论文——碳氧化锌纳米线阵列泡沫石墨烯电化学检测叶酸

碳/氧化锌纳米线阵列/泡沫石墨烯电化学检测叶酸摘 要 通过化学气相沉积法制备三维(3D)泡沫石墨烯(GF)，然后利用水热合成法在GF表面生长氧化锌纳米线阵列(ZnO NWAs)，再利用化学气相沉积法在其表面沉积碳(C)，得到碳/氧化锌纳米线阵列/泡沫石墨烯(C/ZnO NWAs/GF)复合材料。采用该复合材料做电极，采用电化学方法检测叶酸(FA)。结果表明，3D GF具有和模板泡沫镍一样的三维孔状结构，ZnO NWAs均匀且垂直地生长在GF表面，碳沉积在ZnO NWAs表面。在线性范围为0-60 M内，C/ZnO NWAs/GF电极检测FA时，灵敏度为0.13 AM-1，且在尿酸 (UA)干扰下检测FA具有良好的选择性。C/ZnO NWAs/GF电极有良好的稳定性和重复性。关键词 泡沫石墨烯；氧化锌纳米线阵列；碳；电化学生物传感器；叶酸；尿酸分类号 TB332 TP212.3Electrochemical determination of folic acid using carbon/ZnO nanowire arrays/graphene foamABSTRACT Three-dimensional (3D) graphene foam (GF) was synthesized by chemical vapor deposition (CVD). Then, ZnO nanowire arrays (ZnO NWAs) were grown on the 3D GF by hydrothermal synthesis. Finally, carbon (C) was deposited on the surface of ZnO NWAs by CVD to obtain the hybrid of C/ZnO NWAs/GF. This composite was used to detect folic acid. The results showed that the GF inherited the 3D macroporous structure of nickel foam and ZnO NWAs were uniformly and vertically grown on the 3D GF. The length and diameter of ZnO nanowires were 50 nm and 2 m, respectively. Carbon was deposited on the surface of ZnO NWAs. C/ZnO NWAs/GF was used as a working electrode to detect folic acid (FA) using the electrochemical method. The results showed that the sensitivity of the electrode for FA is 0.13 AM-1 and it has good selectivity to detect FA in the presence of uric acid (UA). The electrode also has excellent reproducibility and stability.KEY WORDS graphene foam; ZnO nanowire arrays; carbon; electrochemical biosensor; folic acid; uric acid叶酸(FA)是一种水溶性维生素，它是人体中不可缺少的成分1。人体缺少叶酸会引起贫血，怀孕的女性如果体内缺少FA，会导致胎儿出生时体重过轻，唇裂，心肌缺陷等。但是过量的FA会使人出现缺锌，出现恶心，厌食等消化疾病。所以准确灵敏的检测FA具有重要的意义。目前检测叶酸的方法主要包括高效液相色谱法2-4、分光光度法5, 6、化学发光法7、荧光计法8和酶联配体吸着剂测试法9。这些检测方法复杂，耗时长且成本高。电化学方法在检测生物分子时具有价格低廉、操作方便、稳定性高、灵敏度和选择性高等优点10。石墨烯是一种二维单原子厚度的碳材料，具有高光透性，高电导性以及优异的机械性能11。三维（3D）泡沫石墨烯（GF）是以泡沫镍为模板，用化学气相沉积法(CVD)制备具有空间连通的三维结构石墨烯，具有大的比表面积，同时石墨烯平面为金属氧化物的生长提供了巨大的平台。氧化锌纳米线阵列(ZnO NWAs)具有好的生物兼容性、在生理环境中具有化学稳定性、高催化效率以及大的比表面积12-15。本课题组曾将ZnO NWAs生长在GF表面用于检测帕金森病生物标记物12，能够简单、方便地测到标记物浓度。但是ZnO是半导体II-VI族材料16，电导性较差，响应信号较低，限制了其应用。此外，由于ZnO NWAs具有较高的等电点(9.5)，在检测生物分子的时候，表面容易吸附生物分子，影响电化学性能17。为了改善ZnO NWAs 的这一缺陷，采用CVD法将碳(C)沉积在其表面18，改善复合电极的电化学性能。本文以泡沫镍为模板，通过CVD制备3D GF，然后通过水热合成的方法在GF表面生长ZnO NWAs。最后，再利用CVD在ZnO NWAs表面沉积碳，制备出C/ZnO NWAs/GF。使用该复合材料为电极，检测溶液中FA的浓度，并研究尿酸(UA)的存在下，FA浓度与峰电流之间的关系。