Analytical Methods (v.5, #12)

Front cover (2901-2902).

Contents list (2903-2913).

What causes most errors in chemical analysis? by Analytical Methods Committee, AMCTB No 56 (2914-2915).
In the early 1980s, emerging evidence of poor inter-laboratory precision in routine analysis for trace contaminants led to major changes in the management of analytical chemistry. The adoption of the CODEX and later VAM principles encouraged laboratories to participate regularly in proficiency test (PT) schemes; to adopt validated analytical methods; to rely increasingly on certified reference materials for calibration and validation of their test methods; to implement effective internal QC; and (more recently) to obtain accreditation to standards such as ISO 17025.

Carotenoids extraction and quantification: a review by Héctor Arvayo-Enríquez; Iram Mondaca-Fernández; Pablo Gortárez-Moroyoqui; Jaime López-Cervantes; Roberto Rodríguez-Ramírez (2916-2924).
In recent years carotenoids have represented a good alternative for the pharmaceutical and food industries and especially for the human health, they prevent different diseases, such cancer, macular degradation and cataracts. The use of post-harvest residues represents a good alternative for obtaining analytes of interest. The application of enzymatic pretreatment over plant matrices has resulted in an increase in the level of extraction and lowered operating costs. Today, the use of organic solvents such as hexane in this process is frequent, however, new investigations with novel environmentally friendly solvents (non-toxic) promise new cleaner technologies. The aim of this review is to present some methodologies and technologies used today to obtain carotenoids from plant residues, industrial and post-harvest materials.

Two types of isotope-labelled reference compounds are typically used for the passive sampling of polycyclic aromatic hydrocarbons. One type is added before the exposure of the sampler and is called a performance reference compound (PRC). The other is the laboratory internal standard, which is spiked into the sample after extraction (dialysis) from the membrane and is used for quantification. Generally, PRCs are deuterium-labelled while internal standards are 13C-labelled. Interference originating from the internal standards occurs when the molecular ions of the PRCs gradually lose deuterium in the ion source to generate fragments that overlap with the molecular ions of the 13C-labelled internal standards. This can cause significant systematic errors in quantification. Gas chromatography-high resolution time-of-flight mass spectrometry (GC-HRTOF-MS) was used to investigate the degree of interference at varying virtual resolutions of the MS instrument, and it was shown that many of the spectral interferences can be avoided by using high MS resolutions (35 000 or better).

A ratiometric fluorescent sensor for Ag+ based on 8-hydroxyquinoline platform in aqueous media by Huajie Xu; Zhaodi Liu; Liangquan Sheng; Deqian Huang; Hong Zhang; Chongfu Song; Mingming Chen; Wuyang Hua (2929-2932).
The ratiometric fluorescent sensor L based on 8-hydroxyquinoline has been designed and synthesized. In aqueous media, L exhibits a ratiometric response to Ag+ in the presence of competing metal ions with a large blue-shift of 76 nm. The good linear relationship is easily obtained between the fluorescence changes and the concentrations in the range of 0–80 μM. The complexation mode has been proposed by fluorescence titration, 1H NMR, IR and MS mass spectrometer experiments.

A fast extraction technique involving dispersive liquid–liquid microextraction (DLLME) coupled with magnetic nanoparticle-based dispersive solid-phase microextraction (D-μ-SPE) was developed. For preliminary study, this method was applied for the extraction of 4-n-nonylphenol (4-NP) in water. In this method, n-octyl-triethoxysilane surface-modified magnetic iron oxide nanoparticles (C8MNPs) were first synthesized. The DLLME was performed by injecting 1 mL of methanol (disperser) containing 10 μL of 1-octanol (extraction solvent) into 15 mL of water sample. To this solution, C8MNPs were immediately added for the retrieval of the extraction solvent and of the extracted 4-NP by sonication. Subsequently, a magnet was held next to the vial to attract and isolate the C8MNPs. Then, the water sample was decanted, the nanoparticles were freeze-dried, and the 4-NP was extracted from nanoparticles by sonication with methanol. The detection of 4-NP was performed using a HPLC coupled with a fluorescence detector. Under optimal extraction conditions, a low method of detection limit (13.9 ng L−1), good linearity (R2 = 0.9958), and repeatability (1.7–2.2%) were achieved during 4-NP extraction. This method was also successfully used to analyze the real water samples such as drinking water and secondary wastewater. Generally, this sample preparation method offered an alternative to the conventional DLLME, because, in this method, centrifugation is not required and the retrieval of the extraction solvent is based on the adsorption process by magnetic nanoparticles (MNPs). Also, this two-step method improved the versatility of DLLME because the selection of the extraction solvent was no longer limited to the high density solvents. Because MNPs can be surface-modified for the extraction of different organic solvents, this method could have great prospects in the future.

