CN108760855B - 一种石墨烯-聚吡咯-金纳米粒子复合材料的制备方法及应用 - Google Patents
一种石墨烯-聚吡咯-金纳米粒子复合材料的制备方法及应用 Download PDFInfo
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Abstract
本发明公开了一种石墨烯‑聚吡咯‑金纳米粒子复合材料，采用原位化学聚合和静电吸附的相结合的方法，将金纳米粒子负载与石墨烯‑聚吡咯复合材料上。其制备方法包括以下步骤：1）溶液的配置；2）溶液的混合反应制备聚吡咯‑石墨烯米复合材料；3）金纳米粒子溶液的制备；4）金纳米粒子的吸附。石墨烯‑聚吡咯‑金纳米粒子复合材料的应用，用于阻抗型大肠杆菌生物传感器修饰电极的应用，检测大肠杆菌的线性范围为1×102~1×107 CFU/mL，最低检出限为100 CFU/mL。本发明所制备的阻抗型大肠杆菌生物传感器还具有操作简单、成本低廉、使用方便、选择性高等优点，因而在食品安全和临床分析等领域中具有巨大的潜在应用价值。
Description
技术领域
本发明涉及基于石墨烯的纳米复合材料的制备技术领域，具体涉及石墨烯-聚吡咯-金纳米粒子复合材料的制备方法及应用。
背景技术
大肠杆菌O157:H7型（Escherichia coli O157:H7）是一种肠道出血性大肠杆菌，是食物中毒的原因之一。感染者通常发生出血性腹泻，尤其在年轻儿童和年长者中，有时导致肾衰竭。传染通过粪口途径，很多病例与吃未煮熟或污染的牛肉和猪肉、游泳、喝被污染的水、吃被污染的蔬菜有关。因此，对食源性大肠杆菌O157:H7的快速准确的鉴定, 对于预防疾病的传播具有重要的意义。
现有的传统检测方法，如PCR和ELISA 等，尽管能在复杂背景中进行检测，且检测限较低，但存在以下技术问题：1、对操作人员要求高；2、耗时长；3、无法实现在线检测等。因此，传统检测方法不适用于工业应用的实验室。
近年来，生物传感器在食品安全领域致病菌检测上发挥出越来越重要的作用。到目前为止，大部分检测致病菌的生物传感器都是依赖于标记进行放大信号的免疫传感器，该传感器通过抗体抗原之间的反应转换成能够检测到的电化学信号。但电化学大肠杆菌生物传感器存在响应信号低，稳定性差等问题，需要通过设计纳米复合材料对电化学信号进行放大，另外材料生物相容性的复合材料对大肠杆菌抗体进行固定，可以有效提高传感器的稳定性。
发明内容
本发明的目的是提供一种石墨烯-聚吡咯-金纳米粒子复合材料的制备方法及应用。
为达到上述目的，本发明采用原位化学聚合法制备了石墨烯-聚吡咯复合材料，由于石墨烯-聚吡咯复合材料表面带正电荷，可以和带负电荷的金纳米粒子发生静电相互作用，从而使得金纳米粒子很好地分散在石墨烯-聚吡咯表面。克服了石墨烯化学活性不高，金纳米粒子其表面很容易团聚的问题。而且石墨烯、聚吡咯、金纳米粒子具有良好的导电性、高的比面积和良好的生物相容性，可以吸附尽可能多的大肠杆菌抗体，然后吸附尽可能多大肠杆菌，大肠杆菌吸附在电极表面后，由于大肠杆菌导电性差，在[Fe(CN)6] 3-/4-作为氧化还原对存在的情况下，使电极的电化学反应电阻明显增大，利用大肠杆菌浓度变化和电极的电阻变化之间的对应关系，可以得到大肠杆菌检测的工作曲线，从而实现大肠杆菌的检测。
根据上述机理，本发明采用如下技术方案：
一种石墨烯-聚吡咯-金纳米粒子复合材料，采用原位化学聚合和静电吸附的相结合的方法，将金纳米粒子负载与石墨烯-聚吡咯复合材料上。
