Study on UV-VIS absorption spectra of hydrogen sulfide by probe DMT
In order to evaluate the spectral properties of probe DMT on hydrogen sulfide, the UV-visible absorption spectra after the interaction between probe DMT and S2− were tested, as shown in Fig. 1.
As can be seen from Fig. 1, the maximum absorption wavelength of probe DMT is at 480 nm. After the addition of S2−, the maximum absorption peak at 480 nm gradually decreased, and the peak at 378 nm also gradually decreased, and a new strong absorption peak appeared at 600 nm, indicating that S2− reacted with the probe DMT, changing the original conjugate structure of the probe molecules leading to these phenomena.The methylquinoline group has fluorescence quenching effect on the probe, and the reaction of S2− with the probe causes the probe to restore the intramolecular ICT effect, and the intramolecular charge distribution occurs a new layout. When the concentration of S2− reached 500 µM, the absorption peak intensity did not change. After the probe reacted with S2−, the color of the solution changed from yellow to blue, and the change could be directly observed by the naked eyes.
Study on fluorescence spectra of hydrogen sulfide by probe DMT
The fluorescence spectra of the interaction between probe DMT and S2− are shown in Fig. 2a. Probe DMT shows almost no fluorescence in PBS buffer, while the probe showed an emission band centered at 680 nm (λex = 590 nm) after interacting with S2−. With the increase of S2− concentration, the fluorescence intensity at λem = 684 nm gradually increased by about 60 fold, which was attributed to the recovery of intramolecular ICT effect. When the concentration of S2− reached 500 µM, the fluorescence intensity hardly changed. As can be seen from the small figure in the upper right corner, the product solution after the reaction of probe DMT and S2− was purple red fluorescence. The relationship between fluorescence intensity and concentration of S2− is shown in Fig. 2b, and the concentration of S2− below 500 µM has a good linear relationship (R2 = 0.9926, K = 2.1706).
Stability of probe DMT for S2-
In order to explore the stability effect of the probe in buffer solution (EtOH:PBS = 1:1), the fluorescence intensity of DMT in different pH environments was measured, as shown in Fig. 3. The probe was almost unaffected by the pH of the environment, and the fluorescence intensity of the solution was enhanced with the addition of S2−. As can be seen from Fig. 3, in a strongly acidic environment, the fluorescence intensity of the probe did not increase significantly after adding S2−, but showed obvious fluorescence enhancement in an environment with pH = 6.5–10.5. This indicates that probe DMT can be used as a detection tool for in vivo cell imaging and biological imaging applications.
Reactivity of probe DMT to hydrogen sulfide
Time is a key factor to evaluate the properties of fluorescent probes. In order to study the reaction performance of probe DMT to S2−, the response time of probe and its reaction was measured, as shown in Fig. 4. It can be seen from Fig. 4 that the probe can respond to S2− in a very short time and reach an equilibrium state at 10 s.
Selectivity of probe DMT to hydrogen sulfide
Selectivity is one of the important parameters to evaluate the performance of fluorescent probes. In this paper, Leu, Val, Phe, Ala, Met, Arg, Pro, Gly, CaCl2, MgCl2, KCl, NaCl, NaBr, Na2CO3, NaNO2, NaF, Na2SO4, BaCl2·2H2O, Na2S·9H2O, Na2S2O4, NaHSO3, GSH, Cys and Hcy were analyzed to explore the selectivity of probe DMT and its interaction, as shown in Fig. 5.
As can be seen from Fig. 5, after adding S2−, the probe DMT solution showed obvious fluorescence enhancement at 684 nm, and the fluorescence intensity increased slightly after adding GSH. Since the probe DMT solution added with GSH has no significant change under visible light and 365 nm UV lamp, and there is a significant difference in fluorescence enhancement ratio, it can be well distinguished between the two. The addition of other anions, cations, amino acids and other biological mercaptans had no obvious effect. The results show that probe DMT has good selectivity for S2−.
The anti-interference of probe DMT to S2-
As shown in the Fig. 6, there was no obvious fluorescence change in probe DMT solution after adding other analytical species except S2−. After adding 500 µM S2− solution to different solutions, the fluorescence intensity of probe DMT at 684 nm showed obvious enhancement, and the fluorescence intensity was only slightly different from that of only 500 µM S2− solution. Therefore, it can be shown that the probe DMT has excellent selectivity to S2− and strong anti-interference to other analytical substances.
Sensing mechanism study
Through theoretical analysis and literature investigation, it is speculated that the nucleophilic addition reaction occurs between H2S and the double bond on the pyridine ring in the probe molecule DMT (Fig. 7a), and the fluorescence is turned on [33, 34]. The proposed mechanism was verified by measuring the molecular weight of the probe DMT reacting with H2S. As shown in the figure, after adding 250 eq H2S to fully combine it with the probe molecule, the mass spectrum peak of the reaction product [DMT + H2S + Na+] was measured to be m/z = 448.0657, and its theoretical value was 448.0918, both values were basically consistent (Fig. 7b).