High specific phosphopeptides enrichment by titanium silicalite with post-treatment of desilication by Bo Xu; Chen Chen; Fangjun Wang; Yangyang Bian; Kai Cheng; Hongqiang Qin; Chunxia Song; Jun Zhu; Jing Liu; Mingliang Ye; Hanfa Zou (2939-2946).
The titanium silicalite-1 (TS-1) with post-treatment of chemical selective desilication (T-TS-1) has been synthesized, characterized and applied as a potential adsorbent for selective capture of phosphopeptides from complex biological samples prior to mass spectrometry analysis. A T-TS-1 material-based successive phosphopeptide enrichment strategy has also been established for real biological sample analysis, which exhibited high enrichment selectivity and is comparable to existing materials.

Coupling nanomaterials with biomolecular recognition events represents a new direction in nanotechnology toward the development of novel molecular diagnostic tools. A novel aptamer biosensor based on multi-walled carbon nanotube (MWCNT) long-range energy transfer has been developed for sensitive, selective and multicolor fluorescent detection of Hg2+, Ag+ and Pb2+ ions in homogeneous solution. The fluorescent dye-labeled aptamer is adsorbed on MWCNTs due to their strong π–π stacking interactions, leading to quenching of fluorescence of the dye. Desorption of the dye-labeled aptamer from MWCNTs, through the specific binding of the aptamer with its target, resulted in the restoration of the fluorescence signal of the dye. Due to the non-covalent assembly between the aptamer and MWCNTs, highly efficient long-range energy transfer from the dyes to MWCNTs takes place. In the presence of metal ions, the binding between the aptamer and metal ions will disturb the interaction between the aptamer and MWCNTs, and release the dye-labeled aptamer from the MWCNTs' surface, resulting in restoration of the fluorophore's fluorescence. Importantly, the high specific surface area of MWCNTs allows the quenching of multiple aptamer probes labeled with different fluorescent dyes, leading to a multicolor nanosensor for simultaneous detection of multiple metal ions in the same solution. As a proof of concept, we demonstrated that a three-color nanosensor can rapidly and simultaneously detect three metal ions (i.e. Hg2+, Ag+ and Pb2+) in a single solution. This MWCNT-based sensing platform exhibits high sensitivity and selectivity toward Hg2+, Ag+ and Pb2+versus other metal ions, with a limit of detection of 15 nM for Hg2+, 18 nM for Ag+ and 20 nM for Pb2+.

Real-time diesel particulate monitor for underground mines by James Noll; Samuel Janisko; Steven E. Mischler (2954-2963).
The standard method for determining diesel particulate matter (DPM) exposures in underground metal/nonmetal mines provides the average exposure concentration for an entire working shift, and several weeks might pass before results are obtained. The main problem with this approach is that it only indicates that an overexposure has occurred rather than providing the ability to prevent an overexposure or detect its cause. Conversely, real-time measurement would provide miners with timely information to allow engineering controls to be deployed immediately and to identify the major factors contributing to any overexposures. Toward this purpose, the National Institute for Occupational Safety and Health (NIOSH) developed a laser extinction method to measure real-time elemental carbon (EC) concentrations (EC is a DPM surrogate). To employ this method, NIOSH developed a person-wearable instrument that was commercialized in 2011. This paper evaluates this commercial instrument, including the calibration curve, limit of detection, accuracy, and potential interferences. The instrument was found to meet the NIOSH accuracy criteria and to be capable of measuring DPM concentrations at levels observed in underground mines. In addition, it was found that a submicron size selector was necessary to avoid interference from mine dust and that cigarette smoke can be an interference when sampling in enclosed cabs.

Orange II, an azo dye, is not permitted in food preparations as it can cause many adverse health effects when consumed. Therefore, the monitoring of Orange II is quite important. Herein, an electrochemical method for the sensitive and rapid analysis of Orange II was reported using mesoporous TiO2 decorated graphene nanocomposite as the sensing film. In pH 4.4 HAc–NaAc buffer, an irreversible oxidation peak at 0.68 V was observed for Orange II. Compared to the unmodified electrode, the graphene–mesoporous TiO2 modified electrode remarkably increased the oxidation signal of Orange II. The detection conditions such as pH value, amount of modifier, accumulation potential and time were optimized and the electrochemical reaction mechanism of Orange II was discussed. The limit of detection for Orange II is 0.92 nM. Finally, the new method was successfully employed to detect Orange II in chilli products and ketchup samples, and the results were tested by high performance liquid chromatography-tandem mass spectrometry.

A novel, sensitive and inexpensive supramolecular dispersive liquid–liquid microextraction method based on the solidification of floating organic drops (SM-DLLME-SFO) has been proposed for the speciation and preconcentration of trace quantities of chromium as a precursor to its determination by UV-Vis spectrophotometery for the first time. The chromium ions are micro-extracted with coacervates composed of reverse micelles formed using decanoic acid and dispersed in tetrahydrofuran–water mixtures. THF plays a double role, as a dispersing solvent and also in the self-assembly of decanoic acid. The method involves the partitioning of the metal chelates, produced from the reaction of Cr(vi) with diphenylcarbazide and sodium dodecyl sulfate in an acidic medium and a combination of SM-DLLME with the solidification of floating organic drops. It combines the advantages of dispersive liquid–liquid microextraction with those based on coacervation and reverse micelles and solidification. All the critical parameters affecting the analytical performance were studied. Under the optimum conditions, the enhancement factor was 50. The detection limit and precision (RSD) were 0.23 μg L−1 and 3.8% (n = 6), respectively. The accuracy of the developed method was evaluated by analyzing a certified reference material and applied successfully to the analysis of several water samples.

