Analytica Chimica Acta (v.663, #1)

Hydrodynamic gating valve for microfluidic fluorescence-activated cell sorting by Pu Chen; Xiaojun Feng; Rui Hu; Jian Sun; Wei Du; Bi-Feng Liu (1-6).
Microfluidic cell sorter allows efficient separation of small number of cells, which is beneficial in handling cells, especially primary cells that cannot be expanded to large populations. Here, we demonstrate a microfluidic fluorescence-activated cell sorter (μFACS) with a novel sorting mechanism, in which automatic on-chip sorting is realized by turning on/off the hydrodynamic gating valve when a fluorescent target is detected. Formation of the hydrodynamic gating valve was investigated by both numerical simulation and flow visualization experiment. Separation of fluorescent polystyrene beads was then conducted to evaluate this sorting mechanism and to optimize the separation conditions. Isolation of fluorescent HeLa-DsRed cells was further demonstrated with high purity and recovery rate. Viability of the sorted cells was also examined, suggesting a survival rate of more than 90%. We expect this sorting approach to find widespread applications in bioanalysis.
Keywords: Microfluidic chip; Hydrodynamic gating valve; Fluorescence-activated cell sorting; Mammalian cell;

Ant colony optimization (ACO) is a meta-heuristic algorithm, which is derived from the observation of real ants. In this paper, ACO algorithm is proposed to feature selection in quantitative structure property relationship (QSPR) modeling and to predict λ max of 1,4-naphthoquinone derivatives. Feature selection is the most important step in classification and regression systems. The performance of the proposed algorithm (ACO) is compared with that of a stepwise regression, genetic algorithm and simulated annealing methods. The average absolute relative deviation in this QSPR study using ACO, stepwise regression, genetic algorithm and simulated annealing using multiple linear regression method for calibration and prediction sets were 5.0%, 3.4% and 6.8%, 6.1% and 5.1%, 8.6% and 6.0%, 5.7%, respectively. It has been demonstrated that the ACO is a useful tool for feature selection with nice performance.
Keywords: Naphthoquinone; Quantitative structure property relationship; Ant colony optimization; Maximum wavelength;

In the present study a second-order calibration strategy for high performance liquid chromatography with diode-array detection (HPLC-DAD) has been developed using parallel factor analysis (PARAFAC) and has been applied for simultaneous determination of aflatoxins B1, B2, G1 and G2 in pistachio nuts in the presence of matrix interferences. Sample preparation was based on solvent extraction (SE) followed by solid phase extraction (SPE) on Bond Elut C18 cartridges. Since the sample preparation procedure was not selective to the analytes of interest, exploiting second-order advantage to obtain concentrations of individual analytes in the presence of uncalibrated interfering compounds seemed necessary. Appropriate pre-processing steps have been applied to correct background signals and the effect of retention time shifts. Transferred calibration data set obtained from standardization of solvent based calibration data has been used in prediction step. The results of PARAFAC on a set of spiked and naturally contaminated pistachio nuts indicated that the four aflatoxins could be successfully determined. The method was validated and multivariate analytical figures of merit were calculated. The advantages of the proposed method are using a low-cost SPE step relative to standard method of aflatoxin analysis (immune affinity column assay), a unique and simple isocratic elution program for all samples and a calibration transfer for saving both chemicals and time of analysis. This study show that coupling of SPE-HPLC-DAD with PARAFAC as a powerful second-order calibration method can be considered as an alternative method for resolution and quantification of aflatoxins in the presence of unknown interferences obtained through analysis of highly complex matrix of pistachio samples and cost per analysis can be reduced significantly.
Keywords: Parallel factor analysis; High performance liquid chromatography; Photodiode-array detection; Pistachio nuts; Aflatoxins; Second-order calibration;

