2.2.1. Surface morphology and wettability of the SERS active substrate

The fabrication process of the SERS substrate was prepared by the electrodeposition on a preformed monolayer polystyrene (PS) colloidal crystal (2 μm in PS sphere-diameter) in the HAuCl4 aqueous solution at room temperature, as previously described. [38]. Figure 2(a) shows the typical morphology. The SERS substrate is the gold array consisting of the hexagonally arranged bowl-like pores with 2 μm in period. The skeleton among the pores in the array is built of nearly vertical quasi rod-shaped nanoparticles. The static contact angle of such substrate surface is about 105, exhibiting the slightly hydrophobic surface (see Figure 2b).

Figure 2. The morphology and wettability of a gold micro-/nanostructured array. (a) The scanning electron microscopic (SEM) image. The inset: A local magnified image and the scale bar = 500 nm. (b) and (c) The photos of the water droplets on the Au array before and after surface modification with thiol, respectively. The water droplets are 2 and 5 μL in volume for (b) and (c), respectively [33].

#### 2.2.2. Raman spectral measurements

The DMMP aqueous solutions with different concentrations were firstly prepared and stored in a refrigerator before use. A droplet of the DMMP aqueous solution was then dropped on the SERS substrate or the gold array and spread out to form the thin water film. After evaporation at room temperature for different time intervals, the Raman spectra for the thin water film were measured on a confocal microprobe Raman spectrometer (Renishaw inVia Reflex) with a laser beam of 632.8 nm in wavelength. The Raman spectral integral time is 10 s.

peaks of DMMP molecules in the aqueous solution are usually difficult to be detected by the SERS effect. No characteristic Raman peak from DMMP was observed after the water film was completely evaporated (see curve (III) in Figure 3). Obviously, such method is quite valid to

Figure 3. The Raman spectra of DMMP molecules. Curves (I), (II) and (III) are for a droplet of 10<sup>2</sup> M DMMP aqueous solution on the SERS substrate after evaporation for a very short time, a certain time and sufficiently long time

(completely dried), respectively. Curve (IV): The Raman spectrum of the pure DMMP liquid. [33].

Spectral evolution with the evaporation. Further, the Raman spectral evolution of the DMMPcontained water film on the substrate with the evaporation time was measured. Figure 4(a) shows the spectra after evaporation for different durations for the water film with 10<sup>2</sup> M DMMP in initial concentration. During initial stage of the evaporation, no characteristic Raman signals were detected and the film is opaque in the field of optical microscope, as shown in Figure 4(b). With the evaporation, the water film gets thinner and thinner. When evaporation is for t0, which varies from several minutes to few 10 min, depending on the ambient conditions (temperature and relative humidity), the water film on the substrate was slightly transparent, and the characteristic peaks at 2936 and 2966 cm<sup>1</sup> are very weak but detectable (see curve 1) in Figure 4(a). Timing begins at this moment. The further evaporation leads to appearance of the

Raman peaks [see curves 2–6 in Figure 4(a)]. When the evaporation duration was up to 300 s after timing, the intensity of the characteristic Raman peaks were enhanced to the maximum, as illustrated in curve 7 in Figure 4(a). The corresponding photo of the water film is given in Figure 4 (c), showing translucency or semitransparence. The gold pattern on the substrate looms up. However, the longer the evaporation induced, the rapid is the decrease of the Raman signals and vanishing within 20 s, corresponding to complete evaporation or drying. At this moment, the gold array on the substrate is clearly seen [Figure 4(d)]. Representatively, Figure 5(a) shows intensity of the dominant peak at 2936 cm<sup>1</sup> as a function of evaporation time [the data from

) and continual enhancement of the four

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other two characteristic peaks (at 2861 and 3006 cm<sup>1</sup>

Figure 4(a)] and clearly shows such evolution of the peak intensity.

detect them.