In this work, a commercially available portable tungsten coil electrothermal atomic absorption spectrometer (W-coil ET-AAS) was applied and investigated. The portable AAS instrument was developed based on a W-coil atomizer and a charge coupled device detector, and has the characteristics of compactness and portability. In this research, ionic liquid based liquid phase microextraction (IL-LPME) was coupled with a portable AAS instrument to expand its application and improve its analytical performance. Ultra trace lead and copper were investigated as the target elements in the established coupling. The factors influencing both IL-LPME and the portable spectrometer were optimized. Under the optimal conditions, the limits of detection for lead and copper were 0.25 μg L−1 and 0.10 μg L−1, with sensitivity enhancement factors of 38 and 26, respectively. The relative standard deviation (RSD) for seven replicate measurements of 20 μg L−1 of lead was 4.9%, and 10 μg L−1 of copper was 3.4%. The recoveries for the spiked samples were in the acceptable range of 92.7–105%.

Characterization of currently marketed heparin products: composition analysis by 2D-NMR by David A. Keire; Lucinda F. Buhse; Ali al-Hakim (2984-2994).
Each type of heparin has a unique pattern of signals in their NMR spectra because of the effects of the process specific chemical reactions used to manufacture them. Thus, for heparin sodium, 1D-1H-NMR identification tests are part of the United States Pharmacopeia (USP) and European Pharmacopeia (EP) monographs. Previous work has shown that two-dimensional (2D) NMR data can be used to identify heparin types and be used to calculate heparin composition. Therefore, in this study, we applied a 2D 1H–13C-heteronuclear single quantum coherence (HSQC) spectroscopy experiment to characterize the normal variation of intact heparin sodium or low molecular weight heparin (LMWH) molecular composition in USP grade sample lots which were available on the US market in the summer of 2009. We tested 7 heparin sodium active pharmaceutical ingredient (API) samples by the 2D method representing lots from 6 manufacturers and the USP heparin sodium identification standard. In addition, we tested 10 LMWH API samples from 3 manufactures of different types of LMWH and the USP enoxaparin identification standard. Using the integrated volumes of HSQC cross-peaks assigned to specific heparin species the monosaccharide or disaccharide percent compositions were calculated. These data establish the composition and normal range of variability for each of the heparin types in the 2D assay across manufacturers supplying the US market. The values obtained from the NMR test were similar to those obtained from mass spectrometric analysis of heparin digests on the same sample set. The robustness of the assay was tested by varying the acquisition time from ∼3 to ∼68 h by changing the number of transients co-added or the relaxation delay; across these changes the percent composition values obtained did not vary significantly. We conclude that the manner in which the calculations are performed minimizes experimental errors that arise from differences in spectral signal-to-noise, heparin 1JCH through bond coupling constants or relaxation times.

Malondialdehyde (MDA) is a presumptive biomarker for lipid peroxidation in plasma, live organisms, and cultured cells. This study presents an accurate and reproducible method to measure MDA as its hydralazine (HDZ) derivative in plasma using high-performance liquid chromatography (HPLC). A reference curve was prepared by mixing a 250 μL volume of each concentration of standard MDA (0.05–10.0 nmol mL−1) with 25 μL HDZ 5 mM solution in 2 M hydrochloric acid and the mixture was incubated for 60 min at room temperature in the dark. An aliquot of 50 μL of this reaction mixture was injected into the HPLC system. This method was also used for the determination of MDA in phosphate buffer and plasma. The standard addition method was used to determine MDA in plasma. In order to compare this method with another method for the determination of MDA, a HPLC method was applied using derivatization of MDA with 2,4-dinitrophenylhydrazine (DNPH). Results showed that this method was valid, linear, simple, highly sensitive and reproducible for MDA determination in plasma. The limit of detection (LOD) and limit of quantitation (LOQ) were found to be 0.03 and 0.1 nmol mL−1, respectively. This method has a shorter run time, and a lower LOQ in comparison with other HPLC methods described for MDA determination. Also, it can serve as a cost-beneficial method for determination of MDA in plasma as a biomarker for lipid peroxidation.

