Detection of pH and Enzyme-Free H2O2 Sensing Mechanism by Using GdOx Membrane in Electrolyte-Insulator-Semiconductor Structure

A 15-nm-thick GdOx membrane in an electrolyte-insulator-semiconductor (EIS) structure shows a higher pH sensitivity of 54.2 mV/pH and enzyme-free hydrogen peroxide (H2O2) detection than those of the bare SiO2 and 3-nm-thick GdOx membranes for the first time. Polycrystalline grain and higher Gd content of the thicker GdOx films are confirmed by transmission electron microscopy (TEM) and X-ray photo-electron spectroscopy (XPS), respectively. In a thicker GdOx membrane, polycrystalline grain has lower energy gap and Gd2+ oxidation states lead to change Gd3+ states in the presence of H2O2, which are confirmed by electron energy loss spectroscopy (EELS). The oxidation/reduction (redox) properties of thicker GdOx membrane with higher Gd content are responsible for detecting H2O2 whereas both bare SiO2 and thinner GdOx membranes do not show sensing. A low detection limit of 1 μM is obtained due to strong catalytic activity of Gd. The reference voltage shift increases with increase of the H2O2 concentration from 1 to 200 μM owing to more generation of Gd3+ ions, and the H2O2 sensing mechanism has been explained as well.


Background
Recently, hydrogen peroxide (H 2 O 2 ) is a major intermediate of biological cycles which has been used as a potential biomarker for oxidative stress diagnosis as well as a major catalyst for immune sensing [1,2]. On the other hand, it is also an essential compound of bleach industries and waste water treatment. H 2 O 2 has a major role in modulating mitochondrial function by inhibiting activities of the mitochondrial enzyme in a fully reversible fashion [3,4]. The H 2 O 2 sensing assay relies on the use of the enzyme horse radish peroxidase (HRP) to oxidize its substrates and detection using spectrophotometer [5]. H 2 O 2 sensing in a simple way, with a short time detection with high specificity, is demanded for future disease diagnosis of the human body, and enzyme-free electro-catalytic methods have gained the attention for H 2 O 2 sensing. Therefore, various catalysts such as metal, metal oxides, and redox polymers have been reported to detect H 2 O 2 [6][7][8][9][10][11][12]. Huang et al. [13] have used the glassy carbon electrode modified by Si nanowire-dispersed CuO nanoparticle. Maji et al. [14] have demonstrated an amperometric H 2 O 2 sensor based on reduced graphene oxide-coated silica modified with Au nanoparticles. Wang et al. [15] have developed a H 2 O 2 sensor by using MoS 2 nanoparticles. Sun et al. [16] have reported a dumbbell-like Pt-Pd-Fe 3 O 4 nanoparticle-modified glassy carbon electrode which shows electro-catalytic reduction. Liu et al. [17] have reported an amperometric H 2 O 2 sensor based on a Si substrate modified with carbon nanotube microelectrode coated by Pd nanoparticles. Kong et al. [18] have reported a non-enzymatic H 2 O 2 sensor based on a Co 3 O 4 nanowire grown over a reduced graphene oxide sheet. Hao et al. [19] have developed an amperometric H 2 O 2 sensor based on Fe 2 O 3 nanoparticles. Bai et al. [20] have reported a sensor based on carbon dot-decorated multi-walled carbon nano-composites. Silver (Ag) nanowire [21] and nanoparticle-decorated graphene [22] have been also reported for H 2 O 2 sensing. Most of the above groups have used different materials using cyclic voltammetry/amperometric methods to sense H 2 O 2 (ranging from few nanomolars to millimolars) due to different oxidation states in the presence of H 2 O 2 . On the other hand, high-k materials such as Al 2 O 3 [23], Ta 2 O 5 [24], and HfO 2 [25] in an electrolyte-insulator-semiconductor (EIS) structure have been reported for pH sensing only; however, the Gd 2 O 3 materials that have been reported are few [26,27], and even then, there is no report for enzyme-free H 2 O 2 sensing by using a GdO x (x < 1.5) material in a simple EIS structure. In this paper, detection of a pH and enzymefree H 2 O 2 sensing mechanism has been investigated by using a GdO x membrane in a simple EIS structure for the first time. Polycrystalline grain, Gd content, and oxidation states (Gd 2+ /Gd 3+ ) have been confirmed by transmission electron microscope (TEM), X-ray photo-electron spectroscopy (XPS), and electron energy loss spectroscopy (EELS) on grain and boundary regions. The 15-nm-thick GdO x membrane detects H 2 O 2 whereas both 3-nm-thick GdO x and bare SiO 2 membranes do not sense H 2 O 2 . Due to the strong catalytic activity of Gd, a low detection limit of 1 μM is obtained. Both time-and concentrationdependent H 2 O 2 sensing and its mechanism have been investigated.
