Analytical Methods (v.8, #32)

Front cover (6123-6124).

Contents list (6125-6130).

A graphene oxide-based fluorescent aptasensor for alpha-fetoprotein detection by Ya Zhang; Yunfeng Bai; Feng Feng; Shaomin Shuang (6131-6134).
A simple, rapid and sensitive method for the detection of alpha-fetoprotein was developed based on FAM-labeled aptamers and the fluorescence quenching ability of graphene oxide.

A 3D printed device for quantitative enzymatic detection using cell phones by G. Comina; A. Suska; D. Filippini (6135-6142).
A disposable device for quantitative enzymatic detection capable of coupling illumination and image readouts from cell phones is demonstrated. The device integrates a calibration range for glutamate detection, utilizes the phone screen as a light source, and provides the necessary actuation for autonomous operation. Custom made optics required to couple to the cell phone camera is accomplished using affordable stereolithography (SLA) 3D printers. The described method does not involve polishing, requires only two steps from design to implementation, and can be locally applied to 3D printed lab-on-a-chip (LOC) prototypes, using the same materials. Optical finishing and dimensional variability within 2% were achieved, supporting entirely arbitrary geometries for elements larger than 400 μm in radius. Representative fabrication times and costs were 20 min and $0.50 USD per prototype.

Asimina triloba, commonly known as paw paw, is one of approximately 2100 species in the Annonaceae family, scores of which are known to biosynthesize bioactive secondary metabolites, termed Annonaceous acetogenins. Even with over 400 acetogenins identified, a high-throughput screening protocol for these compounds does not exist. Advances in direct ambient ionization mass spectrometry have opened the door to many metabolite profiling methodologies, but for acetogenins, this is often complicated by the abundance of isomers that are present. A droplet-liquid microjunction-surface sampling probe coupled to UPLC-PDA-HRMS/MS system was employed to detect acetogenins in situ from A. triloba. The seeds, fruit pulp, twigs, leaves, and flowers of A. triloba were all examined for acetogenins. Additionally, lithium was infused post-column to increase the sensitivity of the fragments, thus allowing for characterization of the structural classes, and mass defect filtering was used to mine the data for the various acetogenin analogues. This surface sampling system allowed for the rapid identification and differentiation of Annonaceous acetogenins directly from the various organs of A. triloba, including the never before studied flowers.

Accurate determination of arsenic and selenium in plant food samples by using ICP-MS/MS by Xianqiao Hu; Zhaoyun Cao; Weihua Sun; Huan Yang; Ping Xu; Zhiwei Zhu (6150-6157).
Arsenic (As) and selenium (Se) in food have caused public concern. Determination of As and Se in plant food samples encounters mass spectral interferences. The influence of polyatomic and doubly charged ions on the determination was evaluated by monitoring 75As, 77Se, 78Se, 80Se, and 82Se in four modes. The result showed that serious interferences were observed by using “no gas” mode (octopole vented, single quadrupole mode). Most but not all interferences could be removed when using He mode (He used as the collision gas, single quadrupole mode) or H2 mode (H2 used as the reaction gas, MS/MS mode), while O2 mass-shift mode (O2 used as the reaction gas, MS/MS mode) enabled the effective removal of all interferences. In the O2 mass-shift mode configuration, the ions of interest were filtered twice by using the mass-to-charge ratio, enabling the removal of all interfering species. As and Se concentrations could be determined interference-free by using mass-shift mode with O2 as reaction gas. The performance of O2 mass-shift mode was investigated for the determination of As and Se in five plant reference materials. The obtained results were consistent with the certified values, indicating that the O2 mass-shift mode was an effective method to determine As and Se in plant food samples. The detection limits of the method for 75As, 77Se, 78Se, 80Se, and 82Se were 0.5, 0.7, 1.1, 0.3 and 0.6 ng g−1, respectively.

