Analytical Methods (v.9, #48)

Front cover (6727-6727).

Inside front cover (6728-6728).

Contents list (6729-6733).

Graphene based nanocomposites have successfully been employed for high yield and stable immobilization of oxidoreductive and hydrolytic enzymes. Enzymes immobilized both via adsorption and covalent attachment on graphene nanocomposites were found highly stable against various types of denaturants and were noticed less sensitive to inhibition mediated by their specific inhibitors. The oxidoreductases and hydrolases immobilized through graphene nanocomposites have resulted in the formation of robust biocatalysts which have easily been applied in various biosensors on their repeated uses. These nanocomposite bound enzymes have shown very high specificity, selectivity, broad ranges of linearity and sensitivity and very low detection limit. By using such types of techniques several enzymes have successfully been exploited. The results have demonstrated that in many cases the activity of enzymes was remarkably enhanced upon immobilization on such supports. Graphene mediated immobilized enzymes on electrodes have successfully been employed as biosensors for the analysis of various compounds in clinical, environmental, fuel and food samples. The compounds that have been detected by using such kinds of technology are acetylcholine, ethanol, H2O2, c, histamine, catechol, phenol, 4-CP, 3,4-DMP, 3-MP, 4-MP, 4-Cl-3-MP, 4,-Cl-2-MP, caffeic acid, 17β-estradiol, artesunate, CEA, H2S, BPA, captopril, glucose, urea, clenbuterol, Con A, OPs pesticides, paraoxon-ethyl and chlorpyrifos.

A simple amino-thiacalix[4]arene modified graphitic carbon has been prepared by a microwave-irradiation process and employed for the quantification of Hg(ii) ions immobilized on screen-printed graphite macroelectrodes (SPEs) as a novel voltammetric sensor. The synthesized molecule has been characterized by FT-IR, 1H-NMR and XPS. The proposed sensor exhibits linearity in the range of 2–20 pM with a detection limit of (3σ) 1 pM. Furthermore, the modified graphitic material has been used as a sensing platform for the measurement of Hg(ii) ions in real sample matrices.

TiO2/CdS nanorod array-based photoelectrochemical sensing of Cu2+ in human serum samples by Kejun Feng; Minghui Yang; Fang Xie; Guiqiang Diao; Mingming Ou; Huanfeng Huang (6754-6759).
In this study, we report a sensitive and selective photoelectrochemical sensor for the detection of Cu2+ in human serum samples based on a TiO2/CdS nanorod array. To prepare the sensor, a TiO2 nanorod array was initially deposited onto a fluorine-doped tin oxide (FTO) conductive glass, and then, CdS nanoparticles were deposited onto the TiO2 nanorod array surface. The well-matched energy level between TiO2 nanorod and CdS efficiently suppressed the recombination of photogenerated electron and hole (e/h+) pairs; this led to improved photon-to-current conversion efficiency. Experimental results demonstrated increased photoelectrochemical current of TiO2/CdS as compared to that of TiO2 alone. Utilizing the interaction between Cu2+ and CdS, the constructed photoelectrochemical sensing platform shows selective response towards Cu2+. Increased Cu2+ ion concentration resulted in the decreased photoelectrochemical current intensity, and a linear range of 1 nM to 1 μM was obtained with a detection limit of 0.5 nM. Due to its good performances, the sensor was successfully applied for the direct detection of Cu2+ in human serum samples, and the results suggested the potential of this sensor for practical clinical applications.

Effective and sensitive microRNA detection is a great challenge. Most microRNA assays use an amplification strategy to obtain high sensitivity, which makes them complicated. Herein, we present a simple and amplification-free solid-state electrochemiluminescence (ECL) biosensor for the detection of miRNA based on high effective quenching by ferrocene and using N-butyldiethanolamine (BDEA) as a more effective co-reactant. Initially, the mixture of Nafion and Au nanoparticles (AuNPs) was cast onto the glassy carbon electrode (GCE) to form a Nafion–AuNPs film. The resulting electrode was then immersed into a solution of Ru(bpy)32+, where the solid-state Nafion–AuNPs/Ru(bpy)32+ could be obtained via electrostatic adsorption. The DNA molecular beacon, which was modified with thiol at one distal and the other was labeled with ferrocene (i.e., Fc-MB), was covalently attached to AuNPs on the ECL film through a Au–S bond. Afterwards, the remaining active sites of nonspecific adsorption were blocked with 6-mercapto-1-hexanol (MCH). When the target miRNA was present, it hybridized with the complementary part in the loop of Fc-MB and formed a double-stranded structure. This conformational change led the quencher away from the electrode, resulting in a remarkable increase in ECL intensity. Under the optimum condition, the ECL signal had a linear relationship with miRNA concentration ranging from 0.01 pM to 1 × 103 pM, with a limit of detection of 10 fM. Moreover, our biosensor showed remarkable specificity, outstanding reproducibility, good stability and excellent performance in real sample investigation with no amplification. Therefore, this developed strategy has great potential in early miRNA-related cancer diagnosis.

