Enhanced Doubly Activated Dual Emission Fluorescent Probes for Selective Imaging of Glutathione or Cysteine in Living Systems

Abstract

The development of novel fluorescent probes for monitoring the concentration of various biomolecules in living systems has great potential for eventual early diagnosis and disease intervention. Selective detection of competitive species in biological systems is a great challenge for the design and development of fluorescent probes. To improve on the design of fluorescent coumarin-based biothiol sensing technologies, we have developed herein an enhanced dual emission doubly activated system (DACP-1 and the closely related DACP-2) for the selective detection of glutathione (GSH) through the use of one optical channel and the detection of cysteine (Cys) by another channel. A phenylselenium group present at the 4-position completely quenches the fluorescence of the probe via photoinduced electron transfer to give a nonfluorescent species. Probes are selective for glutathione (GSH) in the red region and for cysteine/homocysteine (Cys/Hcy) in the green region. When they were treated with GSH, DACP-1 and DACP-2 showed strong fluorescence enhancement in comparison to that for closely related species such as amino acids, including Cys/Hcy. Fluorescence quantum yields (Φ) increased for the red channel (<0.001 to 0.52 (DACP-1) and 0.48 (DACP-2)) and green channel (Cys) (<0.001 to 0.030 (DACP-1) and 0.026 (DACP-2)), respectively. Competing fluorescent enhancements upon addition of closely related species were negligible. Fast responses, improved water solubility, and good cell membrane permeability were all properly established with the use of DACP-1 and DACP-2. Live human lung cancer cells and fibroblasts imaged by confocal microscopy, as well as live mice tumor model imaging, confirmed selective detection.

A fluorescent probe for the dual detection of mercury ions and thiols based on a simple coumarin derivative

“…Thus the conclusion was reached that the E1 probe is useful for fluorogenic detection of Hg(ii) over a relatively wide pH range of pH 4-11. Changes in the fluorescence emission spectra of the E1 probe (20 µmol/L) upon addition of different amounts of Hg(ii) (0, 1,2,4,8,12,16,20,40 and 60 µmol/L) were measured ( Figure 5). It was found that with increasing amounts of Hg(ii) added to E1 that the intensity of the fluorescence emission band at 466 nm decreased gradually, and that the intensity of the emission peak at 395 nm increased.…”

“…However, the fluorescence emission spectra of the E1 probe changed in accordance with the nature of each amino acid which was added: Cys, Phe, Asn, Val, His, Glu and Gly all led to a decrease in intensity of the fluorescence emission band at 466 nm. The Changes in the fluorescence emission spectra of probe E1 (20 µmol/L) upon the addition of different amounts of Cys (0, 2,4,6,8,10,12,14,16,18,20 and 40 µmol/L) were measured ( Figure 8). The strongly fluorescent E1 probe gradually turned weakly fluorescent as increasing amounts of Cys were added to E1.…”

“…The fluorescence quenching effect of a thiol on probe E1 was further demonstrated by measurement of the change in the fluorescence emission spectrum of probe E1 upon the addition of one equivalent of p-toluenethiol in HEPES buffer (0.1 mol/L, pH = 7.16) with water/ethanol (9:1, v/v) as solvent(Figure 7). The results indicated that E1 can serve as a fluorescence turn-off sensor for the detection of thiols, in contrast to most other reported probes which were capable of sensing thiols by fluorescence enhancement [1][2][3][4][5][6][7].…”

“…Biothiols represented by cysteine (Cys), homocysteine (Hcy) and glutathione (GSH) play fundamental roles in living systems, and the design of fluorescent probes for the selective detection of biothiols has attracted growing interest. [1][2][3][4][5][6][7] Thiophenols, which are also called benzenethiols, are extensively utilised in the synthesis of various products including pharmaceuticals, pesticides and polymers. However, thiophenols are a highly toxic and pollutant species.…”

Simultaneous Visualization of Endogenous Homocysteine, Cysteine, Glutathione, and their Transformation through Different Fluorescence Channels

“…To understand the reaction mechanism between probe 1 and Cys/Hcy/GSH, several control compounds (C5‐C9) were synthesized (Supporting Information, Scheme S2), and characterized (Supporting Information, Figures S31–S40). Judging from the LC‐MS and HRMS spectra from the reactions between probe 1 and Hcy/Cys/GSH and the absorption spectra of these control compounds (Supporting Information, Figures S8 and S14–S16), mechanisms were proposed for the reactions between probe 1 and Hcy/Cys/GSH (Supporting Information, Scheme S2) . The isolation of the products of these reactions failed, which may be due to their instability.…”