A label free colorimetric sensor based on silver nanoparticles directed to hydrogen peroxide and glucose

ORIGINAL ARTICLEA label-free colorimetric sensor based on silver nanoparticles directed to hydrogen peroxide and glucose Nghia Duc Nguyena, Tuan Van Nguyena, Anh Duc Chua, Hoang Vinh Tra

ORIGINAL ARTICLE

A label-free colorimetric sensor based on silver

nanoparticles directed to hydrogen peroxide and

glucose

Nghia Duc Nguyena, Tuan Van Nguyena, Anh Duc Chua, Hoang Vinh Trana,*,

a

Department of Inorganic Chemistry, School of Chemical Engineering, Hanoi University of Science and Technology (HUST), 1st Dai Co Viet Road, Hanoi, Viet Nam

Received 7 November 2017; accepted 31 December 2017

Available online 7 January 2018

KEYWORDS

Graphene quantum dots;

Silver nanoparticles;

Hydrogen peroxide (H 2 O 2 ),

Glucose detection;

Human urine;

Colorimetric sensor

Abstract A simple method has been developed for preparation of silver nanoparticles (AgNPs) based on the use of graphene quantum dots (GQDs) as a reducing agent and a stabilizer The syn-thesized nanocomposites consisting of silver nanoparticles and graphene quantum dots (AgNPs/ GQDs) has been characterized by X-ray diffraction (XRD), Transmission Electron Microscopy (TEM), Ultraviolet–visible spectroscopy (UV–Vis), Fourier-Transform Infrared spectroscopy (FT-IR), Energy Dispersive X-ray spectroscopy (EDX) and Dynamic Light Scattering (DLS) Results indicate that monodisperse of AgNPs has been obtained with particles size ca.
40 nm and specific plasmon peak of silver nanoparticles at 425 nm by UV–Vis spectrum Using AgNPs/ GQDs nanocomposite, we have constructed a colorimetric sensor for hydrogen peroxide (H2O2) and glucose sensors based on the use of AgNPs/GQDs as both probes: capture probe and signal probe The fabricated sensors perform good sensitivity and selectivity with a low detection limit

of 162 nM and 30lM for H2O2and glucose sensing, respectively Moreover, the biosensors have been successfully applied to detect glucose concentrations in human urine

Ó 2018 The Authors Production and hosting by Elsevier B.V on behalf of King Saud University This is

an open access article under the CC BY-NC-ND license ( http://creativecommons.org/licenses/by-nc-nd/4.0/ ).

1 Introduction

Diabetes, as well known, is a serious health problem, which has been declared as a global epidemic by World Health Orga-nization (WHO) owing to its unprecedented growth worldwide (Jia et al., 2015; Vashist, 2012) The glucose level in blood is used as a clinical indicator of diabetes (Su et al., 2012; Baghayeri et al., 2016; Lu et al., 2015; Ensafi et al., 2016; Gao et al., 2017) However, drawing blood from vein or finger-tip causes discomfort and pricking sensation Compared with

* Corresponding author.

E-mail address: hoang.tranvinh@hust.edu.vn (H.V Tran).

Peer review under responsibility of King Saud University.

Production and hosting by Elsevier

King Saud University Arabian Journal of Chemistry

www.ksu.edu.sa

www.sciencedirect.com

https://doi.org/10.1016/j.arabjc.2017.12.035

1878-5352 Ó 2018 The Authors Production and hosting by Elsevier B.V on behalf of King Saud University.

This is an open access article under the CC BY-NC-ND license ( http://creativecommons.org/licenses/by-nc-nd/4.0/ ).

blood, urine is another informative body fluid as well and

more importantly, it can be obtained noninvasively The

glu-cose level in urine is also a good indicator for preliminary

screening of patients with high level diabetes or having renal

glycosuria (Jia et al., 2015; Radhakumary and Sreenivasan,

2011) In order to avoid the inconveniences caused by drawing

blood intravenously or by hand pricking, a preliminary

screen-ing of the patients with high level diabetes can be done

instantly by checking their urine glucose levels When the

con-centration of glucose in urine is more than 500–1000 mg/L

(2.8–5.6 mM), the urine test is positive (Su et al., 2012;

Radhakumary and Sreenivasan, 2011; Fine, 1965; Lankelma

et al., 2012; Urakami et al., 2008; Zhang et al., 2017)

