Low-cost Au nanoparticle-decorated cicada wing as sensitive and recyclable substrates for surface enhanced Raman scattering

Sensors and Actuators B 209 (2015) 820–827

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Sensors and Actuators B: Chemical journal homepage: www.elsevier.com/locate/snb

Low-cost Au nanoparticle-decorated cicada wing as sensitive and recyclable substrates for surface enhanced Raman scattering Ming Yang Lv a,b , Hai Yan Teng a,b , Zhao Yang Chen a,b,∗ , Yong Mei Zhao c , Xin Zhang a,b , Luo Liu a , Zhenglong Wu d , Li Min Liu b , Hai Jun Xu a,b,∗ a

Beijing Key Laboratory of Bioprocess, Beijing University of Chemical Technology, Beijing 100029, China College of Science, Beijing University of Chemical Technology, Beijing 100029, China c Engineering Research Center for Semiconductor Integrated Technology, Institute of Semiconductors, Chinese Academy of Sciences, Beijing 100083, China d Analytical and Testing Center, Beijing Normal University, Beijing 100875, China b

a r t i c l e

i n f o

Article history: Received 29 July 2014 Received in revised form 6 December 2014 Accepted 12 December 2014 Available online 22 December 2014 Keywords: Surface enhanced Raman scattering Au nanoparticle Cicada wing Recyclability Quantitative detection

a b s t r a c t A large-scale, low-cost and three-dimensional biomimetic surface enhanced Raman scattering (SERS) substrate with vast and regular ‘hot spots’ was prepared by the deposition of gold (Au) nanoparticles on the bioscaffold arrays of cicada wings by the electron beam evaporation. Characterizations of the formed three-dimensional Au/cicada wing nanostructure proved that it owns outstanding SERS properties of highly active sensitivity, good uniformity and repeatability. The SERS limit of detection (LOD) for Rhodamine 6G reaching as low as 10−7 M, and a Raman enhancement factor being as large as 105 with a relative standard deviation of 10.1% were obtained. More importantly, this substrate has a good recyclability by the simple NaBH4 washing. For the application of Au/cicada wing, thiram was also detected rapidly and quantitatively with a LOD as low as 10−7 M. Our findings are beneficial in the application of SERS substrate, allowing quantitative and recyclable detection of trace organic contaminants and making it very promising in the label-free detection of biological molecules. © 2014 Elsevier B.V. All rights reserved.

1. Introduction Raman spectrum, a powerful and sensitive analytical technique known as the ‘fingerprints’ of molecules, is finding increased application in analytical chemistry, biosensing field and environmental evaluation with its integration of rapid response, non-destructive analysis and weak signal of water [1–4]. However, Raman scattering is very weak due to the small cross-section of molecules. The surface-enhanced Raman scattering (SERS) effect can overcome this drawback. SERS is the phenomenon whereby Raman signals are strongly increased when molecules are attached to nanorough metallic structures. The enhancement effects observed in SERS have been attributed to the electromagnetic (EM) mechanism and the chemical mechanism. The EM mechanism, which results from an increased local electromagnetic field (hot spot) at a metallic nanoparticle surface where the interaction of the incident laser

∗ Corresponding authors at: Beijing Key Laboratory of Bioprocess, Beijing University of Chemical Technology, Beijing 100029, China. Tel.: +86 10 64442357; fax: +86 10 64435170. E-mail addresses: [email protected] (Z.Y. Chen), [email protected] (H.J. Xu). http://dx.doi.org/10.1016/j.snb.2014.12.061 0925-4005/© 2014 Elsevier B.V. All rights reserved.

