Isotope Dilution Analysis Research Articles

Isotope dilution quantification of ultratrace gamma-glutamyl-Se-methylselenocysteine species using HPLC with enhanced ICP–MS detection by ultrasonic nebulisation or carbon-loaded plasma

A method for the accurate determination of ultratrace selenium species of relevance to cancer research, such as gamma-glutamyl-Se-methylselenocysteine (gamma-glutamyl-SeMC), using species-specific double isotope dilution analysis (IDA) with HPLC-ICP-MS is reported for the first time. The (77)Se-enriched gamma-glutamyl-SeMC spike was produced in-house by collecting the fraction at the retention time of the gamma-glutamyl-SeMC peak from a chromatographed aqueous extract of (77)Se-enriched yeast, pooling the collected fractions and freeze-drying the homogenate. The Se content of this spike was characterised using reverse isotope dilution mass spectrometry (IDMS) and the isotopic composition of this spike was checked prior to quantification of the natural abundance dipeptide species in garlic using speciated IDA. The extraction of the gamma-glutamyl-SeMC species in water was performed in a sonication bath for 2 h after adding an appropriate quantity of (77)Se-enriched gamma-glutamyl-SeMC to 50 mg of garlic to give optimal (78)Se/(77)Se and (82)Se/(77)Se ratios of 1.5 and 0.6, respectively. The effect of ultrasonic nebulisation, in comparison with the loading of the ICP with carbon (through the addition of methane gas on-line), on the detection of Se associated with gamma-glutamyl-SeMC using collision/reaction cell ICP-MS with hydrogen as collision gas was investigated. Sensitivity enhancements of approximately fourfold and twofold were achieved using USN and methane mixed plasma, respectively, in comparison with conventional nebulisation and conventional Ar ICP-MS. However, an approximately twofold improvement in the detection limit was achieved using both approaches (42 ng kg(-1) for (78)Se using peak height measurements). The use of species-specific IDMS enabled quantification of the dipeptide species at ng g(-1) levels (603 ng g(-1) Se) in the complex food matrix with a relative standard deviation (RSD, n = 4) of 4.5%, which was approximately half that obtained using standard addition as a confirmatory technique. Furthermore, good agreement was found between the gamma-glutamyl-SeMC species concentrations obtained using both calibration methods.

Use of Isotope-Labeled Aflatoxins for LC-MS/MS Stable Isotope Dilution Analysis of Foods

Aflatoxins are a group of very carcinogenic mycotoxins that can be found on a wide range of food commodities including nuts, cereals, and spices. In this study, the first LC-MS/MS stable isotope dilution assay (SIDA) for the determination of aflatoxins in foods was developed. The development of this method was enabled by easily accessible isotope-labeled (deuterated) aflatoxins B2 and G2, which were synthesized by catalytic deuteration of aflatoxin B1 and G1, purified, and well-characterized by NMR and MS. All four aflatoxins of interest (B1, B2, G1, and G2) were quantified in food samples by using these two labeled internal standards. The response factors (RF) of the linear calibrations were revealed to be matrix independent for labeled aflatoxin B2/aflatoxin B2 and labeled aflatoxin G2/aflatoxin G2. For labeled aflatoxin B 2/aflatoxin B 1 and labeled aflatoxin B2/aflatoxin G1 matrix-matched calibration was performed for the model matrices almonds and wheat flour, showing significant differences of the RFs. Limits of detection (LOD) were determined by applying a statistical approach in the presence of the two model matrices, yielding 0.31 microg/kg (aflatoxin B1), 0.09 microg/kg (aflatoxin B2), 0.38 microg/kg (aflatoxin G1), and 0.32 microg/kg (aflatoxin G2) for almonds (similar LODs were obtained for wheat flour). Recovery rates were between 90 and 105% for all analytes. Coefficients of variation (CV) of 12% (aflatoxin B1), 3.6% (aflatoxin B2), 14% (aflatoxin G1), and 4.8% (aflatoxin G2) were obtained from interassay studies. For further validation, a NIST standard reference food sample was analyzed for aflatoxins B1 and B2. The method was successfully applied to determine trace levels of aflatoxins in diverse food matrices such as peanuts, nuts, grains, and spices. Aflatoxin contents in these samples ranged from about 0.5 to 6 microg/kg.