Cellulose-g-oxolane-2,5-dione nanofibers were prepared as an adsorbent for simultaneous separation and preconcentration of trace amounts of Cd, Cu, Fe, Pb and Zn ions in gasoline samples prior to ICP-OES detection. The nanofibers were chemically and morphologically characterized by FTIR, solid-state 13C NMR, BET and SEM techniques. The influences of experimental parameters such as pH, HNO3 concentration on metal ion elution from the nanofibers, flow rate and sample volume were investigated. The metal ions were retained on 0.5 g of the adsorbent at pH 6 and recovered with 5.0 mL of 2.0 mol L−1 HNO3. The adsorption capacities for the adsorbent were 273.1, 183.6, 195.5, 236.2 and 182.4 mg g−1 for Cd, Cu, Fe, Pb and Zn, respectively. The relative standard deviation was <3% (n = 15), limits of detection and quantification were 0.13–0.68 μg L−1 and 0.42–2.2 μg L−1, respectively, and the maximum preconcentration factor was 60. It was observed that cellulose-g-oxolane-2,5-dione nanofibers can be used for more than 30 adsorption–elution cycles without decreasing the extraction efficiency. The accuracy of the method was confirmed by analyzing a certified reference material and by performing the spike recovery test. The accuracy and recovery for different metal ions were in the range of 97–102% and 96–99%, respectively. The optimized method was applied for the separation and preconcentration of metal ions in gasoline samples.

Molecularly imprinted fluorescence sensors operate on the basis of the recognition of imprinted sites to guest and the resultant changes of fluorescence emission have been studied. However, the origin of guest-induced fluorescence enhancement and the function of host molecule are still unclear in theory. In this work, we have first designed three isomers, 2-acrylamidoquinoline, 3-acrylamidoquinoline and 8-acrylamidoquinoline, with weak fluorescence emission, and used them as both functional monomers and signaling units in molecularly imprinted fluorescence sensors. Quantum chemical calculation within the density functional theory (DFT) framework has been introduced to accurately evaluate and predict the hydrogen bonding interaction between these monomers and the analyte melamine. As a result, the as-synthesized 2-acrylamidoquinoline exhibits a highest hydrogen bonding ability and the ideal molar ratio of monomer to template is 3 : 1 in molecularly imprinted polymers, which can greatly enhance the fluorescence emission of functional monomer after guest-host binding due to the strong hydrogen bonding restriction to the transformation of monomer conformations. The prediction is in good agreement with the experimental observation. Moreover, the imprinted nanoparticles display significant fluorescence enhancement upon titration with different concentrations of melamine in methanol. The fluorescence sensors can be applied to detect the melamine in dairy products with a low limit of quantification of 0.5 μM. The results reported herein supply an excellent model for the design of molecularly imprinted fluorescence sensors and their prediction of chemical sensitivity to nonfluorescent compounds.

A novel metal–dye system for urinary phytate detection at micro-molar levels in rats by A. Costa-Bauza; F. Grases; S. Fakier; A. Rodriguez (3016-3022).
At present, beneficial properties of phytate have been observed. However, due to the special characteristics of this molecule and its low concentrations in physiological samples, the analytical determination of phytate in these samples becomes a complex and tedious problem. Therefore, the aim of this paper was to develop a simple and reliable methodology for phytate analysis in rat urine. In this study, two diets were administered to Wistar rats, namely: UAR-A03 (standard diet containing 1% phytate) and AIN-76A (purified diet with undetectable phytate). 24 h urine samples were collected and analysed for phytate by employing a procedure based on the purification and preconcentration of phytate by solid-phase extraction prior to detection with Al(iii)–xylenol orange. All variables for both the solid phase extraction and colorimetric assay were optimized. The working linear range for colorimetric detection was 0–12 μM phytate. The limit of detection was 0.044 μM and the limit of quantification was 0.188 μM. The relative standard deviation (RSD) for 1 μM phytate was 1.46%. When the standard rat chow was replaced by the phytate-poor rat chow, the urinary phytate concentrations fell to very low values after 15 days. These concentrations were restored to normal values when the standard diet was newly supplied. A recovery study was conducted with satisfactory results (88%). The results obtained in the present study, using the new metal–dye system, are comparable to the results obtained with an alternative method of detection. All these results demonstrate the suitability of the new method for phytate analysis in rat urine.

A friendly environmental and low-cost preparation of water-soluble photoluminescent carbon nanodots (C-Dots) utilizing the rhizome of the plant Giant Knotweed Rhizome, a traditional Chinese medicine, as a carbon source via hydrothermal method is developed for the first time. The as-prepared photoluminescent C-Dots show favorable blue color photoluminescence at the peak wavelength of 430 nm with a quantum yield of approximately 11.5%. It has been successfully applied as a novel probe for the detection of Hg2+. This novel probe exhibits excellent sensitivity and selectivity toward Hg2+ in the broad linear range from 50 nM to 100.0 μM with a limit of detection as low as 8.2 nM. The practical use of this system was tested in river water by adding 10.0 μM and 50.0 μM Hg2+. The recoveries ranged from 101.0% to 104.0% and from 99.2% to 101.2%, respectively. These stable and economical photoluminescent C-Dots show promise as a powerful tool for environmental analysis.

