Downconversion Luminescence-Based Nanosensor for Label-Free Detection of Explosives.

We report a selective and sensitive nanosensor probe based on polyethylenimine (PEI)-capped downconverting nanophosphors β-NaYF4:Gd3+,Tb3+@PEI for the detection of 2,4,6-trinitrotoluene (TNT), both in water and buffer media. These downconverting phosphors were synthesized via a hydrothermal route and are known to show excellent chemical, thermal, and photostability. They emit sharp emission peaks centered at ∼488, 544, 584, and 619 nm, among which the peak at ∼544 nm was remarkably quenched (∼90%) by the addition of TNT without giving any new emission peak. The sensing mechanism is based on the formation of a Meisenheimer complex between the electron-rich amine-functionalized β-NaYF4:Gd3+,Tb3+ nanophosphors and electron-deficient TNT molecule, which was prominently visualized by the change in the color of the solution from whitish to brownish yellow, enabling visual detection, followed by luminescence resonance energy transfer between the nanophosphors and the complex. A linear range for TNT detection was obtained from 0.1 to 300 μM with a limit of detection as low as 119.9 nM. This method displayed excellent selectivity toward TNT over other nitroaromatic compounds, which had no influence on the detection. Moreover, various other classes of analytes, viz., amino acids, pesticides, and sugars, did not quench the luminescence intensity of the nanophosphors. This developed nanosensor probe possesses high, stable fluorescence brightness and capability for the selective and sensitive on-site recognition of TNT molecules in aqueous media, avoiding complicated strategies and instruments. Thus, this work promises to pave ways to many applications in the detection of ultratrace analytes.

Introduction

The riveting optical features of lanthanide ions such as narrow and sharp emission bandwidth, large Stokes and anti-Stokes shift, tunable emission spectra, long-lived emission generally in milliseconds, low autofluorescence, and reduced blinking arise from their electronic transitions within the 4f orbitals, hence giving them an edge over conventional fluorophores and quantum dots.[1] Having these advantages, recently, rare earth ion-doped phosphors have been emerging as a potential material in various applications such as solid-state lasers,[2] light-emitting devices,[3] solar cells,[4] sensing,[5] cell imaging,[6] drug delivery,[7,8] and so forth. One of the most propitious applications of these luminescent phosphors is their use as the optical probes for the identification of molecules such as glutathione,[9] glucose,[10] avidin,[11] and ions such as cyanide,[12] mercury,[13] and so forth for the sensing applications. Profound efforts have been devoted to the development of novel, innovative, and implicit sensors. Thus, these lanthanide ion-doped phosphors can be applied to the highly selective and sensitive real-time and on-site detection of the explosives.

Identification and quantification of explosives has constituted an emerging and important topic of interest. Reliable detection of trace amount of explosive substances is of critical importance concerning homeland security threats, military applications, mine-field analysis, forensic investigations, and so forth. Moreover, these compounds are known to have toxicity, carcinogenicity, mutagenicity, and their release into the environment from military sites and ammunition plants causes the contamination of water and soil.[14] Because of the risk associated with the environment, human and wildlife, the detection has gained increasing attention. The detection could help in reducing the fatalities among the civilians and health risk hazards.

The chemical structures of the commonly used nitrocompounds are shown in the Supporting Information in Figure S1. In recent years, a variety of analytical techniques and detection methods of explosives have been developed including gas chromatography,[15] ion mobility spectrometry,[16] surface-enhanced Raman spectroscopy,[17,18] conductivity-based techniques,[19] and so forth. However, these methods are usually limited by several intrinsic shortcomings; some require complicated instrumentation or synthetic processes, whereas others require labeling procedures, which are typically time-consuming, expensive, and require specialized personnel. Thence, to meet this need, the fluorescence quenching approach has been explored for the analysis of nitro-based explosives. Fluorescence-based sensing methods have gained immense attention because of the relatively lower operational costs than the conventional methods, simplicity, higher sensitivities, portability, short response times, and its pertinence in both solution and the solid phase. Various nanomaterials were used to develop a fluorescence-sensing platform for explosive sensing.[20−23] Although these various nanostructured materials are reported to construct a fluorescent probe for the selective detection of the explosives, there is still a challenge to utilize them as sensors because they possess low chemical and thermal stability, poor aqueous solubility, require time-consuming synthetic methods, are receptive toward photobleaching, and thus are insufficient for the detection of analytes in aqueous samples. Therefore, other classes of compounds based on the lanthanide ions have garnered attention for sensing applications in the past few years.