with the electrons occurs, is the commonly accepted origin of SERS [5,6]. Noble metals such as gold (Au), silver (Ag) and copper (Cu) show the superior performances in supporting the EM enhancement of SERS substrates, and are widely used as SERS-active substrates with or without decoration. Ag is extremely efficient in enhancing the Raman scattering, but it is easily oxidized, leading to a very limited lifetime of an Ag substrate. Compared with Ag, Au has several unique features, such as excellent resistance against oxidation and biocompatibility as well as extremely strong and tunable surface plasmon resonance in visible and near-infrared spectral regions [7]. More importantly, the Au modificated substrates possess an effective function of recyclability by the simple NaBH4 washing, reducing the waste of novel metals and environmental pollution efficiently. Various Au nanostructures, such as nanowire [8], nanorod [9] and nanopillar [10] arrays, have been utilized as highly sensitive SERS substrates. These three-dimensional (3D) nanoarchitectures with nanogap-rich Au nanoparticles (NPs) were considered to improve the SERS effect more obviously than onedimensional and two-dimensional nanostructures [11]. Actually, the 3D SERS substrates not only generate more effective, adequate, and regular ‘hot spots’, but also increase plenty of binding sites for probing molecules within the laser footprint, and both of them could create strong and tunable EM field. Preparation of 3D SERS

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substrate with high density of ‘hot spots’ by straightforward and economical methods is the key to the widespread adoption of SERS as a general sensing technique. Recently, many researchers have paid attention to utilizing the natural creatures (e.g. plant leaves, insect wings and legs) with non-smooth 3D nanostructures and super-hydrophobic surfaces after noble metal NPs deposition as SERS substrates [12]. Interestingly, these super-hydrophobic surfaces allow a significant accumulation of analyte molecules in a tiny area through the evaporation of a water droplet, showing tremendous potential [12–14]. For example, butterfly wings, modified with Ag NPs have been chosen as a SERS substrate, which can detect Rhodamine 6G (R6G) at a concentration as low as 10−9 M [15]. Although the Ag NPs substrate possesses a lower limit of detection (LOD), it is neither durable nor recyclable, resulting in waste of resources and pollution of environment. Therefore, it is worth seeking a better SERS substrate to avoid these problems. In this paper, the periodical 3D nanostructures and superhydrophobic surface of the cicada wing inspired us to use it as a regular substrate, and a large-scale 3D SERS substrate with high sensitivity, good stability and recyclability was prepared by the electron beam evaporation (EBE) of Au NPs onto cicada wings. Obviously, in comparison with other kinds of SERS substrates, the fabricated Au/cicada wing is low-cost, green, soft, easy to obtain and has extensively potential application in biological sensing, and the formed hierarchical nanostructure could produce many SERS ‘hot spots’ at the formed 3D arrays. Based on vast comparative analyses, the Au/cicada wing with a deposited thickness of 50 nm was adopted for the best enhancement, sensitivity and reproducibility. The Au/cicada wing substrate was used to detect R6G of concentration as low as 10−7 M and the single molecule detection could be realized. More importantly, the formed SERS substrates are recyclable for many organothiols and other Au surface adsorbates such as R6G, methyl orange (MO), paminothiophenol (PATP), Crystal violet (CV) and Nile blue A (NBA), which can all be rapidly and completely removed from Au/cicada wing surface. This recyclability reveals that the unique recyclable property not only paves a new way to solve the single-use problem of traditional SERS substrates but also provides more SERS platforms for multiple detections of other organic molecular species. Additionally, for the application of this Au/cicada wing substrate, thiram was detected quantitatively and the LOD was also down to 10−7 M.

2. Experimental 2.1. Materials and instruments Cryptotympana atrata fabricius were purchased from Jia Ying Art Museum of Entomology. R6G, MO, CV, NBA, PATP, thiram and NaBH4 were procured from J&K Scientific LTD. Deionized water (18 M) which was obtained from Beijing Chemical Works was used for all experiments. All chemicals, unless mentioned otherwise, were of analytical grade and were used as received. The typical morphologies and microstructures of cicada wings were investigated by field emission scanning electron microscopy (FE-SEM) (JEOL JSM-6700F) and X-ray diffractometer (XRD) (Rigaku Ultima III). The UV–vis absorption spectra were monitored by a Shimadzu UV-3600 UV–vis spectrophotometer. The wings were coated by Au NPs in EBE system (Tri-Axis) and the rates of deposition were 1 nm/s to fabricate Au nanofilms with different thicknesses. The contact angle (CA) was measured with OCA20 machine (Data Physics, Germany).

