Electrochemical Determination of Acetaminophen Using Gold Nanoparticles–Multiwall Carbon Nanotubes Modified Glassy Carbon Electrode

Since their discovery in 1991, carbon nanotubes (CNTs) have been extensively studied for use in emitting devices, energy storage, sensor field, and biological and biomedical applications.1 Gold nanoparticles are well known for being biocompatible materials with strong adsorption ability and good conductivity; thus they strongly interact with biomaterials. A film containing both CNTs and gold nanoparticles (AuNPs) on the electrode surfaces will lead to synergism between the properties of these nanoparticles.2 Carbon nanotubes combined with gold nanoparticles (AuNPs), resulting in AuNPs/CNTs hybrid materials, combine the high conductivity of the CNTs with the size-dependent optothermal properties of the AuNPs. Potential direct applications include use as gas sensors, catalysts and especially as structural components of electrochemical sensors.3,4

Acetaminophen (ACT) is an important medicine used widely both in its pure form and in pharmaceutical preparations.5 It is often self-prescribed, without medical control, for relief of moderate pain, fever, migraine and even nonspecific indications. ACT has reportedly been useful in osteoarthritis therapy. However, an overdose of ACT can result in the accumulation of toxic metabolites, which may cause severe and sometimes fatal hepatotoxicity.6-8 It can also cause liver disorders, skin rashes and inflammation of the pancreas.9

Traditionally, techniques like spectrophotometry, titrimetry, fluorimetry, and high-performance liquid chromatography have been widely employed to determine ACT. These methods require a tedious extraction process prior to detection and therefore are unsuitable for routine analysis. Recently, use of the electrochemical technique has attracted attention to sense drug molecules due to its direct rapid response, simple operation and high sensitivity.10,11

In this study, an AuNPs/MWCNTs hybrid modified glassy carbon electrode (AuNPs/MWCNTs/GCE) was fabricated using the electrochemical deposition method. AuNPs were homogeneously monodispersed on the surface of the MWCNTs. At the modified electrode, remarkable peak current enhancement and negative shift of oxidation peak potential of ACT occurred compared with that of the bare GCE. The experimental results indicate that the AuNPs/MWCNTs hybrid has the ability to increase the electroactive surface area and enhances the electrontransfer between the electrode and the analyte. In addition, the analytical performance of this sensor for determination of ACT in human serum, human urine and actual pharmaceutical preparation samples was evaluated.

Experimental

Chemicals

MWCNTs with diameters of 10–30 nm and lengths of 1–2 μm were obtained from Shenzhen Nanotech Port Co. Ltd. (China). Acetaminophen (≥98%) was purchased from the National Institutes for Food and Drug Control (Beijing, China). N,N-dimethylformamide (DMF) and HAuCl4·3H2O were purchased from Sigma-Aldrich (St. Louis, MO). A 0.05-mol/L phosphate buffer at pH 7.0 was prepared from KH2PO4 and K2HPO4·3H2O in an appropriate proportion. All other reagents were of at least analytical reagent grade, and double-distilled water was used for all solutions.

Apparatus

Electrochemical experiments, such as cyclic voltammetry (CV) and linear sweep voltammogram (LSV), were performed on a CHI 800C workstation (ChenHua Instruments Co., Shanghai, China) with a conventional three-electrode system. A bare or modified glassy carbon electrode served as the working electrode, and a saturated calomel electrode and platinum wire electrode were used as the reference and counter electrodes, respectively. Solutions were deaerated (using prepurified nitrogen) for 10 min before the electrochemical experiment. The morphologies of MWCNTs and AuNPs electrodeposited on MWCNTs were obtained using an S-4800 scanning electron microscope (SEM) (Hitachi, Tokyo, Japan).

Preparation of AuNPs/MWCNTs modified electrodes

An appropriate amount of pure MWCNTs was functionalized under a concentrated nitric acid treatment process at 120 °C for 4 hr to obtain more edge sites and better dispersion of nanotubes by the creation of carboxylate groups. Prior to modification, the surface of the bare GCE was carefully hand-polished with a 0.3- and 0.05-μm alumina–water slurry using a polishing cloth in sequence, thoroughly ultrasonically rinsed with ethanol and double-distilled water for 5 min in turn.