1 实验部分 1.1 C/ZnO NWAs/GF的制备 以泡沫镍为模板，采用CVD法制备3D GF 12, 19。将3D GF转移到氧化铟锡(ITO)玻璃上，在其表面滴加ZnO种子层，通过水热合成的方法制备ZnO NWAs，具体的实验操作如文献12，与文献12不同的是，反应物中氨水的浓度增加为0.5 M，以降低ZnO NWAs的密度，制备出ZnO NWAs/GF。将ZnO NWAs/GF放入真空管式炉中，在Ar(500 sccm)保护下，以10 /min的升温速率将管式炉加热至700 。将CH4通入管式炉20 min，停止通入CH4后，炉温降至室温，取出产物即为C/ZnO NWAs/GF复合材料。1.2 材料表征采用荷兰飞利浦公司的FEI-Sirion200型热场发射扫描电镜(SEM)配备有能谱（EDS）组件，对C/ZnO NWAs/GF的形貌进行表征并对元素进行分析。拉曼光谱(Raman) 分析采用英国RENISHAW 公司的RM-1000 型显微共焦激光拉曼光谱仪，激光波长为514 nm。1.3 电化学性能测试电化学实验在VMP3 型电化学工作站(Biologic Science instrument，法国) 上进行。采用三电极检测体系，以C/ZnO NWAs/GF为工作电极，Ag/AgCl 为参比电极，Pt丝为对电极。采用循环伏安测试法(CV)和差分脉冲伏安(DPV)检测C/ZnO NWAs/GF 对FA 的电化学性能。检测均在0.01 M的磷酸缓冲盐溶液（PBS，20 mL，pH 7.4) 中进行。2结果与讨论2.1 C/ZnO NWAs/GF的表征图1a-d为不同放大倍数的C/ZnO NWAs/GF的扫描图。可以看出：GF保留着泡沫镍的三维结构骨架，平均孔径为300 m。ZnO NWAs均匀且垂直生长在GF表面，ZnO NWAs的直径和长度分别为50 nm和2 m。图1e为C/ZnO NWAs/GF的拉曼光谱图，可以看出复合材料有ZnO和GF组成，GF为多层的（G/2D大于1）且无明显的缺陷(无D带)。图1f 为C/ZnO NWAs/GF的能谱图，表明C沉积在ZnO NWAs表面。图1 C/ZnO NWAs/GF的扫描形貌与结构Fig. 1 SEM images and structure of C/ZnO NWAs/GF. a-d, SEM images of C/ZnO NWAs/GF with different magnifications, e, Raman spectrum of C/ZnO NWAs/GF, f, EDS of C/ZnO NWAs/GF.2.2 C/ZnO NWAs/GF的电化学性能测试2.2.1 CV测试图2为C/ZnO NWAs/GF电极在50 M FA和UA中扫描速率为50 mVs-1的循环伏安测试曲线。从图中可以明显看出，FA和UA的氧化峰电位分别为0.7 V和0.45 V，修饰后的C/ZnO NWAs/GF电极的氧化峰电流值明显高于裸ITO玻璃电极，这主要因为C和GF都具有高的导电性，同时ZnO NWAs也为生物分子的检测提供了更多的活性点。两曲线的氧化峰与还原峰并不是完全的对称图形，说明FA与UA在 C/ZnO NWAs/GF电极上的氧化和还原反应不是可逆反应。图2 C/ZnO NWAs/GF电极在50 M 的FA和UA中的CV曲线Fig. 2 CV curves of C/ZnO NWAs/GF electrode in 50 M FA and UA. a, FA; b, UA图3为C/ZnO NWAs/GF电极在50 M的FA中不同扫描速率下的循环伏安曲线。扫描速率由下至上分别为10, 25, 50, 75, 100 mVs-1。由图3a可以看出，随着扫描速率的增加，FA氧化峰电流逐渐升高。图3b是FA的氧化峰电流与扫描速率的拟合曲线。拟合方程为Ip = (5.931.44) + (0.280.02) (R2 =0.973)。由此得知，该电极反应机理为吸附型20。图3C/ZnO NWAs/GF电极在50 M FA中不同扫描速率的CV曲线Fig. 3 CV curves of C/ZnO NWAs/GF electrode in FA with different scanning rates.a, CV curves of C/ZnO NWAs/GF at different scanning rates, b, linear relationship of oxidation peak current and scanning rate.2.2.2 DPV测试与CV测试相比，DPV可以降低背底电流的干扰而得到更强的分析信号，从而获得更好的灵敏度21。为了得到更精确的检测结果，进一步采用DPV测试方法对C/ZnO NWAs/GF电极进行电化学性能测试。图4a可以看出，在浓度为0-60 M的范围内，随着FA浓度的增加，氧化峰电流值逐渐升高。FA浓度与氧化峰电流的拟合曲线为Ip = (1.110.36) + (0.130.01)CFA (R2 =0.9765)。所以，C/ZnO NWAs/GF电极检测FA时的检测限为1 M，灵敏度为0.13 AM-1 (图4b)。图4C/ZnO NWAs/GF电极对不同浓度FA的DPV曲线Fig. 