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2.2. Sample preparation All cicada wings were first ultrasonically cleaned in acetone, ethanol and deionized water in turn for 5 min each. Then the wings were modified by Au NPs with different deposition thicknesses in the EBE system. When the system working, electron beams from the electron gun bombarded the Au disk which was the target of evaporation and brought about a high temperature to the disk, leading to the gasification of Au, and escaping of amounts of Au atoms from the Au disk to the surface of substrates. Then, Au NPs were solidified and adhered to the surface by the lower temperature of the substrates. Clearly, the interaction force of the Au NPs and cicada wing is the van der Waals force, which is the intermolecular interaction force. During the process of coating, the voltage of electron gun was 6.5 kV, the vacuum was 2.8 × 10−6 Pa, the temperature of the substrates (cicada wings) was about 22 ◦ C and the rate of deposition was 1 nm/s. Inficon crystal oscillator was used to monitor the thickness of films. The contrast analysis indicated that 50 nm Au nanofilms on Au/cicada wing have the best SERS sensitivity and therefore the Au/cicada wing with a deposited thickness of 50 nm Au films was used in all tests. 2.3. SERS measurements The Raman signals of R6G and thiram were obtained after the droplet evaporated naturally and all of the signals were obtained at room temperature on the LabRAM ARAMIS Raman system with the 785 nm laser as excitation. This was because excitation lasers with different wavelengths all generate fluorescent signals with R6G, which may influence the SERS signals, but among the 532, 633 and 785 nm lasers, we found that the fluorescent signals with R6G generated by the last one are the weakest. The diameter of the light spot area was ∼1 ␮m and the incident power was 0.325 mW. Choosing 0.325 mW as the laser power was to get the strongest Raman spectra without burning substrate. It is well known that the substrate temperature will be increased because of the laser energy depositing onto the substrate. The higher temperature may cause the substrate to burn and the lower temperature will reduce the SERS sensitivity. We also noticed that both cicada wing and Au NPs had no obvious changes of structure or performance in a reasonable temperature range. Thus in this paper we use the fixed laser power but not study in detail the temperature effect. The spectra were recorded with the accumulation time of 15 s and the spectral resolution was 1 cm−1 . The accumulation time and the laser power were the same for all Raman spectra in the case of no special instructions. All the data were averaged over 20 randomly selected positions. 3. Results and discussion 3.1. Characterization Fig. 1(a) and (b) shows top-view and sloping-view FE-SEM images of the cicada wing surfaces. Clearly, they exhibited a largescale protrusion nanostructure which possesses a high degree of regularity. The protrusions presented a shape of cone contributing to the hydrophobicity of cicada wing [16]. Meanwhile, as shown in Fig. 1(b), the average values of the basal diameter, the top diameter, the top spacing and the height of the cone-shaped protrusions were estimated to be about 100, 50, 100 and 240 nm respectively. Fig. 1(c) and (d) presents the images of the cicada wings deposited with 50 nm Au NPs. Obviously, the Au/cicada wing substrate was successfully constructed without damaging the structure of cicada wing, but both the tops and the sides of the protrusions were covered with Au NPs. The average diameters of Au NPs were ∼65 nm

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Fig. 1. FE-SEM micrographs of (a), (b) cicada wing and (c), (d) Au/cicada wing respectively, and the insets in (a) and (c) are the corresponding optical images of water droplets on surfaces. (e) XRD patterns of cicada wing and Au/cicada wing. (f) UV–vis spectra of cicada wing and Au/cicada wing.