Liquid–liquid extraction and separation of lead(ii) by using N-n-octylcyclohexylamine as an extractant: analysis of real samples by Arjun Kokare; Vishal Suryavanshi; Sunil Zanje; Gurupad Kore; Mansing Anuse (6158-6167).
A method for the determination of micro amounts of lead(ii) is described. N-n-Octylcyclohexylamine (N-n-OCA) was employed as an ion-pair forming a neutral [N-n-OCAH+PbCl3] complex in hydrochloric acid medium. The quantitative extraction of lead(ii) was observed with N-n-OCA (0.03 to 0.055 M) in a dichloromethane (DCM) and xylene mixture (1 : 4), from hydrochloric acid medium (3.0 to 5.0 M). The extracted ion-pair complex was back stripped with 0.5 M nitric acid and determined spectrophotometrically with PAR. The quantitative extraction of lead(ii) was found in the DCM : xylene ratio of 1 : 4 as a mixed solvent system. The various parameters studied, such as concentration of acid, N-n-OCA concentration, equilibrium time, solvent study, back stripping agents and loading capacity were optimized for the quantitative extraction of lead(ii). The stoichiometry of the extracted ion-pair complex was determined on the basis of the slope analysis method, and it was found to be 1 : 3 : 1 (metal : chloride : extractant). The proposed method was successfully applied to the analysis of diverse ions, binary mixtures of associated metal ions, ternary mixtures, alloys, ayurvedic samples and water samples, by using N-n-OCA and lead(ii) was determined using PAR and the results of analysis were confirmed by ICP-OES.

In this paper, a magnetic graphene framework (MGF) material was prepared and used as a magnetic solid-phase extraction adsorbent for the preconcentration of flunitrazepam from beverage samples prior to high resolution mass spectrometric determination. The MGF nanocomposite was characterized by scanning electron microscopy, transmission electron microscopy, vibrating sample magnetometry, and N2 adsorption. The high surface area and specific framework structure of the MGF endows it with a high adsorption capacity. Several experimental parameters affecting the extraction efficiencies, such as the amount of the adsorbent, extraction time, sample pH and desorption conditions were optimized. Under the optimum conditions, a good linearity was achieved in the concentration range of 0.01–1.0 nmol L−1 for ginger ale and 0.05–5.0 nmol L−1 for the Pure tea sample with the correlation coefficients larger than 0.9875. The limits of detection of the method were 0.001 nmol L−1 (0.3 ng L−1) for ginger ale and 0.005 nmol L−1 (1.5 ng L−1) for Pure tea, which is about 3–4 orders of magnitude lower than those of literature reports. The recoveries of the method for the analytes were in the range of 92.0–102.5%. The results indicated that the developed method was efficient for the preconcentration of trace levels of flunitrazepam from beverage samples. The MGF adsorbent would have a significant application potential for the enrichment of other abused drugs and organic pollutants in different samples.

In this work, azo-coupling derivatization was performed for propofol analysis. The derivatization produced a red shift in the maximum absorbance of propofol, thus preventing signal interference from endogenous substances in plasma. An automated method was developed for the detection of propofol based on the reaction of propofol with a diazonium salt via sequential injection (SI). The resulting coloured product had an absorbance maximum at 483 nm. The method was compared to a traditional spectrophotometric method performed manually. Both methods provided comparable validation parameters, with linear working ranges of 3–18 μg mL−1 and 6–21 μg mL−1 of propofol in plasma (R = 0.998, 0.996); recoveries of 89.3–99.6% and 102–104% and RSDs of 4.25–5.58% and 4.08–5.52% were obtained at three different concentrations for the SI-spectrophotometric and spectrophotometric methods, respectively. A sample throughput of approximately 46 injections per h was achieved for the SI-spectrophotometric method. This automated method provides a foundation for the rapid measurement of propofol in plasma samples.

A novel carbon paste electrode modified with a magnetic core–shell Fe3O4@SiO2/MWCNT nanocomposite and an ionic liquid (n-hexyl-3-methylimidazolium hexafluoro phosphate) was fabricated. The electrochemical study of the modified electrode, as well as its efficiency for the simultaneous voltammetric oxidation of amlodipine and hydrochlorothiazide is described. The electrode was also employed to study the electrochemical oxidation of amlodipine and hydrochlorothiazide, using cyclic voltammetry, chronoamperometry and square wave voltammetry as diagnostic techniques. Square wave voltammetry exhibited linear dynamic ranges from 2.5 × 10−7 to 5.0 × 10−4 M and 1.0 × 10−6 to 6.0 × 10−4 M for amlodipine and hydrochlorothiazide, respectively. Finally, the modified electrode was used for the determination of amlodipine and hydrochlorothiazide in real samples.