Simultaneous determination of bisphenol A and tetrabromobisphenol A in tea using a modified QuEChERS sample preparation method coupled with liquid chromatography-tandem mass spectrometry by Guanwei Gao; Hongping Chen; Li Zhu; Yunfeng Chai; Guicen Ma; Chen Wang; Zhenxia Hao; Xin Liu; Chengyin Lu (6769-6776).
A rapid and sensitive method for the simultaneous determination of bisphenol A (BPA) and tetrabromobisphenol A (TBBPA) in tea using liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed and validated. Sample preparation was based on a modified QuEChERS procedure through extraction of the target analytes using acetonitrile with 0.7% acetic acid followed by a dispersive solid phase extraction (d-SPE) clean up procedure using C18, MWCNTs and SCX as the adsorbent mixture to eliminate tea co-extracts. The linearity of the method with correlation coefficients (R2) higher than 0.99 was obtained. The limits of detection (LODs), and limits of quantitation (LOQs) were 8.0 and 20.0 μg kg−1 for BPA and 0.4 and 1.0 μg kg−1 for TBBPA, respectively. Satisfactory mean recoveries of BPA and TBBPA at three fortification levels ranged from 88% to 109% and 77% to 99%, respectively, while intra-day and inter-day precisions were below 15%. The developed method was successfully applied to determine BPA and TBPA in 136 tea samples.

DES-based microwave hydrodistillation coupled with GC-MS for analysis of essential oil from black pepper (Piper nigrum) and white pepper by Guo-Wei Yu; Qiang Cheng; Jing Nie; Peng Wang; Xia-Jun Wang; Zu-Guang Li; Maw-Rong Lee (6777-6784).
Deep eutectic solvents (DESs) have been gaining much attention in recent years due to their various novel properties. The present study develops a DES-based pretreatment followed by microwave-assisted hydrodistillation for the extraction of essential oil (EO) from dry fruits of white and black peppers. The process is optimized by single-factor experiments taking improvement in the EO yield of white pepper as the target. The results indicate that by using the reaction product of choline chloride and fructose as the DES with a 3 : 1 DES/powder mass ratio, the optimal conditions are (1) pretreatment stage: 600 W microwave power, 80 °C temperature, and 10 min duration; (2) fast heating stage: 600 W microwave power, 110 °C temperature, and 5 min duration; (3) hydrodistillation stage: 300 W microwave power, 110 °C temperature, and 35 min duration. Under these conditions, the EO yields of white and black peppers are 1.78% and 1.77%, respectively. The EOs are analyzed by using a gas chromatography-mass spectrometer (GC-MS), with a total of 54 compounds being identified. The new method brings about more identified compounds than hydrodistillation. Thus the DES-based approach has been proved superior in the enhancement of EO production, and has the potential of being extended to the analysis of volatile compounds from other plant resources.

A novel dual target simultaneous chemiluminescence signal amplification strategy for enhancing sensitivity of multiple biomolecule detection by Jian Li; Jingjin Zhao; Shuting Li; Liangliang Zhang; Chuanqing Lan; Yong Huang; Shulin Zhao (6785-6790).
Disease marker detection plays an important role in clinical practice; however, rapid, low-cost and simultaneous detection of multiple trace disease markers remain a challenge because of low abundance of related strategies. Herein, we report a G-quadruplex DNAzyme and nicking enzyme-assisted multiplex chemiluminescence (CL) signal amplification (EMCSA) strategy based on a microchip electrophoresis (MCE) platform for simultaneous detection of two trace target molecules. In this study, two target molecules coupled with their hairpin aptamer probes (H-probes), and the open H-probes hybridized with two different hairpin DNAs containing the G-quadruplex sequence (HG4) to form two different DNA duplexes. These DNA duplexes are then selectively cleaved by the nicking enzyme Nb.BbvCI, which forms the target–H-probe complexes and G-rich DNA segments. The obtained target–H-probe complexes hybridize again with another HG4 to form more DNA duplexes, which initiate continuous enzyme cleavage reaction to form more G-rich DNA segments. Free G-rich DNA segments combine with hemin and K+ to form G-quadruplex DNAzymes, which catalyze the CL reaction between hydrogen peroxide (H2O2) and luminol. Finally, G-quadruplex DNAzyme and streptavidin (SA)-functionalised G-quadruplex DNAzyme are quickly separated and detected by the MCE–CL platform. The proposed EMCSA-based MCE–CL method has been applied for simultaneous detection of two significant biomarkers, namely, thrombin (Tb) and carcinoembryonic antigen (CEA), in quantities as low as 2.4 pM and 0.11 ng mL−1 (0.62 pM), respectively. This method for simultaneous detection of two trace disease markers holds great potential in early clinical diagnosis of diseases.