on the top and ∼35 nm on the side of the protrusions, respectively, and these Au NPs deposited on the regular protrusions could create plenty of uniform SERS ‘hot spots’ to promote the SERS effect [11]. Meanwhile, as shown in the inset of Fig. 1(a) and (c), the high values of CA (149◦ and 144.4◦ ) indicate that the cicada wing always possesses a super-hydrophobic surface before and after the Au NPs deposition [17]. These super-hydrophobic surfaces will provide an accumulation of analyte molecules to enhance the SERS effect [14]. Moreover, the large-scale and orderly distribution of protrusions coated with Au NPs on the surfaces of the cicada wings also indicate that the substrate might have a good reproducibility during the SERS detection. For the elemental analysis, the XRD patterns of the cicada wing and Au/cicada wing are depicted in Fig. 1(e), and four characteristic peaks of Au are clearly observed at 2 = 38.3◦ , 44.6◦ , 64.7◦ and 77.5◦ , which were indexed to the (1 1 1), (2 0 0), (2 2 0) and (3 1 1) crystallographic planes of the face centered cubic (fcc) Au crystal, respectively (Au XRD Ref. No. 40-1139) [18]. According to the Debye–Scherrer equation D = K/ˇ cos , where K was the Scherrer constant (0.9),  was X-ray wavelength (0.15418 nm), ␤ was the width of the XRD peak at half height and  was the Bragg angle, the (2 0 0) diffraction peak was selected to calculate the average size of Au NPs and the estimated value was ∼31 nm. As shown in Fig. 1(f), Au/cicada wing exhibited obvious enhanced capability of light absorption in the range of 300–700 nm in comparison to the unprocessed cicada wing. Furthermore, upon the

deposition of Au NPs, a board plasmon band centered at ∼500 nm appeared, indicating the formation of Au NPs onto the cicada wing [19]. Although there was a wavelength mismatch between the surface plasmon band (500 nm) and the exciting source (785 nm), the formed SERS substrate showed strong Raman signals, which may be because UV–vis absorption spectra provide only a rough indication on matching between incident laser and surface plasmon resonance [20]. 3.2. SERS performances and EF calculation R6G, which has been extensively used for SERS study previously due to its well-established vibrational features, was used as the probe molecule to demonstrate the SERS performance of Au/cicada wing substrate [21]. Fig. 2(a) shows the Raman spectra of R6G droplets in the range of 10−3 –10−7 M, which was detected after being evaporated naturally onto the Au/cicada wing substrate. The Raman bands at 611, 780 and 1187 cm−1 can be assigned to C C C ring in-plane bending, C H out-of-plane bending and C H in-plane bending, respectively. And the other features at 1311, 1361, 1510, 1063 and 1647 cm−1 all stemmed from the aromatic C C stretching vibrations [22]. It can be found that these Raman peaks of R6G are still quite clear even at low concentration of 10−7 M. Apart from the 3D structure, the lower LOD may be due to the surface super-hydrophobicity, arising from hierarchically

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Fig. 2. (a) SERS spectra of R6G solution at different concentrations. (b) The response curve of the peak intensities centered at 1361 cm−1 of R6G, and the inset is the quantitative logarithmic relation curve. (c) SERS spectra of R6G from 10−5 to 10−1 M, and the inset is the variation of the peak intensities centered at 1361 cm−1 . (d) SERS spectra of 10−3 M R6G collected on the 100 spots from 10 Au/cicada wing substrates. (e) The intensity distribution of the peaks centered at 1361 cm−1 corresponding to (d) with the corresponding RSD of 10.1%, and the green line indicates the average intensity and the violet zones represent ±5% intensity variation. (f) SERS mapping (step size 1 ␮m, 10 × 10 = 100) of one Au/cicada wing substrate.

structured surface roughness [23]. The surface of the cicada wing shows this property due to the presence of air pockets under the liquid drop [24]. The Au/cicada wing substrates possess superhydrophobic surfaces as well. When a drop of liquid was dripped onto the substrate, the droplet was easy to form a nearly spherical shape (as shown in the inset of Fig. 1(c)). As the droplet evaporated naturally, the analyte molecules in solution would be condensed into a certain region, improving the LOD of the Au/cicada wing substrate [25]. Fig. 2(b) shows the relationship between the SERS integrated intensity of the peaks centered at 1361 cm−1 and the concentration of R6G. When logarithm-scale axes are used, the response between the logarithmic integrated signal intensity (log I) and the logarithmic concentration (log C) is linear, as shown in the inset in Fig. 2(b). The linear relationship between log I and log C was previously confirmed in a publication, when the adsorption of analyte molecules follows Langmuir isotherm [26]. Obviously, the linear relationship implemented the determination of unknown concentrations of R6G solution with this substrate. The SERS enhancement factor (EF) for the 3D substrate was calculated using the accepted formula from the previous literatures [27]: EF = (ISERS /IRaman ) × (NRaman /NSERS ), where ISERS and IRaman