Because of good chemical, thermal, and photostability, the lanthanide ion-doped phosphors are highly desirable as sensors.[24] Moreover, long-lived luminescence of lanthanide ions could allow them to be potentially used for analyte detection in strongly autofluorescent biological matrices. These have been used in sensing of various hazardous analytes of environmental and clinical importance.[25−27] These phosphors have also been employed for sensing explosives via fluorescence-based detection up to a level of nanomolar concentration. In 2014, Ma and Wang reported label-free detection of 2,4,6-trinitrotoluene (TNT) using upconverting nanoparticles (NPs) NaYF4:Yb3+,Er3+ at pH = 12 with a detection limit of 9.7 ng/mL.[28] In the same year, a miniaturized device was fabricated using NaYF4:Yb3+,Er3+@PEI, which detected the TNT explosive by the formation of the Meisenheimer complex and quenched the green luminescence.[29] Although the mentioned methods utilize the selectivity of these materials over other analogous analytes, they involved multistep and time-consuming synthetic processes. Also, these sensors worked in the high pH range (12 and above), thus limiting their applicability as sensors. Nevertheless, β-NaYF4;Gd3+,Tb3+ downconverting nanophosphors, being highly luminescent, have rarely been explored for their potentiality as a fluorescent probe for the selective and sensitive detection of explosives.

In this contribution, we have developed a label-free method for the selective detection of TNT, in the aqueous solution of nitrocompounds, characteristic to explosives based on the green-emitting phosphors. These lanthanide ion-doped phosphors are functionalized with amine groups (NH2) using PEI via a hydrothermal route. The detection is based on photoluminescence (PL) method by observing the change in intensity at 544 nm of the green light which was selectively and dramatically quenched via luminescence resonance energy transfer (LRET)-based energy transfer from nanophosphors to the Meisenheimer complex, detailed mechanism of which is explained. Meanwhile, the intensity was not influenced by the addition of other nitrocompounds in the aqueous solution. Furthermore, no drastic change in intensity was observed with other category of analytes such as amino acids, pesticides, and sugars. The current developed sensing probe is also applicable in a wide pH range from 7 to 13. Combined with good water and chemical stability, photostability and wide pH adaptability will facilitate the application of this system as a potential nanosensor probe.

Results and Discussion

The composition, crystallinity, and phase purity of the NaYF4 nanophosphors were first checked by powder XRD as shown in Figure . The sample diffraction peaks can be indexed to NaYF4 with lattice parameters as a = 5.9 Å, and c = 3.5 Å, that is in good agreement with standard JCPDS data (JCPDS no. 16-0334), confirming the hexagonal phase of the as-synthesized product. The detailed structure analysis was carried out with the Rietveld refinement method using the general structure analysis system, GSAS–EXPGUI suit of programs.[30] The refinement proceeded smoothly with the Na1.5Y1.5F6 model (ICSD collection code no. 51917) and the corresponding profile is displayed in the Supporting Information in Figure S2. The lattice parameters and reliability factors are summarized in Table S1. The background was fitted well with a shifted Chebyschev function and all the parameters including unit cell parameters, occupancy, Gaussian–Lorentzian factors were refined. The transmission electron microscopy (TEM) and field emission scanning electron microscopy (FESEM) images (Figure a,b) show that the β-NaYF4:Gd3+,Tb3+@PEI particles prepared at 180 °C are grown in rod-shaped particles and exhibit uniform shapes and are monodispersed with an average diameter of ∼92 nm and length of ∼280 nm. The contrast in the TEM image observed was due to the different orientation of the crystalline nanoparticles with respect to the electron beam, which changed the elastic scattering diffraction and thus appeared in different contrast. Further, the chemical composition of the β-NaYF4:Gd3+,Tb3+ nanophosphors was characterized by energy-dispersive X-ray analysis (EDXA) where all the elements in the nanophosphors could be detected including doped Gd3+ and Tb3+ ions, confirming the presence of the lanthanide ions (Figure S3). Furthermore, the detailed structure analysis was done through high-resolution TEM (HRTEM) analysis explained in Figure S4a, which shows the HRTEM image of the pristine sample, which was subsequently analyzed by a combination of fast Fourier transform (FFT) followed by inverse FFT (IFFT) image to check crystal lattice periodicity. Figure S4b shows the IFFT image of the selected region (marked as the red square) in the HRTEM image, which was constructed after the masking of the (110) plane using the Digital Micrograph software as shown in the FFT image in the inset. The IFFT image clearly shows the presence of an array of ordered planes whose interplanar spacing was measured from the line profile and was 0.30 nm, corresponding to the (110) plane of the hexagonal NaYF4 lattice. The SAED pattern shown in Figure S4d revealed the crystalline nature of β-NaYF4:Gd3+,Tb3+@PEI NPs. The diffraction pattern obtained matched well with the crystal planes of hexagonal NaYF4 and have been assigned to the (110), (101), and (111) planes. Thus, d-spacing calculated in HRTEM and SAED are in agreement with those given in the standard JCPDS data no. 16-0334, confirming the hexagonal phase of the as-prepared samples.

