The Reaction Kinetics of Glutathione Capped Quantum Dots for the Detection of Hydrogen Peroxide

Research Article

Austin J Biosens & Bioelectron. 2015;1(2): 1010.

The Reaction Kinetics of Glutathione Capped Quantum Dots for the Detection of Hydrogen Peroxide

Imtiaz Ahmad¹, Haife Zhang² and King-Tong Lau¹*

¹Department of Chemistry, Xi’an Jiaotong-Liverpool University, China

²Department of Chemistry, University of Liverpool, UK

*Corresponding author: Kim King Tong Lau, Department of Chemistry, Xi’an Jiaotong-Liverpool University, Suzhou, China

Received: October 13, 2014; Accepted: February 14, 2015; Published: March 31, 2015


Glutathione capped cadmium tellurium quantum dot was investigated as the fluorescence probe for the detection of hydrogen peroxide. In this detection regime, we regard QDs as only the signal transducer for the pseudo first-order reaction between hydrogen peroxide and glutathione. In this way the relationship between the rate of QD fluorescence decay and the rate of change in hydrogen peroxide concentration during the reaction were established and experimentally validated. Hence, the rate constants for the reaction were evaluated. The application of kinetic measurement to chemical analysis was presented and the results were comparable to those obtained from normal measurement at equilibrium.

Keywords: Quantum dots; Hydrogen peroxide sensing; Optical sensors; Fluorescence; Reaction kinetics


Hydrogen peroxide is an important metabolic by product of various physiochemical and pathological processes that involve molecular oxygen. There are many circumstances where the concentration of hydrogen peroxide needs to be measured. Cellular metabolism process involves reduction of molecular oxygen to produce hydrogen peroxide. High concentration of H2O2 can produce reactive oxygen species, which potentially causes DNA impairment and propagate cancer [1-3]. Enzymes such as oxidases produce hydrogen peroxide as a byproduct, [4] hence it is also used as a principal indicator for the detection of a range of important biomolecules, including glucose, [5] cholesterol [6] and triglyceride [7].

Many different sensing techniques have been used for the detection of H2O2 [2,8]. Among these the fluorescence approach [9,10] offers rapid measurement and excellent sensitivity, and the technique has been reported to be used for single molecule tracking [9,11]. The recent emergence of colloidal semiconductor quantum dots has brought significant advancement in fluorescence based sensing [12,13]. QDs have unique size dependent fluorescence emission with narrow emission peak as a result of the quantum confinement effect. Contrast to organic dyes, QDs offer higher photo stability, high fluorescence quantum yield and they can be excited by broad excitation wavelengths [14]. Further, the physical and chemical properties of QD nanocrystals can be tuned by a selection of capping molecules with specific physical and chemical properties [15] which, among other things, provide the QDs with water solubility, biocompatibility and high quantum yield [16].

Glutathione is a natural antioxidant that regulates the redox state of biological system [17] Owing to its biocompatibility, glutathione has been used as capping agent for QDs for use in bio-imaging and sensing applications [18,19] QDs capped with thiols are sensitive to oxidizing agents such as hydrogen peroxide; hence they can be used as chemical sensor to detect these oxidants. The detection of H2O2 by QDs is attributed to the reaction between the surface bound glutathione molecules and hydrogen peroxide. The surface bound glutathione is oxidized to the corresponding disulphide as the main product; this change in surface characteristics disrupts the electronhole recombination process of the QD, resulting in the fluorescence quenching [12].

Mono-dispersed QDs sample is rather like a polymer, consists not of truly mono dispersed particles but of a range of particle sizes or molecular weights. Although the stoichiometry of the reaction between H2O2 and free glutathione in solution to give the corresponding disulphide is well understood, there are still many uncertainties regarding to using QDs as a reagent for H2O2measurement. Further, the degree of QD surface disruption (i.e. the amount of glutathione reacted) to effect measurable fluorescence change has not been established. Hence the reaction stoichiometry between H2O2 molecules and QD particles is not known. Therefore, it is necessary to establish the relationship between the measured QD fluorescence change and the change in H2O2 concentration to validate the use of QDs for the detection of H2O2.