are the Raman peak intensities in the SERS spectra and in bulk Raman spectra, respectively. Here, we choose the Raman band centered at 1361 cm−1 to calculate I values. NRaman is the number of bulk molecules probed in a bulk sample, and NSERS is the number of molecules constituting the first monolayer adsorbed on the substrate surface under the laser spot area. The conventional Raman spectra were collected when a droplet of 10−3 M R6G loaded on a cicada wing and the integrated peak intensity IRaman of the 1361 cm−1 Raman band was measured to be ∼104 after the droplet evaporated naturally. NRaman was determined based on 10−3 M R6G solution and the illuminated volume (Villu ) of the Raman system. For our Raman setup, the illumination focus had a diameter of ∼1 m and the penetration depth of 785 nm laser beam was ∼3 mm. The calculated Villu was ∼2.36 × 103 ␮m3 . Thus, the estimated value for NRaman was ∼1.42 × 109 . When determining NSERS in Villu of our Raman setup, we assumed that R6G molecules were absorbed as a monolayer on the surface of Au/cicada wing substrate. One R6G molecule’s surface area is the product of its length (1.37 nm) and width (1.43 nm) and equals ∼2.0 nm2 [28]. Fig. 2(c) presents the SERS spectra of R6G molecules collected from the Au/cicada wing with different concentrations from 10−5 to 10−1 M. Clearly, the

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Table 1 Calculated EF of R6G solution (10−3 M) on Au/cicada wing substrate to R6G solution (10−3 M) on cicada wing when the Raman bands centered at 611, 780, 1361 and 1510 cm−1 were chosen, respectively. Peak (cm−1 )