Consid-ering its convenience, painlessness, and affordability, urine

glucose monitoring should not be completely given up,

espe-cially in low-income regions (Su et al., 2012)

The sensing of glucose is usually based on electrical signal

or color change generated by the specific reaction of active

spe-cies (e.g., glucose oxidase or phenylboronic acid) with glucose

(Jia et al., 2015) Most glucose sensors have been structured

based on natural enzymes (e.g., horseradish peroxidase

(HRP)) Natural enzymes in organisms are proteins composing

of hundreds of amino acids that can catalyze chemical

reac-tions It has been widely applied in various fields because of

their high substrate specificity and catalytic efficiency

How-ever, their catalytic activity can be easily affected by

environ-mental conditions such as acidity, temperature and

inhibitors Furthermore, high costs of preparation,

purifica-tion and storage also restrict their widespread applicapurifica-tions

(Ding et al., 2016; Tran et al., 2018; Zhang et al., 2014; Xing

et al., 2014) In the light of this, exploiting stable enzyme

mimetics is an urgent need Nowadays, many nanomaterials

with unique peroxidase-like activity have been discovered,

including magnetic nanoparticles and its composite (Ding

et al., 2016; Wei and Wang, 2008; Dong et al., 2012), cerium

oxide nanoparticles (Zhao et al., 2015), silver nanoparticles

(Tran et al., 2018), carbon-based nanomaterials (Wang et al.,

2015; Nirala et al., 2015; Wang et al., 2016); exfoliated Co–

Al layered double hydroxides (Co-Al ELDHs) (Chen et al.,

2013) and manganese selenide nanoparticles (MnSe NPs)

(Qiao et al., 2014) These nanostructured materials as

peroxidase mimetics show unparalleled advantages of low

cost and stability over natural enzymes (Ding et al., 2016)

Among them, carbon-based nanomaterials, such as graphene/

graphene oxide; carbon nanotubes and graphene quantum

dots are the most widely studied enzyme mimics (Shu and

Tang, 2017)

In this work, we synthesized nanocomposites consisting of

silver nanoparticles and graphene quantum dots (AgNPs/

GQDs) by a simple and green method Using AgNPs/GQDs,

we constructed a directed colorimetric method for the direct

detection of hydrogen peroxide (H2O2) A colorimetric glucose

sensor has been designed and developed based on combining

with glucose oxidase (GOx) The fabricated sensors perform

excellent sensitivity and selectivity for hydrogen peroxide and

glucose sensing Moreover, the proposed sensor has been

suc-cessfully applied to detect of glucose concentrations in human

urine samples Based on the good performances, the proposed

colorimetric glucose sensor becomes a great promising

candi-date for glucose level sensing as a without blood needing and

needle-free approach

2 Experimental 2.1 Chemical

Citric acid (C6H8O7H2O); urea ((NH2)2CO); ammonia (NH3) solution 28%wt.; acetic acid (CH3COOH) solution 99%wt.; sodium hydroxide (NaOH); silver nitrate (AgNO3); glucose; ascorbic acid; galactose; fructose; lactose; scructose; hydrogen peroxide solution 30% (H2O2); phosphate buffered saline tablets (PBS); and glucose oxidase (GOx) were purchased from Sigma Aldrich Human urine samples were collected from a local hospital

2.2 Synthesis of graphene quantum dots (GQDs)

3.44 g citric acid and 3.005 g urea were dissolved into 100 mL distilled (D.I) water The solution was transferred to an auto-clave and heated at 160°C for 4 h Then, the mixture was cen-trifuged at 5000 rpm for 20 min to remove the big carbon particles The supernatant containing graphene quantum dots (GQDs) was collected

2.3 Synthesis of silver nanoparticles (AgNPs) using GQDs as reducing reagent and stabilizer

100mL of GQDs stock solution was diluted by 3 mL of D.I water Then 0.1 M NaOH and 1 M CH3COOH solutions were used to control pH of GQDs solutions from 3 to 11 After that,

20lL of 0.1 M AgNO3 solution was added into the GQDs solutions The mixtures were heated to 90°C for 3 h to com-plete reduction of silver cation (Ag+) to silver nanoparticles (AgNPs) process to form nanocomposites consisting of silver nanoparticle and graphene quantum dots (AgNPs/GQDs) as the results AgNPs/GQDs solutions then were cooled to room temperature (RT) and stored at 4°C for use

