Quantitative Detection of Acetylcholine in Mouse Brain Tissues Using Zeolitic Imidazolate Frameworks/ Graphitic Carbon Nitride Quantum Dots/ Acetylcholinesterase Complex
摘 要
以二水合乙酸锌、2-甲基咪唑、石墨相氮化碳量子点(g-CNQDs,7 mg)和乙酰胆碱酯酶(AchE)为起始原料,采用一锅法制备了沸石咪唑酯骨架材料(ZIFs)包载g-CNQDs和AchE的酸度敏感型复合物ZIFs@g-CNQDs@AchE,并和患阿尔茨海默病(AD)小鼠的脑组织样品在pH 7.4磷酸盐缓冲溶液中室温反应2.5 h,在380 nm激发波长下测量体系在475 nm处的荧光强度,用于测定乙酰胆碱(Ach)的含量。结果显示,合成的复合物呈十二面体结构,平均粒径约300 nm,加入Ach后,Ach渗透进入复合物,被AchE催化生成乙酸,体系酸度增加,复合物结构瓦解,g-CNQDs释放,475 nm处出现明显的荧光发射峰。Ach的浓度在5.5~82.5 mmol·L-1内和对应的体系荧光强度呈线性关系,检出限(3S/N)为1.83 mmol·L-1;常见的干扰物质(葡萄糖、多巴胺)以及干扰离子(Fe2+、Fe3+、Cu2+、Mg2+、Zn2+)对Ach的检测无干扰,复合物的特异性较高;按照标准加入法对阳性实际样品进行回收试验,成功检出了Ach,回收率为93.2%~112%,测定值的相对标准偏差(n=5)为0.82%~3.4%。构建的酸度敏感型复合物可作为一种潜在的载体,用于AD诊疗。
Abstract
Using zinc acetate dihydrate, 2-methylimidazole, graphitic carbon nitride quantum dots (g-CNQDs, 7 mg) and acetylcholinesterase (AchE) as starting materials, the acidity sensitive complex of zeolitic imidazolate frameworks (ZIFs) containing g-CNQDs and AchE (denoted as ZIFs@g-CNQDs@AchE) was prepared by one pot method. In order to determine Ach, the complex and brain tissues samples of mouse with AD were reacted in the pH 7.4 phosphate buffer solution at room temperature for 2.5 h, and the fluorescence intensity of the system was measured at 475 nm with excitation wavelength of 380 nm. It was shown that the synthesized complex had a dodecahedron structure with an average particle size about 300 nm. After adding Ach, Ach penetrated into the complex and was catalyzed by AchE to generate acetic acid. The acidity of the system increased, the complex structure was collapsed, g-CNQDs were released, and an obvious fluorescence emission peak appeared at 475 nm. The linear relationship between values of the concentration of Ach and the corresponding fluorescence intensity of the system was kept in the range of 5.5-82.5 mmol·L-1, with detection limit (3S/N) of 1.83 mmol·L-1. Common interfering substances (glucose and dopamine) and interfering ions (Fe2+, Fe3+, Cu2+, Mg2+ and Zn2+) did not interfere with the detection of Ach, indicating that the specificity of the complex was high. Test for recovery was made according to standard addition method on the positive actual samples, and Ach was successfully detected, giving recoveries in the range of 93.2%-112% and RSD (n=5) of the determined values in the range of 0.82%-3.4%. The constructed acidity sensitive complex can serve as a potential carrier for the diagnosis and treatment of AD.
中图分类号 O657.34 DOI 10.11973/lhjy-hx202308001
所属栏目 试验与研究
基金项目 苏州市科技发展计划民生科技-医疗卫生应用基础研究项目(sysd2020192)
收稿日期 2022/1/4
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备注杨渊,硕士研究生,主要从事利用化学分析法对生物标志物检测的研究
引用该论文: YANG Yuan,ZHU Liang,HU Yuanyuan,YU Yanyan. Quantitative Detection of Acetylcholine in Mouse Brain Tissues Using Zeolitic Imidazolate Frameworks/ Graphitic Carbon Nitride Quantum Dots/ Acetylcholinesterase Complex[J]. Physical Testing and Chemical Analysis part B:Chemical Analysis, 2023, 59(8): 869~874
杨渊,朱亮,胡园园,于妍妍. 沸石咪唑酯骨架材料/石墨相氮化碳量子点/乙酰胆碱酯酶复合物定量检测小鼠脑组织中的乙酰胆碱[J]. 理化检验-化学分册, 2023, 59(8): 869~874
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【2】KANG T W, JEON S J, KIM H I, et al. Optical detection of enzymatic activity and inhibitors on non-covalently functionalized fluorescent graphene oxide[J]. ACS Nano, 2016,10(5):5346-5353.
【3】LUO Y, KIM E H, FLASK C A, et al. Nanosensors for the chemical imaging of acetylcholine using magnetic resonance imaging[J]. ACS Nano, 2018,12(6):5761-5773.
【4】ALBISHRI H M, ABD EL-HADY D. Hyphenation of enzyme/graphene oxide-ionic liquid/glassy carbon biosensors with anodic differential pulse stripping voltammetry for reliable determination of choline and acetylcholine in human serum[J]. Talanta, 2019,200:107-114.
【5】CHEN W H, LIAO W C, SOHN Y S, et al. Stimuli-responsive nucleic acid-based polyacrylamide hydrogel-coated metal-organic framework nanoparticles for controlled drug release[J]. Advanced Functional Materials, 2018,28(8):1705137.
【6】CHEN W H, YU X, CECCONELLO A, et al. Stimuli-responsive nucleic acid-functionalized metal-organic framework nanoparticles using pH- and metal-ion-dependent DNAzymes as locks[J]. Chemical Science, 2017,8(8):5769-5780.
【7】DENG J J, WANG K, WANG M, et al. Mitochondria targeted nanoscale zeolitic imidazole framework-90 for ATP imaging in live cells[J]. Journal of the American Chemical Society, 2017,139(16):5877-5882.
【8】ZHANG Y, MENG S, DING J H, et al. Transition metal-coordinated graphitic carbon nitride dots as a sensitive and facile fluorescent probe for β-amyloid peptide detection[J]. The Analyst, 2019,144(2):504-511.
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