611

780

1361

1510

ISERS IRAMAN NSERS NRAMAN ISERS /IRAMAN NRAMAN /NSERS EF

3.75 × 103 45 3.93 × 105 1.42 × 109 83 3.61 × 103 3 × 105

2.96 × 103 28 3.93 × 105 1.42 × 109 105 3.61 × 103 3.8 × 105

9.89 × 103 104 3.93 × 105 1.42 × 109 95.1 3.61 × 103 3.4 × 105

1.1 × 104 85 3.93 × 105 1.42 × 109 129 3.61 × 103 4.7 × 105

corresponding intensities (ISERS ) of the 1361 cm−1 Raman bands are significantly enhanced with increasing R6G concentration until 10−3 M. The values of ISERS for the concentration of 10−1 and 10−2 M are very close to that for 10−3 M. Therefore, the surface of Au/cicada wing is presumed to be fully adsorbed with R6G molecules when the concentration reaches 10−3 M. Through dividing the illuminated area by the surface area of one R6G molecule, the calculated NSERS is 3.93 × 105 , and ISERS corresponding to the 1361 cm−1 Raman band is 9.89 × 103 . The ratios of NRaman /NSERS and ISERS /IRaman could thus be estimated to be ∼3.61 × 103 (1.42 × 109 /3.93 × 105 ) and ∼95.1 (9.89 × 103 /104), respectively. Accordingly, the 3D Au/cicada wing substrate exhibits a large EF of ∼3.4 ×105 . As shown in Table 1, the same EF magnitude was obtained for other fingerprint Raman peaks of R6G. The high EF value indicates that the Au/cicada wing substrate had a higher SERS efficiency. For evaluating substrate-to-substrate reproducibility, SERS signals of R6G were measured at 100 randomly chosen spots from 10 substrates, as shown in Fig. 2(d). To obtain a statistically meaningful result, the relative standard deviation (RSD) of the intensity at the 1361 cm−1 peak of 10−4 M R6G was calculated to be 10.1%, as shown in Fig. 2(e). Meanwhile, to assess the spotto-spot reproducibility further, the substrate was functionalized with 10−4 M R6G and Raman-mapped using a random selected 10 ␮m × 10 ␮m = 100 ␮m2 area with a step size of 1 ␮m. As illustrated in Fig. 2(f), the intensity of the 1361 cm−1 peak from R6G was plotted to demonstrate uniformity across the entire SERS substrate, where each pixel represents the intensity of the Raman peak at the spatial position on the Au/cicada wing substrates. These results show a good uniformity and reproducibility over the entire area of the Au/cicada wing substrates. 3.3. 3D finite-difference time-domain simulation 3D finite-difference time-domain (FDTD) method [29] was used to study the spatial distribution of the electric fields. Fig. 3(a) presents the profile of the local model Au/cicada wing, where 65 nm and 35 nm Au NPs decorated on the top and the side of the two rows protrusions respectively and a basal spacing of 10 nm were adopted. A rectangular-shaped continuous wave laser with a wavelength of 785 nm propagating along the z direction was input into the structure, and the k direction was along the z direction too. Fig. 3(b) exhibits the calculated spatial distributions of the electric field intensities for the y–z plane shown in Fig. 3(a), which passes through the big Au NPs on the top. Fig. 3(c) and (d) presents the spatial distributions for x–y plane defined in Fig. 3 (a), which passes through the small Au NPs on the top and bottom respectively. Two typical kinds of ‘hot spots’ are formed: one presented between the neighboring Au NPs on the side surface of a single protrusion and the other from the Au NPs on the sidewall of two adjacent protrusions. The local electric fields for the model Au/cicada wing were calculated at the maximum of 20.2 V m−1 . The calculated EF is ∼1.7 × 105 according to the relationship of the Raman enhancement scales, roughly as the fourth power of the local field [30]: 4 GSERS ≈ [Eloc (ωexc )/Einc (ωexc )] , where Eloc (ωexc ) and Einc (ωexc ) are

the E and E0 in the FDTD calculations respectively. This value is on the same order of magnitude of the experimental EF (∼3.4 × 105 ). The FDTD calculations clearly show that the SERS enhancement of the substrate is mainly due to the extremely strong electric fields at the nanogaps and, therefore, can be explained by the EM mechanism. 3.4. Recyclability and stability Both recyclability and stability of the SERS substrates can reduce the waste of novel metals and environmental pollution effectively. Fig. 4(a) and (b) shows the results for the analytes collected at initial SERS detection and after washing with NaBH4 aqueous solution several times. Two kinds of analytes with (PATP) and without (R6G, MO, NBA and CV) mercapto group ( SH) were used as the probe molecules. In all cases, the substrate was first immersed in a solution containing one corresponding analyte and characterized by SERS laser, then washed with NaBH4 aqueous solution for 5 min. Subsequently, the substrate was washed with water 3 times to remove residual ions and molecules and then dried at 30 ◦ C under vacuum. Here all the data were averaged from 20 randomly selected positions. Fig. 4(a) shows the spectra for 10−5 M R6G dripped on the Au/cicada wing substrate before and after cleaning with NaBH4 solution. Repeating this cycle for 3 times, it was observed that the fingerprint peaks of R6G were similar to that of the initial one and almost disappeared after washing with NaBH4 aqueous solution. As shown in Fig. 4(b), the substrate was immersed into the solutions of MO, CV, PATP and NBA consecutively with 10−5 M and the SERS signals were detected. Lines i, ii, iii, iv and i , ii , iii , iv represent the SERS signals before and after washing with NaBH4 aqueous solution, respectively. The results show the phenomenon similar to that in previous recyclable detection of R6G. Actually, this could be because the hydride derived from NaBH4 has a higher binding affinity to Au NPs than Au S bond or the interaction of N atom and Au NPs, and replaces them completely [31]. This commutative SERS analysis demonstrates that the Au/cicada wing substrate has an effective function of recyclability. Fig. 4(c) shows the three-month stability test of the SERS substrate. The line i represents the Raman spectrum of 10−5 M R6G solution detected on a fresh Au/cicada wing substrate and the line i gives the spectrum under the same conditions except on an Au/cicada wing substrate prepared three months before. It is clear that the intensity of the Raman peaks change slightly, verifying the long-time stability of the Au/cicada wing substrate. This may owes to gold’s resistance against oxidation and the steady adhesion between Au NPs and cicada wing. 3.5. Quantitative detection of thiram In recent years, the abuse of pesticides on fruits, vegetables and crops has been a threat to the food safety. Especially, thiram is widely used to protect crops from downy mildew, blight, anthracnose and cereal smut as a kind of pesticide [32]. The excess of the thiram residue remained on the surface of foods not only pollutes the environment, but also damages humans’ health.

