Abstract
1. INTRODUCTION
Surface-enhanced Raman scattering (SERS) can perform label-free detection of analyte at trace or even single molecule levels, with high sensitivity and selectivity [1]. As a nondestructive vibration testing model, SERS can provide the molecular fingerprint that has been widely used in a variety of scientific fields such as environmental monitoring, food safety testing, drug inspection, and disease diagnosis [2–6]. Noble metal SERS substrates such as gold and silver nanostructures with rough surfaces have been extensively studied by means of surface plasmon resonance excited local electromagnetic field amplification [7–12]. However, the noble metal-based SERS substrates lack repeatability and controllability due to the elaborate and complex synthesis processes. With the development of nanotechnology, SERS research has recently shifted from noble metals mainly based on the electromagnetic mechanism (EM) to novel two-dimensional (2D) nanomaterials based on the chemical mechanism (CM) due to the cheap sources and simple preparation methods [13–16]. However, since CM is a short-range process that relies on the charge transfer (CT) during the resonance electron transition at the interface between the substrate and the adsorption molecule, its enhancement effect in the entire SERS is still limited to some extent [17,18]. In addition, the limits of detection (LODs) and the enhancement factors (EFs) are still much lower than that of metal nanostructures. Therefore, exploring promising 2D materials suitable for SERS analysis becomes more attractive. -phase transition metal tellurides (TMTs) have recently been investigated as plasmon-free 2D SERS substrates for their brilliant physicochemical properties, which have a flat surface where probe molecules can be uniformly chemisorbed and abundant energy states near the Fermi energy level [19,20]. In addition, telluride nanosheets can also be prepared by facile ultrasonic peeling and hydrothermal reaction.
Uric acid (UA), the final product of purine metabolism, is one of the most important biomarkers in body fluid. The equilibrium concentration of UA in urine is determined to be 0.952 to 5.948 mM (1 M = 1 mol/L) per 24 h and normouricemia is 0.208 to 0.416 mM (male) or 0.149 to 0.357 mM (female) [21–23]. Abnormal UA content is closely related to many diseases such as gout, kidney stones, hypertension, and cardiovascular disease [24–26]. Various methods such as high-performance liquid chromatography, isotope dilution mass spectrometry, colorimetric chemosensors, capillary electrophoresis, and electrochemical biosensors have been applied to detect UA. These techniques, however, are often hindered by complicated sample pretreatment, expensive instruments, and sophisticated instrumentation and equipment, and often lack sensitivity [27–30]. The SERS technique can provide a fast, ultrasensitive, and conventional diagnostic test method for real-time detection of UA. The SERS technique that uses various metal-based SERS substrates such as Au nanofibers, composites, Ag-paper, and Ag-modified graphene oxide nanosheets, has been reported for the detection of UA-related diseases, which shows good detection sensitivity [31–34]. However, these types of SERS substrates have certain practical limitations, including sophisticated preparation procedures, high cost due to the use of noble metals, and relatively poor SERS reproducibility caused by the complex nanostructural components.
In this work, a 2D SERS platform based on hafnium ditelluride () nanosheets is proposed to fabricate an effective SERS detection system for the quantitative analysis of uric acid (Fig. 1). Few-layered nanosheets are prepared by a facile liquid exfoliation plus a hydrothermal method. The obtained nanosheets exhibit outstanding SERS activity to analytes with low LODs and high EFs, which can be ascribed to the electronic transition between the -phase layered structure and the detection molecule after theoretical study. The SERS performance based on the substrate also exhibits superior reproducibility and uniformity, which is further utilized as a new label-free SERS platform to detect trace UA at different conditions.
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Figure 1.Schematic illustration of the SERS detection of uric acid based on
2. EXPERIMENT
A. Materials
Hafnium ditelluride was purchased from SixCarbon Technology Co., Ltd. (Shenzhen, China). Uric acid, urea, rhodamine 6G (Rh6G), crystal violet (CV), malachite green (MG), and methylene blue (MB) were purchased from Sigma-Aldrich (now MilliporeSigma, St. Louis, MO, USA). All reagents were of analytical grade and used directly without further purification. Deionized water was used throughout the study (Milli-Q System, MilliporeSigma).
