A near-infrared fluorescent probe for H₂S based on tandem reaction to construct iminocoumarin-benzothiazole and its application in food, water, living cells

Highlights

• A novel NIR fluorescent probe for H₂S based on tandem reaction to construct iminocoumarin-benzothiazole.

• TCF-DNs detects H₂S with a large Stokes shift, a high signal-to-noise ratio, a low detection limit.

• TCF-DNs can detect H₂S in red wine, natural waters, and image H₂S living cells.

• TCF-DNs was prepared for test strip and was used for the qualitative detection of H₂S.

Abstract

With the merits of non-destructive, high penetration ability and minimizing autofluorescence, near-infrared (NIR) fluorescent probes have attracted much attention. In this paper, a NIR emission fluorescent turn-on probe THQ-L for H₂S was synthesized by the knoevenagel condensation between tetrahydroquinoxaline-6- formaldehyde derivative and 2-benzothiazoleacetonitrile. THQ-L can recognize H₂S through tandem reaction triggered by HS⁻ to construct 1,4-diethylpiperazine-modified iminocoumarin-benzothiazole, which produces a strong red fluorescent signal. THQ-L displayed an excellent selectivity toward H₂S, a large stokes shift (126 nm), a high signal-to-noise ratio (200-fold), the detection limits of 38.3 nM in PBS (10 mM, pH 7.4, 30% THF). The application study indicates that THQ-L can sensitively detect H₂S in red wine, natural waters, living cells and can be prepared for a test paper strip for the qualitative detection of H₂S.

Introduction

Hydrogen sulfide (H₂S) is considered as an environmental pollutant from industry, and can cause harm to the body after prolonged exposure to high concentrations of H₂S [1,2]. The acceptable ceiling concentration of H₂S permitted in workplace air in China is 10 ppm (15 mg/m³), a level established by National Health Commission of the People’s Republic of China (GBZ2.1-2019). On the other hand, H₂S is also an important endogenous signaling molecule and plays a crucial role in human health [[3], [4], [5]]. If the H₂S levels in the brain and blood are too high, it will cause related diseases such as hyperglycemia, down syndrome and Alzheimer’s disease, etc. [[6], [7], [8]] Therefore, there is great interest in developing specific detecting methods for H₂S in environmental and biological specimens.

In recent years, the methods for detecting H₂S have been widely reported. Compared with traditional methods, the fluorescence method becomes powerful tools to detect H₂S because of its high sensitivity, non-invasiveness and ease of operation [[9], [10], [11], [12], [13], [14]]. Until now, four strategies are commonly used in designing H₂S fluorescent probe including the reduction of azide or nitroso [[15], [16], [17], [18], [19], [20]], nucleophilic addition of H₂S [[21], [22], [23], [24]], thiolysis of leaving groups by H₂S [[25], [26], [27], [28], [29], [30]], Cu²⁺ removal of Cu²⁺ complexes [[31], [32], [33]]. Various fluorophores have been used to develop fluorescent probes, among which iminocoumarin-benzothiazole (ICBT) fluorophore has attracted much attention because of its high photostabilities and high fluorescence quantum yields [34]. Since the first ICBT scaffold-based probe was developed by Ghosh et al. in 2011 [35], various ICBT-based probes have been developed for recognition of different species including F⁻ [36], ONOO⁻ [37,38], Hg²⁺ [39], CO [40], biothols [41], alkaline phosphatase [42], hydrazine [43], diethyl chlorophosphate [44], F⁻ and Cu²⁺ [45]. To our knowledge, only two probes have been developed to recognize H₂S using ICBT framework. In 2015, Sun et al. reported the first fluorescent probe with high specificity for H₂S based on diethylamino-ICBT scaffold utilizing a H₂S-induced tandem reaction containing reduction, elimination and cyclization (Scheme 1, top) [46]. Although the probe showed good sensing properties toward H₂S and images H₂S in living cells, shorter emission wavelength is still an obvious defect, which is also a common problem in fluorescent probes constructed with ICBT framework. It is well known that near-infrared (NIR) fluorescent probe is more favourable on account of the deep tissue penetration, low autofluorescence background and less photo-damage to biological samples [[47], [48], [49], [50], [51], [52], [53]]. Therefore, the development of H₂S fluorescent probe with NIR emission has been the focus of our attention [54].

In 2019, Feng et al. reported the NIR fluorescent probe for H₂S based on 1,4-diethylpiperazine-modified ICBT skelecton utilizing strategies of protection and deprotection on imino group [55]. This work provides a good idea to achieve NIR emission by embedding 1,4-diethylpiperazine segment. However, this method still holds some limitations, such as the dissatisfactory variability of probe limited by the reaction activity of imino group. It is unfavorable to make full use of the NIR structure to develope more probes for meeting the needs of multiple detecting objects. Since a large number of reactive recognition groups are available to develop fluorescent probes by etherifying or esterifying on phenolic hydroxyl groups, such as allyl, tert-butyldimethylsilyl, (4-azidophenyl)methylene, 2,4-dinitrobenzenesulfonyl, 2,4-dinitrophenyl, 4-(bromomethyl)benzeneboronic acid pinacol ester, diethyl chlorophosphate etc. If incorporate a reactive recognition group on the phenolic hydroxyl group in advance, and the tandem reaction caused by the targeted species can lead to the generation of ICBT skelecton, the strategy will be very advantageous for the deep utilization of ICBT-based fluorophore.

Based on this view, we synthesized a novel non-fluorescent compound THQ-L and utilized two steps tandem reaction (thiolysis and cyclization) triggered by HS⁻ to build 1,4-diethylpiperazine-modified ICBT fluorophore (Scheme 1, down), which showed the significant fluorescence enhancement. This probe detects H₂S not only with fluorescent turn-on responding, NIR emission (652 nm), a large Stokes shift (126 nm) and a high signal-to-noise ratio (200-fold), but also it enables effective detection of H₂S in water, wine sample, and visualization of H₂S in living cells. We believe that the strategy of constructing NIR fluorophore by tandem reaction is more universal, and through replacing the protecting group, more different NIR fluorescent probes can be developed [56].