Selective and low potential electrocatalytic oxidation and sensing of l-cysteine using metal impurity containing carbon black modified electrode by Sundaram Sornambikai; Mohammed Rafiq Abdul Kadir; Annamalai Senthil Kumar; Nagamony Ponpandian; Chinnuswamy Viswanathan (6791-6800).
The detection of thiol-containing amino acids, particularly l-cysteine (l-CySH), without any interference from other biochemicals is a challenging research interest in electroanalytical chemistry. Amongst various electrodes, the nanogold modified electrode has been reported to be effective for low potential electrochemical oxidation (∼0 V vs. Ag/AgCl) and sensing of l-CySH. Herein, we report a conductive carbon black (CB, CL-08) modified glassy carbon electrode (GCE/CL-08) with a high surface area (1000 m2 g−1), prepared by a simple drop-casting technique for efficient electrocatalytic oxidation at the lowest oxidation potential, −0.13 V vs. Ag/AgCl, and sensing of l-CySH in a pH 6.65 phosphate buffer solution. The trace metal impurities such as Ni (0.18 wt%) and Fe (0.42 wt%) in CL-08 are found to be key for such unique and unusual electrocatalytic property observed in this study. Compared with the multiwalled carbon nanotube and graphite powder modified electrodes, the GCE/CL-08 showed about 400 mV reduction in the oxidation potential and twenty times enhancement in the current signal for the CySH. A chronoamperometry detection of l-CySH on the GCE/CL-08 at an applied potential = −0.13 V vs. Ag/AgCl yielded a current linearity from 50 to 700 μM with a LOD = 45.87 nM. There are no interferences by common biochemicals such as ascorbic acid, dopamine, uric acid, xanthine, hypoxanthine and homo-cysteine on the l-CySH oxidation potential. Selective chronoamperometric detection of l-CySH in the blood serum demonstrated ∼100% recovery value as a validation for the present protocol.

The key issue in efficient electrochemical detection of trace heavy metal ions (HMIs) is to design hierarchical nanostructure electrodes with high sensitivity and low detection limit. In this study, bimetallic PtAu alloy nanoparticles (PtAuNPs) with uniform size and distribution were constructed in electrospun carbon nanofibers (CNFs) by combining an electrospinning procedure and an in situ thermal reduction process. The PtAu/CNF membrane can be directly used as a sensor electrode for simultaneous detection of trace Cd2+, Pb2+, and Cu2+ by square wave anodic stripping voltammetry (SWASV). The prepared PtAu/CNF membrane is capable of simultaneously detecting Cd2+, Pb2+, and Cu2+ with a sensitivity of 0.10 μM and correlation coefficients of 0.976, 0.993, and 0.976, respectively, indicating high sensitivity and good linear relation. The excellent sensitivity and low detection limit for HMI detection were ascribed to the high conductivity of CNFs, fast response of PtAu alloy NPs, and high specific surface area of the hybrid structure. The present research provides a convenient and efficient way to construct new sensors for simultaneous trace detection of HMIs.

In this work, bread-derived carbon foam (CF) was synthesized and used as a sorbent for solid-phase microextraction (SPME) of pollutants from environmental water samples. CF fabricated with flour and yeast via fermentation and carbonization is stiff, light weight, high in specific area, and porous. SPME coupled with gas chromatography-negative chemical ionization mass spectrometry was developed for determining polybrominated diphenyl ethers from environmental samples. Under optimal conditions, the developed method showed high sensitivity with limits of detection of 0.25–2 ng L−1 and S/N = 3 : 1, good linearity (R2 > 0.98) in the concentration range of 1–500 ng L−1, and satisfactory extraction repeatability (RSD < 12%, n = 6) and recoveries (71.8–114.4%). Furthermore, tap, pond, snow, and river water samples were used to test the applicability of the developed method. The satisfactory analysis performance demonstrated that the bread-derived CF could be used as a green, safe, and efficient sorbent for SPME of pollutants from environmental water samples.

Development of fluorescence polarization immunoassays for parallel detection of pesticides carbaryl and triazophos in wheat grains by Anna Yu Boroduleva; Jing Wu; Qingqing Yang; Hui Li; Qi Zhang; Peiwu Li; Sergei A. Eremin (6814-6822).
Fluorescence polarization immunoassays (FPIA) were developed for the determination of pesticides triazophos and carbaryl in wheat grains using the portable FPIA device Sentry 200 (Ellie). FPIA of carbaryl was developed first. Tracers for FPIA were synthesized and their structures were confirmed via mass spectrometry. The influence of tracer cross-linking bridge length on the assay sensitivity was estimated. The use of tracers labeled with ethylenediamine fluorescein thiocarbamyl (EDF) resulted in the best assay sensitivity with minimal reagent consumption. A rapid and easily performed sample preparation technique that allowed the parallel determination of both analytes in aliquots of the same sample was developed. The detection limits for triazophos and carbaryl were 40 and 20 μg kg−1, respectively, and the measurement ranges were 40–200 and 60–650 μg kg−1, respectively. A recovery test was performed using FPIA and high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), and the results obtained showed good correlation (r2 = 0.9843 for triazophos, r2 = 0.9916 for carbaryl).