B. Synthesis of
A liquid exfoliation method including probe sonication and bath sonication was combined with hydrothermal reaction to prepare nanosheets. The stripped procedures were as follows.
C. Characterization
The surface morphology of nanosheets was characterized by a 200 kV transmission electron microscope (TEM, JEM-2010HR, JEOL Ltd., Tokyo, Japan), equipped with an energy-dispersive X-ray (EDX) spectrum. A scanning electron microscope (SEM, SU8010, Hitachi, Ltd., Tokyo, Japan) was used to observe the morphology and size of materials. The height of the nanomaterials was measured by an atomic force microscope (AFM, FSM-Nanoview, Fishman, Suzhou, China). X-ray diffraction (XRD) spectrum was measured by a D8 focus X-ray diffractometer (Bruker Corp., Billerica, MA, USA) by Cu Ka radiation (, 1 Å = 0.1 nm). X-ray photoelectron spectroscopy (XPS) profile of nanosheets was measured by a photoelectron spectrometer (Escalab 250 Xi, Thermo Fisher Scientific Inc., Waltham, MA, USA). The ultraviolet-visible-near infrared (UV-Vis-NIR) absorbance spectrum of the nanosheets was recorded on an absorption spectrometer (UV-6100S, Shanghai Mapada Instruments Co., Ltd., Shanghai, China). Raman spectra were collected using a microspectrometer (inVia, Renishaw plc, Wotton-under-Edge, UK) under a 785 nm diode laser excitation.
D. SERS Experiments
Rh6G, CV, MB, and MG were chosen as the Raman reporters for the SERS study. 4 μL of the nanosheets solution was firstly deposited on the Si substrate by the spin-coating method, followed by dropping of 4 μL of dye molecules. Then the samples were placed under a Renishaw inVia Raman microspectrometer for SERS detection equipped with a 785 nm laser. The laser power on the sample was 1 mW. Raman spectra were recorded in the static mode for a 5 s laser exposure (10 accumulations) in the range of . All experiments were independently conducted six times. To study the reproducibility, the SERS test was repeated for 20 times. For practical application, we finally performed the SERS detection of uric acid based on substrate (using urea as the interfering material).
3. RESULTS AND DISCUSSION
A. Characterization of Nanostructures
Figure 2(a) shows the schematic diagram of the synthetic process of lamellar nanosheets. The morphological characteristics of the as-prepared 2D nanosheets were studied by TEM, SEM, and AFM analysis, respectively. The TEM image reveals the transparency of the flakes to the electron beam of , confirming the obvious multilayer morphology after liquid peeling, as shown in Fig. 2(b). The high-resolution TEM (HRTEM) image shows the crystalline structure of nanosheets with a lattice spacing of 0.33 nm [Fig. 2(b) inset]. As displayed in Fig. 2(c), the sizes and thicknesses of nanosheets become significantly smaller after 180°C hydrothermal reaction, and the selected-area electron diffraction (SAED) pattern reveals the crystalline nature of [Fig. 2(c) inset]. Moreover, the SEM image in Fig. 2(d) further demonstrates the layered structure of . The elemental maps of Te and Hf elements are well overlapped with the high-angle annular dark field (HAADF) image of nanosheets, as shown in Fig. 2(e), indicating the element composition of nanosheets that is also discerned by EDX spectroscopy [Fig. 2(f)]. The AFM image clearly shows the thickness of nanosheets in Fig. 2(g), which displays the desirable monodispersity and fairly well-defined dimensions with the average thickness concentrated at 1–2 nm, as shown in the inset of Fig. 2(g). Figure 2(h) demonstrates two representative topographic plots of nanosheets in Fig. 2(g), proving the ultrathin layer. The Raman features of are shown in Fig. 2(i). It can be noted that the main Raman peaks of are located at , which is far from the Raman fingerprint region of ordinary analytes (); thus, unnecessary Raman interference can be largely avoided. The Raman bands of few-layered nanosheets experienced a slight blue shift compared to that of bluk , which may be ascribed to the layer-dependent band structure of the 2D material nanostructures [20,35,36].