A novel biocompatible composite film based on a water-insoluble surfactant, didodecyldimethylammonium bromide (DDAB), and a hydrophobic room-temperature ionic liquid (RTIL), 1-hexyl-3-methyl-imidazolium hexafluorophosphate (HIMIMPF6), for the immobilization of biocatalytical proteins was reported. Differential scanning calorimetry (DSC) showed that the DDAB–HIMIMPF6 composite film has higher thermal stability than the DDAB film alone. SEM images indicated that different microstructures existed between the DDAB film and the composite film, indicating the interaction between DDAB and RTILs. This composite can be used as the immobilization matrix of proteins and other biomacromolecules. Heme-proteins, including hemoglobin (Hb), myoglobin (Mb) and horseradish peroxidase (HRP), were used as model proteins for studying the electrochemical behaviors of the resulting biocatalytical composite films. In the case of Hb, a pair of well-defined quasi-reversible redox peaks was obtained when the composite film containing Hb was modified on a glassy carbon electrode. The formal potential (E°′), the surface coverage (Γ *) and the electron transfer rate constant (k s) were calculated as −0.308 V, 1.32 × 10−11  mol cm−2 and 11.642 s−1, respectively. While, these parameters for Hb on DDAB films alone were −0.309 V, 7.20 × 10−12  mol cm−2 and 2.748 s−1, respectively. Therefore, the composite are more suitable for the direct electron transfer between Hb than DDAB alone. The native conformation and bioactivity of Hb adsorbed on the composite film was proved to be maintained, reflected by the unchanged ultraviolet–visible (UV–vis) as well as the catalytic activity toward hydrogen peroxide (H2O2) and nitric oxide (NO) compared with the free Hb molecules. Furthermore, Hb on the composite film are more sensitive for the detection of hydrogen peroxide (H2O2) and nitric oxide (NO) than that on DDAB film alone. The linear range of H2O2 on Hb/DDAB–RTILs/GC electrode is from 0.5 to 57.5 μM with linear regression equations I(μA) = 0.149 + 0.00904C(μM), while, the linear range of H2O2 on Hb/DDAB/GC electrode is from 0.5 to 57.5 μM with linear regression equations I(μA) = 0.0938 + 0.00553C(μM). For NO, its linear range on Hb/DDAB–RTILs/GC electrode is from 1.8 to 21.6 μM with linear regression equations I(μA) = 0.0937 + 0.0232C(μM). But its linear range on Hb/DDAB/GC electrode is from 1.8 to 21.6 μM with linear regression equations I(μA) = 0.0285 + 0.0167C(μM). Similar results were observed for Mb and HRP in the DDAB–HIMIMPF6 composite film.
Keywords: Direct electrochemistry; Room-temperature ionic liquid; Composite film; Hemoglobin; Bioelectrocatalysis;

Pressurized Hot Water Extraction of anthocyanins from red onion: A study on extraction and degradation rates by Erik V. Petersson; Jiayin Liu; Per J.R. Sjöberg; Rolf Danielsson; Charlotta Turner (27-32).
Pressurized Hot Water Extraction (PHWE) is a quick, efficient and environmentally friendly technique for extractions. However, when using PHWE to extract thermally unstable analytes, extraction and degradation effects occur at the same time, and thereby compete. At first, the extraction effect dominates, but degradation effects soon take over. In this paper, extraction and degradation rates of anthocyanins from red onion were studied with experiments in a static batch reactor at 110 °C. A total extraction curve was calculated with data from the actual extraction and degradation curves, showing that more anthocyanins, 21–36% depending on the species, could be extracted if no degradation occurred, but then longer extraction times would be required than those needed to reach the peak level in the apparent extraction curves. The results give information about the different kinetic processes competing during an extraction procedure.
Keywords: Anthocyanins; Subcritical water extraction; Degradation; Rates; Red onion;