Figure 1

XRD pattern of as-prepared β-NaYF4:Gd3+,Tb3+@PEI. The data have been compared with standard data of hexagonal NaYF4, JCPDS 16-0334 as the reference.

Figure 2

(a) TEM and (b) FESEM images showing the morphology of the β-NaYF4:Gd3+,Tb3+@PEI phosphor nanoparticles, which are monodispersed in nature, (c) FTIR spectra indicating the capping of PEI on the surface of the rods. Inset: Zoomed view in the range of 3000–2500 cm–1, (d) TGA results confirming the presence of PEI on the terminus. The decomposition temperature obtained for branched PEI when coated on the surface is ∼510 °C, as shown in the derivative curve of TGA in the inset. Bare β-NaYF4:Gd3+,Tb3+ phosphors without the capping of PEI, showed no weight loss at this temperature.

The functionalization of PEI was investigated by zeta potential of β-NaYF4:Gd3+,Tb3+@PEI NPs, which was +43 mV, suggesting the successful capping of PEI on the surface, rendering the rods to be positively charged. The presence of amino groups on the surface of rods was further verified by Fourier transform infrared (FTIR) spectroscopy (Figure c). The β-NaYF4:Gd3+,Tb3+@PEI nanophosphors exhibited an absorption band at 3446 cm–1 because of typical N–H stretching vibration of the amino groups, which overlaps with the O–H stretching band and its presence is supported by the peak ascribed to the bending mode of amino groups (−NH2) at 1656 cm–1, thus revealing the abundance of electron-rich amino groups on the terminal of the rods.[7] Moreover, the absorption band at 1398 cm–1 is attributed to the stretching vibrations of the C–N bond, whereas the bands at 2857 and 2937 cm–1 are attributed to the methylene symmetric and asymmetric C–H stretching vibrations as shown in inset of Figure c. The PEI capping is further confirmed by thermo gravimetric analysis (TGA) results, which were carried out in nitrogen atmosphere at a heating rate of 10 °C/min. As shown in Figure d, the initial weight loss of 5.5% occurred in the temperature range 300–550 °C due to the degradation of bound PEI present in the system and no further loss was observed till 650 °C, confirming the stability of the fluorides. The inset shows the first derivative of the TGA curve, confirming the presence of PEI in the system and degradation at 510 °C. The TGA spectra were recorded for bare β-NaYF4:Gd3+,Tb3+ nanophosphors, which showed a slight weight loss of 2.1 wt % in the 300–550 °C temperature range. It is believed that the decomposition temperature of branched PEI is situated at around 400 °C in nitrogen atmosphere,[31] which can be verified in the TGA results shown in Figure S5. The observed increase in the decomposition temperature of PEI in the β-NaYF4:Gd3+,Tb3+@PEI as compared to free PEI is attributed to the fact that the capping agent is protected from degradation because of its binding to the rods.[32] This confirms that the PEI is indeed bound to the nanorods.

The composition and chemical state of the β-NaYF4:Gd3+,Tb3+@PEI were ascertained by X-ray photoelectron spectroscopy (XPS) measurements. As shown in Figure a, the survey scan spectrum confirmed the presence of Na, Y, F, C, N, and lanthanides (Gd, Tb) in the material. It should be mentioned here that the presence of C 1s and N 1s in the spectrum was due to the capping of the PEI on the surface. The high-resolution peak analyses of Gd 3d and 4d core level spectra revealed that these energy levels are split into doublet because of spin-orbit coupling (Figure b,c). The energy levels 3d5/2 and 3d3/2 appeared at 1189.2 and 1220.9 eV, respectively, with spin–orbit splitting of 31.9 eV, whereas the peaks at 144.8 and 150.4 eV can be assigned to the binding energy of levels 4d5/2 and 4d3/2, respectively. The presence of Gd 3d and 4d peaks showed that the Gd ion exists as Gd3+ in the crystal lattice of hexagonal NaYF4. As shown in Figure d, the energy level of Tb 3d also split into doublet corresponding to 3d5/2 and 3d3/2 at 1243.7 and 1278.4 eV, respectively. Also, a satellite appeared between the two main peaks. These results suggested that the Tb ion substituted Y ion in NaYF4 appeared in only one oxidation state, i.e., +3. The magnified XPS spectra of other elements such as Na, Y, and F in β-NaYF4:Gd3+,Tb3+@PEI is shown in Figure S6, displaying the energy levels; Na 1s, Y 3d, and F 1s core levels. The surface functionalization of the downconverting nanorods conducted by XPS was in accordance with the FTIR results. The presence of these functional hydrophilic groups improved the water dispersibility and stability of the nanorods. To explore the local structure of β-NaYF4:Gd3+,Tb3+ crystals, Raman spectroscopy was recorded to analyze the vibrational properties of the sample under the laser 325 nm (Figure S7). The Raman spectra of β-NaYF4:Gd3+,Tb3+ crystals revealed the strongest phonon bands at 308, 367, 416 cm–1, which are in fair agreement with reported data of the NaYF4 lattice.[33] These bands are attributed to the host lattice vibrations of Y–F and Na–F distances and bond strength features.