Figure 2.Synthesis and characterization of
crystal possesses a stable -phase layered structure with weak interlayer interactions, corresponding to the type triangular structure with a P– space group [37,38]. In each layer, Hf atoms are sandwiched by Te atom layers with reverse symmetry at the top and bottom, as shown in Figs. 3(a) and 3(b). In the unit cell, Hf atoms are distributed at eight apex angles, and Te atoms are distributed in a regular trigonal column composed of three upper and lower Hf atoms, spaced at the center of the upper half and the center of the lower half, as shown in Fig. 3(c). The XRD pattern of the synthesized is shown in Fig. 3(d), and the positions of the diffraction peaks found in the XRD pattern are consistent with the results of in the standard JCPDS card No. 26-0736, with standard lattice parameters (, ) [39].
Figure 3.Atomic structure of monolayer
XPS analysis was used to validate the stoichiometry of 2D nanosheets. Figure 4(a) illustrates the XPS spectra of the bulk and nanosheets, which can discern the variation of the surface chemical composition during the peeling process. Comparing the peak positions of before and after the exfoliation, the results remain consistent across the basic, which proves that the chemical state of the material remained in a stable state during the preparation process. The high-resolution XPS spectra of Hf 4f are illustrated in Figs. 4(b) and 4(c), where the peaks in the spectral line of few-layered sheets are slightly wider than that of bulk, indicating mild oxidation occurred during the preparation process [40,41]. In addition, an absorption band at around 587 nm with decreasing absorption intensity is mainly observed in the UV-Vis-NIR absorption spectrum of the prepared nanosheets, as shown in Fig. 4(d). The optical bandgap of was evaluated by its absorption spectrum, and its band gap was estimated by
Figure 4.(a) XPS analysis of
B. SERS Activity of Hafnium Telluride
Figure 5.(a) Raw Raman spectra of Rh6G (
The ultrathin 2D nanosheets have smooth surfaces and can produce uniform SERS signals better than rough surfaces [50]. For SERS reproducibility study, we randomly acquired 20 SERS spectral lines of Rh6G on substrate. It is strikingly apparent that the Raman signals of Rh6G can be clearly displayed on the substrate with predominant reproducibility, as shown in Fig. 6(a). Then the relative standard deviations of the Raman characteristic peaks at , , and are calculated to be 4.201%, 4.459%, and 7.198%, respectively, as shown in Figs. 6(b)–6(d), indicating better SERS uniformity than that of other telluride SERS substrates [46,48]. Similar data are also discerned in the SERS analysis of MB and CV. These results obviously demonstrate that nanosheets can be used as excellent SERS substrate with uniform SERS signals.
Figure 6.(a) Cluster of SERS spectra of Rh6G samples randomly collected at 20 sites on the
Raman mapping was further carried out in a randomly selected region (, step size 1 μm) to evaluate the SERS reproducibility and uniformity of nanosheets. The laser exposure time on the sample was 3 s under 785 nm laser excitation. Figure 7(a) shows the SERS image of Rh6G molecules on nanosheets using the Raman peak at , which indicates a relatively uniform distribution of SERS signals. Then 170 Raman spectral lines were collected from the SERS image, and the contour map is plotted in Fig. 7(b). It can be seen that the characteristic Raman peaks of Rh6G (, , , and ) have favorable continuity and uniformity. Moreover, we reconstructed a Raman spectrum along the green diagonal line in Fig. 7(b), which exhibited almost identical spectral pattern compared to the average SERS spectrum, as shown in Fig. 7(c), corroborating the good SERS uniformity of nanosheets. The same scenario also emerges in the SERS mapping of CV molecules.