A novel molecularly imprinted polymer-coated polypropylene hollow fiber tube (MIP-HFT) was photoinitiated for the copolymerization of diethylstilbestrol (DES) as a template molecule, α-methacrylic acid as a functional monomer, and ethylene glycol dimethacrylate as a crosslinking agent. The characteristics and applications of the MIP-HFT were investigated. In order to compare its characteristics with those of a non-imprinted polymer-immersed hollow fiber tube, the selectivity of the MIP-HFT was investigated using dienestrol and hexestrol as the structural analogues of a DES template, and phenol and methylbenzene were taken as reference compounds. The MIP-HFT was employed in the HPLC analysis of spiked milk samples. The detection limits of the method were found to be in the range 2.5–3.3 μg L−1 for DES, dienestrol and hexestrol and the RSD% were in the range 6.4–8.9. The limits of quantitation were found to be in the range 8.7–9.4 μg L−1 in milk for DES, dienestrol and hexestrol, and their average recoveries were 83.7–90.6% in the spiked milk samples. The experimental results revealed that the MIP-HFT provides a good carrier for the selective adsorption of DES and its chemical structure analogs, and can be used for the preconcentration of these compounds in complicated samples.
Keywords: Molecularly imprinted polymer; Hollow fiber tube; Diethylstilbestrol; Synthesis;

Coal and fly ash contain many elements. These elements exist in different forms which may change throughout the coal combustion process. There are several processes, including X-ray techniques and leaching techniques by which studies have attempted to assess the form of a particular element in a sample. This work focuses on determining the leachability of selected elements sequentially leached in four extraction solutions: water, 1 M ammonium acetate, 3 M hydrochloric acid and 50% hydrofluoric acid. The emphasis is on evaluating the steps involved in the leaching process with the mass recovery for each element being the basis for evaluation. The total amount of each element that will leach out under the given extraction condition is presented as a fraction of the total present in the material. The materials evaluated were NIST coal and fly ash standards. The elements measured in this study include aluminum, barium, beryllium, calcium, chromium, cobalt, iron, magnesium, manganese, nickel, potassium, sodium, strontium, vanadium and zinc.
Keywords: Leaching; Coal; Fly ash; Inductively coupled plasma optical emission spectrometry;

Three commonly applied extraction techniques for persistent organic chemicals, Soxhlet extraction (SE), accelerated solvent extraction (ASE) and microwave-assisted extraction (MAE), were applied on soil and fish samples in order to evaluate their performances. For both PCBs and PBDEs, the two more recent developed techniques (ASE and MAE) were in general capable of producing comparable extraction results as the classical SE, and even higher extraction recoveries were obtained for some PCB congeners with large octanol–water partitioning coefficients (K ow). This relatively uniform extraction results from ASE and MAE indicated that elevated temperature and pressure are favorable to the efficient extraction of PCBs from the solid matrices. For PBDEs, difference between the results from MAE and ASE (or SE) suggests that the MAE extraction condition needs to be carefully optimized according to the characteristics of the matrix and analyte to avoid degradation of higher brominated BDE congeners and improve the extraction yields.
Keywords: Soxhlet extraction; Accelerated solvent extraction; Microwave-assisted extraction; Polychlorinated biphenyls; Polybrominated diphenyl ethers;

The content of free-base nicotine in cigarette smoke is a controversial subject, partly due to methodological issues. In this investigation, an improved method to measure free-base nicotine in cigarette smoke using headspace solid-phase microextraction (HS-SPME) combined with GC/MS analysis, was developed and validated for this purpose. Cigarette smoke particulate phase (PP) was collected onto a 44 mm glass fiber filter pad. The pad was cut in halves with one half used to determine the concentrations of total nicotine and water. The remaining half was analyzed by HS-SPME for free-base nicotine. The following factors were found to have a significant impact on the responses of free-base nicotine: SPME fiber type, pre-equilibrium time before HS-SPME, extraction time and temperature, PP water content, and the solvent used for the preparation of standards. It was also found that the impact of PP water content on the determination of free-base nicotine from smoke sample could be corrected by a water correction factor calculated based on an experimentally determined reciprocal model. The precision of the method was evaluated with smoke samples of reference cigarettes: Canadian flue-cured monitor and Kentucky reference 2R4F. The RSD values obtained were in the 12.8–16.8% range.
Keywords: Free-base nicotine; Cigarette; Mainstream tobacco smoke; Solid-phase microextraction; Headspace; Gas chromatography;