A method using dissolved carbon dioxide flotation after dispersive liquid–liquid microextraction (DCF-DLLME) was proposed for the determination of sixteen pyrethroids in water samples, coupled with gas chromatography with triple quadrupole mass spectrometry in a homemade glass round flask. The DCF-DLLME method is based on the rapid and simple phase separation of a low density organic solvent from the aqueous phase via introducing a saturated NaHCO3 solution into the acidified sample solution (0.10 mol L−1 CH3COOH) containing the analytes. The optimum extraction conditions used 45 μL of dodecanol and 2.6 mL of acetonitrile as the extraction and dispersive solvent respectively, with an extraction time of 40 s and a demulsification time of 80 s. The LOQ for the sixteen pyrethroids was 5 μg L−1 and the LODs were in a range between 0.87 and 1.39 μg L−1. The mean recoveries (n = 15) ranged from 79.6% to 98.2%, with the RSD (n = 15) data ranging from 2.7% to 8.5%. The method broke through the limit of volume for centrifugation, and the device is easy to clean and the volume of sample can be enlarged. Adequate analytical performance characteristics were obtained in terms of linear range and precision for sixteen pyrethroids in water. Satisfactory results were obtained in authentic samples. The results of analysis revealed that this method is simple, fast and environmentally friendly, being successfully applicable for the determination of sixteen pyrethroids in water samples.

An ultrasensitive label-free immunoassay for C-reactive protein detection in human serum based on electron transfer by Xi Zhang; Rong Hu; Kunlei Zhang; Ruyan Bai; Delei Li; Yunhui Yang (6202-6207).
A rapid, ultrasensitive, and practical label-free electrochemical immunoassay for measuring C-reactive protein (CRP) in real serum samples was developed. Molybdenum disulfide–polyaniline–gold nanoparticles (MoS2–PANI–GNPs) with high conductivity were employed as the substrate to assist the electron transfer. Compared with a bare electrode, an about 10-fold increase in the peak current could be detected for MoS2–PANI–GNPs. Moreover, MoS2–PANI–GNPs which have abundant adsorbing sites and a large surface area could greatly increase loading amounts of antibodies and thus, significantly improve the performance of biosensing. Under optimized experimental conditions, the proposed sensing strategy provides a linear dynamic range from 0.2 to 80 ng mL−1 and a detection limit of 40 pg mL−1. The proposed immunosensor has also been used to determine CRP in human serum with satisfactory results. This possibly makes it an attractive platform in bioanalysis of proteins and other molecules.

This study was conducted to optimize the ultrasonic-assisted extraction (UAE) of higher fatty acids (with aliphatic tails more than or equal to 14 carbons) in grape berries (seed-free fruit sections). Preliminary experiments showed that the extraction solvent, extraction duration, extraction temperature and ultrasound power were the main influencing factors in terms of UAE efficiency. Orthogonal testing was also performed to further investigate the optimal UAE parameters. The results showed that the extraction solvent n-hexane (25 mL, extraction 4 times), extraction duration (30 min), temperature (45 °C) and ultrasound power (396 W) were the optimal conditions. The optimized method took less time and caused less degradation of α-linolenic acid from grape berries compared to traditional Soxhlet extraction. The linearity of this method covered a wide concentration range and the correlation coefficients were satisfactory (99.22% < R2 < 99.88%). The limits of detection (signal/noise = 3), which ranged from 0.002 mg mL−1 to 0.005 mg mL−1, were sufficiently low to meet the detection requirements. The relative standard deviations (RSDs) of the method ranged from 2.10% to 8.50% and the recoveries of the method at 0.5-, 1- and 2-times addition levels were in the range of 81.93% to 118.82%.

Classification of iron ore based on acidity and alkalinity by laser induced breakdown spectroscopy coupled with N-nearest neighbours (N3) by Chunhua Yan; Zhanmei Wang; Fangqi Ruan; Junxiu Ma; Tianlong Zhang; Hongsheng Tang; Hua Li (6216-6221).
Laser induced breakdown spectroscopy (LIBS) coupled with N-nearest neighbours (N3) method was developed for classification and identification of four types of iron ore (acid iron ore, seiili-self fluxing iron ore, self-fluxing iron ore and alkaline iron ore). The parameters included spectral pretreatment methods and spectral range selection and the model parameter α was optimized at the same time by 5-fold cross validation and evaluated by average classification error rate. The region of 400–600 nm was normalized by maximum integrated intensity and used to construct the N3 and KNN (K nearest neighbor) models. The N3 and KNN models were evaluated and applied to discriminate iron ore. The classification accuracy is 100% for the N3 model, which shows better predictive capabilities than the KNN model for the classification of iron ore. Therefore, LIBS technique combined with N3 could be a promising method for real-time online, rapid analysis in mining and mineral processing industries.