2.4 Characterization

UV–Vis spectra were measured using Agilent 8453 UV–Vis spectrophotometer system with the wavelength in a range of 200–1200 nm Morphology and crystal structure of nanoparti-cles were characterized using Transmission Electron Micro-scopy (TEM: JEM1010 - JEOL) Particles size distribution was analysed by Dynamic Light Scattering (DLS) on the Nano Partica SZ-100 (HORIBA Scientific, Japan) XRD pattern of AgNPs/GQDs was measured using D8 ADVANCE - Bruker Chemical composition of samples was determined by JEOL Scanning Electron Microscope/Energy Dispersive X-ray (SEM/EDS) JSM-7600F Spectrometer

2.5 Direct detection of hydrogen peroxide

200lL of H2O2 solutions with different concentrations was added into a 1.5 mL eppendorf Then, 1000lL of AgNPs/ GQDs solution was added into the eppendorf and the mixture was stirred by vortex machine The mixture was then incu-bated at 40°C in a water bath for 30 minutes Then the UV–Vis spectra of the solutions were recorded The optical densities at 425 nm (OD425) of the AgNPs/GQDs solution before and after addition of various HO quantities were used

to draw a calibration curve, i.e.DA/A0vs [H2O2] the following

equation:

DA

A0 ð%Þ ¼A0 Ac

Here, A0and ACare OD425of the AgNPs/GQDs solution

before and after H2O2addition, respectively

2.6 Detection of glucose

100lL of glucose solutions with the different concentrations

(from 0.5 mM to 8 mM) in PBS buffer (pH = 7) were added

into eppendorfs, then after, 100lL of GOx (2 mg mL1 in

0.001 M PBS solution) solution was added The solution was

mixed and incubated in a 37°C water bath for 30 min Then,

1000lL of the AgNPs/GQDs solution was added to the above

eppendorfs Finally, the mixed solutions were incubated in a

40°C water bath for 30 min and then they were transferred

to cuvettes for UV–Vis absorbance measurement and the

opti-cal density at wavelength of 425 nm was recorded The optiopti-cal

densities at 425 nm (OD425) of the AgNPs/GQDs solution

before and after addition of various glucose quantities and

GOx were used to draw a calibration curve, i.e.DA/A0 (Eq

(1)) vs Cglucose (here, DA = A0 AC where A0 and AC are

OD425of the AgNPs/GQDs solution before and after adding

the mixture of glucose and GOx, respectively)

3 Results and discussions

3.1 Characterization of AgNPs/GQDs hybrid

The simple method has been developed for the preparation of

nanocomposites consisting of silver nanoparticles and

gra-phene quantum dots (AgNPs/GQDs) First, the small sized

graphene quantum dots (GQDs) with abundant oxygen

con-taining functional groups have been synthesized by the

hydrothermal method Then GQDs adsorbed Ag+ ions and

reduced them into silver nanoparticles (AgNPs) without

add-ing any reducadd-ing reagents, while the oxygen containadd-ing

func-tional groups were partially removed from the GQDs Thus,

GQDs were coated on the surfaces of the resultant AgNPs,

leading to the formation of AgNPs/GQDs The residual

oxygen-containing groups on the GQDs made the obtained

AgNPs/GQDs be excellent dispersive and long-term stable in

water (Tetsuka et al., 2012)