Internal correction of spectral interferences and mass bias in ICP-MS using isotope pattern deconvolution: Application to the determination of selenium in biological samples by isotope dilution analysis

The internal and simultaneous correction of mass bias and spectral interferences (BrH+ and SeH+) in the determination of total selenium in biological samples by isotope dilution analysis is evaluated here using isotope pattern deconvolution (IPD). The procedure takes the measured intensities at masses 76 to 82 in the spiked samples (using 77Se as spike) and calculates the isotopic composition of selenium. This isotopic composition would be a linear function of the isotopic composition of natural abundance selenium and that of enriched 77-selenium if no mass bias or spectral interferences were present or if they were adequately corrected. So, the molar fractions of natural abundance selenium and isotopically enriched selenium are calculated using IPD along with the variance of the multiple linear regression model. The variance of the regression is first calculated when no correction is applied (either mass bias or spectral interferences) and then this value is minimised by applying different factors for mass bias and the ratios SeH/Se and BrH/Br. It is demonstrated that, using the SOLVER application of Excel, the regression variance can be minimized and the correction factors converge to those factor which could have been calculated by external correction. Hence, this procedure eliminates the need for any external correction of mass bias or isobaric interferences in isotope dilution analysis provided that enough isotopes of the element are measured. To illustrate the applicability of this method, raw data obtained by Hinojosa Reyes et al. in a previous paper (J. Anal. At. Spectrom., 2003, 18, 11) were recalculated using the proposed methodology with satisfactory results.

Acceleration of enzymatic hydrolysis of protein-bound selenium by focused microwave energy

This paper demonstrates for the first time that enzymatic hydrolysis of protein-bound selenium can be significantly accelerated by microwave energy. Extraction of selenomethionine from selenised yeast, used as the model sample, was performed using two-step enzymatic hydrolysis with protease/lipase/driselase in a focused microwave reactor. HPLC-ICP-MS with species-specific isotope dilution analysis (IDA) was used for species quantitation. To investigate enzymatic hydrolysis assisted by microwave energy the effect of parameters such as microwave power and extraction time on the extraction efficiency of SeMet from the Se-yeast CRM SELM-1 was investigated. The main chemical variables affecting enzymatic extraction of SeMet, such as pH, enzyme type, sample to enzyme mass ratio and sample to solvent mass ratio were set at the conditions used for conventional enzymatic hydrolysis (in a hybridisation oven). Using a power of 60 W and two consecutive extraction cycles, each of 15 min (total extraction time of 30 min) at 37 °C, the recovery of SeMet obtained by the proposed methodology and calculated based on the certified value of 3389 ± 173 mg kg−1SeMet was 99.6 ± 1.9% (n = 3). The major advantage of the newly proposed method is the dramatic reduction of extraction time in comparison with that of conventional enzymatic hydrolysis, which requires a two-step extraction, each cycle of approximately 20 h. This was achieved without compromising extraction efficiency. Application of the proposed method to the accurate quantification of SeMet in pharmaceutical yeast tablets with a total Se concentration of approximately 300 mg kg−1 was also addressed. The recovery of SeMet, which was calculated based on the mean of results [561.5 mg kg−1 (n = 11, with a standard deviation of 44.3 mg kg−1)] obtained for the yeast tablets in the international intercomparison CCQM-P86,1 was found to be 100.1 ± 3.3%.

Analysis of mercury-containing protein fractions in brain cytosol of the maternal and infant rats after exposure to a low-dose of methylmercury by SEC coupled to isotope dilution ICP-MS

The mercury-containing protein fractions in the brain cytosol of the maternal and infant rats after exposure to low-dose of methylmercury chloride (0.3 mg Hg kg−1 day−1) were analyzed by a SEC-ICP-MS method with the postcolumn isotope dilution analysis. The enriched spiking including 34S, 65Cu, 67Zn and 198Hg was continuously added into the eluate from the HPLC column and then the isotope-diluted fractions were on-line measured by ICP-MS. Therefore, the absolute amounts of sulfur, mercury, zinc and copper in the eluted protein fractions could be attained after calculation of the corresponding peak areas in the mass flow chromatogram. Five mercury-containing protein fractions were monitored in the maternal sample, whereas only one was in the infant sample. The mercury content in the infant sample was about 10 times lower than the one in the maternal sample. In the meantime, two copper- and four zinc-containing fractions were found in both maternal and infant samples. The detection limit for S, Cu, Zn and Hg are 11, 0.1, 1.5 and 0.2 ng, respectively. The results demonstrate that different mercury-containing protein fractions may exist in brain cytosol between maternal and infant rats and the quantitative calculation may be helpful for the toxicological study.