一种石墨烯-聚吡咯-金纳米粒子复合材料的制备方法，包括以下步骤：
步骤1）溶液的配置，以盐酸、吡咯、石墨烯和水的质量比为0.1: 1:（1-3）:100，将盐酸和吡咯溶解到水溶液中，然后将石墨烯超声分散到该溶液中，记为溶液M，以过硫酸铵和水的质量比为2:10，将过硫酸铵加入到水溶液中，记为溶液N，所述步骤1盐酸、吡咯、石墨烯和水的质量比为0.1: 1:（1-3）:100；
步骤2）溶液的混合反应制备聚吡咯-石墨烯复合材料，将步骤1所述的溶液N缓慢地向溶液M中滴加，并用磁力搅拌器对进行搅拌，滴加完成后，再让溶液反应2-4小时，然后经过滤、洗涤、干燥，得到的聚吡咯-石墨烯复合材料；
步骤3）金纳米粒子溶液的制备，以氯金酸、柠檬酸三钠、NaBH4和水的质量比为（0.1-0.3）:1:0.1:100，将氯金酸加入水中，搅拌，然后加入柠檬酸三钠，再加入NaBH4还原，得到所需要的金纳米粒子溶液；
步骤4）金纳米粒子的吸附，将步骤2所得到的聚吡咯-石墨烯复合材料加入到步骤3所制备的金纳米粒子溶液中，搅拌，然后过滤、洗涤、干燥，得到石墨烯-聚吡咯-金纳米粒子复合材料。
一种石墨烯-聚吡咯-金纳米粒子复合材料的应用，用于阻抗型大肠杆菌生物传感器修饰电极的应用，检测大肠杆菌的线性范围为1×102~1×107 CFU/mL，最低检出限为100CFU/mL。
本发明的一种石墨烯-聚吡咯-金纳米粒子复合材料的制备方法及应用对于现有技术，具有以下优点：
1．采用静电吸附法在石墨烯-聚吡咯表面吸附金纳米粒子，有效提高了金纳米粒子在石墨烯表面的分散性，克服了金纳米粒子在石墨烯表面易团聚的问题，方法简单；
2．石墨烯-聚吡咯-金纳米粒子具有良好的导电性、高的比表面积和良好的生物相容性，可以有效提高抗体分子在电极上的负载量，并保持良好的活性；
3. 高分散的金纳米粒子对生物分子有良好的亲和力，可以直接对抗体分子进行吸附，无需交联剂的参与；
4．本发明石墨烯-聚吡咯-金纳米粒子复合材料的制备方法工艺简单，产品性能稳定，适合大批量的制备，而且后处理工艺简单。
因此，本发明在生物传感器领域具有广阔的应用前景。
附图说明
图1为本发明实施例制备石墨烯-聚吡咯-金纳米粒子复合材料的透射电镜图；
图2为本发明实施例制备石墨烯-聚吡咯-金纳米粒子复合材料修饰电极对大肠杆菌O157：H7检测的交流阻抗图；
图3为本发明实施例制备的石墨烯-聚吡咯-金纳米粒子复合材料修饰电极检测大肠杆菌O157：H7的工作曲线。
具体实施方式
本发明通过实施例，结合说明书附图对本发明内容作进一步详细说明，但不是对本发明的限定。
实施例
一种石墨烯-聚吡咯-金纳米粒子复合材料的制备方法：
步骤1）溶液的配置，将0.1 g和 1 g吡咯溶液加入到100 mL水溶液中，然后将1 g石墨烯加入到该溶液中，超声分散1 h，记为溶液M，将2 g过硫酸铵溶解到10 mL水溶液中，记为溶液N；
步骤2）溶液的混合反应制备聚吡咯-石墨烯复合材料，将步骤1所述的溶液N缓慢地向溶液M中滴加，并用磁力搅拌器对进行搅拌，滴加完成后，再让溶液反应2小时，然后经过滤、洗涤、干燥，得到的聚吡咯-石墨烯复合材料；
步骤3）金纳米粒子溶液的制备，将0.1 g氯金酸加入100 mL水中，搅拌，然加入0.5g柠檬酸三钠，再加入0.1g NaBH4还原，得到所需要的金纳米粒子溶液；
步骤4）金纳米粒子的吸附，将步骤2所得到的聚吡咯-石墨烯复合材料加入到步骤3所制备的金纳米粒子溶液中，搅拌，然后过滤、洗涤、干燥，得到石墨烯-聚吡咯-金纳米粒子复合材料。