Figure 3

(a) XPS survey spectra of as-synthesized β-NaYF4:Gd3+,Tb3+@PEI and high-resolution spectra of (b) Gd 3d, (c) Gd 4d, and (d) Tb 3d showing the binding energy of the core levels.

The optical emission spectra of β-NaYF4:Gd3+,Tb3+@PEI are displayed in Figure a, where the photon emission originates from the transition within the 4f electronic transitions of Tb3+ ions. The emission spectra of Tb3+ ions can be obtained at the excitation wavelength, λex = 375 nm.[34] The efficient emission is difficult to realize in the system containing Tb ions under direct excitation at 375 nm due to intraconfigurational parity-forbidden transitions. Hence, Gd3+ ions act as a sensitizer (i.e., light-harvesting antenna) to enhance the luminescence of Tb3+ ions.[7] As Gd3+ ions exhibit a strong absorption band at 273 nm because of their 8S7/2 → 6I11/2 transition corresponding to f–f transitions, nanophosphors containing Gd3+, Tb3+ ions together exhibit a very intense excitation band at 273 nm followed by a nonradiative (nr) energy transfer to Tb3+ ions. The emission peak of Gd3+ is observed at 311 nm in the UV region, which is assigned to the 6P7/2 → 8S7/2 transition under 273 nm irradiation.[35] Additionally, the obtained emission spectra excited at λex = 273 nm (Figure a) yielded emissions from Tb3+ ions in the range of 480–680 nm. Four prominent emission peaks centered at ∼488, ∼544, ∼584, and ∼619 nm originate from the transitions of 5D4 → 7F6, 5D4 → 7F5, 5D4 → 7F4, and 5D4 → 7F3, respectively. It is clear that among the above mentioned transitions, the green emission 5D4 → 7F5 at ∼544 nm is the most intense emission. The PL decay curve for the luminescence of Tb3+ in β-NaYF4:Gd3+,Tb3+@PEI nanorods is shown in Figure b. Here, we displayed the PL decay curve fitted over first 20 ms with biexponential decay functions because of the variation in decay rates of Tb3+ ions in the nanoparticles. This emissive biexponential decay is ascribed to the inhomogeneous distribution of Tb ions close to the surface, giving the short lifetime, and inside the nanoparticles with a long lifetime.[36] The curve was well fitted by decay equation: I = Io + A1(e–) + A2(e–), where I and Io are the intensities at times x and 0, respectively, A1 and A2 are constants, τ1 and τ2 are the decay times for the two exponential components, which were 0.35 and 2.50 ms for 5D4 → 7F5 electronic transition of Tb3+ under excitation at 375 nm. The average lifetime of these nanophosphors is in milliseconds, indicating the usability of these phosphors in biological applications also where the autofluorescence and background fluorescence can be suppressed offering high signal to noise ratio.[37]

Figure 4

(a) PL emission spectra of PEI-capped β-NaYF4:Gd3+,Tb3+ at an excitation wavelength of 273 nm yielding emission in the visible region corresponding to the Tb3+ ion transitions, (b) luminescence decay kinetics behavior for the emission of the 5% Tb3+ ion corresponding to transition 5D4 → 7F5 at 544 nm in the hexagonal NaYF4 lattice under excitation of 375 nm.

