A novel copper(ii) complex as a nitric oxide turn-on fluorosensor: intracellular applications and DFT calculation

Abstract

We report, herein, the development of an easily synthesizable novel dansyl-based turn-on NO sensor L2. The UV-Vis titration data of L2 with Cu²⁺ display a gradual increase in absorbance at 418 nm with [Cu²⁺], which were analyzed by using a non-linear least-squares computer-fit program yielding K = (1.16 ± 0.36) × 10⁶ M⁻¹ and n = (1.28 ± 0.03) indicating a 1 : 1 complexation. The ground state geometries of L2 as well as its complex [Cu(L2)Cl]⁺ (1) were optimized by DFT calculations which showed that in complex 1 the central metal ion is in distorted tetrahedral geometry with bond distances very close to those found in analogous Cu²⁺ complexes. The fluorescence of L2 was dramatically quenched (∼60-fold) through complexation with paramagnetic Cu²⁺ to form [Cu(L2)Cl]⁺ in MeCN–H₂O (9 : 1, v/v) at pH 7.2 in HEPES buffer, which on further treatment with Angeli's salt (Na₂N₂O₃) restores its fluorescence property by ∼15-fold due to the reduction of Cu²⁺ to Cu⁺ by NO generated in solution from Na₂N₂O₃. The lifetime measurements displayed a substantial decrease in the lifetime of free ligand L2 (τ₀ = 12 ns) on complexation with Cu²⁺ (τ₀ = 2.1 ns). The detection limit of NO calculated by the 3σ method gives a value of 1.6 nM. The NO induced fluorescence enhancement of [Cu^II(L2)Cl]⁺ was due to the reduction of [Cu^II(L2)Cl]⁺ (1) to [Cu^I(L2)]⁺ (2) and is supported by the disappearance of the d–d transition band at 850 nm as well as the X-band EPR signal of 1. The selective “turn on” fluorogenic behavior of L2 was examined on HeLa cells of human cervical cancer origin by fluorescence microscopy which showed very intense intracellular fluorescence that was strongly suppressed by the addition of Cu²⁺ but it regains its fluorescence property on further incubation with Angeli's salt (Na₂N₂O₃). The existence of [Cu^II(L2)Cl]⁺ and [Cu^I(L2)]⁺ in solution was confirmed by ESI-MS⁺ (m/z) analysis. The effect of different biologically relevant cations and anions on the fluorescence property of L2 indicates that it was only the [Cu^II(L2)Cl]⁺ which displayed high selectivity for NO, indicating its suitability for intracellular application without much worry about its cytotoxicity in a specified dose.