Stable isotope labelling and FPLC–ICP-SFMS for the accurate determination of clinical iron status parameters in human serum

Iron is involved in the function of all living cells and, in fact, many diseases arise from imbalances in iron homeostasis. Therefore, the development of analytical methodologies to improve and automate the measurement of clinical indices of iron status has increased tremendously over the years. This work describes the development of two complementary methodologies to evaluate transferrin (Tf) saturation, total iron-binding capacity (TIBC), unsaturated iron-binding capacity (UIBC) and serum iron based on the use of iron-selective monitoring by inductively coupled plasma mass spectrometry (ICP-MS). The first methodology is based on the saturation of transferrin (Tf) with natural Fe3+ followed by separation of the different sialoforms in an anion exchange column (Mono-Q) to quantify the iron in each Tf sialoform and total Tf by ICP-MS using post-column isotope dilution analysis. In the second part, the saturation is done with an iron tracer (57Fe) and the application of pattern deconvolution analysis permits the direct quantification of the Tf saturation, the serum iron and the unsaturated iron-binding capacity. These strategies are validated by using a reference serum certified for total Tf and tested also in serum samples from individuals suffering from hemochromatosis and Fe-supplemented patients. The results obtained for all the parameters related to Fe status were in good agreement with those obtained by clinical tests. The use of stable isotope labelling in connection with the on-line coupling of fast protein liquid chromatography (FPLC) to ICP-MS allows the accurate determination of several parameters of great clinical relevance in iron homeostasis by means of two independent chromatographic runs. The main advantage of the proposed methodology is the number of parameters that can be simultaneously obtained.

Isotope pattern deconvolution for internal mass bias correction in the characterisation of isotopically enriched spikes

Enriched spikes of 53Cr, 57Fe, 67Zn, 95Mo, 111Cd, 143Nd, 150Sm, 199Hg and 207Pb were characterised on a multi-collector ICP-MS instrument both in terms of isotopic composition and concentration. The isotopic composition of the spikes was determined using either a natural abundance or certified standard for mass bias correction, except for lead where thallium internal correction was used. It was observed that the use of weighted regression lines for mass bias correction provided similar or, in most cases, lower uncertainties in the isotopic composition of the spikes in comparison with unweighted regression lines or single ratio measurements. For the establishment of the spike concentration by reverse isotope dilution analysis, isotope pattern deconvolution was evaluated and mass bias could be corrected internally in each blend by minimising the variance of the multiple linear regression model. It was also observed that isotope pattern deconvolution with internal mass bias correction provided lower uncertainties in the concentration of the spikes in comparison with the usual procedure based on external mass bias correction.

Isotope pattern deconvolution as a tool to study iron metabolism in plants

Isotope pattern deconvolution is a mathematical technique for isolating distinct isotope signatures from mixtures of natural abundance and enriched tracers. In iron metabolism studies measurement of all four isotopes of the element by high-resolution multicollector or collision cell ICP-MS allows the determination of the tracer/tracee ratio with simultaneous internal mass bias correction and lower uncertainties. This technique was applied here for the first time to study iron uptake by cucumber plants using 57Fe-enriched iron chelates of the o,o and o,p isomers of ethylenediaminedi(o-hydroxyphenylacetic) acid (EDDHA) and ethylenediamine tetraacetic acid (EDTA). Samples of root, stem, leaves, and xylem sap, after exposure of the cucumber plants to the mentioned 57Fe chelates, were collected, dried, and digested using nitric acid. The isotopic composition of iron in the samples was measured by ICP-MS using a high-resolution multicollector instrument. Mass bias correction was computed using both a natural abundance iron standard and by internal correction using isotope pattern deconvolution. It was observed that, for plants with low 57Fe enrichment, isotope pattern deconvolution provided lower tracer/tracee ratio uncertainties than the traditional method applying external mass bias correction. The total amount of the element in the plants was determined by isotope dilution analysis, using a collision cell quadrupole ICP-MS instrument, after addition of 57Fe or natural abundance Fe in a known amount which depended on the isotopic composition of the sample.

