Signal-on Type Electrochemical Detection of DNase I in Homogeneous Media Using New Ferrocenylnaphthalene Diimide

A simple DNase I detecting method was achieved under a homogenous medium containing DNA duplex and a newly synthesized ferrocenylnaphthalene diimide derivative (FND) carrying alanine (1) or alanine-lysine residue (2) as an electrochemical indicator. The binding affinity of 1 or 2 with double stranded DNA was 105 M−1 or 106 M−1 order, which was estimated from a Scatchard analysis calculated by its absorption change upon the addition of calf thymus DNA (CT-DNA). It was expected that 1 or 2 intercalated to double stranded DNA with a threading mode and electrostatic interaction of the linker ammonium cations of 1 or 2 with phosphate anions of the DNA duplex backbone. The largest current increase after a DNase I treatment was observed under the mixture in a ratio of [3]:[CT-DNA-bp] = 10 : 1 (bp: DNA concentration per base pair), where the distance between the bound ligands is expected to a 10 bp or 34 Å theoretically. When considering to be ca. 30 Å of DNase I size and decreasing of the current increase under an increased amount of 3, DNase I required over ten base pairs to access and digest with the DNA duplex. The current increase in the case of a mixture of 10 μM 1 or 2 and 100 μM CT-DNA was obtained 340 % or 640 %, respectively, after being treated with DNase I (final concentration: 2.0 × 10−3 U μL−1), which might be derived from an apperarent molecular weight or diffusion coefficient of the ligand bound DNA duplex and the detection limit of DNase I in the case of 1 or 2 was 2.0 × 10−5 or 2.0 × 10−9 U μL−1, respectively, where the detection limit for 2 was 104-times higher than that for 1.