Ruthenium(Ⅱ) Complex Functionalized Nanomaterials And Their Applications In Chemiluminescence Bioassays

In this dissertation, the state of art in the field of chemiluminescence (CL), chemiluminescence functionalized nanomaterials and label-free bioassays were reviewed. The development of chemiluminescence functionalized nanomaterials (CF-NMs) is a new trend with innovative concepts in nanoscience, which makes thousands of CL signal-generating molecules be coated on or encapsulated in a nanoparticle host. However, limited CF-NMs have been reported. Most of the reported CF-NMs focuse on approach which links the luminescent reagent to the surface of nanomaterials by bridge molecules. CF-NMs based on the direct synthesis strategy for functionalized nanomaterials have rarely been reported. In our research group, it was found that luminol and its derivatives could directly reduce chloroauric acid to form CL functionalized gold nanomaterials, which could be used as analytical probe for label-related bioassays. The methodologies have been successfully applied for the detection of DNA, thrombin and cardiac troponin I. However, the research for CF-NMs is limited to luminol and its derivatives. Ruthenium(II) complex with higher light-emitting efficiency has rarely explored for the synthesis of CF-NMs. And CF-NMs were mainly used in label-related detection. Label-free detection has some advantages such as fast, low-cost and simple procedure without labeling, which makes it be very attractive in bioassays. The aim of this dissertation is to explore the synthesis and CL property of new ruthenium(II) complex CF-NMs and their applications in label-free bioassays. Two kinds of ruthenium(II) complex CF-NMs including gold nanoparticles and graphene oxide were synthesized. The CL and electrochemiluminescence (ECL) activities of these nanomaterials were investigated. Novel label-free sensors were designed by using CF-NMs as analytical nano-interface. The main results are as follows:1. Ru(bpy)2(NH2-phen)2+functionalized gold nanoparticles (AuNPs) were successfully synthesized by a simple one-pot method via the reduction of HAuCl4with NaBH4in the presence of Ru(bpy)2(NH2-phen)2+at room temperature. During synthetic process, the Ru(II) complex as a stabilizing reagent was linked to the surface of AuNPs by the weak covalent interaction between nitrogen atoms of its amino-terminal ligands and gold atoms to form luminescence functionalized gold nanoparticles. Such luminescence functionalized AuNPs could directly generate ECL under cyclic voltammetry. This concept is a promising way to obtain amino-ruthenium(II) complex functionalized gold nanoparticles, which could be used as probe in sensors and bioassays.2. An approach was developed for the preparation of graphene oxide covalently functionalized with a Ru(II) complex (Ru-GO). A ruthenium(II) complex designed with a long alkyl amino functional group on one of its diimine ligands, bis(2,2’-bipyridine)(N-(2-aminoethyl)-4-(4’-methyl-2,2’-bipyridine-4-yl)butanamide) ruthenium(II), was covalently grafted onto GO by the amide reaction between the amino group of the Ru(II) complex and the carboxyl group of the GO. The as-prepared Ru-GO showed excellent ECL activity, good solubility and good stability. Using this hybrid material, a reagent-free ECL method was developed for the detection of tripropylamine to demonstrate the applicability of this new functionalized material. Tripropylamine in a range of1.0×10-7～1.0×10-3mol L-1could be detected by use of the ECL intensity with a detection limit of7.5×10mol L-1. This approach is reliable and highly efficient, and might be extended to other chemiluminescent molecules containing an amino group such as luminol and its analogues to functionalize GO with ECL activity.3. A novel homogeneous label-free aptasensor for2,4,6-trinitrotoluene (TNT) detection was developed based on an assembly strategy of electrochemiluminescent graphene oxide (GO) with AuNPs and aptamer. In this sensing strategy, the anti-TNT aptamer was first assembled with AuNPs to form aptamer-AuNPs. Then ruthenium(II) complex functionalized GO (denoted as Ru-GO) was assembled with aptamer-AuNPs by electrostatic interaction. AuNPs could directly quench the ECL emission of ruthenium(II) complex on the surface of Ru-GO due to the energy transfer from luminophore to the AuNPs. Weak ECL signal of the assembly was obtained. In the presence of target molecule TNT, the aptamer-AuNPs would aggregate partly due to the aptamer-target interaction and reduce quenching effect, leading to ECL signal restoration and strong ECL signal was obtained. TNT in a range of0.01-100ng mL-1could be detected by use of the ECL intensity discrepancy with a low detection limit of3.6pg mL-1. The aptasensor also showed high selectivity towards TNT against2,4-dinitrotoluene, p-nitrotoluene and nitrobenzene. The present aptasensor has been successfully applied to the detection of TNT in real water samples. Compared with previous reported sensors, this homogenous aptasensor avoided complicated labeling and purification procedure and showed magnificent sensitivity and high selectivity, which made it not only convenient but also time-saving and applicable. Furthermore, this sensing strategy also provides a promising way to develop new ECL aptasensor for other analytes by virtue of other aptamers.4. Sulphonatocalix[4]arene functionalized gold nanoparticles were synthesized and characterized. A homogeneous label-free sensor for arginine detection was proposed based on sulphonatocalix[4]arene functionalized gold nanoparticles. It was found that guest molecule arginine could react specifically with its host molecule sulphonatocalix[4]arene which was functionalized on the surface of gold nanoparticles, leading to that aggregation of mono-disperse of gold nanoparticles to some extend due to the host-guest interaction. It was also found that sulphonatocalix[4]arene functionalized gold nanoparticles could catalyze the CL reaction between luminol and AgNO3. The dispersity of gold nanoparticles would affect the catalytic activity, resulting a decrease in CL intensity. On this basis, A homogeneous label-free sensor for arginine detection was constructed. Arginine in a range of1.0×10-7～1.0×10-5mol L-1could be detected with a detection limit of4.0×10-8mol L-1. The proposed sensor has some advantages such as no-washing, simple operation, high sensitivity, while using a microplate as the detection container make this methods time-saving and easy automation. It was proved that calixarene functionalized gold nanoparticles could be used as specific recognition and label-free nano-detection interface for the determination of the guest molecules. In view of the diversity and richness of the calixarene, gold nanoparticles functionalized with various calixarene as special molecular recognition would provide a simple, sensitive label-free bioassay methods for the determination of various guest molecules.