A liquid chromatography-electrospray ionization-tandem mass spectrometric method (LC-ESI-MS/MS) was developed and validated for the direct determination of 11-nor-Δ9-tetrahydrocannabinol-9-carboxylic acid (THCCOOH) and its glucuronide (THCCOOH-glu) in urine. The deuterium-labeled compounds were used as internal standards to quantify the analytes. After a simple liquid–liquid extraction with ethyl acetate at pH 4, the dried extracts were reconstituted in the mobile phase followed by injection onto the LC-ESI-MS/MS system. The mobile phase was composed of 0.2% formic acid and 2 mM ammonium formate in water and acetonitrile. A reversed phase column (C8, 150 mm × 2.1 mm i.d., 3.5 μm) was used as the analytical column. The limit of detection and lower limit of quantification values were 0.08 and 0.3 ng mL−1 for THCCOOH and 0.4 and 1.5 ng mL−1 for THCCOOH-glu, respectively. The calibration curve was linear over the concentration range of 0.3–100 ng mL−1 for THCCOOH and 1.5–100 ng mL−1 for THCCOOH-glu with the coefficients of determination (r2) above 0.9955. The intra- and inter-day precisions (% CV) were within 16.6% and 18.3%, respectively. The intra-day accuracies (% bias) ranged from −18.4 to 1.5% while inter-day accuracies were −5.9 to 1.8%. The validated method was successfully applied for the determination of THCCOOH and THCCOOH-glu in urine samples from marijuana users.

In this paper some of the most common methods for overcoming matrix effects in LC/ESI/MS (matrix-matched calibration, standard addition, post-column standard infusion, extrapolative dilution, and post-column flow splitting) are compared according to their ability to give both true and accurate results for pesticide determination in complicated matrices such as onion and garlic. In order to provide a quantitative comparison we use a measure of accuracy that would account for both average trueness and scatter of the results. Extrapolative dilution and standard addition were found to give results statistically insignificantly different from the correct values. In addition extrapolative dilution – a hybrid approach for both reducing and correcting for matrix effects – was found to result in the highest accuracy of the measurements.

Qualitative and quantitative analysis of effective components in Fructus Ligustri Lucidi by near infrared spectroscopy by Shi-sheng Wang; Li-juan Luo; Yue-qing Li; Rui Cai; Wei-jie Zhao (3045-3049).
In an attempt to establish the qualitative and quantitative analysis of Fructus ligustri lucidi, near-infrared spectroscopy (NIRS) was applied. The qualitative model can identify nine alcohol-broiled products from eight different geographical origins and one crude sample with a factorization algorithm and pretreatment of the second-derivative (2-Der). For the quantitative models, a partial least squares (PLS) algorithm with optimum parameters was applied for determination of oleanolic acid (OA) and ursolic acid (UA) in Fructus ligustri lucidi. There is no significant difference for the observed values between the NIRS and HPLC method since the results of the t-tests are 0.590 and 0.441, respectively, both less than t(0.05,19) = 2.093 with a 95% confidence level. The data obtained revealed the proposed NIRS method is viable for the rapid qualitative and quantitative analysis of the active ingredients in Fructus ligustri lucidi.

Nonenzymatic glucose sensor based on three different CuO nanomaterials by Fuying Huang; Yanmei Zhong; Jie Chen; Shunxing Li; Yancai Li; Fei Wang; Shuqing Feng (3050-3055).
A non-enzymatic glucose biosensor was constructed by immobilizing three different nanostructures of CuO (nanoparticles, nanoplatelets, nanorods), respectively, on a glassy carbon electrode (GCE) with Nafion. Compared with the bare GCE, the modified electrodes exhibited excellent catalysis to direct glucose oxidation. An electrochemical study showed that CuO nanoparticles exhibited the highest catalytic effect on glucose oxidation compared to CuO nanorods or nanoplatelets. This can be attributed to the fast electron transfer ability and highly specific surface area of the nanoparticles. Amperometric response showed that the CuO nanoparticle-modified electrode had a good response for glucose with a linear range up to 6 mM CuO and a sensitivity of 1.43 mA cm−2 mM−1 in 0.2 M NaOH. Detection of ascorbic acid (AA), dopamine (DA) and uric acid (UA) was carried out using these CuO modified electrodes, the results indicated good selectivity for glucose detection.

An accurate, rapid and reliable method for the simultaneous determination of 11 pesticide residues in tea by liquid chromatography–electrospray ionization tandem mass spectrometry has been established. Samples were purified by a modified QuEChERS method, followed by analysis by LC–ESI(+)-MS/MS and quantification using isotope internal standards. In this study, different extraction conditions, such as soaking time, extraction time of pesticides from the real tea samples and spiked samples, were optimized and compared. It was found that pesticides in real tea needed more time to be soaked and extracted than those in spiked tea for complete extraction. In order to eliminate the matrix effect and obtain satisfactory recoveries, the dosage of sorbents PSA (125, 250, 400, 600, 800, 1000, 1200 mg) and GCB (0, 25, 50, 250, 500 mg) was optimized in this study, and 800 mg PSA, 25 mg GCB, 750 mg MgSO4 were used in the purification process. In addition, the use of isotope internal standards and blank matrix matched standard calibration are discussed in depth. When the concentration range of 11 pesticides was 1–500 ng g−1, the linear correlation coefficients were 0.9981–0.9999. At the low, medium and high fortification levels of 2–100 ng g−1 (n = 6), recoveries were within 88–103%. The relative standard deviation was between 0.2% and 7.6%, and the limits of detection (LODs) ranged from 0.02 to 0.38 ng g−1 for 11 pesticides. It was found that the method was successfully applied to the qualitative and quantitative analysis of pesticides in actual commercial tea samples.