Benzophenone (BP) and 4-methylbenzophenone (4MBP) are photo-initiators that are generally used to cure ink on carton boards.In this contribution, a fast and reliable method for the determination of BP and 4MBP in breakfast cereals is described. The sample was extracted ultrasonically using a mixture of dichloromethane and acetonitrile (1:1), followed by a clean-up of the extract using solid phase extraction with a silica cartridge. Finally, the extract was analysed by GC–MS n . Benzophenone-d10 was used as internal standard. The presented method is validated in terms of linearity, recovery, repeatability and intra-laboratory reproducibility, specificity, limit of detection and limit of quantification. In conclusion, this method is able to detect both BP and 4MBP at very low concentrations (LOD = 2 μg kg−1) in breakfast cereals.
Keywords: Photo-initiators; UV curing inks; Food; Ultrasonic extraction; Gas chromatography; Tandem mass spectrometry;

The photosynthetic glycerolipids composition of algae is crucial for structural and physiological aspects. In this work, a comprehensive characterization of the photosynthetic glycerolipids of the diatom Stephanodiscus sp. was carried out by ultra performance liquid chromatography–electrospray ionization-quadrupole-time of flight mass spectrometry (UPLC–ESI-Q-TOF MS). By use of the MSE data collection mode, the Q-TOF instrument offered a very viable alternative to triple quadrupoles for precursor ion scanning of photosynthetic glycerolipids and had the advantage of high efficiency, selectivity, sensitivity and mass accuracy. Characteristic fragment ions were utilized to identify the structures and acyl compositions of photosynthetic glycerolipids. Comparing the abundance of fragment ions, it was possible to determine the position of the sn-glycerol-bound fatty acyl chains. As a result, four classes of photosynthetic glycerolipid in the extract of Stephanodiscus sp. were unambiguously identified, including 16 monogalactosyldiacylglycerols (MGDGs), 9 digalactosyldiacylglycerols (DGDGs), 23 sulfoquinovosyldiacylglycerols (SQDGs) and 8 phosphatidylglycerols (PGs). As far as our knowledge, this is the first report on global identification of photosynthetic glycerolipids, including lipid classes, fatty acyl composition within lipids and the location of fatty acids in lipids (sn-1 vs. sn-2), in the extract of marine microalgae by UPLC–ESI-Q-TOF MS directly.
Keywords: Photosynthetic glycerolipid; Stephanodiscus sp.; UPLC–ESI-Q-TOF MS; MSE mode;

Indicator inks, previously shown to be capable of rapidly assessing photocatalytic activity via a novel photo-reductive mechanism, were simply applied via an aerosol spray onto commercially available pieces of Activ™ self-cleaning glass. Ink layers could be applied with high evenness of spread, with as little deviation as 5% upon UV–visible spectroscopic assessment of 25 equally distributed positions over a 10 cm × 10 cm glass cut. The inks were comprised of either a resazurin (Rz) or dichloroindophenol (DCIP) redox dye with a glycerol sacrificial electron donor in an aqueous hydroxyethyl cellulose (HEC) polymer media. The photo-reduction reaction under UVA light of a single spot was monitored by UV–vis spectroscopy and digital images attained from a flat-bed scanner in tandem for both inks. The photo-reduction of Rz ink underwent a two-step kinetic process, whereby the blue redox dye was initially reduced to a pink intermediate resorufin (Rf) and subsequently reduced to a bleached form of the dye. In contrast, a simple one-step kinetic process was observed for the reduction of the light blue redox dye DCIP to its bleached intermediates. Changes in red–green–blue colour extracted from digital images of the inks were inversely proportional to the changes seen at corresponding wavelengths via UV–visible absorption spectroscopy and wholly indicative of the reaction kinetics. The photocatalytic activity areas of cuts of Activ™ glass, 10 cm × 10 cm in size, were assessed using both Rz and DCIP indicator inks evenly sprayed over the films; firstly using UVA lamp light to activate the underlying Activ™ film (1.75 mW cm−2) and secondly under solar conditions (2.06 ± 0.14 mW cm−2). The photo-reduction reactions were monitored solely by flat-bed digital scanning. Red–green–blue values of a generated 14 × 14 grid (196 positions) that covered the entire area of each film image were extracted using a custom-built program entitled RGB Extractor(C). A homogenous degradation over the 196 positions analysed for both Rz (Red colour deviation = 19% UVA, 8% Solar; Green colour deviation = 17% UVA, 12% Solar) and DCIP (Red colour deviation = 22% UVA, 16% Solar) inks was seen in both UVA and solar experiments, demonstrating the consistency of the self-cleaning titania layer on Activ™. The method presented provides a good solution for the high-throughput photocatalytic screening of a number of homogenous photocatalytically active materials simultaneously or numerous positions on a single film; both useful in assessing the homogeneity of a film or determining the best combination of reaction components to produce the optimum performance photocatalytic film.
Keywords: Photocatalysis area; Thin film; Aerosol; Mapping method; Rapid;