Detection of Explosives

Nitro group-based compounds, 1,3-dinitrobenzene (1,3-DNB), 2,4-dinitrophenol (2,4-DNP), 2,4,6-trinitrophenol (TNP), 2,4-dinitrotoluene (2,4-DNT), 2,6-dinitrotoluene (2,6-DNT), TNT, 1,3,5-trinitro-1,3,5-triazinane (RDX), 4-nitrophenol (4-NP), 1,4-dinitrobenzene (1,4-DNB), nitromethane (NM), 4-nitrotoluene (4-NT), and nitrobenzene (NB) were used in this experiment. The designed principle of detection is based on the interaction of the amine groups on the surface of the nanoparticles with the electron-deficient nitrocompounds. For comparison, the PL intensity of nanophosphors was detected in deionized water at ∼544 nm. Equal amounts of analytes were taken in a cuvette for the measurements as discussed in the synthesis section and compared against the PL intensity in the deionized water. It is interesting to note that among these explosive samples (Figure a), only TNT could quench the PL intensity of the nanophosphors. Figure b presents the quenching profile for all the analytes for the emission at 544 nm. One can see in the figure that the quenching efficiency for 300 μM TNT is about 90%, whereas all other analytes contributed little changes in the luminescence intensity. On the basis of the results mentioned, TNT was recognized as a detection target owing to its ability to quench the luminescence dramatically, suggesting the high sensitivity of the sensing probe. As a result of the significant quenching, we next studied the quenching phenomenon with varied concentrations of TNT (Figure c). As shown in Figure d, under the optimum conditions in our experimental procedure, the PL quenching was analyzed using the Stern–Volmer (SV) plot between the change in PL intensity at 544 nm and the concentration of TNT. A significant linear correlation (R2 = 0.998) existed between the relative PL intensity (Io/I) and the concentration of TNT in the range of 0.1–300 μM. The quenching efficiency was investigated using S–V equation, Io/I – 1 = Ksv [Q], where Io is the PL intensity in the absence of analyte, I is the PL intensity in the presence of analyte with the molar concentration [Q], and Ksv is the quenching constant in M–1. The quenching constant was determined to be 3.32 × 104 M–1. The limit of detection (LOD) of TNT can be found using the linear regression method and calculated using the formula 3σ/m, where 3 is the factor of 99% confidence level, σ is the standard deviation of the measured intensity for the blank β-NaYF4:Gd3+,Tb3+@PEI nanophosphors (n = 5), whereas m denotes the slope of the linear calibration curve. It should be mentioned here that the LOD was found to be 119.9 nM (27.2 ppb), revealing higher sensitivity of the as-synthesized nanophosphors compared to the other earlier reported results (Table S2).

Figure 5

(a) Emission spectra and (b) quenching efficiency of the samples containing β-NaYF4:Gd3+,Tb3+@PEI and different nitrocompounds (300 μM) in aqueous solution, (c) emission spectra of β-NaYF4:Gd3+,Tb3+@PEI containing different concentrations of TNT (0–300 μM) in aqueous solution, (d) fitting graph of the linear S–V plot for TNT by equation; Io/I – 1 = Ksv[Q], where Ksv is quenching constant and found out to be 3.32 × 104 M–1. The error bar is calculated from three parallel samples.

Interference Study

To assess the possibility of the analytical application of our phosphor-based nanosensor in terms of the sensitivity, the effect of different mixed samples of analogous compounds of TNT such as 2,4-DNT, 2,6-DNT, 4-NP, 1,3-DNB, TNP, 2,4-DNP, RDX, 1,4-DNB, NM, 4-NT, and NB: 300 μM on the quenching of PL intensity was also studied and the results are shown in Figure a. In spite of the presence of other analytes, quenching of the PL intensity owing to the TNT was observed, having no interference because of the other analytes. Therefore, they show high sensitivity and selectivity for the detection of specific nitrocompounds. The TNT quenching was also observed in the buffer solution, NaH2PO4–Na2HPO4 (pH 7.0). Furthermore, we scrutinized the effects of pH on the PL intensity before and after the addition of TNT. In the wide range of pH 7–13, it showed no variation in the luminescence intensity (Figure S8a), suggesting the stability of the nanophosphors over the wide range of pH in the neutral and alkali environments. In addition, the influence of incubation time on the luminescence intensity was also studied to ensure the quenching solely because of TNT. As seen in Figure S8b, the intensity at 544 nm was quenched with the addition of TNT after 10 min. Meanwhile, by prolonging the incubation time to 48 h, the PL intensity was almost unchanged. Moreover, the PL intensity remained unchanged for almost 8 months, when stored at room temperature. This demonstrates that these nanophosphors can detect analytes over a long period of time, and thus are stable in nature. It should be mentioned here that no quenching was observed when only β-NaYF4:Gd3+,Tb3+ was used for the PL measurements without being capped by PEI (Figure S9). Therefore, we can say that the interaction of the TNT molecule with amino groups plays an important role.