Raman spectral peaks and identification. When a droplet of aqueous solution, with 20 μL in volume and 10<sup>2</sup> M DMMP in initial content, was dropped on the Au bowl-like array without surface modification, it spread out and formed a water film with about 3 mm in thickness. During the initial evaporation, no distinct Raman signal was detected for such water film on the substrate, except the background [see curve (I) in Figure 3(a)]. After evaporation for sufficiently long time (but without completely drying), however, a significant Raman spectrum could be observed. Typically, curve (II) in Figure 3 shows the result corresponding to the evaporation for 300 s after starting timing at which the distinct signals are very weak but detectable, in this experimental condition (see next subsection). At this moment, the water film was enough thin in thickness. There are one dominant Raman peak at 2936 cm<sup>1</sup> and the other three smaller peaks at 2861, 2966 and 3006 cm<sup>1</sup> , respectively. For reference, the Raman spectrum of the pure DMMP (liquid state in a quartz cell) was shown in curve (IV) of Figure 3, which is in good agreement with the previous report [34]. By comparison between curves (II) and (IV), there are about five wavenumbers lower for the water film than the pure DMMP. These four peaks are assigned to the CdH stretching modes of DMMP molecules [39]. Taking into consideration of the possible influence of water on the DMMP, which leads to a slight peak shift, the peaks in curve (II) of Figure 3 can thus be attributed to the DMMP in the water film. It is well known that DMMP molecule is weak Raman scatterer with a very small crosssection on the order of 1 <sup>10</sup>–30 cm<sup>2</sup> under 514.5 nm excitation for the strongest line [40]. Since it is difficult to adsorb on the SERS active metal substrates, the characteristic vibration

Figure 3. The Raman spectra of DMMP molecules. Curves (I), (II) and (III) are for a droplet of 10<sup>2</sup> M DMMP aqueous solution on the SERS substrate after evaporation for a very short time, a certain time and sufficiently long time (completely dried), respectively. Curve (IV): The Raman spectrum of the pure DMMP liquid. [33].

2.2.2. Raman spectral measurements

for (b) and (c), respectively [33].

132 Raman Spectroscopy

three smaller peaks at 2861, 2966 and 3006 cm<sup>1</sup>

The DMMP aqueous solutions with different concentrations were firstly prepared and stored in a refrigerator before use. A droplet of the DMMP aqueous solution was then dropped on the SERS substrate or the gold array and spread out to form the thin water film. After evaporation at room temperature for different time intervals, the Raman spectra for the thin water film were measured on a confocal microprobe Raman spectrometer (Renishaw inVia Reflex) with a

Figure 2. The morphology and wettability of a gold micro-/nanostructured array. (a) The scanning electron microscopic (SEM) image. The inset: A local magnified image and the scale bar = 500 nm. (b) and (c) The photos of the water droplets on the Au array before and after surface modification with thiol, respectively. The water droplets are 2 and 5 μL in volume

Raman spectral peaks and identification. When a droplet of aqueous solution, with 20 μL in volume and 10<sup>2</sup> M DMMP in initial content, was dropped on the Au bowl-like array without surface modification, it spread out and formed a water film with about 3 mm in thickness. During the initial evaporation, no distinct Raman signal was detected for such water film on the substrate, except the background [see curve (I) in Figure 3(a)]. After evaporation for sufficiently long time (but without completely drying), however, a significant Raman spectrum could be observed. Typically, curve (II) in Figure 3 shows the result corresponding to the evaporation for 300 s after starting timing at which the distinct signals are very weak but detectable, in this experimental condition (see next subsection). At this moment, the water film was enough thin in thickness. There are one dominant Raman peak at 2936 cm<sup>1</sup> and the other

spectrum of the pure DMMP (liquid state in a quartz cell) was shown in curve (IV) of Figure 3, which is in good agreement with the previous report [34]. By comparison between curves (II) and (IV), there are about five wavenumbers lower for the water film than the pure DMMP. These four peaks are assigned to the CdH stretching modes of DMMP molecules [39]. Taking into consideration of the possible influence of water on the DMMP, which leads to a slight peak shift, the peaks in curve (II) of Figure 3 can thus be attributed to the DMMP in the water film. It is well known that DMMP molecule is weak Raman scatterer with a very small crosssection on the order of 1 <sup>10</sup>–30 cm<sup>2</sup> under 514.5 nm excitation for the strongest line [40]. Since it is difficult to adsorb on the SERS active metal substrates, the characteristic vibration

, respectively. For reference, the Raman

laser beam of 632.8 nm in wavelength. The Raman spectral integral time is 10 s.

peaks of DMMP molecules in the aqueous solution are usually difficult to be detected by the SERS effect. No characteristic Raman peak from DMMP was observed after the water film was completely evaporated (see curve (III) in Figure 3). Obviously, such method is quite valid to detect them.

