Colorimetric Detection of Sudan Based on the Aptamer and Cationic Compound Induced Gold Nanoparticles Aggregation
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摘要: 本研究以苏丹红Ⅲ适配体为识别元件，以未修饰的纳米金传感信号，以聚二烯丙基二甲基氯化铵（PDDA）作为纳米金聚集的诱导剂，构建了一种简单、经济、快速的苏丹红比色检测方法。在优化条件下评估本方法的检测灵敏度、准确性和特异性，最后应用于食品中苏丹红快速检测，并将检测结果与国标法（GB/T 19681-2005）对比验证。结果显示，在PDDA浓度20 nmol/L、适配体浓度5 nmol/L、反应时间4 min等优化条件下，纳米金吸光度比值（A650nm/A530nm）与苏丹红III浓度呈良好线性关系（R=0.986），线性检测范围为3.13~50 ng/mL，可视化检测限为3.13 ng/mL，检测时间约为5 min。特异性分析显示，本方法对苏丹红I、II、III和IV有高的特异性，与柠檬黄、日落黄、分散橙11等无交叉反应。将本方法应用于食品中苏丹红检测，加标回收率为85.4%~102.5%，相对标准偏差为3.37%~6.75%。本方法具有操作简便、快速、结果易读等优点，适用于批量样品中苏丹红的现场快速检测。Abstract: In this study, a simple, economical and rapid colorimetric assay was developed for the detection of Sudan, in which Sudan-binding aptamer was used as recognition element, unmodified gold nanoparticles (AuNPs) as sensing signal, and poly(diallyldimethylammonium chloride) (PDDA) as inducer for gold nanoparticle aggregation. The sensitivity, accuracy and specificity of developed method were evaluated under optimized condition. Finally, the colorimetric sensor was applied to detection Sudan in food samples, and the results were compared with GB standard method (GB/T 19681-2005). The best procedure for Sudan analysis in our system was: The concentration of aptamer at 5 nmol/L, the concentration of PDDA at 20 nmol/L, and the reaction time was 4 min. The correlation between concentration of Sudan III and absorbance ratio of gold nanoparticles (A650nm/A530nm) was observed to be linear within the range of 3.13 to 50 ng/mL. The limit of visual detection was 3.13 ng/mL by naked-eye observation. The detection time was 5 min. The colorimetric sensor had high specificity for Sudan I, II, III and IV, and no cross-reactivity towards sunset yellow, tartrazine, and 1-amino-2-methylanthraquinone. Further, the colorimetric sensor was applied to measure Sudan Ⅲ in spiked real samples, and the recoveries were in the range of 85.4%~102.5%, with relative standard deviations of 3.37%~6.75%. Our study provides a simple, fast, and easy to read method for Sudan analysis, which can be applied in future on-site detection in food samples.
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Key words:
- Sudan dyes /
- aptamer /
- gold nanoparticles /
- colorimetric assay
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图 1 纳米金紫外吸收光谱图（A）和透射电镜图（B）
Figure 1. Ultraviolet absorption spectra (A) and TEM images (B) of the gold nanoparticles
图 2 基于适配体-PDDA诱导纳米金聚集比色检测苏丹红示意图
Figure 2. Schematic of colorimetric assay base on the aptamer and PDDA induced gold nanoparticles aggregation for detection of Sudan dyes
图 3 不同条件下检测体系紫外吸收光图谱
Figure 3. UV absorption spectra of the detection system under different conditions
注：a：AuNPs；b：AuNPs+适配体5 nmol/L+PDDA 20 nmol/L；c：AuNPs+PDDA 20 nmol/L；d：AuNPs+适配体5 nmol/L+苏丹红III 200 ng/mL+PDDA 20 nmol/L。
图 6 苏丹红与适配体反应时间优化
Figure 6. The optimization of reaction time between Sudan dyes and aptamer
注：对照组1：AuNPs+适配体5 nmol/L+PDDA 20 nmol/L；对照组2：AuNPs+PDDA 20 nmol/L；图8同。
图 7 不同苏丹红Ⅲ浓度条件下体系的紫外光谱图（A）和标准曲线图（B）
Figure 7. UV-Vis spectrum of aptasensor at different concentrations of Sudan III (A) and the calibration plot of the absorbance versus concentration of Sudan III (B)
图 8 苏丹红及结构类似物在相同检测条件下的紫外光谱图
Figure 8. UV-vis spectrum of Sudan and analogue at same detection conditions
表 1 苏丹红检测方法比较
Table 1. Comparison of methods reported for detection of Sudan dyes
分析方法 分析物 识别元件 检测限 线性范围 检测时间 参考文献 HPLC Sudan I~IV - 0.33~2.27 μg/g 0.5~5.0 μg/mL - [3] HPLC SudanI~IV - 10 ng/g - - [18] MIP-HPLC Sudan I~IV MIP 5~9 ng/g 31~4000 ng/g - [20] HPL-MS/MS Sudan I~IV - 0.2~1.0 ng/g 1~100 ng/mL - [21] GC-MS/MS Sudan III - 2~10 ng/g 0.1~4.0 μg/mL - [22] ELISA Sudan III 抗体 0.1~0.8 ng/mL - 105 min [23] 电化学传感器 SudanI - 0.093 μmol/L 0.3~700 μmol/L - [24] 比色 SudanIII 适配体 2.15 ng/mL 3.1~50 ng/mL 5 min 本方法 注：-代表文献未涉及该部分内容。 
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表 2 本方法加标回收率（n=3）
Table 2. The recovery of the proposed method (n=3)
样品 添加量（ng/g） 检测值（ng/g） 加标回收率（%） 相对标准偏差（%） 辣椒粉 10 8.83±0.45 88.3 5.09 50 46.20±1.76 92.4 3.81 100 93.80±5.22 93.8 5.56 辣椒酱 10 9.19±0.62 91.9 6.75 50 45.40±1.53 90.8 3.37 100 102.50±6.35 102.5 6.20 番茄酱 10 8.54±0.38 85.4 4.45 50 44.50±2.03 89.0 4.56 100 94.60±5.57 94.6 5.88
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