Major sources of uncertainties in the analysis of methylmercury using gold amalgamation sampling by Ki-Hyun Kim; Nhu-Thuc Phan; Yong-Hyun Kim; Hye-On Yoon; Richard J. C. Brown (3068-3073).
In an effort to improve the accuracy of measurement techniques for methylmercury (MeHg), the possible experimental bias in its analysis by the gold amalgamation method was investigated with special reference to gaseous elemental mercury (GEM). To this end, a series of calibration experiments were conducted using liquid-phase standards of MeHg prepared at four concentrations (2, 4, 10, and 20 ng μL−1) at each of four injection volumes (between 0.2 and 2 μL). Calibration results of MeHg were then examined mainly in terms of two calibration approaches: a fixed standard concentration (FSC) method (variable volumetric injection of standards prepared at a given concentration) and fixed standard volume (FSV) method (injection of multiple standards with varying concentrations at a given volume). The calibration results of MeHg, when evaluated against those of GEM, indicate that the FSC-based calibration of the former is unreliable due to the deterioration of linearity with increasing loading volumes of liquid standard. Thus it is recommended that the FSV approach should be used as the liquid sample loading method to improve the robustness and to minimize the calibration bias of MeHg.

A new mixed-mode hydrophilic interaction/cation exchange monolithic silica stationary phase was prepared by chemical modification of a monolithic silica skeleton by dextran sulfate (DS) using 3-aminopropyl-triethoxysilane as a cross-linking agent. The characteristics of the DS column were evaluated by the separation of various biomolecules including amino acids, nucleosides and bases, nucleotides, and peptides. Good separations for these polar biomolecules were obtained within 11 min on the column using isocratic mode. The investigation for the separation mechanism indicated that strong hydrophilic and electrostatic interactions combined to play a role in the separation of these biomolecules on the DS column.

The use of a dual mode sample introduction system for internal standardization in the determination of Hg at the ng L−1 level by cold vapor ICP-MS by Heidi Pyhtilä; Matti Niemelä; Paavo Perämäki; Juha Piispanen; Liisa Ukonmaanaho (3082-3088).
A dual mode sample introduction system was applied to simultaneous connection of cold vapor (CV) and liquid nebulization with ICP-MS. Gaseous mercury from a cold vapor system and liquid aerosol generated by pneumatic nebulization were transported into plasma via individual argon gas flow. The purpose of this configuration was to use aqueous internal standards for drift correction in low-level mercury analysis by CV-ICP-MS. Platinum, thallium and bismuth were studied as potential internal standards for mercury. The results indicated that each internal standard studied can be used to correct mercury signal instabilities during CV-ICP-MS analysis. The accuracy and precision of the results obtained using a dual mode system were confirmed by analyzing a certified reference material (ERM-CA615, groundwater). The internal standardization using a dual mode system was then applied to determine mercury concentrations in natural water samples. The results obtained for natural water samples using internal standards were in good agreement with those obtained by an isotope dilution CV-ICP-MS method, which could be considered as a reference method.

A method based on the cloud point extraction-high performance liquid chromatography (CPE-HPLC) has been developed for simultaneous extraction and quantification of the two pesticides, chlorpyrifos and chlorpyrifos-methyl, in water, soils and food vegetables. The non-ionic PEG-6000 was employed as the extraction solvent. Parameters and variables that affected the cloud point extraction (e.g. equilibration temperature and concentration of Na2SO4) were investigated. The optimal conditions for CPE were presented with 6% (w/v) PEG 6000, 3% (w/v) Na2SO4 and equilibration temperature at 65 °C for 65 min. Under these conditions, the recoveries of chlorpyrifos and chlorpyrifos-methyl spiked in water were 83.2–90.3% (RSD, 1.0–1.7) and 83.9–95.2% (RSD, 1.1–2.1), the recoveries in cabbage vegetable were 87.2–92.2% (RSD, 1.2–2.5) and 83.4–88.2% (RSD, 1.2–4.0), and those in soil were 88.8–100.6% (RSD, 1.7–4.1) and 83.9–92.9% (RSD, 0.9–2.4), respectively. The method presented here was proved to be rapid, efficient, and green for extraction and determination of chlorpyrifos and chlorpyrifos-methyl residues from food and multi-environmental media.