The UV–Vis spectra of GQDs and AgNPs/GQDs

solu-tions have been shown in Fig 1A As can be seen in

Fig 1A, curve b, the adsorption band at 425 nm is attributed

to the characteristic surface plasmon absorption of AgNPs,

while this absorption is not observed in the case of the control

sample (solution containing only GQDs, without AgNO3),

where no AgNPs are formed (Fig 1A, curve a) A shoulder

at 357 nm inFig 1A (curve b) can be attributed to the

pres-ence of GQDs in AgNPs/GQDs solution when comparing to

UV–Vis spectra of GQDs (Fig 1A, curve a) Moreover, the

synthesized AgNPs/GQDs have been characterized by DLS

(Fig 1B), XRD (Fig 1C) and TEM (Fig 1D) DLS data

(Fig 1B) have indicated that AgNPs/GQDs have particles size

from 20 nm to 100 nm with mean size at 40 nm Besides that,

TEM analysis (Fig 1D) shows that AgNPs/QGDs are

spher-ical particles with particles size around 40 nm These data are

in a highly agreement with the DLS results The XRD pattern

of the AgNPs/GQDs (Fig 1C) shows three main characteris-tic peaks at 2h = 37.5°, 43.1° and 64.8° which match very well with those of the standard AgNPs (PCPDF card number 40,783) (Mamatha et al., 2017) with Miller indices (1 1 1), (2 0 0) and (2 2 0) Normally, X-ray diffraction of GQDs pre-sents a weak broad peak (0 0 2) centered at 2h
22.7° which indicates the disordered stacking structures of graphene lay-ers; however, this (0 0 2) peak is strongly depend on the degree of oxidation of GQDs because the attached hydroxyl, epoxy/ether, carbonyl and carboxylic acid groups can increase the interlayer spacing of GQDs (Tetsuka et al., 2012) In Fig 1C, no specific XRD peak of GQDs can be seen, possibly this peak is too weak and overlapped by the background sig-nal The EDS spectra of GQDs (Fig 1E, curve a) showed the peaks of C, O and N, which were three major constituents of GQDs EDX spectra of AgNPs/GQDs hybrid (Fig 1E, curve b) presented new appearing peaks, corresponding to Ag The EDS spectra provided an evidence for silver metal forming by GQDs: strong peak values at 2.99 and 3.17 keV were due to forming of AgNPs These results confirmed that AgNPs were efficiently formed onto surface of GQDs.Fig 1F shows FTIR spectra of GQDs (curve a) and AgNPs/GQDs hybrid (curve b) Fig 1F (curve a) shows that the bands at 3100–3500

cm1 belong to t(OAH) and t(NAH), which is important

to facilitate the hydrophilicity and stability of the GQDs in aqueous state The absorption bands at 1641 cm1is attribu-ted tot(C ‚ O), demonstrating that carboxylic acid may be used as Ag+ binding site These peaks indicate that GQDs have abundance of amino (ANH2), carboxyl (ACOOH) and hydroxy (AOH) groups on their surface and edges responsible for the excellent hydrophilicity of GQDs Interestingly, com-pared with the GQDs, the absorption bands of the OAH group at 1064 cm1almost disappear in the FT-IR spectrum

of the AgNPs/GQDs hybrid (Fig 1F, curve b) These results indicate that Ag+ can be reduced to form AgNPs by OAH groups on the GQD/AgNP hybrid’s surface, resulting in

OAH groups being converted into –COOH groups after the reaction

3.2 Hydrogen peroxide detection 3.2.1 Spectrometric assay for hydrogen peroxide detection and effect of pH

The colorimetric H2O2sensor was constructed basing on the reaction of AgNPs with H2O2, which leaded the change of the color of AgNPs/GQDs solutions from yellowish to color-less, depending on H2O2 concentration As can be seen in Fig 2, the presence of AgNPs/GQDs in the solution results

in a strong absorption band at 425 nm (Fig 2a to Fig 2g, curve (i)), corresponding to the yellowish color The optical density of the AgNPs/GQDs solution at 425 nm (OD425) decreases after addition 200mL of H2O2 50 mM (Fig 2a to Fig 2g, curve (ii)), corresponding to the color changing of the solution from yellowish to colorless This result is explained by the oxidation of AgNPs in the presence of

H2O2 The standard potential of Ag+/Ag is lower than that

of H2O2/H2O (E0

Ag þ =Ag= 0.8 V < E0

H 2 O 2 =H 2 O= 1.77 V) in water at pH = 7 The following reaction will occur (Eq.(2)): ðGQDsÞAg0þ H O ! ðGQDsÞAgþþ 2HO ð2Þ

Therefore, AgNPs in AgNPs/GQDs hybrid will be etched

from Ag0to Ag+ So the concentration of Ag0will decrease,

leading to the fading of the AgNPs/GQDs solution after

add-ing H2O2 The above behaviour provides a potential for quan-titative detection of H2O2 by measuring the decrease in the AgNPs surface plasmon resonance at 425 nm

0.0

0.2

0.4

0.6

0.8

(b)

Wavelength (nm)

420 nm

343 nm

(a)

357 nm

(A)

(a) (b)

0.0 2.0 4.0 6.0 8.0 10.0

Particle size (nm)

(B)

0

20

40

60

80

100

120

2 / degree

Ag Ag

C

O

N

(b)

Energy (keV)

(a) C

O

N

(E)

AgNPs/GQDs

(b)

Wavenumber (cm-1)

(a)

(F)

GQDs

(D)

