Study of Fluorescence Quenching by Bilirubin of a Carbocyanine Dye in Complex with DNA. Effect of Cu2+ Additives

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The effect of bilirubin on the spectral fluorescence properties of the cationic thiacarbocyanine dye Cyan 2 in the presence of DNA was studied. The Cyan 2 dye forms a non-covalent complex with DNA, which leads to an increase in the fluorescence of the dye. Interaction with bilirubin leads to effective quenching of dye fluorescence in complex with DNA (static mechanism), which can be used to construct a spectral-fluorescent sensor for bilirubin. The results of in vitro experiments are illustrated by in silico molecular docking experiments. The effect of Cu2+ ion additives can further enhance the quenching of dye fluorescence by bilirubin. Effective quenching constants and detection limits of bilirubin using the Cyan 2–DNA system (LOD and LOQ) are determined.

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作者简介

P. Pronkin

Emanuel Institute of Biochemical Physics of the Russian Academy of Sciences

编辑信件的主要联系方式.
Email: pronkinp@gmail.com
俄罗斯联邦, Moscow

A. Tatikolov

Emanuel Institute of Biochemical Physics of the Russian Academy of Sciences

Email: pronkinp@gmail.com
俄罗斯联邦, Moscow

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2. Fig. 1. Structures of the studied compounds - Cyan 2 dye and bilirubin (BR).

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3. Fig. 2. a - Normalised absorption spectra of Cyan 2 dye (cCyan 2 = 1.5 μM) in phosphate buffer solution at different concentrations of DNA, BR and Cu2+: cDNA = 0 (curve 1), 20 (2, 4, 5, 6), 140 μM (3); cBR = 0 (1-3), 3 μM (4-6); cCu2+ = 0 (1-4), 6.3 μM (5 and 6); spectrum 5 corresponds to the deaerated sample; b - dependences of the molar extinction coefficient εabs (1), relative width of spectral curves in the absorption spectra Wh/2 (2), position of the maximum λabs (3), fluorescence intensity Ifl, (4) of Cyan 2 dye on the cDNA/cCyan 2 ratio in the absence of quenching; c - normalised fluorescence spectra of Cyan 2 (1-4; λex = 540 nm) at different concentrations of BR: cBR = 0 (curve 1), 0.11 (2), 1.2 (3), 3.3 μM (4). Inset shows the dependence of fluorescence quenching of Cyan 2 dye (cCyan 2 = 1.5 μM, λex = 505 nm, λreg = 565 nm) on BR concentration in Stern-Folmer coordinates (1) and linear analytical dependence using the Stern-Folmer equation (2).

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4. Fig. 3. Results of molecular docking of Cyan 2 dye (a, b) and bilirubin (c, d) with double-stranded DNA: complex in the small trough (a, c; 1BNA.pdb) and intercalation complex (b, d; 198D.pdb).

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