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Fig. 3. (a) Shape of the FDTD model of Au/cicada wing substrate. (b) Calculated spatial distribution of the electric field intensity for the y–z plane defined in (a). (c) and (d) Calculated spatial distributions of the electric field intensity of the top and bottom for the x–y plane defined in (a), respectively.

Fig. 4. Recyclable SERS behaviors and stability of the Au/cicada wing substrate: (a) R6G with four cycles. (b) The alternating analysis of MO, CV, PATP and NBA. (c) Raman spectrum of R6G detected on the Au/cicada wing substrate now (i) and 3 months later (i ). In all cases, every analyte’s concentration was 10−5 M.

According to the USA Food Standards, the maximum residue limit of thiram in the most farm produces is limited to about 0.8 ppm. Therefore, a rational and convenient approach to detecting thiram with high sensitivity is essential. As shown in Fig. 4(a), the SERS

spectra of thiram with different concentrations from 10−3 to 10−7 M were detected, and the LOD was as low as 10−7 M. Although the experiments for R6G and organic molecule thiram each with a concentration of 10−8 M were carried out, their measured Raman

Fig. 5. (a) SERS spectra of thiram solution at different concentrations and (b) the linear relationship between logI and logC for the band peaking at 1368 cm−1 .

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spectra were hard to distinguish from that of the substrate. When we detected them with higher concentration (e.g., 10−7 M), their characteristic Raman spectra were distinguished from that of the substrate. Therefore we adopted 10−7 M as the approximate LOD for both R6G and thiram. The highest peak (at ∼1368 cm−1 ) in the spectra was the result of CN stretching mode and symmetric CH3 deformation mode work together [33]. The other peaks were at 936, 1138, 1434 and 1486 cm−1 , which were attributed to the C S and CH3 N stretching vibrations, the CN stretching vibrations, the antisymmetric n(CH3 ) stretch and rocking CH3 mode, respectively [34,35]. Similar to the case for the R6G detection, Fig. 4(b) revealed a linearly relation between log I of the integrated signal intensities centered at 1368 cm−1 and log C of thiram concentration. The results manifested that the Au/cicada wing substrates have great potential for rapid and quantitative detection of trace organic molecules or even single molecule (Fig. 5). 4. Conclusion The 3D Au/cicada wing substrate was used as a low-cost, uniformed SERS substrate for trace detection of the organic molecules. The substrate with large-scale, regular protrusion nanostructures and hydrophobicity showed an excellent signal reproducibility (10.1%), high EF (3.4 × 105 ) and low LOD for R6G (10−7 M) and thiram (10−7 M). Meanwhile, our results revealed that NaBH4 could be used as a ‘detergent’ for removal of surface adsorbates from the Au/cicada wing surface to realize the recyclability of SERS substrate. This application provides not only a method to solve the waste of resources of traditional SERS substrates but also more possibilities for multiple detections of other organic molecular species. Additionally, the substrate was successfully applied in thiram detection, realizing a rapid and quantitative detection by combining the linear relation between the logarithmic integrated intensity and the logarithmic concentration of the targeting molecules. Due to the good biocompatibility and stability of Au, the Au/cicada wing substrate may also be easily used in label-free chemical/biomolecule detection. Besides, FDTD calculation results clearly show that the SERS enhancement of the substrate is mainly due to EM mechanism. Finally, we would like to emphasize that Au/cicada wing substrates are overall much better than the Ag NPs modified substrates because the former ones are recyclable and time stable and can save much resources as well as reduce the waste of noble metals, although the LOD of the latter ones are slightly lower than that of the former ones. Acknowledgments This work was supported by the National 973 Basic Research Program of China (2014CB745100), the National Natural Science Foundation of China (21390202, 11104008), the Beijing Natural Science Foundation (4142040), and the Beijing Higher Education Young Elite Teacher Project. References [1] S. Nie, S.R. Emory, Probing single molecules and single nanoparticles by surfaceenhanced Raman scattering, Science 275 (1997) 1102–1106. [2] D. Tsoutsi, J.M. Montenegro, F. Dommershausen, U. Koert, L.M. Liz-Marzán, Quantitative surface-enhanced Raman scattering ultradetection of atomic inorganic ions: the case of chloride, ACS Nano 5 (2011) 7539–7546. [3] Z.Y. Lv, L.P. Mei, W.Y. Chen, J.J. Feng, J.Y. Chen, A.J. Wang, Shaped-controlled electrosynthesis of gold nanodendrites for highlyselective and sensitive SERS detection of formaldehyde, Sens. Actuators B 201 (2014) 92–99. [4] C.X. Zhang, L. Liu, H.J. Yin, H. Fang, Y.M. Zhao, C.J. Bi, H.J. Xu, Recyclable surface-enhanced Raman scattering template based on nanoporous gold film/Si nanowire arrays, Appl. Phys. Lett. 105 (2014) 011905. [5] A. Otto, I. Mrozek, H. Grabhorn, W. Akemann, Surface-enhanced Raman scattering, J. Phys.: Condens. Mater. 4 (1992) 1143–1212.