A novel ion imprinted polymer as a highly selective sorbent for separation of ruthenium ions from environmental samples by Elżbieta Zambrzycka; Urszula Kiedysz; Agnieszka Z. Wilczewska; Barbara Leśniewska; Beata Godlewska-Żyłkiewicz (3096-3105).
A complex of Ru(iii) with benzaldehyde thiosemicarbazone was used for the preparation of new ruthenium ion imprinted polymers. The polymers were synthesized using 4-vinylpyridine and styrene as functional monomers, divinylbenzene as a cross-linking agent, and ethanol or chloroform as a porogen in the presence of 2,2′-azobisisobutyronitrile as an initiator via the bulk polymerization technique. The polymers were used in dynamic mode as a selective sorbent for solid phase extraction of Ru(iii) ions. The analyte was efficiently retained on the polymers at pH 8.0 ± 0.1 and eluted with 2 mL of 0.3 mol L−1 thiourea in 0.3 mol L−1 HCl. The influence of the type of porogen (ethanol and chloroform) and column size on the separation of analyte on the prepared material was also studied. The efficiency of retention and elution and sample flow rates were higher on columns of smaller diameter and the polymer prepared in ethanol. The analytical performance of the polymer prepared in ethanol such as the recovery of the Ru(iii) ions, the reproducibility of the results (<3%), the limit of detection (0.26 ng mL−1), the selectivity in the presence of Pt(iv), Pd(ii), Rh(iii), Co(ii) ions and the sorption capacity (237.4 μg g−1) was superior in comparison to other ruthenium imprinted polymers. The developed separation procedure was successfully applied to electrothermal atomic absorption spectrometric determination of trace amounts of ruthenium in tap and river water, municipal sewage, road runoff, and grass.

Determination of dronedarone in the pharmaceutical dosage form by a stability-indicating micellar electrokinetic chromatography method by Ana Isa Pedroso Marcolino; Maximiliano S. Sangoi; Martin Steppe; Andréa Inês Horn Adams; Clarice Madalena Bueno Rolim (3106-3113).
In this study, a micellar electrokinetic chromatography method was developed and validated for the analysis of dronedarone in film-coated tablets. Electrophoretic conditions were investigated by changing factors such as pH, buffer concentration, SDS concentration, capillary temperature, injection time and applied voltage. Separation was performed using a bare fused-silica capillary of 40.0 cm effective length (48.5 cm total length; 50 μm internal diameter) maintained at 30 °C and detection was set at 216 nm. Optimal conditions were obtained using 40 mM borate buffer and 50 mM SDS at pH 9.2 as running buffer with an applied voltage of 28 kV (positive polarity) and using hydrodynamic injection at 50 mbar for 7 s. The method was validated by evaluating typical validation characteristics such as specificity, linearity, accuracy, precision, the limit of detection, the limit of quantitation and robustness. The analytical curve was linear in the concentration range of 25 to 150 μg mL−1 (r = 0.9995). The accuracy was 99.9% and the relative standard deviations of repeatability and intermediate precision were lower than 2%. The limit of detection and limit of quantitation were 0.88 μg mL−1 and 2.66 μg mL−1, respectively. The method proved to be robust by a fractional factorial design evaluation. Forced degradation studies were performed by exposing dronedarone sample solution to stress conditions (acidic, basic, oxidative, thermal and photolytic) in order to verify the stability-indicating capability of the method. The MEKC method was successfully applied for the quality control of dronedarone hydrochloride in commercial film-coated tablets.

Laser Induced Breakdown Spectroscopy (LIBS) has been applied to inked linen textile that belongs to a mummy's linen wrapping dated back to the Ptolemaic period (330 BC: 30 AD). The rarity of the ancient archeological piece introduced in this study has suggested the use of a model sample of currently manufactured linen for comparison purposes and optimization of the experimental conditions. The Nd:YAG laser operating at both wavelengths 532 and 1064 nm as our excitation source along with an Echelle spectrometer with an intensified charge-coupled device detector has been employed. Under the experimental conditions adopted throughout this work, the use of the visible 532 nm laser produced poorer S/B when compared to that produced by a 1064 nm laser which suggests the production of colder plasma leading to less atomization of the ablated material. Additionally, the 532 nm wavelength shows a negative behavior in ablating ink writings which was visually clear. Although, LIBS qualitative results are so comparable to that of SEM-EDX, some elements were detected only by LIBS which could be attributed to the irregularity of ink on linen. Thus, the capabilities of LIBS should be extensively exploited to the in situ measurements and analysis of archeological ink and fabrics.

Molecularly Imprinted Polymers (MIPs) against imiquimod, a highly potent immune response modifier used in the treatment of skin cancer, were synthesised using a template analogue strategy and were compared with imprints of the drug itself. An investigation of the complexation between the functional monomer and the template analogue revealed an association constant of 1376 ± 122 M−1, significantly higher than previously reported values for similar systems. The binding characteristics of the synthesised imprinted polymers were evaluated and extremely strong binding for imiquimod was observed while imprinting factors as high as 17 were calculated. When applied as sorbents in solid-phase extraction of imiquimod from aqueous, urine and blood serum samples, clean extracts and recoveries up to 95% were achieved, and it is concluded that while imiquimod imprints exhibited higher capacity for the drug, template analogue imprints are more selective. The results obtained suggest potential applications of imiquimod imprints as sorbents in rapid extraction and monitoring of undesirable systemic release of the drug.

