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共振光散射技术在原小檗碱类生物碱分析中的应用研究
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摘要
共振光散射(Resonance Light Scattering, RLS)技术是上世纪九十年代发展起来的新型光谱分析技术,因其操作简单、快速、灵敏度高而受到分析工作者的广泛关注。目前该技术已应用于生命科学、环境科学、材料科学及分析化学等多个领域的分析研究工作。本文利用RLS技术对小檗碱类药物的含量进行分析测定并对其RLS光谱进行了深入研究。主要研究内容如下:
     (1)在pH1.81Britton-Robinson (BR)缓冲介质中,盐酸小檗碱(Berberine Hydrochloride, BH)与全氟辛烷磺酸(Perfluorooctanesulfonate, PFOS)相互作用,在290nm处产生显著增强的RLS信号。其增强RLS信号强度与盐酸小檗碱浓度在0.044~7.8μg/mL范围内成线性关系,据此建立了盐酸小檗碱的RLS测定方法,检出限达4.4ng/mL。方法用于药片含量测定,RSD≤4.37%。
     (2)在pH0.91HCl-NaAc缓冲介质中,盐酸巴马汀(Palmatine Hydrochloride, PaH)与硅钨酸(Silicotungstic acid, STA)通过静电相互作用导致RLS信号显著增强,最大RLS波长为297nm,增强的RLS信号强度与PaH浓度在0.091~3.2μg/mL范围内呈线性关系,据此建立了PaH的RLS分析方法,检出限为9.1ng/mL。实验考查了pH、离子强度对体系的影响,研究了体系紫外吸收光谱及荧光光谱,讨论了共振光散射增强机理。该方法用于黄藤素片、黄藤素胶囊及血清样品中盐PaH含量测定,RSD≤4.2%。
     (3)聚A(poly(A))是一种单链腺嘌呤寡居核苷酸,能使13nm左右的金纳米粒子(AuNPs)在高盐浓度下保持分散状态。盐酸巴马汀(Palmatine Hydrochloride, PaH)与poly(A)有强亲和作用可降低poly(A)分散AuNPs的能力而使AuNPs发生聚集,体系RLS信号显著增强。增强的RLS信号与盐酸巴马汀浓度在0.023~2.5μg/mL范围内呈线性关系,据此建立检测盐酸巴马汀的RLS分析法,检出限为2.3ng/mL。研究了体系的吸收分析光谱及颜色变化,表征了扫描电子显微镜、暗场散射成像、动态光散射及圆二色光谱讨论了反应机理。该方法成功用于黄藤素片、黄藤素胶囊以及尿样中盐酸巴马汀含量的测定,RSD≤4.0%。
The resonance light scattering (RLS) technique is developed from the1990s. For its advantages of simple operation, fast response, high sensitivity and it can be completed by ordinary fluorescence spectrophotometers, RLS technique has been arrested extensive attention by analysts. The RLS technique has been used in many fields of analytical research work, such as life sciences, environmental science, materials science and pharmaceutical analysis. In this paper, it has been applied to the analysis of berberine drugs contents and their corresponding RLS spectra have been studied. The main contents are as follows:
     (1) A novel RLS detection method for Berberine Hydrochloride (BH) is established based on the enhanced resonance light scattering signals at290nm which result from BH interacting with Perfluorooctanesulfonate (PFOS) in Britton-Robinson (BR) buffer medium. The enhanced IRLS is proportional to the concentration of BH in a range of0.044-7.8μg/mL. The limit of detection (3a) is4.4ng/mL. This method was successfully applied for the determination of BH in tablets with relative standard deviation (RSD)≤4.37%.
     (2) For the determination of Palmatine Hydrochloride (PaH), a novel RLS method has been developed based on the largely enhanced RLS signals at297nm due to the interaction between Silicotungstic acid (STA) and PaH by electrostatic force in the pH0.91HCl-NaAc medium. The linear range of PaH is0.091~.2μg/mL. The limit of detection (3σ) is9.1ng/mL. In this work, the effects of pH and ionic strength on interaction were investigated. At the same time, the UV spectra, the fluorescence spectra and the reaction mechanism of system were also investigated. This method has been applied successfully to the determination of PaH in pharmaceutical and blood serum samples with relative standard deviation (RSD)≤4.2%.
     (3) Single stranded poly(A), as a kind of Adenine-rich nucleic acids, could stabilize13nm gold nanoparticles (AuNPs) in high ionic strength. It found that Palmatine Hydrochloride (PaH) is strong affinity to poly(A). When poly(A) exposed to PaH, aggregation of AuNPs could be found, which results in the enhanced RLS signals. A novel RLS method for determining PaH has been developed based on the enhanced intensity of RLS with a PaH range of0.023~2.5μg/mL. The limit of detection is2.3ng/mL. The absorption spectra and color changing of PaH were also studied in this work. The reaction mechanism of this system have been further investigated by Scanning Electron Microscope (SEM), Dark-field Light Scattering Images (DLSI), Dynamic Light Scattering (DLS) and Circular Dichroism (CD). The RLS method has been applied successfully for the determination of PaH in pharmaceutical and samples of urine, relative standard deviation (RSD)≤4.0%.
引文
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