The objective of the present study is to optimize the smoke and NOx emissions simultaneously in a diesel engine fuelled with stationary diesel–oxygenate blends using the grey based Taguchi method. Experiments were carried out by adopting the Design of Experiments (DOE) method and tests were conducted based on Taguchi's L9 orthogonal array. The effects of three parameters, namely oxygen content of the additives, oxygenate proportion with diesel and varying injection timing, were investigated. Experimental trials were conducted by blending the various chosen oxygenates in different proportions with diesel and different injection timings. Taguchi's signal-to-noise (S/N) ratio was determined based on performance characteristics. The grey relational grade was obtained from the S/N ratio using the grey relational analysis (GRA). Based on this grade, the optimum level of factors was identified using response tables and response graphs. The individual effects of factors are estimated using analysis of variances (ANOVA). The results of the experiments reveal that diglyme blended with 10% diesel and injected at −21° crank angle is the optimum combination for the simultaneous reduction of smoke and NOx with a less significant impact on performance. This combination shows a smoke reduction of 28.33% with a 17.4% reduction in NOx emissions simultaneously with the best possible performance increase of 6.7% when compared to diesel. The combination of GRA and Taguchi parametric design can be effectively used to obtain the optimal combination of the chosen parameters. Experimental results also show that the response variables can be improved effectively through this approach.

A high level of folate receptor (FR) expression is found frequently in numerous human cancerous cells, and monitoring of the FRs level is important for the diagnosis and treatment of cancers and chronic inflammatory diseases. In this study, a simple but sensitive fluorescent biosensor for FR detection in cancer cells has been proposed, which combines the desirable specificity of terminal protection and high amplification efficiency of hyperbranched rolling circle amplification (HRCA). Single-stranded DNA (ssDNA) terminally tethered to folic acid (FA) can specifically bind with FR, and therefore be protected from degradation by exonuclease I (Exo I). The protected ssDNA can hybridize with the padlock probe and initiate the HRCA reaction. In contrast, ssDNA will be degraded by Exo I and no probe is left to trigger the HRCA reaction without the protection of the target FR. The products of HRCA contain a large amount of double-strand DNA (dsDNA) and a strong fluorescence signal can be detected after the addition of SYBR Green I. The enhanced fluorescence intensity has a linear relationship with the logarithm of FR (standard solution) concentration ranging from 0.18 fM to 22 pM with a detection limit of 0.06 fM. Moreover, the proposed biosensor has been applied to detect FR in cancer cells (HeLa cells as an example). The enhanced fluorescence intensity has a linear dependence on the logarithm of cell concentration in the range of 50–5000 cell per mL. This biosensor has potential applications in the area of early cancer diagnosis.

A novel method for leucomalachite green (LMG) hapten synthesis was presented. This method could introduce a pure alkyl chain at the para-position of LMG to form a spacer arm for hapten preparation. It also could be expanded to prepare a series of haptens with different lengths of spacer arms by increasing the lengths of alkyl chains. This synthesis is easy to operate with high yield. In order to find out the best match coating antigens, three other haptens were synthesized. Polyclonal antibodies against LMG were developed by using the above synthesized antigens. The sensitivity and specificity were examined by indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) by optimizing the types of coating antigens and the concentrations of the antigens and antiserum. We demonstrate that the antibody developed from the hapten with a pure alkyl chain gives the best performance. We also reported that heterogenous coating antigen gives better sensitivity in this ELISA method development. The ELISA assay was further used to analyze the LMG residue in fish tissue with an IC50 value of 2.2 ng mL−1 and a limit of detection (LOD) of around 0.06 ng mL−1. Recoveries ranged from 79.8–92.4%, indicating that the IC-ELISA could be used for effective and low-cost analysis of LMG in fish samples.

Back cover (6245-6246).