Spectral evolution with the evaporation. Further, the Raman spectral evolution of the DMMPcontained water film on the substrate with the evaporation time was measured. Figure 4(a) shows the spectra after evaporation for different durations for the water film with 10<sup>2</sup> M DMMP in initial concentration. During initial stage of the evaporation, no characteristic Raman signals were detected and the film is opaque in the field of optical microscope, as shown in Figure 4(b). With the evaporation, the water film gets thinner and thinner. When evaporation is for t0, which varies from several minutes to few 10 min, depending on the ambient conditions (temperature and relative humidity), the water film on the substrate was slightly transparent, and the characteristic peaks at 2936 and 2966 cm<sup>1</sup> are very weak but detectable (see curve 1) in Figure 4(a). Timing begins at this moment. The further evaporation leads to appearance of the other two characteristic peaks (at 2861 and 3006 cm<sup>1</sup> ) and continual enhancement of the four Raman peaks [see curves 2–6 in Figure 4(a)]. When the evaporation duration was up to 300 s after timing, the intensity of the characteristic Raman peaks were enhanced to the maximum, as illustrated in curve 7 in Figure 4(a). The corresponding photo of the water film is given in Figure 4 (c), showing translucency or semitransparence. The gold pattern on the substrate looms up. However, the longer the evaporation induced, the rapid is the decrease of the Raman signals and vanishing within 20 s, corresponding to complete evaporation or drying. At this moment, the gold array on the substrate is clearly seen [Figure 4(d)]. Representatively, Figure 5(a) shows intensity of the dominant peak at 2936 cm<sup>1</sup> as a function of evaporation time [the data from Figure 4(a)] and clearly shows such evolution of the peak intensity.

Figure 4. Raman spectra and photos of the gold array with a droplet of 10�<sup>2</sup> M DMMP aqueous solution during evaporation. (a) The spectra after evaporation for different intervals. Curves 1–9 correspond to the interval Δt = 0, 112, 155, 199, 243, 279, 301, 323 and 348 s, respectively. (b)–(d) The photos of the gold array covered with water film in the initial stage, microsized-thickness and completely dried stage, respectively [33].

#### 2.3. Concentrating-enhanced Raman scattering (CERS) effect

#### 2.3.1. A quantitative description

The above successful observations of Raman characteristic peaks for the DMMP molecules in an aqueous solution are easily understood. This is mainly attributed to the thin water film confinement and subsequent evaporation-induced DMMP concentrating or enrichment, in addition to the electromagnetic enhancement mechanism from the Au bowl-like array. Because of the space confinement of the thin film, water evaporation induces the concentrating of DMMP within the thinner and thinner film. That is to say, more and more DMMP molecules in the water film are confined within the region with significant local electromagnetic field enhancement above the substrate, exhibiting ever-increasing Raman signal with the increasing evaporation duration, which we could call the concentrating-enhanced Raman scattering (CERS) effect, as demonstrated in Figure 5(a). Obviously, after the water film is completely evaporated, the confinement effect thus vanishes. At this moment, the DMMP molecules cannot stay on the substrate due to the weak interaction, and the corresponding Raman peaks disappear. Besides, the evaporation-induced reorientation of the DMMP molecules could also induce an additional enhancement of the Raman signal owing to the charge-coupling between the molecules and the metallic surface [37].