Figure 6

(a) Histogram demonstrating the sensitivity of the phosphors over other analogous compounds of TNT, (b) UV–visible absorbance spectra of all the nitrocompounds in the presence of β-NaYF4:Gd3+,Tb3+@PEI in aqueous medium. The Meisenheimer complex formed between TNT- and PEI-capped phosphors shows the broad absorbance in the visible range till 650 nm, (c) spectral overlap of emission spectra of β-NaYF4:Gd3+,Tb3+@PEI and absorbance spectra of β-NaYF4:Gd3+,Tb3+@PEI before and after the addition of TNT, (d) time-resolved decay dynamics of 5D4 → 7F5 transition of Tb3+ ions in β-NaYF4:Gd3+,Tb3+@PEI after excitation at 375 nm in the presence of TNT (1, 10, 100, 300 μM), (e) photograph showing the complex formation, which is confirmed by the change in the color of the solution to brownish yellow containing TNT, whereas other samples look white or pale yellow.

Mechanism of Highly Selective Detection

To unravel the origin of the high selectivity of β-NaYF4:Gd3+,Tb3+@PEI nanophosphors toward TNT, the quenching mechanism was investigated. A plausible mechanism for the detection could be proposed to be based on the donor–acceptor interaction between amino groups of phosphor nanorods and TNT. As TNT is an electron-deficient compound because of the presence of three electron-withdrawing nitro groups, it acts as an electron–acceptor, whereas amino groups are electron donors. Thus, an electron transfer mechanism takes place between TNT and amino groups of the PEI on the surface of nanoparticles, which leads to the formation of the Meisenheimer complex (Scheme ). The three electron-withdrawing nitro groups present at the ortho- and para-position in TNT enables the nulceophile amino groups to attack at position 1 (where the methyl group is attached) in the molecule, which leads to the formation of this stable anionic σ-complex where the negative charge is delocalized over the ring.[38] The formation of such a complex with amino groups is typical of nitrocompounds, especially TNT. This complex formation can be confirmed by the change in the color of the solution of nanophosphors containing TNT to brownish yellow (Figure e), whereas there was no color change in the case of 2,4-DNT, 2,6-DNT, 4-NT, 1,3-DNB, 1,4-DNB, NM, and RDX. A pale yellow color appeared in the case of 2,4-DNP, 4-NP, NB, and TNP, which can be easily distinguished from the brownish yellow color of TNT. The UV–vis spectra of the explosives in the presence of PEI-capped nanophosphors were recorded and are shown in Figure b. As it can be seen, all other nitrocompounds have a strong absorbance before 400 nm, and hence do not interfere in the quenching. Meanwhile, it should be mentioned that the Meisenheimer complex formed with the TNT and nanophosphors has a broad absorbance in the range 400–650 nm, which overlaps with the emission of the Tb3+ ions in the phosphors (Figure c). As the emission from the nanoparticles overlaps with the absorbance of the complex formed, it gives the possibility of LRET-based energy transfer, thereby leading to the quenching of the green emission. As mentioned above, when lanthanide ion-based nanoparticles are used as the energy donor, the mechanism is named as LRET, whose principles are similar to fluorescence resonance energy transfer and defined as an nr optical energy transfer between a donor D in its excited state and proximal ground state acceptor A through long-range dipole–dipole interactions.

Scheme 1

Schematic Representation of the Formation of the Meisenheimer Complex Due to the Interaction between the Amino Group Present on the Surface of the Phosphor Nanoparticles and TNT through the Charge-Transfer Mechanism in Aqueous Medium

For effective energy transfer, an acceptor absorption spectrum should overlap with a donor emission spectrum and the donor and acceptor should be linked in close proximity.[39] In Figure b, it is observed that the absorption spectrum of β-NaYF4:Gd3+,Tb3+@PEI + TNT largely overlaps with the emission spectra of the β-NaYF4:Gd3+,Tb3+@PEI phosphors, whereas there is no overlap with other analytes, thereby avoiding energy transfer. Although there is a suitable spectral overlap between the nanophosphor emission and the complex absorption in the UV–vis spectra, LRET cannot occur unless the donor and acceptor are close enough. Because of the formation of the complex between TNT and amine groups, it is brought close enough for the energy transfer to take place. Thus, the spectral overlapping between downconversion emission of β-NaYF4:Gd3+,Tb3+@PEI and absorbance of the Meisenheimer complex, and the close distance between the two manifests the LRET efficiency of the sensor. Meanwhile, in order to ascertain the above mentioned reason, we explored the lifetime of nanophosphors with varied concentrations of TNT. As displayed in Figure d, the lifetime of the β-NaYF4:Gd3+,Tb3+@PEI phosphors decreased with increasing concentrations of TNT, and average lifetimes were considered for consistency in results. The decay was fitted using a biexponential decay function to yield average lifetimes of 2.11, 1.91, 1.72, 1.49, and 0.98 ms under the TNT concentrations of 0, 1, 10, 100, and 300 μM, respectively. Table S3 shows the lifetime parameters, viz. τ, rel %, and average lifetime of the samples. These results excluded the possibility of inner filter effect; thus, the lifetime reduction indicated that LRET is the major process[40,41] between nanophosphors and the TNT complex as it offers the additional relaxation pathway. As mentioned above, the nanosensor exhibited high selectivity for the target TNT over other nitrocompounds with similar structures such as TNP, DNT, DNP, NP, and DNB. Other nitrocompounds are much weaker electronic acceptors when compared with TNT because of lack of electron-withdrawing nitro groups, and thus are not likely to form an effective Meisenheimer complex with the amine groups present on the surface of the nanoparticles.[42] Although weak interactions may lie between the amine groups and these compounds, no significant absorption peaks are detected in the UV–vis spectra in the spectral range of 400–700 nm; thus, they do not interfere with the emission of the downconverting nanophosphors. The proposed method demonstrates the feasibility of the PEI-capped β-NaYF4:Gd3+,Tb3+ phosphor nanoparticles’ application for preferentially detecting TNT as the analyte.