In this work we have synthesized glutathione capped CdTe quantum dots for the study of the fluorescence degradation of QDs by H2O2. A kinetic model is proposed for a pseudo first order reaction with respect to H2O2; in which the rate of change of QD fluorescence intensity is related to the rate of diminishing H2O2. Experimental data were then used to verify the model and evaluate the rate constants.


Chemical s and materials

All the chemicals used were of analytical grade. Tellurium (v) Oxide 99%, citric acid trisodium salt dihydrate 99%, cadmium chloride 99%, sodium borohydride 98%, hydrogen peroxide 30%, glutathione,dipotassium hydrogen phosphate, dihydrogen potassium phosphate and sodium hydroxide were purchased from Acros Chemicals, USA. Milli-Q water (Millipore Co., Billerica, MA, USA) was used for all experiments.

QD Synthesis

Glutathione (GHS) capped cadmium tellurium Quantum Dots (QDs) with red emission was synthesized in-house following reported procedure with modification [20]. In brief, eight mL of 0.04 M cadmium chloride was added to in a round bottomed flask containing trisodium citrate dihydrate (0.2 g), glutathione (0.1 g), TeO2 (0.01 M, 2 mL) in 65 mL of water.NaBH4 (0.1 g) were added with stirring. The mixture was reacted at 90 oC under open-air conditions for a certain period of time. The obtained QDs were precipitated with 1-propanol and the precipitates were separated by centrifugation and were redissolved in50 mM phosphate buffer solution (pH 7.2). The precipitation was repeated three times in order to eliminate the free glutathione ligands from the CdTe QDs colloids.

Characterization of QDs

Fluorescence spectra were obtained with Fluoromax-4 Spectrofluorimeter, Horiba Scientific; uv-vis spectra were obtained with Cary 300 UV-VIS Spectrometer, Agilent Technologies; infrared spectra were obtained with Cary 600 Series FTIP Spectrometer, TEM data were obtained with HR-STEM Transmission Electron Microscope Tecnal G2 F20, FEI Company, USA.

Equilibrium state H2O2 measurement

Buffer solution and hydrogen peroxide solutions were made up in Milli-Q water. QDs solution (0.238 g/L) was made up in 10 mM potassium phosphate buffer adjusted to pH 7.2. 100 mM H2O2 stock solutions were used for analysis. Aliquots of 5, 10, 15, 20, 30, 40, 50 and 60 μL of 100 mM freshly prepared H2O2 stock solution were added into 25 mL QDs solution in a 50 mL conical flask with continues stirring. After each addition of H2O2 the reaction was allowed to proceed for 10 minutes before fluorescence analysis. An excitation wavelength of 420 nmwas used to obtain the all emission spectra.

Kinetic measurement

In brief, a series of H2O2 standards with concentrations of 1.5, 3, 6, 12 and 24 mM were made up in Milli-Q water from a 0.6 M H2O2 stock solution. 10 μL of H2O2 standard was added into a 1 mL cuvette, followed by 1 mL of QDs solution (0.238 g/L). The change of fluorescence intensity at 595 nm was monitored by using an excitation wave length of 420 nm. A sampling rate of 10 data points per second was used and the data were exported to ExcelTM for analysis.

Results and Discussion

Synthesis and characterization of QDs

The one pot synthesis method allows the control of the QDsize simply by monitoring the colour and emission spectrum of the product. Hence, QDs of different sizes were obtained with different reaction time. The uv-vis absorption and emission spectra of the products are presented in Figure 1 & 2 respectively. Table 1 summarized the absorption and emission characteristics of the QDs obtained with different reaction time. The size of the QD products was estimated from the uv-vis absorption spectra using the method described in reference [21].