According to the evolution of the Raman intensity with the evaporation duration shown in Figure 5(a), we could semiquantitatively describe the concentrating kinetics of DMMP molecules in the water film during evaporation. First, under a given ambient condition (temperature and humidity) and a certain volume of solution droplet on the substrate with hydrophilic surface, we have the water film thickness (H) as a function of the water evaporation duration t:

$$H = H\_0 - V \cdot t \quad (t < t\_S) \tag{4}$$

tS <sup>¼</sup> <sup>H</sup><sup>0</sup>

Figure 5. (a) Intensity of the Raman peak at 2936 cm�<sup>1</sup> versus evaporation duration [data from Figure 4(a)]. Point A: The maximal measured value. Point B: The peak vanishes. (b) Plot of the reciprocal Raman intensity versus evaporation

During evaporation of solvent (water), volatilization of the solute or DMMP in water film inevitably takes place. Here, it can be assumed that its volatile rate is directly proportional to

where the volatile rate F is defined as the mole number volatilized in unit time and area

ðt

0

where A is the surface area of the water film. Finally, from Eq. (7), we can quantitatively establish the relation between the solute concentration and evaporation duration of the water film, or

Obviously, if p < < 1 or the solute is nearly involatile in the water, and/or the evaporation speed V value is relatively large enough, the volatile loss ΔM ≈ 0. In this case, Eq. (8) can be approx-

from Eq. (6), the volatile-loss ΔM of the solute (DMMP) in the film can be described by

ΔM ¼ A � p

<sup>A</sup> � <sup>H</sup> <sup>¼</sup>

<sup>C</sup> <sup>¼</sup> <sup>C</sup><sup>0</sup> � <sup>A</sup> � <sup>H</sup><sup>0</sup> � <sup>Δ</sup><sup>M</sup>

) and the parameter p is the proportional constant. So, after evaporation for t,

C<sup>0</sup> � H<sup>0</sup> � p

Ðt 0 C � dt

independent of t, and ts is the duration from the initial to complete evaporation.

where V is the water evaporation speed (m�s

duration [data from (a)]. The solid line is the linear fitting result [33].

its concentration C in the water film, or

(mol s�<sup>1</sup> m�<sup>2</sup>

imately written as

<sup>V</sup> (5)

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�<sup>1</sup> in dimension) and should be a constant

F ¼ p � C (6)

C � dt (7)

<sup>H</sup><sup>0</sup> � <sup>V</sup> � <sup>t</sup> (8)

in which

Figure 5. (a) Intensity of the Raman peak at 2936 cm�<sup>1</sup> versus evaporation duration [data from Figure 4(a)]. Point A: The maximal measured value. Point B: The peak vanishes. (b) Plot of the reciprocal Raman intensity versus evaporation duration [data from (a)]. The solid line is the linear fitting result [33].

$$t\_{\mathbb{S}} = \frac{H\_0}{V} \tag{5}$$

where V is the water evaporation speed (m�s �<sup>1</sup> in dimension) and should be a constant independent of t, and ts is the duration from the initial to complete evaporation.

2.3. Concentrating-enhanced Raman scattering (CERS) effect

initial stage, microsized-thickness and completely dried stage, respectively [33].

The above successful observations of Raman characteristic peaks for the DMMP molecules in an aqueous solution are easily understood. This is mainly attributed to the thin water film confinement and subsequent evaporation-induced DMMP concentrating or enrichment, in addition to the electromagnetic enhancement mechanism from the Au bowl-like array. Because of the space confinement of the thin film, water evaporation induces the concentrating of DMMP within the thinner and thinner film. That is to say, more and more DMMP molecules in the water film are confined within the region with significant local electromagnetic field enhancement above the substrate, exhibiting ever-increasing Raman signal with the increasing evaporation duration, which we could call the concentrating-enhanced Raman scattering (CERS) effect, as demonstrated in Figure 5(a). Obviously, after the water film is completely evaporated, the confinement effect thus vanishes. At this moment, the DMMP molecules cannot stay on the substrate due to the weak interaction, and the corresponding Raman peaks disappear. Besides, the evaporation-induced reorientation of the DMMP molecules could also induce an additional enhancement of the Raman

Figure 4. Raman spectra and photos of the gold array with a droplet of 10�<sup>2</sup> M DMMP aqueous solution during evaporation. (a) The spectra after evaporation for different intervals. Curves 1–9 correspond to the interval Δt = 0, 112, 155, 199, 243, 279, 301, 323 and 348 s, respectively. (b)–(d) The photos of the gold array covered with water film in the

signal owing to the charge-coupling between the molecules and the metallic surface [37].

