光分析法。方法的线性范围为 二刀X
10-’一5刀X 10‘g/mL,检狈限门口)为 6.7X 10-’g/mL,相对标准偏差为 1.1%
(尿酸 1.0 XIO、/mL,n=11人用于血清及尿样中尿酸的分析,结果令人满
意。
(4)以盐酸四环素为例,利用微透析一流动注射一化学发光联用系统建
立了研究药物与蛋白相互作用的新方法。实验发现盐酸四环素对铁氰化钾一
鲁米诺一亚铁氰化钾体系的化学发光强度有增强作用,在本实验条件下,此
体系对盐酸四环素的测定极限为 0.2 g/mL,线性范围为 l—100 g/mL。将
所研究的药物与牛血清白蛋白按不同的摩尔比混合门℃),然后用微透析
探针在 5 pL加i n的灌流速度下进行取样,微透析探针对盐酸四环素的回收
_、,___。,_.回__… 口口二匕凶、归羹二 厂_____--.口口二口色、W
e 刀 dg.O h:+u m 6catcnarQ 刀 o——=nA 一rA co tlotz 刀 7t
士 上=上+一上一二一得到的盐酸四环素与牛血清白蛋白的结合参数值与文献值相
r n uK D。
Q 吻合,这说明简单、快速的微透析一流动注射一化学发光联用技术适用于药物
与蛋白的相互作用研究。
Chemiluminescence analysis, based on the emission during some chemical
reactions, is a technique for the determination of trace-amount species. Due to its
high sensitivity, wide linear range, rapidity, simplicity and relatively low cost,
chemiluminescence analysis has found application in assay of trace amounts of
metal ions, inorganic anions, biological molecules and cancer-causing species, and
in pharmaceutical chemistry, environmental science as well as clinical medicine.
The thesis consists of two parts. The first one is a review dealing with the
development of the application of chemiluminescence in pharmaceutical analysis.
Emphasis is placed on the development in the recent five years.
The other part is research reports. The kinetic properties of two
chemiluminescence systems, K3Fe(CN)6uminol system and
K3Fe(CN)6uminol4Fe(CN)6 system, were studied in detail. Various factors
were also studied that have effect on the chemiluminescence intensity and/or on
the ratio of signal to noise of the two systems. It was found that the aqueous
solution of Vitamin K3 has depressing effect on the chemiluminescence intensity
of the K3Fe(CN)6uminol system, while isoniazid has enhancing effect on it, and
uric acid has enhancing effect on the CL intensity of the other system. Based on
these discoveries, a series of experiments were conducted to optimize the
conditions in point and to make clear the mechanisms of the three reactions.
Sensitive, simple and fast methods were proposed for the determination of VK3,
isoniazid and uric acid.
A microdialysis samplinglow inj ectionhemiluminescence detection
system was set up and has been successfully applied to the study of interaction
between tetracycline hydrogen chloride and bovine serum album(BSA). It
comprised several steps. Firstly, various factors of the
K3Fe(CN)6uminol4Fe(CN)6 chemiluminescence system, such as oxidant
concentration, flow rate and repressor concentration were studied in order to
obtain optimum experiment conditions. As a result, a flow-injection
chemiluminescence analysis method for the determination of tetracycline
hydrogen chloride was developed. Secondly, the dialysis probe was calibrated and
its recoveries at different perfusion rates were measured. Finally, the interaction
between tetracycline hydrogen chloride and BSA was studied in vitro under the
optimized conditions. The result compared well with that in the literature,
indicating that the proposed system is suitable for the study of drugrotein
interaction.
A chemiluminescence (CL) flow system is described for the determination of
menadione sodium bisulfite based on its repression on the chemiluminescence
emission produced upon mixing a hexacyanoferrate(III) solution with an alkaline
luminol solution in the absence of co-oxidizer. The system responds linearly to
menadione sodium bisulfite concentration in the range 0 ?1 pg/mL with a
detection limit (3cr) of 0.01 pg/mL. Relative standard deviation (RSD) of 0.16%
for 0.4 pg/mL menadione sodium bisulfite (n1 1). The system has been
successfully applied to the determination of menadione sodium bisulfite in tablets
and injections.
A chemiluminescence(CL) flow system is described for the determination of
isoniazid based on its enhancement on the chemiluminescence emission produced
upon mixing a hexacyanoferrate(III) solution with an alkaline luminol solution.
The system responds linearly to isoniazid concentration in the range 0?1 pg/mL
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