Figure 7.(a) SERS image of Rh6G molecules (
C. Chemical Mechanism of
Chemical enhancement mechanism plays a leading role in SERS of transition metal dichalcogenides [51]. The possible Raman enhancement mechanism of substrate to Rh6G probe is represented in Fig. 8. There are four possible types of CT resonances in this SERS system: (i) exciton resonance of from VB to CB state; (ii) molecular resonance in dye from the highest occupied molecular orbit (HOMO) to the lowest unoccupied molecular orbit (LUMO) level; (iii) exciton electron transfers from molecule ground state to surface transition detection state, followed by photon-induced electron transition that occurs from the surface transition detection state to CB state; and (iv) light-induced electron transfer occurs from VB state to molecular excited state [52–54]. Rh6G is a traditional SERS probe with HOMO and LUMO levels of and , respectively [55], while the VB and CB of are calculated as and , respectively [Figs. 4(e) and 4(f)]. Therefore, among the possible CT resonances mentioned above, when exciton resonance (i) and molecular resonance (ii) occur, the required excitation energies are 4.93 eV and 2.30 eV, respectively. However, the excitation energy of a 785 nm laser is only 1.58 eV [56]. Consequently, since the energy provided is much smaller than the energy required, these two processes of CT resonance can be excluded [55,57]. Similarly, photon energy of 5.65 eV is needed to directly transfer electron from the HOMO energy level of Rh6G molecule to the CB state of . Even in the presence of a surface transition detection state, photon-induced CT resonance cannot occur. In type (iv), the energy required for the excitation transition of the electron is 1.58 eV between the LUMO and VB state, which matches the laser excitation energy. A similar result is also provided after mechanism deduction in the -based SERS analysis of MB and CV dyes. Therefore, the CT resonance process between the VB state of 2D nanosheet and the LUMO level of dye molecule probably dominates the Raman enhancement of nanosheets.
Figure 8.Schematic diagram of the photo-induced charge transfer process between
D. SERS Screening of Uric Acid
The excessive UA level is prone to cause gout and even uremia. The detection of UA content in urine has momentous predictive significance for the onset of gout disease. For clinical practice, urea is the biggest interference factor for the SERS detection of UA. We first measured the SERS signals of pure uric acid, urea, and their mixture on substrate. As shown in Fig. 9, the SERS spectral patterns of UA and urea are in remarkable agreement with what was reported in the literature [22,58]. The characteristic SERS peaks of UA at , , and can be attributed to ring vibrations, skeletal ring deformation, and C-N vibrations, respectively [31]. In the SERS spectrum of the mixture of UA and urea, the typical SERS peaks of UA and urea can be obviously distinguished with little mutual interference. Then we performed SERS analysis of UA at concentrations ranging from 100 μM to 1 mM to study the limit of detection. Figure 10(a) shows a concentration-dependent SERS effect that the intensity of the SERS signal increases when the concentration of UA improves. The fitting curve of the peak at indicates that the SERS intensity is directly proportional to the amount of UA adsorbed on the nanosheets [Fig. 10(b)], and the LOD of -based SERS analysis to UA is 100 μM. It has been reported that the lowest normal uric acid level is of 149 μM
Figure 9.Mean SERS spectra of pure uric acid, urea, and their mixture on
Figure 10.(a) SERS detection of UA at different concentrations on
4. CONCLUSIONS
In summary, we have prepared few-layered nanosheets via a facile liquid exfoliation combined with hydrothermal reaction. We believe nanosheets can serve as a novel 2D TMT SERS substrate due to the outstanding SERS activity and reproducibility. Compared to some existing SERS platforms, the SERS analysis based on does not suffer from the background interference from the substrate. The charge transfer from the VB state of to the LUMO level of dye molecule may contribute to the enhancement mechanism in a -based SERS system, which leads to the maximum EF of . For practical application, nanosheets have successfully been used for the SERS detection of UA, the important biomarker for gout disease, which demonstrated a reliable LOD of 100 μM. The study of the -based SERS platform opens up bright prospects for nanodiagnostics.
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