According to the evolution of the Raman intensity with the evaporation duration shown in Figure 5(a), we could semiquantitatively describe the concentrating kinetics of DMMP molecules in the water film during evaporation. First, under a given ambient condition (temperature and humidity) and a certain volume of solution droplet on the substrate with hydrophilic surface, we have the water film thickness (H) as a function of the water evaporation duration t:

H ¼ H<sup>0</sup> � V � t tð Þ < tS (4)

2.3.1. A quantitative description

134 Raman Spectroscopy

in which

During evaporation of solvent (water), volatilization of the solute or DMMP in water film inevitably takes place. Here, it can be assumed that its volatile rate is directly proportional to its concentration C in the water film, or

$$F = p \cdot \mathbb{C} \tag{6}$$

where the volatile rate F is defined as the mole number volatilized in unit time and area (mol s�<sup>1</sup> m�<sup>2</sup> ) and the parameter p is the proportional constant. So, after evaporation for t, from Eq. (6), the volatile-loss ΔM of the solute (DMMP) in the film can be described by

$$
\Delta M = A \cdot p \int\_0^t \mathbb{C} \cdot dt \tag{7}
$$

where A is the surface area of the water film. Finally, from Eq. (7), we can quantitatively establish the relation between the solute concentration and evaporation duration of the water film, or

$$\mathbf{C} = \frac{\mathbf{C}\_0 \cdot A \cdot H\_0 - \Delta M}{A \cdot H} = \frac{\mathbf{C}\_0 \cdot H\_0 - p \int\_0^t \mathbf{C} \cdot dt}{H\_0 - V \cdot t} \tag{8}$$

Obviously, if p < < 1 or the solute is nearly involatile in the water, and/or the evaporation speed V value is relatively large enough, the volatile loss ΔM ≈ 0. In this case, Eq. (8) can be approximately written as

$$\mathbb{C} \approx \frac{\mathbb{C}\_0 \cdot H\_0}{H\_0 - V \cdot t} \tag{9}$$

measured after evaporation for an optimal duration, for the water film with 10<sup>3</sup> M DMMP in initial concentration. For the substrate without modification, the characteristic peaks of DMMP are very weak [Figure 6(a)]. However, the surface modification induced much stronger Ramanshift peaks of DMMP [Figure 6(b)], and the intensity of the dominant peak at 2936 cm<sup>1</sup> was one order of magnitude (12 times) higher than that without modification. Such Raman enhancement should be mainly attributed to the superhydrophobic surface-induced much higher CF

Figure 6. Raman spectra of 10<sup>3</sup> M DMMP aqueous solution on the substrates without (a) and with (b) surface modifica-

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Evaporation conditions. As mentioned in Section 2.1.1, only when the thickness of the water film is reduced to the nanoscale by evaporation, the concentrating effect, that is, the CERS effect can reach the maximum. However, for the measurements in the normal ambient conditions, as shown in Figure 4, the thickness of the water film, corresponding to the maximal measured Raman signals could be estimated to be in micron scale ( 8–30 μm) according to its initial thickness and the whole time for evaporating the water droplet on the substrate. In the normal ambient conditions, it is too late to measure the Raman spectra when the water film was reduced to the nanoscale in thickness, because such thin water film would be completely evaporated or dried within one millisecond. This is the reason why the Raman peaks disappear immediately after the maximal measured value, as demonstrated in Figure 5(a) (points A and B). It means that the maximal measured value could be much lower than the real maximal one. Although we can obtain the CF value about 10<sup>3</sup> in order of magnitude when the water film was reduced to micron scale in thickness according to Eq. (2) and enough strong CERS

effect, the real maximal or optimal effect is far from reached in the normal ambience.