Thiourea based novel chromogenic sensor for selective detection of fluoride and cyanide anions in organic and aqueous media by Vinod Kumar; M.P. Kaushik; A.K. Srivastava; Ajay Pratap; V. Thiruvenkatam; T.N. Guru Row (77-84).
Novel chromogenic thiourea based sensors 4,4′-bis-[3-(4-nitrophenyl) thiourea] diphenyl ether 1 and 4,4′-bis-[3-(4-nitrophenyl) thiourea] diphenyl methane 2 having nitrophenyl group as signaling unit have been synthesized and characterized by spectroscopic techniques and X-ray crystallography. The both sensors show visual detection, UV–vis and NMR spectral changes in presence of fluoride and cyanide anions in organic solvent as well as in aqueous medium. The absorption spectra indicated the formation of complex between host and guest is in 1:2 stoichiometric ratios.
Keywords: Chromogenic sensor; Naked-eye detection; Fluoride and cyanide detection; Thiourea; Aqueous solution;

A highly selective fluorescent probe for Hg2+ based on a rhodamine–coumarin conjugate by Qiu-Juan Ma; Xiao-Bing Zhang; Xu-Hua Zhao; Zhen Jin; Guo-Jiang Mao; Guo-Li Shen; Ru-Qin Yu (85-90).
A fluorescent probe 1 for Hg2+ based on a rhodamine–coumarin conjugate was designed and synthesized. Probe 1 exhibits high sensitivity and selectivity for sensing Hg2+, and about a 24-fold increase in fluorescence emission intensity is observed upon binding excess Hg2+ in 50% water/ethanol buffered at pH 7.24. The fluorescence response to Hg2+ is attributed to the 1:1 complex formation between probe 1 and Hg2+, which has been utilized as the basis for the selective detection of Hg2+. Besides, probe 1 was also found to show a reversible dual chromo- and fluorogenic response toward Hg2+ likely due to the chelation-induced ring opening of rhodamine spirolactam. The analytical performance characteristics of the proposed Hg2+-sensitive probe were investigated. The linear response range covers a concentration range of Hg2+ from 8.0 × 10−8 to 1.0 × 10−5  mol L−1 and the detection limit is 4.0 × 10−8  mol L−1. The determination of Hg2+ in both tap and river water samples displays satisfactory results.
Keywords: Fluorescent probe; Mercuric ions; Rhodamine derivative; Fluorescence enhancement;