Methods p-type 4-in. Si (100) wafer was cleaned by the Radio Corporation of America (RCA) process. Prior to thermal growth of SiO 2 , HF dip was used to remove native oxide from the surface. After the cleaning process, a 40-nmthick SiO 2 layer was grown as an insulating layer by dry oxidation process at 950°C. Then, the back-side-grown SiO 2 layer was removed by using a buffer oxide etching (BOE) solution. To fabricate the EIS chip, a 300-nmthick Al film was deposited on the back side of the Si wafer. The sensing membrane area was defined by standard photolithography process using a negative photoresist-SU8. Then, EIS devices were attached on a printed circuit board having copper lines. An epoxy layer was used to encapsulate the EIS structure and the copper line. Therefore, a sensor (S1) using SiO 2 membrane was fabricated. Our fabrication process of EIS structure can be found elsewhere [28]. This SiO 2 sensing membrane was modified by deposition of 3-nm-(S2) and 15-nm-thick (S3) GdO x films. The GdO x film was deposited by electron beam evaporation. The Gd 2 O 3 granules were used during deposition, and the deposition rate was 6 nm/min. A schematic view of the Gd 2 O 3 -(or GdO x (x < 1.5)) modified SiO 2 sensor is shown in Fig. 1. To probe the thickness and microstructure of GdO x films, low-voltage spherical aberration corrected field emission TEM (Cs-corrected FE-TEM) was performed. The model number is JEOL JEM-ARM200F with accelerating voltages of 60, 120, and 200 kV. In addition, a Cs-corrected FE-TEM Oxford energy spectrometer (energy-dispersive spectroscopy, EDS) and electron loss EDS (EELS, Model 965 QuantumER TM ) were used to observe the elemental composition on polycrystalline grain and boundaries. The ambient temperature of our laboratory was 21 ± 3°C and relative humidity was 50 ± 10 %. The elemental composition was investigated by XPS analyzing chamber. The vacuum of the XPS chamber was 1 × 10 −9 Torr. The spectra were recorded by using an Al K∝ monochrome X-ray at an energy of 1486.6 eV. The scanning energy range from 0 to 1350 eV was used. All spectra were calibrated by C1s spectrum at a centered peak energy of 284.6 eV. After depositing the GdO x films on the SiO 2 /Si substrates, the samples were transferred immediately to the XPS chamber. The capacitance-voltage (C-V) measurements were performed by using Agilent 4284A LCR meter and an Ag/AgCl reference electrode was used. The measurement frequency was 100 Hz. The sweep voltage was applied on the Ag/AgCl electrode. The reference voltage (V r ) was measured at 50 % of accumulation capacitance. Figure 2 shows the cross-sectional TEM images of the S2 and S3 sensors. The thickness of SiO 2 is 41.2 nm (Fig. 2a), and the thickness of the GdO x film is 3.3 nm (Fig. 2b). The TEM image of the S3 sensor shows that the thickness of SiO 2 is 41.5 nm (Fig. 2c) and the Fig. 1 Schematic view of our pH and H 2 O 2 sensor using Gd 2 O 3 (or GdO x (x < 1.5)) membrane and demonstration of H 2 O 2 sensing mechanism thickness of the GdO x film is 14.8 nm (Fig. 2d). Therefore, the thickness of SiO 2 is 40 ± 2 nm and the thickness of GdO x is 15 ± 0.5 nm. The thicker GdO x film shows clearly polycrystalline grains and its boundary [29,30], which will help to detect H 2 O 2 . Elemental composition of the SiO 2 and GdO x films is observed by XPS, which is shown in Fig. 3. The peak binding energy of Si2p spectra for the S1 sample is 103.35 eV (Fig. 3a), which is similar to the reported value of SiO 2 at 103.58 eV [31]. The spectra are fitted by Shirley background subtraction and Gaussian/Lorentzian functions. The Si2p spectrum shows one characteristic peak after de-convolution. Similarly, one characteristic peak of O1s centered at 531.5 eV is also observed (Fig. 3d). Lower values of full-width half-maximum (FWHM) are found to be 1.84 and 1.64 eV for the Si2p and O1s spectra, respectively. The ratio of O:Si is 1.84, which signifies the stoichiometric SiO 2 . An XPS spectrum of GdO x shows Gd3d 3/2 and Gd3d 5/2 doublet with binding energy of 1220.5 and 1188.3 eV, respectively (not shown here). However, peak binding energies of Gd3d 3/2 and Gd3d 5/2 spin-orbits are reported as 1218 and 1186 eV, respectively [32]. XPS spectra of Gd3d 5/2 core-level electrons are 1189 eV for S2 (Fig. 3b) and 1188.7 eV for S3 (Fig. 3c) samples, which are identified to be Gd 2 O 3 3d 5/2 or Gd 2 O 3 films. Corresponding lower binding energy peaks at 1186.2 and 1185.8 eV indicate the metallic Gd3d 5/2 peaks for the S2 and S3 samples, respectively. The