Obviously, to further increase the CERS effect, we should decrease evaporation speed of the water film, especially, since t = t0 at which the Raman shift peaks are weak but detectable (if the solute volatilization is neglectable). In fact, control of the evaporation speed is easily achieved. For example, we can control the evaporation rate by putting the substrate with water film into

Laser excitation power. Generally, the Raman scattering intensity is directly proportional to the excited laser power. But too high laser power would break down the molecules due to the

value, which led to stronger CERS effect.

tion after an optimal evaporation (the details are given in the text) [33].

a quartz cell with a controllable opening.

or

$$\frac{1}{C} \approx \frac{1}{C\_0} - \frac{V}{C\_0 H\_0} \cdot (t\_0 + \Delta t) \quad t = t\_0 + \Delta t \tag{10}$$

where Δt is the evaporation interval after starting timing at t0 (as mentioned earlier). It means that the reciprocal solute concentration in the water film is nearly directly proportional to the evaporation duration of water. Eqs. (8) and (9) clearly indicate concentrating of the solute during evaporation.

When the solute concentration (C) in the water film is low enough, intensity (I) of its Ramanshift peaks should be directly proportional to C. Here, DMMP is of good water solubility and much heavier molecular weight (124.08) than water [41]. Therefore, Eq. (9) could be a good description of the solute concentration evolution during water evaporation. In other words, from Eq. (10), the reciprocal intensity of the characteristic Raman peaks should be of nearly linear relation with the evaporation duration (t or Δt) of water, which has been confirmed by further spectral analysis. Figure 5(b) shows the results corresponding to the reciprocal Raman intensity of the peak at 2936 cm�<sup>1</sup> versus the evaporation interval Δt [data from Figure 5(a)], showing a good linear relation between them and significant CERS effect. This also implies that the volatile loss of DMMP in water was very small or negligible during water film evaporation at ambient environment. By linear fitting, the plot in Figure 5(b) can be described as

$$\frac{1}{I} = 190.88 - 0.5608 \cdot \Delta t \tag{11}$$

in which the intensity I was multiplied by 10�<sup>5</sup> and Δt is in second.

#### 2.3.2. Factors influencing CERS effect

The CERS effect mentioned earlier would be influenced by some factors such as SERS active substrates, evaporation conditions and volatility of the solute, and so on. Obviously, the highly SERS active substrates, low solute's volatility and appropriate solvent's evaporation rate would be beneficial to exhibiting significant CERS effect. In addition, further experiments have revealed that the surface wettability of SERS substrate and the laser excitation power are also important to induce the strong CERS effect.

Wettability of SERS substrates surface. As mentioned in Section 2.1.2, the substrate with hydrophobic surface should be of stronger concentrating effect than that with hydrophilic surface. For confirmation, the Au micro-/nanostructured bowl-like array shown in Figure 2(a) was surface-modified with thiol. Correspondingly, the static contact angle was increased up to about 160� [Figure 2(c)], exhibiting superhydrophobic surface. Using such modified array as SERS substrate, the CERS effect was really significantly enhanced. Typically, Figure 6 shows the Raman spectrum of DMMP on the gold array with and without surface modification,

<sup>C</sup> <sup>≈</sup> <sup>C</sup><sup>0</sup> � <sup>H</sup><sup>0</sup>

where Δt is the evaporation interval after starting timing at t0 (as mentioned earlier). It means that the reciprocal solute concentration in the water film is nearly directly proportional to the evaporation duration of water. Eqs. (8) and (9) clearly indicate concentrating of the solute

When the solute concentration (C) in the water film is low enough, intensity (I) of its Ramanshift peaks should be directly proportional to C. Here, DMMP is of good water solubility and much heavier molecular weight (124.08) than water [41]. Therefore, Eq. (9) could be a good description of the solute concentration evolution during water evaporation. In other words, from Eq. (10), the reciprocal intensity of the characteristic Raman peaks should be of nearly linear relation with the evaporation duration (t or Δt) of water, which has been confirmed by further spectral analysis. Figure 5(b) shows the results corresponding to the reciprocal Raman intensity of the peak at 2936 cm�<sup>1</sup> versus the evaporation interval Δt [data from Figure 5(a)], showing a good linear relation between them and significant CERS effect. This also implies that the volatile loss of DMMP in water was very small or negligible during water film evaporation

at ambient environment. By linear fitting, the plot in Figure 5(b) can be described as

