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Electronic Supplementary Information

Laser-induced TiO2-Decorated Graphene Photoelectrode for Sensitive

Photoelectrochemical Biosensing

Lei Ge, Qing Hong, Hui Li, and Feng Li*

College of Chemistry and Pharmaceutical Sciences, Qingdao Agricultural University, Qingdao,

266109, People’s Republic of China

*Corresponding author: Feng Li

E-mail: lifeng@qau.edu.cn

Telephone: +86-532-86080855

Electronic Supplementary Material (ESI) for ChemComm.This journal is © The Royal Society of Chemistry 2019

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Experimental Section

Reagents. 4,4-Diaminodiphenyl ether (ODA), pyromellitic dianhydride (PDA), titanium(IV)

oxyacetylacetonate (Ti4+-OAA), N,N-dimethylformamide (DMF), ascorbic acid (AA),

acetylthiocholine chloride (ATCl), and analytical standard chlorpyrifos (CP) were obtained from

Sigma-Aldrich Chemical Co. (St. Louis, MO, USA). Acetylcholine esterase (AChE, 200 U/g)

was obtained from Beijing Solarbio Science & Technology Co., Ltd. ITO slices (1.5 × 3.0 cm)

were obtained from China Southern Glass Holding Co., Ltd (ITO coating thickness of 180±20

nm with sheet resistance of 8.1 ± 0.6 cm-2). The ITO slices were cleaned by ultrasonic rinsing

thoroughly in acetone, ethanol, and water, respectively, for 30 min each, followed by washing

copiously with ultrapure water and dried under a stream of N2 gas. Working solutions for AChE

catalyzed hydrolysis of ATCl were freshly prepared (not exceeding 3 h) in 0.01 M phosphate

buffered saline (PBS, pH 7.4) containing 2.0 mM ATCl. Unless other indicated, all the chemicals

used were of analytical reagent grade or above and used without further purification. All buffers

and aqueous solutions were prepared using ultrapure water (≥ 18 MΩcm) obtained from a Milli-

Q Gradient System (Millipore, Bedford, MA).

Apparatus. All photoelectrochemical and electrochemcial experiments were carried out on

a Zahner PEC measurement system (Zahner, Germany) and an Autolab electrochemical

workstation (Metrohm, The Netherlands), respectively, at room temperature with a conventional

three-electrode system comprising a laser-scribed ITO (LITG@ITO, LIG@ITO, or LI-

TiO2@ITO) as working electrode (electrode pattern: square with a width of 3 mm), a Pt wire as

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counter electrode, and an Ag/AgCl as reference electrode. Electrolyte solutions for all

electrochemical characterizations is 1.0 M KCl solution containing 5.0 mM [Fe(CN)6]3−/4−.

HITACHI S3400 scanning electron microscope was used to record the morphologies of the as-

prepared LITG@ITO electrode at an accelerating voltage of 1.0 kV. Transmission electron

microscopic (TEM) images were taken using a Hitachi HT7700 type transmission electron

microscope at an accelerating voltage of 100 kV. High-resolution transmission electron

microscopy (HRTEM) was conducted on a FEI microscope (TF30). Powder X-ray diffraction

(XRD) patterns were obtained on a D8 Advance diffractometer using Cu Kα radiation (λ =

0.15416 Ǻ, Bruck, Germany). Raman spectra were acquired in the range of 50−3400 cm−1 on a

DXRxi Raman spectrometer (Thermo Scientific, America). The diffuse reflectance absorption

spectra were measured by using a U-3900 UV-Vis spectrophotometer (Hitachi, Japan).

Fabrication of LITG@ITO Photoelectrode. First, the synthesis of Ti4+-containing

poly(amic acid) (PAA) mixture was started by dissolving 1.578 g ODA and 0.4 g Ti4+-OAA in 20

mL DMF under magnetic stirring at room temperature, followed by adding 1.745 g PDA, in five

batches with a time interval of 20 min, into the above solution under vigorous stirring.

Subsequently, the resulting flaxen solution was continuously stirred overnight at room

temperature to obtain a homogeneous and viscous Ti4+-containing PAA (Ti4+-PAA) mixture,

which was then spin-coated onto the surface of freshly cleaned ITO glass substrates at 6000 rpm

for 100 s. The resulting uniform Ti4+-PAA layer on ITO glass were then heated in a vacuum oven

at 50 °C, 100 °C, 200 °C, and 300 °C, sequentially with the same time interval of 1 h, to

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dehydrate PAA and form a Ti4+-containing polyimide layer (Ti4+-PI) on the surface of ITO glass.