area ratios of Gd/Gd 2 O 3 are found to be 0.64:1 and 0.69:1 for the S2 and S3 samples, respectively, which show higher percentage of Gd in the S3 samples owing to polycrystalline grains. However, the O1s core-level spectra show three distinct peaks for the S2 (Fig. 3e) and S3 (Fig. 3f) samples. The strong peaks at 531.5 eV correspond to the oxygen in the Gd 2 O 3 film, whereas lower (O1s A) and higher (O1s B) binding energy peaks centered at 529 and 532.9 eV are attributed to the hydroxyl (OH − ) and carbonate groups in Gd 2 O 3 films, respectively [33,34]. Moreover, the lower binding energy peak corresponds to Gd-O bonding or GdO x [35]. The area ratios of O1s A and O1s B with respect to O1s are 0.04:1 and 0.48:1 for the S2 samples whereas those values are 0.08:1 and 0.1:1 for the S2 samples, respectively. Therefore, the S2 samples show higher percentage of O1s B owing to higher carbonate groups in the GdO x films, which is insensitive to H 2 O 2 sensing. On the other hand, the S3 samples have higher percentage of O1sA owing to higher OH − and higher Gd content in Gd 2 O 3 film, i.e., GdO x film. So, oxygen can be bonded loosely with Gd on a polycrystalline grain boundary as well as a thicker GdO x film will help to sense H 2 O 2 , which will be explained below. Figure 4a shows the C-V characteristics with pH values from 6 to 10 for the S2 and S3 sensors. The V r values of the S2 sensors are −0.84, −0.75, and −0.63 V for pH 6, 8, and 10, while those values are 0.01, 0.1, and 0.23 V for the S3 sensors, respectively. The V r values of the S3 sensor are shifted towards the positive direction and are lower than the V r values of the S2 sensors. This is due to lower oxide charges for the thicker GdO x membrane (55 vs. 43 nm [36]) and polycrystalline grains with higher OH − ions (Fig. 3f ). The pH sensitivity values are found to be 51.2 and 54.2 mV/pH for the S2 and S3 sensors, respectively, which are higher than the pH sensitivity of approximately 35 mV/pH from pH 2 to 10 [28, 37] and 42 mV/pH from pH 6 to 10 for the S1 sensors. The pH sensitivity of a 30-nm-thick GdO x membrane is approximately 51.7 mV/pH (not shown C-V curves), which is slightly lower than the S3 sensors. The pH sensitivity value of our GdO x membrane is comparable with other reported values of 48.29 mV/pH by Wang et al. [27], 64.78 mV/pH by Chang et al. [38], and 55 mV/pH by Yang et al. [39]. However, the S3 sensors show the lowest drift rate as compared to the S1 and S2 sensors (2.12 mV/h vs. 3.12 mV/h and 2.16 mV/h), as shown in Fig. 4b. The drift characteristics were measured a long time up to 500 min at pH 7 buffer solution. Considering a   3 XPS characteristics of Si2p for a S1, b S2, and c S3 samples. Corresponding O1s spectra of d S1, e S2, and f S3 samples are shown. The S3 film shows higher Gd/Gd 2 O 3 ratio or oxygen deficient and higher OH group which helps to sense H 2 O 2 Fig. 4 a C-V characteristics of the S1, S2, and S3 sensors are shown. b Drift rate characteristics of the S1, S2, and S3 sensors. c Time-dependent response of H 2 O 2 and d reference voltage shift vs. H 2 O 2 concentration for all sensors low drift rate (2.12 mV/h), the pH detection limit of the S3 sensors is 0.039 pH, which is due to high pH sensitivity. It is interesting to note that the GdO x membrane will detect H 2 O 2 . Figure 4c shows the time-dependent response of H 2 O 2 for the S3 sensors. A negligible V r shift is observed for pH 7 buffer solution up to 10 min. By including H 2 O 2 with a concentration of 1 μM, a good V r shift of approximately 40 mV is observed because of Gd 1+ , Gd 2+ , and Gd 3+ oxidation states (https://en.wikipedia.org/wiki/ Work_function) [40]. On the other hand, both S1 and S2 sensors do not show H 2 O 2 sensing. When in contact with H 2 O 2 , the Gd 2+ changes to the Gd 3+ oxidation state and provides electrons for the reduction of H 2 O 2 . H 2 O as a byproduct is observed (Fig. 1). However, the pH value is unchanged by adding H 2 O 2 in the buffer solution. A short response time of <2 min is needed without enzyme. After washing out, the sensor does not show any V r shift owing to the reduction from the Gd 3+ to Gd 2+ states. Therefore, this sensor can be used repeatedly for H 2 O 2 sensing. Based on our knowledge, this is the first ever report of H 2 O 2 detection with a polycrystalline GdO x membrane. Basically, the oxidation/reduction of the GdO x material in contact with H 2 O 2 with buffer solutions is responsible for the V r shifting, which is shown by chemical reactions below.