Development and validation of methods for the determination of copper and iron in serum of dogs with canine visceral Leishmaniasis using multivariate optimization and GF AAS by Carolina Carvalho de Souza; José Henrique Ferraz Fabrino; Mark A. Beinner; Waldomiro Borges Neto; Silvia Dantas Cangussu; Wagner Luis Tafuri; José Bento Borba da Silva (3129-3135).
In this work efficient methods to determine copper and iron in dog serum samples by graphite furnace atomic absorption spectrometry were developed. The samples were diluted at a 1 : 9 or 1 : 19 (for Cu and Fe, respectively) ratio with 1% (v/v) nitric acid containing a 0.1% Triton X-100 solution. Rhodium for copper and no modifier for iron proved to be the better permanent modifier. Optimization included fractional factorial planning using Pareto and the CCD designs. For both analytes the working linear range was 0–100 mg L−1 (r2 > 0.99). The obtained LOQ was 19.3 ± 2.8 μg L−1 for Cu and 16.5 ± 0.2 μg L−1 for Fe. Aqueous and matrix matching calibration curves had average angular coefficients that were not statistically different, i.e. the matrix effect was absent for both analytes. The accuracy was checked by recovery tests with an average of 101 ± 4% (n = 45) for Cu and 90 ± 3% for Fe (n = 45). The certified reference material Seronorm™ Trace Elements Serum L-1 obtained was 1705 ± 20 μg L−1 for Cu (certified: 1691 ± 84 μg L−1) and 1.40 ± 1.22 mg L−1 for Fe (certified: 1.43 ± 0.08 mg L−1). Cu and Fe levels in 39 canine serum samples – 9 uninfected, 19 symptomatic and 11 asymptomatic dogs naturally infected with Leishmania chagasi– ranged from 374 to 913 μg L−1, and 1103 to 4260 μg L−1, respectively. Cu was higher in symptomatic than in asymptomatic and control dogs, and Fe was significantly lower in symptomatic than in either asymptomatic or control dogs.

Quantitative LC-MS/MS analysis of azide and azidoalanine in in vitro samples following derivatisation with dansyl chloride by Lieve Dillen; Luc Sips; Ronald de Vries; Carine Borgmans; Maarten Vliegen; Laurent Leclercq; Filip Cuyckens (3136-3141).
An LC-MS/MS method for the simultaneous quantification of azide and azidoalanine in bacterial and mammalian cellular systems was developed to support mutagenicity evaluation of azido containing nucleoside drugs. With the aim to enhance ionization efficiency and chromatographic retention, derivatisation with dansyl chloride was optimized; study samples (diluted 1/10) were derivatised for 2 h at room temperature and subsequently analysed with reversed phase liquid chromatography in combination with electrospray mass spectrometry in the positive ion mode. Good linearity and accuracy were obtained with a lower limit of quantification of 10.0 ng mL−1 for both analytes. The method was applied in mutagenicity experiments and showed time-dependent release of azide, but no formation of azidoalanine could be detected.

A new analytical method was developed to determine trace arsenic in soil by coupling sequential injection multichannel ultrasonic extraction and online pre-reduction with slurry hydride generation atomic fluorescence spectrometry. Significantly enhanced sampling throughput, reduced sample and reagent consumption, and minimized potential contamination and analyte loss were achieved. Slurry samples spiked with different concentrations of As(iii) and hydrochloric acid were pumped into the ultrasonic extraction chamber for 20-minute irradiation to efficiently extract As, which were subsequently merged with a mixture of 1% (m/v) thiourea and 0.5% (m/v) ascorbic acid to pre-reduce As(v) species to As(iii), followed by the generation of AsH3via using KBH4. Upon optimization of experimental parameters, limits of detection (LODs) ranging from 30 to 70 ng g−1 were obtained, and the precisions (RSDs) of this method were better than 2.0%. The proposed method was used for the determination of trace arsenic in several Certified Reference soil samples, with the obtained results in excellent agreement with certified values based on a simple and fast standard addition method.

Feasibility of energy dispersive X-ray fluorescence determination of gold in soft tissue for clinical applications by Isidro B. Magaña; Pratik Adhikari; Megan C. Smalley; Sven E. Eklund; D. Patrick O'Neal (3148-3151).
The feasibility of using EDXRF for a rapid quantitative analysis of gold in tumor tissue has been investigated. The protocol described here demonstrates that sample biopsies can be analyzed in as little as 8 hours, with minimal sample preparation. Samples were prepared by drying a 35 μL aliquot of tissue dissolved in KOH in a standard EDXRF cup on a Prolene® support, producing a thin sample. Calibration curves of XRF peak intensity (cps per mA) to the gold concentration (0–50 ppm) were prepared for liver, tumor, and a mix of tissue types. A linear regression demonstrated an R2 correlation of 0.93. Analysis of experimental samples showed that gold accumulation in tumors (5.8–41.3 ppm) was in agreement with previous studies, where samples were processed using INAA or ICP-MS. This report provides guidance for elemental analysis of gold in tissue in a shorter time span, showing potential for its use in a clinical setting.

Back cover (3153-3154).