阻抗型大肠杆菌生物传感器修饰电极制备和检测方法：
（1）取0.1 g石墨烯-聚吡咯-金纳米粒子超声分散到1mL N,N-二甲基甲酰胺溶液中，取该纳米复合材料的悬浮液10 μL滴在玻碳电极的表面，在红外灯下烤干；
（2）取溶液3 μL 0.1 mg/mL 的大肠杆菌O157：H7抗体滴在步骤7的修饰电极上。然后将该修饰电极放置在4 ℃的冰箱中60 min，然后用磷酸缓冲溶液洗涤。即可得到大肠杆菌生物传感器工作电极；
（3）配制浓度为1×102，1×103，1×104，1×105，1×106，1×107 CFU/mL的大肠杆菌O157：H7溶液，将（2）制备的大肠杆菌生物工作电极浸在上述溶液中1h， 然后以铂电极为对电极，Ag/AgCl电极为参比电极，在5 mM Fe(CN)6 3-/4- 的溶液中，测试工作电极的交流阻抗，利用大肠杆菌浓度对交流阻抗的电化学反应电阻作图，可得到大肠杆菌生物传感器的工作曲线。
石墨烯-聚吡咯-金纳米粒子复合材料的透射电镜如图1所示，可以看出所得的金纳米粒子很好地分散在石墨烯-聚吡咯基体上。
石墨烯-聚吡咯-金纳米粒子复合材料修饰电极对大肠杆菌检测的交流阻抗图，如图2所示，可以看出大肠杆菌浓度的增加，电极的电化学反应电阻明显增大。
图3为本发明实施例制备的石墨烯-聚吡咯-金纳米粒子复合材料修饰电极检测大肠杆菌的工作曲线。在浓度范围为1×102～1×107 CFU/mL时，表现出良好的线性关系，最低检出限位1×102 CFU/mL。发明构建的一种基于石墨烯-聚吡咯-金纳米粒子复合材料的电化学阻抗大肠杆菌生物传感器，利用石墨烯-聚吡咯-金纳米粒子复合材料对抗体亲和力强的特点，有效地对电化学检测信号增强，对大肠杆菌进行了快速、灵敏的检测，具有广泛的应用前景。
Claims (2)
1.一种石墨烯-聚吡咯-金纳米粒子复合材料的制备方法，其特征在于包括以下步骤：
步骤1）溶液的配置，以盐酸、吡咯、石墨烯和水满足一定质量比，将盐酸和吡咯溶解到水溶液中，然后将石墨烯超声分散到该溶液中，记为溶液M，以过硫酸铵和水满足一定质量比，将过硫酸铵加入到水溶液中，记为溶液N；所述步骤1）盐酸、吡咯、石墨烯和水的质量比为0.1: 1:（1-3）:100，过硫酸铵和水的质量比为2:10；
步骤2）溶液的混合反应制备聚吡咯-石墨烯复合材料，将步骤1）所述的溶液N缓慢地向溶液M中滴加，并用磁力搅拌器对进行搅拌，滴加完成后，再让溶液反应2-4小时，然后经过滤、洗涤、干燥，得到的聚吡咯-石墨烯复合材料；
步骤3）金纳米粒子溶液的制备，以氯金酸、柠檬酸三钠、NaBH4和水满足一定质量比，将氯金酸加入水中，搅拌，然后加入柠檬酸三钠，再加入NaBH4还原，得到所需要的金纳米粒子溶液；
步骤4）金纳米粒子的吸附，将步骤2）所得到的聚吡咯-石墨烯复合材料加入到步骤3）所制备的金纳米粒子溶液中，搅拌，然后过滤、洗涤、干燥，得到石墨烯-聚吡咯-金纳米粒子复合材料；所述步骤4）氯金酸、柠檬酸三钠、NaBH4和水的质量比为（0.1-0.3）:1:0.1:100。
2.根据权利要求1所述制备方法所制得的石墨烯-聚吡咯-金纳米粒子复合材料的应用，其特征在于：所述制得的石墨烯-聚吡咯-金纳米粒子复合材料用于阻抗型大肠杆菌生物传感器修饰电极的应用，检测大肠杆菌的线性范围为1×102~1×107 CFU/mL，最低检出限为100 CFU/mL。
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