The role of sulfur and sulfur isotope dilution analysis in quantitative protein analysis

The element sulfur is almost omnipresent in all natural proteomes and plays a key role in protein quantification. Incorporated in the amino acids cysteine and methionine, it has been served as target for many protein-labeling reactions in classic quantitative proteomic approaches based on electrospray or MALDI mass spectrometry. This critical review discusses the potential and limitations of sulfur isotope dilution analysis (IDA) by inductively coupled plasma-mass spectrometry (ICP-MS) for absolute protein quantification. The development of this approach was made possible due to the improved sensitivity and accuracy of sulfur isotope ratio measurement by ICP-MS in recent years. The unique feature of ICP-MS, compound-independent ionization, enables compound (species)-unspecific sulfur IDA. This has the main advantage that only one generic sulfur standard (i.e., one isotopically labeled sulfur spike) is required to quantify each peptide or protein in a sample provided that they are completely separated in chromatography or electrophoresis and that their identities are known. The principles of this approach are illustrated with selected examples from the literature. The discussion includes also related fields of P/S and metal/S ratio measurements for the determination of phosphorylation degrees of proteins and stoichiometries in metalloproteins, respectively. Emerging new areas and future trends such as protein derivatization with metal tags for improved sensitivity of protein detection in ICP-MS are discussed.

New ways in qualitative and quantitative protein analysis: Nano chromatography coupled to element mass spectrometry

The potential of inductively coupled plasma-mass spectrometry (ICP-MS), which allows element-specific detection of heteroelements (e.g. Se and S) incorporated in protein structures, is highlighted for sensitive qualitative and quantitative protein analysis. ICP-MS coupled to separation techniques such as size exclusion chromatography and gel electrophoresis (via laser ablation) can be employed at different steps in the proteomic workflow. Special emphasis is made on the couplings of capillary and nanoHPLC to ICP-MS that required the development of dedicated interfaces. Element-specific peptide mapping by nanoHPLC–ICP-MS has turned out to be a key technique in combination with peptide sequencing via nanoHPLC–electrospray MS. This could impressively be demonstrated for the identification of selenium-containing proteins in selenium-rich yeast. Furthermore the potential of sulfur isotope dilution analysis in nanoHPLC–ICP-MS is presented as generic tool for highly accurate, absolute protein quantification.

Ion exchange in birnessite

Ion exchange isotherms for the sodium form of birnessite, a layered manganese oxide common in soils, were determined using isotope dilution analysis, by employing 22Na to pre-label the solid. The results were represented by plotting the equivalent ionic fraction of an entering cation in the ion exchanger versus the equivalent ionic fraction of the entering cation remaining in solution. Studying these ion exchange isotherms for sodium-univalent (i.e. NH4-Na, Li-Na, K-Na, and Cs-Na) and sodium-divalent pairs (Mg-Na, Ca-Na, Sr-Na and Ba-Na) on 22Na-birnessite made it possible to obtain a clear picture of the exchangeable sites available in birnessite. The isotherms also gave an estimation of the effect of cations likely to be present in aqueous environments on the uptake of the fission product radioisotopes of caesium and strontium onto soils.

Evaluating the potential and limitations of double-spiking species-specific isotope dilution analysis for the accurate quantification of mercury species in different environmental matrices

A new double-spiking approach, based on a multiple-spiking numerical methodology, has been developed and applied for the accurate quantification of inorganic mercury (IHg) and methylmercury (MeHg) by GC-ICPMS in different environmental matrices such as water, sediments and a wide range of biological tissues. For this purpose, two enriched mercury species (201MeHg and 199IHg) were added to the samples before sample preparation in order to quantify the extents of the methylation and demethylation processes, and thereby correct the final species concentrations. A critical evaluation of the applicability of this methodology was performed for each type of matrix, highlighting its main advantages and limitations when correcting for the conversion reactions of the species throughout the whole sample preparation procedure. The double-spike isotope dilution (DSIDA) methodology was evaluated by comparing it with conventional species specific isotope dilution (IDA) when analysing both certified reference materials and environmental samples (water, biotissues and sediment). The results demonstrate that this methodology is able to provide both accurate and precise results for IHg and MeHg when their relative concentrations are not too different (ratio MeHg/IHg > 0.05), a condition that holds for most natural waters and biotissues. Significant limitations on the accurate and precise determination of the demethylation factor are however observed, especially for real sediment samples in which the relative concentrations of the species are substantially different (ratio MeHg/IHg < 0.05). A determination of the sources of uncertainty in the methylation/demethylation factors has demonstrated that the accurate and precise measurement of the isotope ratios in the species involved in the transformations is crucial when quantifying the extents of these reactions. Although the double-spike methodology is established as a reference approach that permits the correction of most analytical biases and the accurate quantification of Hg species, some limitations have been identified for the first time in this work.