Prior to the deposition of MWCNTs, the bare GCE was cyclic potential scanned in the potential range from 0 to +0.8 V in 1 mmol/L·K3[Fe(CN)6] solution containing 0.1 mol/L KCl supporting electrolyte until a pair of well-defined redox peaks was observed. A 2.0-mg portion of the functionalized MWCNTs was dispersed in 2.0 mL DMF and homogenized ultrasonically for 10 min. Then, 8 μL of the suspension was placed on the GCE surface by micropipet, and the suspension was allowed to desiccate by keeping the electrode in the open at room temperature (25 ± 2 °C). To electrodeposit AuNPs on the surface of the MGCE, MWCNTs modified GCE was immersed in a 0.2 mol/L Na2SO4 solution containing 1 mmol/L HAuCl4. Modification of AuNPs was conducted by CV scanning from –0.2 to 1.0 V with a scan rate of 50 mV s–1 for five cycles. The AuNPs/MWCNTs hybrid modified GCE was taken out and rinsed with double-distilled water.

Figure 1 – SEM images of a) MWNTs /GCE and b) AuNPs/MWNTs/GCE.

Sample preparation

Fresh human serum samples were obtained from the 180th Hospital (Quanzhou, Fujian Province, China). The serum and urine sample were filtered and diluted 100 times with 0.05 mol/L PBS (phosphate buffered saline) at pH 7 and checked for the determination of the recovery by spiking with ACT.

Results and discussion

Morphologies of AuNPs/MWCNTs/GCE

The microscopic structure of AuNPs immobilized on MWCNTs was explored using SEM imaging. As shown in Figure 1a, the MWCNTs layer without aggregation was observed on the electrode surface, indicating that the MWCNTs were homogeneously dispersed on the surface of the GCE. As can be seen in Figure 1b, the AuNPs deposited on the MWCNTs were spherical with an average diameter of approx. 40 nm (30–50 nm).

Electrochemical behavior of ACT on AuNPs/ MWCNTs/GCE

Figure 2 – Cyclic voltammograms of a) GCE, b) MWNTs/GCE and c) AuNPs/MWNTs/GCE in 5 μmol/L ACT (N2-saturated PBS (0.05 mol L–1, pH 7.0). Scan rate: 50 mV s –1.

CV was used to investigate the electrochemical behavior of acetaminophen on AuNPs/ MWCNTs/GCE and bare GCE in a 0.05-mol/L phosphate buffer (pH 7.0) at a scan rate of 50 mV s−1 . At the bare GCE (Figure 2a), acetaminophen shows an irreversible behavior with relatively weak redox current peaks at Epa (anodic peak potential) at 0.463 V and Epc (cathodic peak potential) at 0.219 V. As can be seen from Figure 2b, acetaminophen exhibits a pair of well defined redox peaks on the MWCNTs modified GCE with Epa at 0.434V and Epc at 0.382 V. The increase in the current of the CV for ACT is due to the electrocatalytic effect of the MWCNTs and the increase in the electroactive area.

It can be seen from Figure 2c that there is a large background current at the AuNPs/MWCNTs/GCE, which is caused by a larger surface area of the nanocomposite film on the GCE. The redox performs a quasi-reversible process because the nanocomposite film can accelerate the electrochemical reaction.

Figure 3 – Cyclic voltammograms of AuNPs/ MWNTs/GCE in 5 μmol/L ACT at different pH (N2-saturated PBS, 0.05 M). Scan rate: 50 mV s–1.

Effect of buffer pH

The effect of buffer pH on the electrochemical response of the AuNPs/MWCNTs/GCE toward ACT was investigated using CV. Variations of peak current with respect to pH of the electrolyte in the pH range from 3 to 9 are shown in Figure 3. The anodic peak currents of ACT increase with solution pH until the pH reaches 7. However, at higher pH the ACT oxidation peak current starts to decrease. A pH value of 7.0, which is close to the biological pH value, was chosen as an optimum solution pH for further experiments.