θ

Fig 1 (A) UV–Vis spectra of (a) GQDs and (b) AgNPs/GQDs (Inset: color of the corresponding samples); (B) Particles size distribution

of AgNPs/GQDs by DLS method; (C) XRD pattern of AgNPs/GQDs; (D) TEM image of AgNPs/GQDs; (E) EDX of (a) GQDs and (b) AgNPs/GQDs; (F) FT-IR of (a) GQDs and (b) AgNPs/GQDs

300 400 500 600 0.0

0.2 0.4 0.6 0.8 1.0

Wavelength (nm)

pH = 11

(i)

(ii)

(a)

A

0.0 0.2 0.4 0.6 0.8 1.0

Wavelength (nm)

pH = 9

A

(b)

(ii) (i)

0.0 0.2 0.4 0.6 0.8 1.0

Wavelength (nm)

pH= 7.5

(c)

(i) (ii)

A

0.0 0.2 0.4 0.6 0.8 1.0

Wavelength (nm)

pH = 7

(d)

A

(i)

(ii)

0.0 0.2 0.4 0.6 0.8 1.0

Wavelength (nm)

pH = 5

(e)

A

(i)

(ii)

0.0 0.2 0.4 0.6 0.8 1.0

Wavelength (nm)

pH = 4

(f)

A (i)

(ii)

Δ

Δ

Δ

Δ

Δ Δ

0.0 0.2 0.4 0.6 0.8 1.0

(ii)

Wavelength (nm)

pH = 3

(i)

(g)

A

2 3 4 5 6 7 8 9 10 11 12 20

40 60 80 100

A0

pH

(h)

Δ

Fig 2 UV–vis spectrum of AgNPs/GQDs solutions: (i) before and (ii) after addition of 50lM hydrogen peroxide at room temperature and reaction time was 15 min at different pH: (a) pH = 11, (b) pH = 9, (c) pH = 7.5, (d) pH = 7, (e) pH = 5, (f) pH = 4, (g) pH = 3; (h) summarization of effect of pH on response signal of hydrogen peroxide sensors based on AgNPs/GQDs

UV–vis spectra of AgNPs/GQDs solutions at different pH

values without (curve i) and with (curve ii) 50 mM H2O2are

shown in Fig 2a–g, which corresponding with pH from 11

to 3 It can be seen, when H2O2was adding, the adsorption

at 425 nm (OD425) was decreased The decreasing of OD425

was strongly depended on pH of solution Fig 2h presents

the summarizing of the effect of pH on response signal of

H2O2 sensors based on AgNPs/GQDs solution, which was

given on the graphDA/A0vs pH (here,DA = A0 ACwhere

A0and ACare OD425of the AgNPs/GQDs solution before and

after H2O2addition, respectively) As can be seen inFig 2h,

the value ofDA/A0is higher in acid environment than that

in base environment This result is explained by the

disintegra-tion of H2O2in base environment according to the following

equation (Eq.(3)):

Therefore, in base environment, the decreasing of the

con-centration of Ag0according to Eq (1) is lower than that in

acid environment The maximum value ofDA/A0was obtained

(ca 96.61%) at pH = 7, so that following experiments will be performed in neutral environment

3.2.2 Sensitivity of the sensor The UV–Vis spectra of samples containing different H2O2 con-centrations are shown inFig 3A When the concentration of

H2O2 increases from 0.5lM to 100 lM, the optical density

of the AgNPs/GQDs solution at 425 nm (OD425) decreases from 0.6 (a.u) to 0.07 (a.u) It can be seen that, a shoulder at

357 nm appears more clearly which can be attributed to speci-fic plasmon peak of free GQDs in solution This phenomenon can be explained following: when H2O2 is added, H2O2 will react with AgNPs (Eq (2)), therefore, GQDs from AgNPs/ GQDs will be released to free GQDs in solution

Using OD425as the recorded signal, the calibration curve of hydrogen peroxide detection was generated under optimum conditions has been shown inFig 3B byDA/A0vsH2O2 con-centration (Eq.(1)) In the calibration, the linear relationship

ofDA/A0vsH2O2 concentration is in range from 0.5lM to

50lM with the regression equation DA/A0= (1734 ± 72.58) CH2O2 (mM) + (2.74412 ± 1.79846) with R2= 0.98615 Based on the calibration curve and the blank samples, the limit of H2O2detection (LOD) of the sensor is estimated of

to be 162 nM) Moreover, it is able to monitor the color chang-ing of the AgNPs/GQDs solution by naked eyes in the case of immediate and qualitative H2O2detection (Fig 3B, insert)