Interference from Other Analytes

In order to illustrate the good selectivity, sensitivity, and feasibility of the developed nanosensor probe for the detection of TNT, the effects of other analytes such as pesticides, sugars, and amino acids were also checked on the luminescence quenching profile and the results are shown in Figure . The stock solution of 300 μM of the following amino acids was prepared in water: phenylalanine, isoleucine, tryptophan, glutamine, glutamic acid, and histidine. The structures of all the analytes are given in Figure S10. Interestingly, among the amino acids, none of them showed any obvious quenching effects in the intensity of the nanophosphors. This is due to the groups present in the structure of the amino acids, which did not interact with the amine group in the PEI and thus are not able to form a complex with the nanorods. Similarly, stock solutions of some pesticides such as chloropyrifos, malathion, cypermethrin, copper oxychloride, fenvalerate, and carbendazim, and sugars such as glucose, fructose, sucrose, maltose, lactose, and sorbitol (Figure S10) were also prepared and incubated for 10 min before the PL measurements. The findings in Figure indicate that these analytes did not show any significant change in the intensity of the nanophosphors. Thence, it can be seen that this nanoprobe sensor promised to be an intriguing indicator for the existence of any risks and threats from the explosives, thus assuring us of our security.

Figure 7

Histogram showing the selectivity of the phosphors over other analytes such as amino acids, pesticides, and sugars. There was no obvious change observed in the PL intensity, suggesting the selectivity toward TNT.

Conclusions

In summary, an approach has been reported for selective detection of TNT over a number of other analogous nitrocompounds using β-NaYF4:Gd3+,Tb3+@PEI nanophosphors based on fluorimetric sensing technique. The results from various characterization methods suggested the formation of β-NaYF4:Gd3+,Tb3+ nanophosphors capped with PEI. The excellent selectivity toward TNT is ascribed to the interactions of the amine group present in PEI and nitro groups in TNT forming the Meisenheimer complex, which has caused a decrease in the PL signal because of the LRET-based energy-transfer mechanism between the nanophosphors and the complex formed. The PL intensity variations have displayed that the nanophosphors were sensitive and efficient for the explosive detection with a LOD of 119.9 nM (27.2 ppb). To further note the advantages of our work, we investigated the PL intensity of the nanophosphors with other analytes and no obvious effects were observed. Thus, the proposed method demonstrates the feasibility of the PEI-capped β-NaYF4:Gd3+,Tb3+ phosphor nanoparticles’ application for detecting TNT as the analyte independent of complicated instruments and immunoassays.

Methods

Warning! Nitrocompounds, viz., TNT, TNP, and RDX, are highly explosive and therefore should be handled carefully in small amounts with proper safety precautions. In Figure S1, we have shown the chemical structure of some of the nitro explosives.