Combination of an e-nose, an e-tongue and an e-eye for the characterisation of olive oils with different degree of bitterness by C. Apetrei; I.M. Apetrei; S. Villanueva; J.A. de Saja; F. Gutierrez-Rosales; M.L. Rodriguez-Mendez (91-97).
An electronic panel has been used to characterise the organoleptic characteristics of twenty-five extra virgin olive oils from varieties Hojiblanca, Picual and Arbequina, with different degree of bitterness. The method consists in the combination of three systems: electronic nose, electronic tongue and electronic eye. The Principal Component Analysis (PCA), where PC1, PC2 and PC3 explained 59% of the total variance between the samples, has demonstrated that the capability of discrimination of the combined system is superior to that obtained with the three instruments separately. This improvement is due to the increased information extracted from each sample. Partial Least Squares-Discriminant Analysis (PLS-DA) has allowed separation of the groups in function of olive variety with a root mean square error of prediction (RMSEP) lower than 0.099.Using PLS1 and PLS2 regression models, good correlations have been found between the signals obtained from the electronic tongue and the polyphenolic content (measured by chromatographic methods) or the bitterness index (scored by a panel of experts) with correlation coefficients higher than 0.9 in calibration and validation.These preliminary results indicate that the combination of an e-nose, an e-tongue and an e-eye can be a useful tool for the analysis of olive oil bitterness.
Keywords: Sensor; Electronic panel system; Virgin olive oil; Bitterness; Polyphenol;

Trend analysis of time-series data: A novel method for untargeted metabolite discovery by Sonja Peters; Hans-Gerd Janssen; Gabriel Vivó-Truyols (98-104).
A new strategy for biomarker discovery is presented that uses time-series metabolomics data. Data sets from samples analysed at different time points after an intervention are searched for compounds that show a meaningful trend following the intervention. Obviously, this requires new data-analytical tools to distinguish such compounds from those showing only random variation. Two univariate methods, autocorrelation and curve-fitting, are used either as stand-alone methods or in combination to discover unknown metabolites in data sets originating from target-compound analysis. Both techniques reduce the long list of detected compounds in the kinetic sample set to include only those having a pre-defined interesting time profile. Thus, new metabolites may be discovered within data structures that are usually only used for target-compound analysis.The new strategy is tested on a sample set obtained from a gut fermentation study of a polyphenol-rich diet. For this study, the initial list of over 9000 potentially interesting features was reduced to less than 150, thus significantly reducing the expensive and time-consuming manual examination.
Keywords: Metabolic profiling; Biomarker discovery; Trend analysis; Autocorrelation; Gut fermentation; Polyphenols;

Glucoraphanin, a glucosinolate, is found naturally in plants and is present in relatively high concentrations in broccoli. Glucosinolates have received much attention as studies have indicated that a diet rich in them may provide some protection from certain cancers. A micellar electrokinetic chromatography (MEKC) method using sodium cholate as the micellar phase has been developed to quantify for glucoraphanin in broccoli (seeds and florets) and Brussels sprouts. The glucoraphanin peak elutes just under 5 min with a theoretical plate number of 380,000 per metre of capillary. The method is suitable for crude extracts of broccoli and Brussels sprouts. Glucoraphanin in broccoli seeds (1330 mg/100 g) broccoli florets (89 mg/100 g) and Brussels sprouts (3 mg/100 g) was determined and agreed with the data obtained by high performance liquid chromatography. The LODs were 10–100 times below the levels typically found in broccoli seeds (4 mg/100 g), broccoli florets (0.9 mg/100 g) and Brussels sprouts (0.1 mg/100 g).
Keywords: Broccoli; Sodium cholate; Capillary electrophoresis;

The influence of non-ionic surfactants on the selectivity and retention in the ligand exchange chromatography for the enantioselective separation of racemic mixtures of the amino acids dl-methionine, dl-leucine, dl-valine and dl-tyrosine applying chiral mobile phases was investigated, whereas five different surfactants were tested as modifiers. The experiments were carried out using a commercially available non-chiral RP-C8 column and the copper (II) complex of N,N-dimethyl-l-phenylalanine as the chiral additive. Varying the surfactant concentrations the retention factors and the selectivity could be controlled and in general no negative influence on the separation (due to surfactant adsorption on the non-chiral stationary phase) occurred. Changing the temperature the van’t Hoff plots were obtained and the thermodynamic parameters calculated. Temperature had influence on the selectivity for each surfactant and lowered the retention times as expected.
Keywords: Amino acid; Chiral mobile phase; Ligand exchange chromatography; Micellar chromatography;