3.3 Glucose detection 3.3.1 Sensitivity of the sensor When glucose and GOx are added into the solution containing AgNPs/GQDs, the following reaction will occur:

Glucoseþ O2þ H2O! D-glucono-1; 5-lactone þ H2O2

After that, H2O2is measured by using the fabricated colori-metric sensor based on AgNPs/GQDs.Fig 4A shows the UV– Vis spectra of samples containing different glucose concentra-tions When the concentration of glucose increases from 0.5

mM to 8 mM, the optical densities of the AgNPs/GQDs solu-tion at 425 nm (OD425) are decreased from 0.61 (a.u) to 0.31 (a.u)

The calibration curve of glucose detection is shown in Fig 4B by DA/A0 vs glucose concentration as mentioned above In the calibration (Fig 4B), the linear relationship of DA/A0vs glucose concentration is in range from 0.5 mM to

8 mM with the regression equation DA/A0= (7.06087 ± 0.40925) Cglucose (mM) + (3.05473 ± 1.49321) with R2= 0.97695 The limit of detection (LOD) was estimated to be

30lM based on three times the standard deviation of the blank tests, which is comparable to those of the previously reported methods (Table 1) The linear range of the sensor is from 0.5 mM to 8 mM and the LOD value (30lM) is lower than the value of the concentration of glucose in a urine sam-ple which is positive for diabetes (2.8–5.6 mM) Thus, the developed colorimetric glucose sensor has great potential for application to a daily glucose test In addition, the above results have indicated that the synthesized AgNPs/GQDs nanostructured material not only has catalytic efficiency as peroxidase mimetics but also shows unparalleled advantages

of low cost and stability over natural enzymes

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

Wavelength (nm)

0 0.5x10 -3

1x10 -3

5x10 -3

10x10 -3

20x10 -3

30x10 -3

40x10 -3

50x10 -3

100x10 -3 (A)

[H 2

0

20

40

60

80

100

0.

m

0.

m

0.1 mM

(B)

0.05 mM 0.0 4 m M

0 mM

0.0005 mM

0.00 1 m M

0.0

05 m M

0.0 1 M

Fig 3 (A) UV–Vis spectra of hydrogen peroxide sensor with

various H2O2 concentrations; (B) Calibration curve for H2O2

detection (inset: color of sensor with corresponding samples in

(A)) Experimental conditions were described in the text

Moreover, as can be seen in Fig 4A, when the concen-tration of glucose increases from 0.5 mM to 8 mM, the shoulder at 357 nm which can be attributed to specific plas-mon peak of free GQDs in solution appears more clearly This result is in good agreement with the result obtained

in the case of H2O2 sensors (Fig 3A) This result is an important clue for suggestion of a glucose detection mecha-nism following two steps (Fig 5): In the first step, glucose is converted to D-glucono-1,5-lactone (which is named as glu-conic acid) and H2O2 following Eq (4) Then, AgNPs/ GQDs are etched by H2O2 (Eq (2)) in the second step Therefore, GQDs from AgNPs/GQDs nanocomposite will

be released to free GQDs in solution, leading to the appear-ance more clearly of the shoulder at 357 nm The above result is a new point in the comparison with previous work (Chen et al., 2014; Xia et al., 2013) In this work, the phe-nomenon of etching AgNPs and releasing GQDs can be seen

by experimental results It is thanks to the excellent disper-sive and the long-term stable in water of the synthesized AgNPs/GQDs nanocomposite

3.3.2 Selectivity of the sensor The selectivity of the glucose sensor was tested by conduct-ing the control experiments in the presence of glucose, galactose, lactose, sucrose and fructose at concentration of

4 mM

It can be seen inFig 6A, a small decreasing of OD425was found when galactose, lactose, sucrose and fructose were added When glucose was added, a strong decreasing of

OD425was obtained TheDA/A0values of sensor when using various saccharides at concentration of 4 mM were summa-rized inFig 6B It can be found that theDA/A0values were 14.59% for presence of galactose; 11.27% for lactose, 17.34% for sucrose, 16.29% for fructose These values are lower than that of the solution containing glucose (DA/A0

= 54.76%) at least 3.16 times at the same concentration These results have indicated the excellent selectivity for glucose

of the developed sensor

0.0

0.2

0.4

0.6

Wavelength (nm)