Linear range and method detection limit

Figure 4a displays the hydrodynamic chronoamperogram response of the rotated modified electrode with successive addition of ACT at an applied potential of 0.41 V in PBS (pH 7). For ACT, the linear dynamic range was 0.05 μmol/ Lto 400 μmol/L. A calibration equation of Ipa (μA) = 8.9158 c (μmol/L) + 0.0123 (R2 = 0.9992) (Figure 4b) and a detection limit of 8 × 10–9 mol/L (S/N = 3) were obtained. It can be seen that the proposed method has a lower limit of detection compared to that obtained on the ethynylferrocene–NiO/MWCNT nanocomposite modified carbon paste electrode9 and cobalt oxide nanoparticles modified carbon ceramic electrode.10

Figure 4 – a) Hydrodynamic amperometric response at rotating AuNPs/MWNTs/GCE in PBS (0.05 mol/L, pH 7.0) for determination of ACT by successive additions of ACT, c (ACT)/(mol/L). a) 5 × 10–8, b) 2 × 10–7,   c) 4 × 10–7, d) 8 × 10–7, e) 1 × 10–6,  f) 2 × 10–6,  g) 4 × 10–6,  h) 8 × 10–6,  i) 2 × 10–5 and  j) 4 × 10–5. Applied potential: + 0.41 V (b); corresponding calibration curve for ACT obtained by I–t curve.

Repeatability and long-term stability of the electrode

The repeatability of the analytical signal was studied. The relative standard deviations (RSDs) of 2.8% for 5 μmol/L ACT over seven consecutive determinations were obtained.

Another benefit of the proposed modified electrode is that it exhibits good long-term stability. Stability of the proposed electrode was tested by measuring the decrease in voltammetric current during repetitive CV measurements of ACT solutions with AuNPs/MWCNTs/GCE for a certain period of time. When the electrode was stored in the atmosphere for seven days, the corresponding current responses fell less than 16% in a solution containing 5 μmol/L ACT.

Effect of interferences

The influences of common interfering species were investigated for solutions of 5 μmol/L ACT under optimum conditions. The results showed that concentrations of tyrosine [100], uric acid [100], ascorbic acid [200], dopamine [100], D-glucose [600], L-glutamic acid [500], and phenol [500] did not significantly affect the height of the peak currents. (The data in the brackets are concentrations of the interfering species in μmol/L.) The tolerance limit was defined as the concentrations that give an error of 5% in the determination of ACT.

Application of the proposed method

The proposed method was utilized for the determination of ACT in real samples including various tablets and syrups having ACT, human serum and human urine. Solution obtained by dissolution of ACT tablets and syrup were subsequently diluted so that ACT concentration was in the range of the calibration plot. I–t curves were then recorded under identical conditions that were employed for plotting the calibration plot.

Table 1 – Determination of ACT in pharmaceutical preparations by the proposed electrochemical sensor (n = 3)
Table 2 – Determination of ACT in human serum and human urine with the proposed electrochemical sensor (n= 3)

Keeping the dilution factor in mind, it was found that the ACT concentration determined using this method was in good agreement with the manufacturers’ stated contents (as shown in Table 1). When used in the analysis of human serum and human urine, the recoveries varied from 92% to 106% (as shown in Table 2). Therefore, the method may be applicable for the detection of ACT in real samples without pretreatment.

Conclusion

Merging the unique properties of AuNPs, such as highly effective surface area, with the intrinsic properties of MWCNTs, an AuNPs/MWCNTs/GCE was prepared for the sensitive determination of ACT. It was demonstrated that AuNPs can be electrochemically synthesized on the MWCNTs substrate at room temperature.

A highly effective microscopic area of the AuNPs/MWCNTs nanohybrid increased the peak current of ACT considerably. The AuNPs/MWCNTs/GCE has remarkable electrochemical advantages, such as antifouling behavior, good reproducibility, excellent repeatability and wide linear dynamic range. Therefore, the authors believe this sensor has the potential to be used for the accurate determination of ACT in pharmaceutical and clinical preparations.

References

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Dr. Peng is a Senior Researcher, and Mr. Zheng is a Young Scholar, Department of Chemistry, Quanzhou Normal University, Bincheng Rd. 28, Quanzhou 362000, Fujian, PR China; tel.: +86 595 2291 9531; fax: +86 595 2291 9530; e-mail: [email protected]. Dr. Peng is grateful for the financial support provided by the Science Foundation of Fujian Province (grant no. 2014J01054).

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