Nitrogen-doped carbon dots for sensitive detection of ferric ions and monohydrogen phosphate by the naked eye and imaging in living cells

Abstract

Nitrogen doped carbon dots (N-CDs) have been prepared via a one-pot hydrothermal method by using formamide and o-phenylenediamine as the carbon precursors. The as-fabricated N-CDs display excellent water dispersibility, good biocompatibility and anti-photobleaching properties. A strong emission band with an emission maximum (λ ^fl _max) of 556 nm is observed under 450 nm excitation, and a large Stokes shift of 106 nm is presented. However, the fluorescence is quenched by the addition of Fe³⁺; a good linearity is shown in the range of 0-65 μM with a detection limit as low as 0.85 μM. Fortunately, the quenched fluorescence could be recovered rapidly by the addition of monohydrogen phosphate (HPO₄ ^2-) due to the formation of the stable [N-CDs-Fe³⁺-HPO₄ ^2-] complex, and a good linearity is exhibited in the range of 0-60 μM with a low detection limit of 0.80 μM for HPO₄ ^2-. A novel "on-off-on" fluorescence response is seen with an obvious color change from yellow-crimson-yellow by the naked eye. In addition, the confocal microscopy images suggest that the as-synthesized N-CDs could serve as a sensitive nanosensor for Fe³⁺ and HPO₄ ^2- detection, implying the diverse potential application of N-CDs in the biomedical field.

Scheme 1. Schematic illustration of the N-CDs as a nanosensor for the detection of Fe³⁺ and HPO₄²⁻ with obvious “on–off–on” fluorescence behaviour in aqueous solution.

Fig. 1. (a) TEM image of the N-CDs (inset: corresponding HRTEM image). (b) The size distribution of the N-CDs.

Fig. 2. (a) Absorption spectra of the N-CDs in pure water (0.04 mg mL⁻¹) with increased concentrations of Fe³⁺ (0–140 μM). The inset shows the image of the N-CDs in pure water with the increased Fe³⁺ from left to right under daylight. (b) Linear relationship between absorbance and the concentration of Fe³⁺.

Fig. 3. (a) Fluorescence emission spectra of the N-CDs in pure water (0.04 mg mL⁻¹) with the different concentrations of Fe³⁺ (0–140 μM) under 450 nm excitation. The inset shows the image of the N-CDs in pure water with the increased Fe³⁺ from left to right under UV lamp irradiation at 365 nm. (b) Linear relationship between [(F₀ − F)/F₀] and the concentrations of Fe³⁺, where F₀ and F are the fluorescence intensities of the N-CDs at 556 nm in the absence and presence of Fe³⁺, respectively.

Fig. 4. (a) Fluorescence emission spectra of the N-CDs–Fe³⁺ complex in the presence of different concentrations of HPO₄²⁻ from 0 to 140 μM at an excitation wavelength of 450 nm. The inset is the image of the N-CDs–Fe³⁺ complex in pure water with the increased HPO₄²⁻ from right to left under UV lamp irradiation at 365 nm. (b) Linear relationship between [(F − F₀)/F₀] and the concentration of HPO₄²⁻, where F₀ and F are the fluorescence intensities of the N-CDs–Fe³⁺ complex at 556 nm in the absence and presence of HPO₄²⁻, respectively.

Fig. 5. (a) Absorption spectra of the N-CDs–Fe³⁺ complex in pure water with the different concentrations of HPO₄²⁻ (0–140 μM). The inset is the image of the N-CDs–Fe³⁺ complex in pure water with the increased HPO₄²⁻ from right to left under daylight. (b) Linear relationship between ratiometric absorbance (A₄₁₅/A₄₅₀) and the concentration of HPO₄²⁻.

Fig. 6. Relative fluorescence intensity of the N-CDs (0.04 mg mL⁻¹) in the presence of 140 μM Fe³⁺, and 500 μM of K⁺, Na⁺, Mg²⁺, Zn²⁺, Ca²⁺, Ba²⁺, Cd²⁺, Cu²⁺, Sr²⁺, Pb²⁺, Hg²⁺, Co²⁺, Mn²⁺, Fe²⁺, Al³⁺ and Cr³⁺ in an aqueous solution (pH 7.0), where F₀ and F are the fluorescence intensities of the N-CDs in the absence and the presence of various metal ions, respectively. The excitation wavelength is 450 nm.

Fig. 7. Confocal fluorescence images of HEp-2 cells. (a) Incubation with 0.50 mg mL⁻¹ of the N-CDs. (b) Followed by incubation with 140 μM of Fe³⁺ solution. (c) Further subsequent incubation with 140 μM of HPO₄²⁻ solution. (d–f) Bright-field images. (g–i) Merged images. (j) Mean fluorescence intensities of HEp-2 cells under different incubation conditions (data are presented as mean ± SD with replicates (n = 3)).