After naturally cooling down to room temperature, the obtained Ti4+-PI@ITO glass was

subjected to CO2 laser engraving in a commercial CO2 laser (10.6 μm) cutter system (Epilog laser,

Mini) to generate LITG on the surface of ITO glass with self-designed patterns (saved as CDR

files) under room temperature and ambient air. The laser engraving parameters are as follows:

laser Z-distance between the laser head and the substrates was fixed at 52 mm; laser DPI (dots

per inch) was set at 1200; the applied laser engraving power was 12.0% of the full laser power

(40 W); the scan rate was set at 9.0% of the full scan speed (1000 mm/s). Starting with the spin-

casting of 20 mL DMF solution containing 0.4 g Ti4+-OAA onto the surface of freshly cleaned

ITO glass substrates, LI-TiO2@ITO electrode was obtained analogously under the same

conditions for comparison.

Detection Procedures for AChE Inhibitor. First, 50 μL freshly prepared AChE solution (5.0

mU/mL) in 0.01 M PBS (pH 7.4) were mixed with different amounts of CP (dissolved in 2.5 μL

acetone, then diluted by PBS solution, 50 μL) for 1 h incubation at room temperature. Next, 100

μL of working solution was added, and the resulting mixture was incubated at 37 °C for 25 min.

Afterward, the LITG@ITO photoelectrode were immersed into a home-made PEC cell (Fig. S1)

containing the above 200 μL mixture solutions. Finally, the photocurrent responses of the mixture

solution were recorded at a fixed potential of 0.0 V (vs Ag/AgCl) under the illumination with a

~470 nm LED light source.

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Fig. S1 Device schematic for the proposed PEC sensor using LITG@ITO photoelectrode.

Fig. S2 The FTIR spectra of (a) commercial Kapton PI sheet and (b) synthesized Ti4+-PI layer on ITO glass.

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Fig. S3 High resolution TEM images showing (A) the distribution and (B) the crystallinity of TiO2 in LITG.

Fig. S4 XRD patterns of LI-TiO2@ITO photoelectrode and bare ITO electrode (: rutile, : ITO).

A B

d=0.219 nm(111)

d=0.248 nm(101)

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Fig. S5 Photocurrent stability tests of the as-fabricated LITG@ITO photoelectrode by repeated on/off illumination cycles in 0.01 M PBS (pH 7.4) containing 0.1 M AA.

Fig. S6 (A) The plot of transformed Kubelka-Munk function versus the energy of light. (B) Nyquist plots of the as-made LI-TiO2@ITO and LITG@ITO. The Nyquist plots of both LITG@ITO and LI-TiO2@ITO show a high-frequency semicircle, the diameter of which is associated with the charge transfer resistance (Rct) at the photoelectrode/solution interface. Compared with its LI-TiO2@ITO counterparts, LITG@ITO shows a significant decrement in the diameter of semicircle, indicating that LITG@ITO has much smaller Rct than LI-TiO2@ITO.

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Table S1. Assay performance comparison of our platform with other biosensors for chlorpyrifos (M=350.59).

Detection method Linear range Detection limit Reference

Electrochemical method 20 μM to 110 μM 3.5 μM 1

Electrochemical method 10 fM to 1.0 μM 10 fM 2

Electrochemical method 10 nM to 1.0 μM 4.0 nM 3

Electrochemical method 0.1 ng/mL to 105 ng/mL 33 pg/mL 4

Electrochemical method 1.5 nM to 40 nM 1.5 nM 5

Electrochromic method 100 fM to 1.0 mM 0.1 pM 6

Chemiluminescence 0.1 ng/mL to 50 ng/mL 33 pg/mL 7

Chemiluminescence 1.0 ng/mL to 60 ng/mL 33 pg/mL 8

Photoelectrochemical method 0.3 ng/mL to 80 ng/mL 10 pg/mL 9

Photoelectrochemical method 0.2 μM to 16 μM 10 nM 10

Photoelectrochemical method 0.1 ng/mL to 50 ng/mL 30 pg/mL 11

Fluorescence 0.1 nM to 10 μM 0.1 nM 12

Microimmunoassay 0.26 ng/mL to 18 ng/mL 0.11 ng/mL 13

Surface-Enhanced Raman Spectroscopy ---- 1.0 μM 14

Surface-Enhanced Raman Spectroscopy 1.0 nM to 10 μM 0.78 nM 15

Photoelectrochemical method 0.01 ng/mL to 10 ng/mL5.4 pg/mL

(c.a. 15.4 pM)This work

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Fig. S7 (A) Selectivity investigation of the proposed PEC sensor using aspartic acid, glycine, glucose, and thiram as interference. The concentration of chlorpyrifos is 0.1 ng/mL. The concentration of each interference is 50 ng/mL. (B) Operational stability of the proposed PEC sensor towards the detection of 0.1 ng/mL chlorpyrifos. (C) Test of storage stability using ten independent sensors. Left: The photocurrent response of five sensors measured at the first day. Right: The photocurrent response of another five sensors measured after 50 days.

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