The CERS effect mentioned earlier would be influenced by some factors such as SERS active substrates, evaporation conditions and volatility of the solute, and so on. Obviously, the highly SERS active substrates, low solute's volatility and appropriate solvent's evaporation rate would be beneficial to exhibiting significant CERS effect. In addition, further experiments have revealed that the surface wettability of SERS substrate and the laser excitation power are also

Wettability of SERS substrates surface. As mentioned in Section 2.1.2, the substrate with hydrophobic surface should be of stronger concentrating effect than that with hydrophilic surface. For confirmation, the Au micro-/nanostructured bowl-like array shown in Figure 2(a) was surface-modified with thiol. Correspondingly, the static contact angle was increased up to about 160� [Figure 2(c)], exhibiting superhydrophobic surface. Using such modified array as SERS substrate, the CERS effect was really significantly enhanced. Typically, Figure 6 shows the Raman spectrum of DMMP on the gold array with and without surface modification,

1

in which the intensity I was multiplied by 10�<sup>5</sup> and Δt is in second.

2.3.2. Factors influencing CERS effect

important to induce the strong CERS effect.

1 <sup>C</sup> <sup>≈</sup> <sup>1</sup> C0 � <sup>V</sup> C0H<sup>0</sup>

or

136 Raman Spectroscopy

during evaporation.

<sup>H</sup><sup>0</sup> � <sup>V</sup> � <sup>t</sup> (9)

� ð Þ t<sup>0</sup> þ Δt t ¼ t<sup>0</sup> þ Δt (10)

<sup>I</sup> <sup>¼</sup> <sup>190</sup>:<sup>88</sup> � <sup>0</sup>:<sup>5608</sup> � <sup>Δ</sup><sup>t</sup> (11)

Figure 6. Raman spectra of 10<sup>3</sup> M DMMP aqueous solution on the substrates without (a) and with (b) surface modification after an optimal evaporation (the details are given in the text) [33].

measured after evaporation for an optimal duration, for the water film with 10<sup>3</sup> M DMMP in initial concentration. For the substrate without modification, the characteristic peaks of DMMP are very weak [Figure 6(a)]. However, the surface modification induced much stronger Ramanshift peaks of DMMP [Figure 6(b)], and the intensity of the dominant peak at 2936 cm<sup>1</sup> was one order of magnitude (12 times) higher than that without modification. Such Raman enhancement should be mainly attributed to the superhydrophobic surface-induced much higher CF value, which led to stronger CERS effect.

Evaporation conditions. As mentioned in Section 2.1.1, only when the thickness of the water film is reduced to the nanoscale by evaporation, the concentrating effect, that is, the CERS effect can reach the maximum. However, for the measurements in the normal ambient conditions, as shown in Figure 4, the thickness of the water film, corresponding to the maximal measured Raman signals could be estimated to be in micron scale ( 8–30 μm) according to its initial thickness and the whole time for evaporating the water droplet on the substrate. In the normal ambient conditions, it is too late to measure the Raman spectra when the water film was reduced to the nanoscale in thickness, because such thin water film would be completely evaporated or dried within one millisecond. This is the reason why the Raman peaks disappear immediately after the maximal measured value, as demonstrated in Figure 5(a) (points A and B). It means that the maximal measured value could be much lower than the real maximal one. Although we can obtain the CF value about 10<sup>3</sup> in order of magnitude when the water film was reduced to micron scale in thickness according to Eq. (2) and enough strong CERS effect, the real maximal or optimal effect is far from reached in the normal ambience.

Obviously, to further increase the CERS effect, we should decrease evaporation speed of the water film, especially, since t = t0 at which the Raman shift peaks are weak but detectable (if the solute volatilization is neglectable). In fact, control of the evaporation speed is easily achieved. For example, we can control the evaporation rate by putting the substrate with water film into a quartz cell with a controllable opening.