4 DPV curves of C/ZnO NWAs/GF electrode in FA with different concentrations.a, DPV curves of FA with different concentrations, b, linear relationship of oxidation peak current and FA concentration表1为不同电极材料检测FA的灵敏度及检测限等电化学性能结果对比。可以看出C/ZnO NWAs/GF电极在检测FA时具有良好的性能指标。表1不同电极通过DPV检测FA的电化学性能结果Table 1 Electrochemical properties of FA by DPV using different electrodes电极灵敏度 (AM-1)线性范围(M)参考文献carbon fibre 0.01 0.02-1 22 MC-CNPE0.03 15-550 23ZONMCPE 0.01 20-2500 24C/ZnO NWAs/GF 0.13 1-60 Ours MC-CNPE: 钼(VI)/碳纳米管修饰碳糊电极ZONMCPE/CPE: ZrO2纳米颗粒修饰碳糊电极UA与FA共同存在于人体体液中，研究UA存在下，采用 C/ZnO NWAs/GF电极对测量FA具有重要的意义。图 5a是在15 M UA干扰下对不同浓度(1, 5, 10, 20, 30, 40，50和60 M ) FA进行的DPV测试。随着FA浓度的增加，氧化峰电流逐渐升高。图5b是FA氧化峰电流与浓度的拟合曲线，曲线方程为Ip =(0.870.19）+(0.120.01) CFA (R2 =0.983)。由此证明，UA的存在并未对FA造成明显干扰。C/ZnO NWAs/GF电极在检测FA时具有优异的选择性。图5 15 M UA干扰下检测FA的DPV曲线Fig. 5 DPV curves of C/ZnO NWAs/GF electrode to detect FA in the presence of UA. a, DPV curves of FA with different FA in the presence of UA, b, linear relationship oxidation peak current and concentration of FA2.2.3 C/ZnO NWAs/GF电极稳定性与重复性测试我们用C/ZnO NWAs/GF电极对50 M的FA进行重复性的DPV测试（如图6a）。共进行了15次重复实验，实验时间间隔为10分钟。结果表明，氧化峰电流值得相对标准偏差(RSD)为1.9%，证明该电极具有优异的重复性。将该电极放置5天后，继续对FA进行DPV测试（如图 6b）每两天进行一次测试，一共测试两周。其RSD为2.1%，表明C/ZnO NWAs/GF电极具有优异的稳定性。图6 C/ZnO NWAs/GF电极的重复性与稳定性测试Figure 6 Reproducibility and stability of the C/ZnO NWAs/GF electrode. a, DPV curves for 15 repeated cycles. The reproducibility was investigated through continuous DPV scanning in 50 M FA at a 10 min interval. b, The long-term storage stability was evaluated by measuring the DPV response of 50 M FA every 2 days. The electrode was tested for 2 weeks and was maintained in a 0.01 M PBS solution after use.2.2.4 C/ZnO NWAs/GF电极在蛋白质及离子环境中检测FA为了测试C/ZnO NWAs/GF电极的实用性，真实生物样本分析是在蛋白质， K+、Na+、Cl-、HCO3-（NaCl，KHCO3）等存在下采用DPV方法对FA进行测量。测试结果如表2中所示。表明C/ZnO NWAs/GF电极在离子及蛋白质存在下对FA测量是可行的。表2 为C/ZnO NWAs/GF电极用于在离子及蛋白质等存在下检测FATable 2 The application of C/ZnO NWAs/GF electrode for simultaneous determination of FA in ionicss and protein样品加入量(M)发现量(M)回收率(%)相对标准偏差(%)蛋白质/氯化钠/碳酸氢钾00-54.998.02.11010.3103.12.22020.2100.82.03. 结论通过CVD制备出3D GF，采用水热合成法将ZnO NWAs垂直生长于GF表面，再次CVD将碳沉积于ZnO NWAs表面。C/ZnO NWAs/GF电极在检测FA时具有优异的电化学性能，在浓度为0-60 M的范围内，灵敏度为0.13 AM-1，具有良好的稳定性及重复性，且在UA的干扰下，C/ZnO NWAs/GF电极具有优异的选择性。C/ZnO NWAs/GF电极也将在检测其它生物分子上具有巨大的潜力。参 考 文 献1. Asaikkutti A, Bhavan P S,Vimala K. 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