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Biographies Ming Yang Lv is currently a student in Physics (graduate year) at Beijing University of Chemical Technology, China. His efforts are devoted to utilizing the surface enhanced Raman scatting spectrum into biosensing application. Hai Yan Teng is currently a student in Physics (senior year) at Beijing University of Chemical Technology. She will join the State Key Laboratory of Chemical Resource Engineering and School of Science in September. Zhao Yang Chen received his B.Sc. degree from Peking University (China) in 1989. He earned two Ph.D. degrees from Lanzhou University (China) and Ruhr University, Bochum (Germany) in 1998 and 2004, respectively. He has been with Beijing University of Chemical Technology as a professor since 2012. Currently, his research interests are focused on the laser-materials interaction, nonlinear optics and dusty plasmas. Yong Mei Zhao received her bachelor’s degree and master’s degree from Zhengzhou University in 2002 and 2005, respectively. From 2005 to 2008, she studied in Institute of Semiconductors, Chinese Academy of Sciences and got her Ph.D. degree in

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2008. And ever since 2008, she has worked as a research assistant professor in Engineering Research Center for Semiconductor Integrated Technology, Institute of Semiconductors. Her interest is focused on the preparation and characterization of new semiconductor materials and related device process. Xin Zhang received his B.Sc. and Ph.D. degrees from the Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, in 2007 and 2010 respectively. He joined in Beijing University of Chemical Technology as a lecturer in 2013, and at present his research is focused on the surface enhanced Raman scattering substrates. Luo Liu received his Dr. rer. nat. in Technical Biology in 2007 under the supervision of Prof. Rolf D. Schmid at University of Stuttgart, Germany. He is currently an associate professor of College of Life Science and Technology at Beijing University of Chemical Technology. His main scientific interests are in the areas of enzyme engineering, metabolic engineering and biosensors, to find out desired properties of enzymes or bacteria strains for production of biofuels and other value-added products. Zhenglong Wu is the Ph.D. and professor of condensed matter physics. His research fields are surface chemical analysis, Raman spectral analysis, material analysis for semiconductor materials, luminescent materials and surface-enhanced Raman scattering (SERS) matrix materials. Li Min Liu received her B.Sc. degree from the School of Science, Beijing University of Chemical Technology in 1986, and then has worked in the Beijing University of Chemical Technology until now. Her research is now focused on the surface enhanced Raman scattering. Hai Jun Xu received his B.Sc. and Ph.D. degrees from the Department of Physics, Zhengzhou University, China, in 2002 and 2008 respectively. He has joined the Beijing University of Chemical Technology since 2008 as an associate professor, and his research is now focused on the physics of semiconductor optoelectronic material and surface enhanced Raman scattering substrates.