0.0 1.0 3.0 4.0

(A)

0 mM

0.5 m M 1.0 m 2.0

m 3.0

M

3.

m M

0

10

20

30

40

50

60

Glucose concentration/ mM

(B)

Fig 4 (A) UV–Vis spectra of glucose sensor with various

glucose concentrations (inset: color of sensor with various glucose

concentrations); (B) Calibration curve for glucose detection

Experimental conditions were described in the text

Table 1 Comparison with some reports based on label-free colorimetric methods for the detection of glucose

immobilization

detection ( lM)

Actual samples Reference

Sreenivasan (2011 )

DNA-embedded Au@Ag

nanoparticles

GOx 0.01  10 3 –0.2  10 3 ; 1 

103–100  10 3 0.01 Fetal bovine serum ( Kang et al., 2015 ) Au@Ag core–shell

nanoparticles

human serum

( Zhang et al., 2016 )

P(DMA-co-PBMA)

copolymer and AuNPs

GOx N/R 50 N/R ( Li et al., 2011 )

N/R-not reported.

3.3.3 Application of the sensor for detection glucose in human urine sample

Human urine sample was firstly diluted because human urine may contain many soluble salts and residues In our previous work on human urine samples (Tran et al.,

2017), we have found that high diluted ratio gives better signal than low diluted ratio However, because of the lim-its of the LOD, the optimized dilution ratio was 1:4 The standard addition method was used to analyse glucose con-centration in the human urine sample using the proposed sensor

Fig 7A shows UV–Vis spectra the glucose sensor in pres-ence of the diluted urine sample and the three spiked urine samples by adding glucose with concentration from 1 mM

to 3 mM The calibration curve for determination of glucose concentration using the standard addition method with the human urine sample is described inFig 7B From these data, glucose concentration in the human urine sample has been determined of 3.68 mM Therefore, it is able to conclude that the above human urine sample is of a diabetic patient This experimental result shows the great potential for application

of the developed colorimetric glucose sensor based on a low-cost, blood-free and needle-free approach to daily glu-cose tests

4 Conclusions

In this work, silver nanoparticles decorated graphene quantum dots carbon (AgNPs/GQDs) hybrids have been synthesized and characterized by DLS, XRD, FT-IR, EDX and TEM methods; and the results indicate that mono-dispersed AgNPs have been obtained with particles size ca.
40 nm Using AgNPs/GQDs as capture probe and signal probe, a spec-troscopy method has been developed for determination of hydrogen peroxide with a low detection limit of 162 nM of

UV-Vis spectra

(a) (b)

H2O2

H2O2

H2O2

H2O2

H2O2

H2O2

Reaction

H2O2

AgNPs Ag +

GQDs

Glucose + O2+ H2O D-Glucono-1,5-Lactone + H2O2

STEP 1

STEP 2

(Gluconic Acid)

Glucose Oxidase-GOx

GOx

without glucose

with glucose

Fig 5 Illustration of detection mechanism of proposed label free and reagentless colorimetric sensor for hydrogen peroxide and glucose using AgNPs/GQDs as capture probe and signal probe

(A)

(B)

Fig 6 (A) UV–Vis spectra of the glucose sensor in presence of

different saccharides at concentration of 4 mM; (B)

Correspond-ing of DA/A0 of (A) (inset: color of sensor with various

saccharides)

H2O2 Combining with the use of glucose oxidase (GOx), a

simple colorimetric method for selective and sensitive detection

of glucose has also been fabricated The above sensors perform

excellent sensitivity and selectivity with a low detection limit of

30lM of glucose concentration In addition, the level of

glu-cose in the real human urine sample can also be measured

accurately by using the AgNPs/GQDs-based colorimetric

sen-sor following the addition standard method Therefore, the

proposed colorimetric glucose sensor has great potential for

application to a daily glucose test based on a low-cost;

blood-free and needle-free approach

Acknowledgments

This research was funded by Vietnam National Foundation

for Science and Technology Development (NAFOSTED)

under grant number 104.99-2016.23

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0.0

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(5i) (4i) (3i) (2i)

Wavelength (nm)

(i) AgNPs/GQDs (2i) = (i) + diluted urine (3i) = (2i) + 1 mM glucose (5i) = (3i) + 3 mM glucose

(A) (i)

0

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[Additional glucose] (mM)

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0.92mM

Fig 7 Method of standard addition for glucose detection in

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