Figure 3 (A) Fluorescence intensity of CDs solutions in the presence of different pH; (B) Fluorescence spectra of CDs with varied pH ranging from 2 to 7; (C) Linear relationship between fluorescence intensity of CDs and pH; (D) Zeta potentials of CDs, (E) UV-vis spectra of CDs and (F) FTIR patterns of CDs with different pH from 2 to 7.
3.5. Establishing the assay strategy of tigecycline by CDs
Based on the favorable fluorescence of CDs prepared here, we addressed that whether they could serve as the fluorescent probe towards the potential targets. As shown in Figure 4A, the fluorescence of CDs was quenched when tigecycline was introduced, proving that these CDs showed the potential for sensing tigecycline. Meanwhile, the fluorescence intensity of CDs gradually decreased with the concentration of tigecycline increasing from 7 nM to 200 μM (Figure 4B), and the linear equation was fitted as F0/F-1 = 3.0642CTGC - 0.0422 with R2 of 0.992, while the detection limit was calculated as 3.56 nM (Figure 4C). These findings suggested that we successfully built up a strategy of detecting tigecycline by CDs described here.
3.6. Optimizing the conditions of CDs detecting tigecycline
To acquire the optimal detection conditions of tigecycline by CDs, the further experiments were designed and performed. As shown in Figure S3A and S3B, the incubation temperature and time scarcely showed the effect on the detection processes, thus 10 minutes of incubation at room temperature were selected as the optimized conditions of CDs assaying tigecycline.
3.7. Anti-interference and selectivity for CDs detecting tigecycline
To investigate the anti-interference ability of the proposed assay, tigecycline were initially added into the paralleled solutions of CDs, and then the substances that might exist in the surrounding environment, including Na+, K+, Ba2+, Ca2+, Mg2+, SO42-, glucose, cysteine, arginine, histidine, lysine, urea and uric acid, were respectively introduced. As shown in Figure 4D, the fluorescence intensity of CDs and tigecycline in the absence or presence of these substances were recorded, and the results described that their fluorescence intensity were hardly affected by other substances, revealing the generally satisfactory anti-interference. In addition, the selectivity of CDs detecting tigecycline was explored by examining the effect of the similar or other classes of antibiotics on their fluorescence intensity. The results showed that the fluorescence of CDs only responded to tigecycline with a concentration of 200 μM rather than any other antibiotic introduced (Figure 4E), demonstrating the favorable selectivity during sensing tigecycline by CDs prepared here.
3.8. Detection of tigecycline in the lake water sample
To evaluate the practicability of CDs detecting tigecycline, the lake water served as the actual samples by the established assay. In particular, the lake water was supplemented with tigecycline with different concentrations, and the concentrations of tigecycline in the lake water were separately calculated according to the proposed equation. Again, the recoveries of tigecycline were examined as 96.0% and 108.0% (Table S1), demonstrating that the fluorescent method we raised here exhibited the satisfactory practicality.
3.9. Detection mechanism of tigecycline by CDs
Additionally, we also explored the specific mechanism of CDs sensing tigecycline. Particularly, the UV spectrum of tigecycline did not overlap with that of CDs (Figure 4F), thus excluding the internal filtration effect. Then, the UV spectra of TGC, CDs and CDs with TGC were obtained, and the absorption band of CDs with TGC differed from that of TGC or CDs only, revealing that there appeared the new substance through the interaction between CDs and tigecycline (Figure 4G). Meanwhile, there appeared no obvious variation for the fluorescence lifetime of CDs with tigecycline compared with that of CDs only (Figure 4H). Taken together, these findings demonstrated that the quenched fluorescence of CDs was mainly associated with the static quenching. In addition, the broadening of the absorption band in the range of 2800-3750 cm-1 was observed after introducing tigecycline into CDs compared with that of tigecycline, suggesting that there may form the new substance between CDs and tigecycline (Figure 4I), which was consistent with our previous findings. Therefore, the static quenching was mainly responsible for their reduced fluorescence intensity of CDs [34,35].