Results and Discussion
By following the above Eqs. (1), (2), (3), and (4), the oxidation state of Gd changes from Gd 2+ to Gd 3+ . The H + ions are supplied by buffer solutions. The V r shift increases with increasing H 2 O 2 concentration from 1 to 200 μM because the generation of Gd 3+ ions increases (Fig. 4d). A moderate sensitivity of 0.13 mV/μM is obtained from a linear range of 1 to 200 μM whereas it is 82 mV/μM from a linear range of 0.5 to 1 μM. Our detection limit of 1 μM is inferior than the published results [9-12, 15, 16, 41-43], comparable with the published results [44][45][46][47], and superior than the published results [13,17,18,20,[48][49][50][51][52] in literature by using different sensing methods, as shown in Table 1. Further study is needed to improve the detection limit in the future. However, our sensing method's surface potential is changed when in contact with H 2 O 2 because of the catalytic activity of Gd. It is known that Gd 2 O 3 material is n-type and the energy difference in between the Fermi level and the conduction band (E c ) is 2.71 eV [53]. The electron affinity of Gd 2 O 3 is 1.45 eV by considering the conduction band offset of 2.6 eV with Si [54]. The work function of Gd increases from 2.9 eV (https://en.wikipe dia.org/wiki/Work_function) to 4.16-4.76 eV [53][54][55] after oxidation. This suggests that the work function of GdO x is modulated by oxidation/reduction or Gd 3+ concentration as well as the energy band bending of Si is changed. In consequence, the V r is needed to bring Si energy bands to be flat. On the other hand, the S1 and S2 sensors do not show H 2 O 2 detection because they do not have redox properties. The thinner GdO x film (S2) has a smaller crystalline grain with less Gd content (Fig. 3), while the S3 sensor has larger crystalline grain (Fig. 5a) with higher Gd content. Figure 5b shows electron energy loss spectroscopy of Gd measured at polycrystalline grain (P 1 ) and amorphous region or grain boundary (P 1 ). The regions of P 1 and P 2 are marked on  peak at both P 1 and P 2 regions are located at 538.5 eV, as shown in Fig. 5c, which is close to the reported value of 536.5 eV [56]. It is interesting to note that another peak of crystalline grain (P 1 ) is located at 532.9 eV, which is shifted downwards to 3.9 eV. Egerton has reported the reduced energy gap of SiO x at the SiO 2 /Si interface with energy shift downwards to 3 eV [57]. In our case, this reduced energy gap is observed in the polycrystalline grain region. Therefore, the crystalline grain is GdO x (or Gd 2+ ) and the amorphous region or grain boundary is Gd 2 O 3 (or Gd 3+ ). When in contact with H 2 O 2 , the oxidation state of the S3 sensor changes from Gd 2+ to Gd 3+ and the crystalline grain takes a major role, which is confirmed by EELS spectra. So, the thicker crystalline GdO x membrane can sense H 2 O 2 repeatedly which will be useful to detect human disease in the near future.

Conclusions
Higher pH sensitivity (54.2 m/pH) and the enzyme-free H 2 O 2 sensing characteristics have been investigated by using 15-nm-thick GdO x membranes for the first time.
The polycrystalline grain and thickness of the GdO x / SiO 2 film have been observed by TEM image. XPS characteristics of the S3 membrane show higher Gd/Gd 2 O 3 The polycrystalline grain corresponds to Gd 2+ and the grain boundary corresponds to Gd 3+ oxidation states ratio than the S2 membrane (0.69/1 vs. 0.64/1). The S3 membrane shows GdO x and higher OH content in the crystalline grain, which help to sense H 2 O 2 whereas both S1 and S2 sensors do not show H 2 O 2 detection. Therefore, a larger polycrystalline GdO x grain has oxidation/reduction properties when in contact with H 2 O 2 , which is confirmed by EELS. During oxidation, the Gd 2+ changes to the Gd 3+ state and the amount of Gd 3+ ions increases with increasing H 2 O 2 concentration from 1 to 200 μM. A low defection limit of 1 μM is obtained owing to the catalytic effect of Gd. The time-dependent response and the sensing mechanism of H 2 O 2 have been explored. Due to the short time detection of H 2 O 2 in the EIS structure, this novel GdO x sensing membrane paves a way to diagnose other diseases of the human body in the near future.