Chemicals and Reagents

All the chemicals were of analytical grade and were used without further purification; yttrium nitrate hexahydrate (Y(NO3)3·6H2O, 99.89%), gadolinium nitrate hexahydrate (Gd(NO3)3·6H2O, 99.89%), and terbium nitrate hexahydrate (Tb(NO3)3·6H2O, 99.89%), PEI (with Mw = 25 000 and Mn = 10 000), TNP, 2,4-DNT, 2,6-DNT, 2,4-DNP, RDX, phenylalanine, cysteine, isoleucine, tryptophan, glutamic acid, aspartic acid, lysine, histidine, and glutamine were purchased from Sigma-Aldrich Inc. TNT and RDX were purchased from HEMRL, Pune. NaCl, Na2HPO4, NaH2PO4, Na2CO3, NaHCO3, and NaOH were received from Thomas Baker. NH4F was received from Merck. Dextrose, glucose, sucrose, fructose, lactose, and maltose were received from Himedia laboratories Pvt. Ltd. 1,3-DNB, 4-NP, 2,4-DNB, NM, 4-NT, NB, copper oxychloride, cypermethrin, malathion, fenvelerate, carbendazim, and chloropyrifos were purchased from a local company and used as received. Ethanol was obtained from Hayman Ltd. Deionized water was used throughout the experiments.

Synthesis

Synthesis of Amine-Functionalized β-NaYF4:15%Gd3+, 5%Tb3+ (β-NaYF4:Gd3+,Tb3+@PEI)

Downconverting NaYF4 nanophosphors were prepared using the hydrothermal method reported earlier.[7,43] Briefly, 10 mL of 0.2 M solutions of Y(NO3)3, Gd(NO3)3, and Tb(NO3)3 were added in a 10 mL solution of NaCl (0.2 M). The solution was continuously stirred for nearly 30 min. Thereafter, 10 mL of PEI (10 wt %) was added followed by the addition of 20 mL of ethanol. In the resultant solution, 0.5 M of NH4F was added dropwise. Finally, the mixture was poured in a Teflon container with 100 mL capacity and reaction was set for 24 h at 180 °C. The autoclave was cooled to room temperature, and the green luminescent nanophosphors were obtained after washing with water and ethanol thrice. Then, the product was dried under vacuum at 70 °C overnight.

Preparation of Stock Solutions

For the PL study, 300 μM stock solution of the analytes was prepared in deionized water and a fixed amount of β-NaYF4:Gd3+,Tb[3]+@PEI was added to the desired concentrations levels in a 1 mL quartz cuvette (path length of 1 cm). The mixture was then incubated at room temperature for 10 min before the spectral measurements. The luminescence spectra of the solutions were measured at an excitation wavelength of 273 nm and emission data were collected in the range of 400–700 nm. Each measurement was repeated at least thrice and consistent results were recorded. The feasibility of the probe was also investigated in buffer solution with pH 7 to pH 13. For pH ranging from 7 to 8, NaH2PO4–Na2HPO4, pH ranging from 9 to 11 NaHCO3–Na2CO3–NaOH and pH ranging from 12 to 13 KCl–NaOH were used.

Characterization Techniques

The phase purity and crystallinity of the as-synthesized samples were characterized by powder X-ray diffraction using a PANalytical X’PERT PRO instrument and the iron-filtered Cu-Kα radiation (λ = 1.54 Å) in the 2θ range of 10°–80° covered in a step size of 0.08° with a count time of 2s. The operating voltage and current were kept at 30 kV and 40 mA, respectively. The specific structural details and morphology were obtained by using an FEI Tecnai T20 transmission electron microscope equipped with a super-twin lens (s-twin) operated at 200 keV voltage. To analyze the shape and size of the samples, FESEM (Hitachi S-4200) was done. EDXA of the samples was performed during FESEM measurements to obtain the elemental composition of the samples. The XPS spectra were recorded on Thermo Fisher Scientific Instruments K Alpha + with monochromatic Al Kα as the X-ray source with 6 mA beam current and 12 kV voltage. A PALS Zeta Potential Analyzer Ver 3.54 (Brookhaven Instrument Corps.) was used to determine the zeta potentials (ζ). Deionized water was the dispersion medium. The FTIR spectrum was recorded by a PerkinElmer spectrum two FTIR spectrophotometer with a resolution of 4 cm–1 and scan speed of 32 scans/min. TGA was done using SDT model Q600 of TA Instruments Inc., USA, at a heating rate of 10 °C/min under continuous flux of nitrogen at 100 mL/min. Raman spectroscopy measurements were recorded at room temperature on an HR 800 Raman spectrophotometer (Jobin Yvon, HORIBA, France) equipped with an achromatic Czerny–Turner type monochromator (800 mm focal-length) with silver-treated mirrors. Monochromatic radiation emitted by a 325 nm laser, operating at 20 mW, was used. PL spectra were acquired using a Cary eclipse fluorescence spectrophotometer, equipped with a 400 W Xe lamp as an excitation source with excitation and emission slit width of 10 nm and a Hamamatsu R928 photomultiplier tube as a detector. The UV–visible spectra were taken on a Cary 300 absorbance spectrometer. PL decay dynamics were carried out on FLS 980 (Edinburgh Instruments).