Ag@SiO2 nanocube loaded miniaturized filter paper as a hybrid flexible plasmonic SERS substrate for trace melamine detection by Menbere Leul Mekonnen; Wei-Nien Su; Ching-Hsiang Chen; Bing-Joe Hwang (6823-6829).
A robust flexible paper-plasmonic hybrid SERS substrate is reported by exploiting the intrinsic properties of filter paper and anisotropic Ag@SiO2 nanocubes. The fibrous structure of paper promotes the assemblage of sharp-edged nanostructures which significantly improved the SERS activity. An inexpensive sensing platform with reasonable distribution and interparticle spacing of nanocubes was obtained by filtering Ag@SiO2 through a miniaturized filter paper. The thin silica shell improved the stability and interparticle spacing of silver nanocubes in the sensing platform, rendering enhanced SERS activity through the plasmon-coupling effect as compared to a conventional rigid substrate. Assessment of the analytical performance of the substrate for melamine quantification showed a good linearity (R2 = 0.9948) up to 1 mg L−1 with a limit of detection of 0.06 mg L−1. The detection limit in liquid milk was down to 0.17 mg L−1, which is below the permissible residue limit signifying adequate sensitivity for real sample analysis with less sample treatment.

A squaraine-based sensor for colorimetric detection of CO2 gas in an aqueous medium through an unexpected recognition mechanism: experiment and DFT calculation by Jianqi Sun; Xiangjuan Zheng; Xinjie Wu; Dong Li; Guomin Xia; Shuxian Yu; Qiming Yu; Hongming Wang (6830-6838).
A cationic squaraine-based chemosensor SQM was synthesized and its sensing behaviors towards nucleophiles followed by CO2 gas were described in detail by interpreting the results from UV-Vis, ESI-MS, and 1H NMR spectral analysis. The results indicated that there were two different paths for the bleaching of SQM, depending on whether water existed or not when the nucleophilic addition reaction between SQM and a superbase, such as TBD or DBU, occurred. More interestingly, the bleached SQM in MeCN–H2O (v/v = 90 : 10) could be colorimetrically restored with bubbling CO2 gas, which enabled it to act as a highly sensitive “naked-eye” CO2 gas detector in an aqueous medium. However, when treated in the same way, the SQM bleached in MeCN remained unchanged, unexpectedly. Combining theoretical analysis and experimental tests, a plausible sensing mechanism was proposed to illustrate the bleaching of SQM and the response of the bleached SQM to CO2 gas in an aqueous medium.

In this study, a rapid and selective method based on magnetic surface molecularly imprinting technology coupled with high performance liquid chromatography was used for the determination of melamine from egg and milk samples. A novel magnetic surface molecularly imprinted polymer (Fe3O4@SiO2@MIPs) with super-paramagnetic properties and high selectivity for melamine was synthesized as a magnetic adsorbent. The extraction, purification, and concentration processes were carried out simultaneously in a single step using an external magnet. The Fe3O4@SiO2@MIPs were characterized by scanning electron microscopy (SEM) and vibrating sample magnetometer (VSM). The adsorption thermodynamics, kinetics, and selectivity were used to investigate the rebinding mechanism of the proposed polymers, which showed better affinity and selectivity as compared to Fe3O4@SiO2@NIPs. In addition, the matrix effect was evaluated. Various parameters, such as extraction solvent, pH of the solution, amount of the polymers, extraction time, washing conditions, and eluent, affecting the extraction efficiency were evaluated. The calibration curves show a satisfactory linear relationship (r = 0.9990–0.9997) in the linear range of 0.1–1000 ng g−1. The limit of detection (LOD) was 0.79 ng g−1 for milk and 0.43 ng g−1 for eggs. Recoveries ranging from 88.4 to 98.2% were obtained with the intra- and inter-day RSDs in the range of 3.5–5.4% and 5.7–7.8%, respectively. The proposed Fe3O4@SiO2@MIPs were successfully applied towards the selective determination of melamine in different real egg and milk samples, and satisfactory recoveries in the range from 87.9% to 102.4% with an RSD of less than 5.40% were obtained. The proposed method simplified the procedures of the classical solid-phase extraction (SPE) method and improved the reliability of the applied method.

Back cover (6849-6850).