Laser excitation power. Generally, the Raman scattering intensity is directly proportional to the excited laser power. But too high laser power would break down the molecules due to the thermal effect, leading to the low Raman signal instead. However, the case here is an exception. The thin water film could protect the target molecules from the laser-induced damage because the water film can remarkably reduce the laser-induced thermal effect, as demonstrated in Figure 7(a), corresponding to the Raman spectra of DMMP in the water film on the substrate, excited with different laser powers under the same evaporation duration. We could use the maximal power (Pmax = 17 mW) of the equipment in this case, while in the conventional measurement only 5 mW or less is usually used. It has been shown that the intensity of the peak at 2936 cm<sup>1</sup> has a good linear relation with the power in whole power range, as indicated in Figure 7(b). The straight line passes through the origin. So, for this thin water film confinement and evaporation concentrating strategy, one can use enough high laser excitation power (>17 mW) to further increase Raman scattering intensity, exhibiting the stronger CERS effect.

3. The surface modification and amidation reaction

MPA in the solution with good selectivity and high sensitivity.

3.1. Surface modification-based SERS detection strategy

3.1.1. Choice of sarin-simulated agent

each other.

In addition to the abovementioned thin water film confinement strategy, here, we introduce another approach to the SERS-based ultrasensitive detection of metal weakly interacted organophosphorus nerve agent sarin based on surface modification of the SERS substrates and amidation reaction [42]. The methanephosphonic acid (MPA) was chosen as the sarin simulation agent (or the target molecule). The Au-coated Si nanocone array was surface-modified with 2-aminoethanethiol molecules and used as the SERS-substrate for detection of MPA. It has been demonstrated that the modified substrate can selectively capture MPA in the solution under the existence of the coupling agent, and hence realize the SERS-based detection of the

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For convenient study of SERS-based detection of sarin, its simulation agent should be chosen. Such simulation agent should be of less or moderate toxicity but the chemical properties and especially the Raman spectrum should be similar to sarin. It has been found that methanephosphonic acid (MPA) is also a suitable simulation agent for sarin, in addition to the commonly used DMMP. The molecular formula of sarin and MPA are (CH3)2CHOOPF(CH3) and CH5O3P, respectively. Both have the C-P bonds and the similar bond length, chemically belonging to the organophosphorus group. Both MPA and sarin can produce amidation reaction with amino compounds [43]. It is expected that these similarities in chemical structures could have similar Raman spectral pattern to

The Raman spectra of sarin and MPA were simulated based on density functional theory (DFT) by means of the Gaussian 09 software [44]. Figure 8(a) is the measured Raman spectrum for the pure MPA. The simulated Raman spectrum is very similar in the primary and minor peaks except the small difference in the peak positions, demonstrating the validity of the spectral simulations. Figure 8(b) shows the simulated Raman spectrum of sarin. Correspondingly, the

Figure 8. The Raman spectra for MPA (a) and sarin (b). (a) The measured Raman spectrum of pure MPA (excited by

785 nm laser). (b) The simulated Raman spectrum of sarin based on DFT calculations [42].

#### 2.4. Suitability of the strategy

Based on the abovementioned text, using the thin water film confinement and evaporation concentrating strategy, one can effectively capture the hydrosoluble and weak affinity molecules within the strong electromagnetic field enhanced space above the SERS substrate and realize the SERS-based detection of them. The thin water film not only confines the target molecules within a limited space but also protects the target molecules from laser-induced damage.

It should be mentioned that the hydrophobic substrate surface, slower evaporation and stronger excitation power can further increase CERS effect. Especially, the slow and controlled evaporation in the anaphase would lead to several orders of magnitude in higher CERS effect. The strategy given here is an effective route to the SERS-based detection of the soluble molecules, which are of small Raman scattering cross-section and hardly adsorbed on the SERS substrates, by choosing proper solvents, but not suitable for the volatile soluble molecules as the liquid film cannot confine these molecules.

Figure 7. (a) The Raman spectra of DMMP aqueous solution droplet on the substrate, under the excitation with different laser powers (P), after the evaporation for the same duration. The maximal excitation power of the equipment Pmax = 17 mW. (b) The plot of the intensity of the peak at 2936 cm<sup>1</sup> versus the laser excitation power [the data are from (a)]. The solid line is the linear fitting results [33].
