The utilisation of n-heterocycles in the development of electrochemical porcine and malaria biosensors, and application as dna and heme binders

Abstract

The N-heterocycle exhibit excellent binding ability due to the presence of nitrogen atoms. This leads to this study aiming to explore N-heterocycle binding ability with biomolecules DNA porcine and β-hematin biomarker using copper tetraaza complex CuTL3 and 1,2,4- triazole-3,5-diamine TZ, respectively. And also leads to study on improvement of binding ability of new nitrogen heterocycles containing amidino indole derivatives 48 – 53, 57 towards the biomolecules such as DNA and heme. The binding ability of copper-tetraaza heterocycle complex CuTL3 towards DNA biomolecule was performed with regards to the development of porcine biosensor. The porcine biosensor was fabricated by assembling the SPCE/AuNPs/MPA/EDC/NHS layer. CuTL3, an excellent electrochemical indicator was applied to obtain the DNA hybridisation response using differential pulse voltammetry. The developed biosensor exhibits a broad linear dynamic range of 1 × 10−13 M - 1 × 10−8 M (R=0.99) and a low detection limit of 5.7 × 10−14 M. The biosensor showed good selectivity toward the complementary DNA and also good reproducibility. The developed biosensor could sensitively detect pork DNA in real meat samples. On the other hand, the binding potential of nitrogen heterocycle of 1,2,4-triazole-3,5-diamine TZ towards biomolecule β- hematin was conducted with respects to the implementation of β-hematin biosensor. The β-hematin biosensor was fabricated by immobilising TZ onto SBA15. The fabricated biosensor was successfully characterized and the optimum conditions were determined. The developed biosensor exhibits a linear range of 1.0 × 10-7 – 2.0 × 10-3 M (R=0.99) with a detection limit (LOD) of 1.5 × 10-8 M. The biosensor showed excellent reproducibility performance with a 1.5 % RSD value and was stable until day 10, about a 20% decrease from the initial current response. The biosensor successfully detects the interference, Salmonella simultaneously without signal disruption. The real samples analysis in plasma culture medium exhibited good recovery of β-hematin. Nonetheless, in order to improve the binding ability of N-heterocycles against biomolecules (DNA and biomarker), new mono- and bis-amidinoindole 48 – 53, 57 were synthesised from their amidoxime unit by reduction, and their structures were characterised. The ligands docking studies against DNA and hemozoin displayed moderate to good interactions. The DNA and hemin binding ability of the ligands 49 and 57 were evaluated using UV-vis and fluorescence spectroscopy. As expected, ligand 57 containing two amidino and indole groups has the highest binding affinity towards ct-DNA, in which the binding constant (Ka) was 1.07 × 104. The binding constant indicated that compounds interact with DNA by groove binding. The binding studies of ligands 49 and 57 against heme exhibited an association constant (log K) of 4.31 and 4.38, accordingly. The binding strength indicated the ligand's possibility to interact significantly with heme and act as a binder to the malaria parasite. The compounds' frontier molecular orbitals (HOMO and LUMO) have been calculated and shown that amidino indole acts as an electron donor and acceptor to the DNA and heme. Collectively, the nitrogen heterocycles, mono- and bis-amidinoindoles showed the potential and capability of the ligands to interact with biomolecules. Therefore, development of biosensing and biological activities are suggested in the future.