Phenylboronic acid derivative-modified (6,5) single-wall carbon nanotube probes for detecting glucose and hydrogen peroxide

Abstract

In this paper, we presented a new method for constructing near-infrared fluorescence probes and their applications in detecting glucose and hydrogen peroxide (H₂O₂). We used purified (6,5) single-wall carbon nanotubes (SWCNTs) separated by a polyethylene glycol/dextran aqueous two-phase system as the basis for near-infrared probes. Different phenylboronic acids were used for non-covalent modification of SWCNTs (6,5). Glucose was detected by the specific binding of the boronic acid group with cis-diol. Hydrogen peroxide was detected by horseradish peroxidase (HRP) combined with phenylboronic-acid-modified SWCNTs. The results revealed that the fluorescence intensity of purified SWCNTs was significantly enhanced without other chiral nanotube interactions compared to that of the raw SWCNT material. The fluorescence responses of 3-carboxy-5-nitrophenylboronic acid-modified purified CNTs could be used to effectively measure glucose in the concentration range from 0.01 mM to 0.50 mM with an interval linear index of R ² = 0.996 (LOD = 1.7 μM, S/N = 3). The detected H₂O₂ with the 3-aminobenzeneboronic acid-modified (6,5) SWCNT with HRP was in the concentration range from 5.0 μM to 40 μM with an interval linear index of R ² = 0.997 (LOD = 0.85 μM, S/N = 3). Moreover, this sensor exhibited a strong anti-interference effect without biological matrix fluorescence effects above 1000 nm wavelength. Thus, the proposed novel near-infrared fluorescence probes employed for reducing the interference of the biomatrix in this region are expected to be used in subsequent biosensor investigations.

Scheme 1. Diagram of glucose and H₂O₂ detecting complex. The phenylboronic acid derivative lies on the sidewalls of the nanotube and initially quenches SWCNT. In the presence of diols, the hydroxyl of the boron can bind with 1,2-diols to form diol–phenylboronic complexes with five-membered rings, which changes the photoluminescence intensity of SWCNT. Besides, HRP can specifically bind H₂O₂, which can also change the fluorescence of the functionalized SWCNT. The change in the concentration of H₂O₂ can be reflected in the change in fluorescence by utilizing this feature.

Fig. 1. UV-VIS-NIR spectrum (a) and fluorescence spectrum (b) that compare the purified SWCNTs (purple) and unsorted SWCNTs (black). In the fluorescence spectrum, the concentration of purified SWCNTs (6,5) is 0.06 mg mL⁻¹ after calculation according to absorbance value (Fig. S1†), and the concentration of unsorted SWCNTs is 0.33 mg mL⁻¹ after calculation according to absorbance value (dispersed in 1% SDS solution). Excitation at 560 nm.

Fig. 2. Representative fluorescence spectra that compare the original spectra of the two kinds of SWCNTs (purple and black), the spectrum after adding 5-nitro-3-carboxyphenylboronic acid to the two SWCNT solutions (red and magenta), and the spectrum after adding glucose to the two PBA-SWCNT complex solutions (blue and dark yellow). Excitation at 560 nm.

Fig. 3. Screening of the reactivity of SWCNTs (6,5) with 17 phenylboronic acid derivatives and the fluorescence response of the 17 phenylboronic acid-functionalized purified SWCNTs (6,5) to glucose. (a) Fluorescence intensity change (red) of purified SWCNTs (6,5) after the addition of 17 different phenylboronic acid derivatives. (b) Fluorescence intensity change (blue) of purified SWCNTs (6,5) after the subsequent addition of 0.2 mM glucose to the 17 phenylboronic acid derivative-functionalized purified SWCNT (6,5) complexes shown in (a). Excitation at 560 nm and emission wavelength at 1010 nm.

Fig. 4. (a) Fluorescence spectra of the 5-nitro-3-carboxyphenylboronic acid-functionalized SWCNTs (6,5) at various glucose concentrations. (b) Linear plots of the reduced intensity of the SWCNT (6,5) fluorescence as linear relationship of glucose concentration, R² = 0.996. I₀ and I are the corresponding fluorescence intensities of SWCNTs in the absence and presence of the analytes. Excitation at 560 nm and emission wavelength at 1010 nm.

Fig. 5. SWCNT-based system for various potential interfering substances. The concentrations of various inorganic salts (from left to right) are 0.16, 5.6, and 0.04 mM. The concentrations of various amino acids (from left to right) are 6, 6, 14, 8, and 30 μM. The concentrations of various nucleosides (from left to right) are 0.2, 200, and 0.1 nM. The concentrations of uric acid and glucose are 12 and 200 μM, respectively. The concentrations of potential interfering substances correspond to 25-fold dilution of those in serum considering the proper concentration of glucose detected by this sensor.

Fig. 6. SWCNT fluorescence responses of five kinds of phenylboronic acids to HRP modification and subsequent detection of H₂O₂. The black column shows the effect of HRP on the fluorescence of five phenylboronic acid-modified SWCNTs (6,5). The purple column shows the effect of H₂O₂ on the fluorescence of five PBA-HRP-modified SWCNTs (6,5).

Fig. 7. (a) Representative fluorescence spectra that compare two kinds of SWCNTs detecting H₂O₂, the spectrum after adding horseradish peroxidase to two SWCNT solutions (purple and black), and the spectrum after adding 40 μM H₂O₂ to the two 3AMBA-HRP-SWCNT complex solutions (red and blue). (b) Fluorescence spectra of SWCNTs (6,5) mixed with 3AMBA, 3AMBA + HRP, 3AMBA + HRP + H₂O₂, and 3AMBA + H₂O₂. The concentration of H₂O₂ was 40 μM.

Fig. 8. (a) Fluorescence spectra of the 3AMBA-HRP-functionalized SWCNTs (6,5) at various H₂O₂ concentrations. (b) Linear plots of the reduced intensity of the SWCNT (6,5) fluorescence as linear relationship of H₂O₂ concentration, R² = 0.997. I₀ and I are the fluorescence intensities of the SWCNTs in the absence and presence of analytes, respectively.