Chemical Preparation of an Isotopically Enriched Superoxide Dismutase and Its Characterization as a Standard for Species-Specific Isotope Dilution Analysis

The development of methods to analyze accurately and precisely individual metalloproteins is of increasing importance. Here we describe for the first time the chemical preparation and characterization of an isotopically enriched metalloenzyme containing two different metal isotopes. Its evaluation as a standard in species-specific isotope dilution analysis by HPLC coupled to inductively coupled plasma mass spectrometry is carefully evaluated. Our model enzyme bovine superoxide dismutase (SOD) contains both Cu and Zn and is remarkably stable at high temperatures and even under denaturing conditions. The enzyme's metal cofactors were removed under a range of different conditions and replaced with isotopically enriched 65Cu and 68Zn. Depending on the conditions, various isotopic ratios differing from the natural Cu and Zn abundances were obtained for the reconstituted enzyme. Both the wild type and isotopically enriched enzyme had the same migration pattern on native 1D-PAGE. Using an enzyme activity test, we showed that the incorporated 65Cu was bound to the right SOD-binding site, since the measured activity correlated directly with the amount of Cu incorporated. Mixing the native and the isotopically enriched enzyme standard with free enriched 65Cu and 68Zn or a metal chelator did not result in any exchange or loss of the metals from the enzyme at neutral pH. This verifies the stability of the enzyme metal center under the chosen conditions. The isotopically enriched enzyme standard was spiked into a wild type SOD solution to evaluate its use for species-specific isotope dilution experiments. To our knowledge, this is the first report of the chemical preparation of a metalloenzyme containing two different isotopically enriched metals. We provide evidence that the incorporated isotopically enriched metals are bound to the right binding site of SOD using an specific enzymatic activity assay.

Stable Isotope Dilution Analysis of the Fusarium Mycotoxin Zearalenone

Zearalenone is a secondary metabolite produced by molds of the Fusarium genus. Beside its nonsteroidal molecular structure, zearalenone has estrogenic activity and can disrupt the function of the endogenous hormone 17beta-estradiol in animals and possibly in humans. It can frequently be found in all major cereal grains as well as in processed food. Because of the estrogenic properties of zearalenone and its metabolites, legal regulations are installed in the European Union setting maximum levels in cereals and cereal products. Routine analysis of zearalenone in various commodities is carried out by HPLC with fluorescence detection, but due to the development of multi-mycotoxin methods and the reduced sample cleanup, HPLC-MS/MS has become a fast and efficient alternative. However, to achieve a reliable quantitation with this technique suitable internal standards are required. This paper reports the synthesis of stable isotope labeled 3,5- d 2-zearalenone (ZON) as internal standard for stable isotope dilution analysis. Furthermore, a method for the analysis of zearalenone by HPLC-MS/MS using 3,5- d 2-zearalenone as IS has been developed. Fifteen cereal products from the German retail markets were analyzed, of which seven contained ZON in levels from 4.9 to 45.0 microg/kg.

Iodine measurements by isotope dilution analysis in drinking water in Western Turkey

This study was designed to evaluate iodine concentrations in drinking water samples using isotope dilution analysis (IDA) in the Aegean region of Turkey A total of 76 drinking water samples from rural and urban areas in regional cities were analyzed. The mean iodine concentration was 78±27 μg/l and iodine concentration ranges were within 69±26 μg/l and 103±6 μg/l.

Optimization of GC–ICPMS system parameters for the determination of butyltin compounds in Hungarian freshwater origin sediment and mussel samples

The optimization and application of gas chromatograph coupled with inductively coupled plasma mass spectrometer (GC–ICPMS) (equipped with a commercially available interface) for the speciation of butyltin compounds in freshwater origin sediment and mussel samples is described. Optimization focused on the system parameters that have the greatest effect on signal intensity such as plasma power, ion lenses and make up gas flow (in the interface). Xenon (Xe) containing argon gas (Ar) was applied as tuning gas providing continuous Xe signal for the optimization of system parameters. It was found that plasma power and make up gas are interrelated variables and provide a set of paired optimum values at each power settings. The absolute optimum values obtained at 800 W plasma power and 1.2 L min −1 make up gas flow rate when 7 mm sample depth was adjusted. The optimum settings obtained were then checked by means of a test solution (tetraethyltin dissolved in hexane). Same optimum conditions were found when tin (Sn) transient signals were monitored. Detection limits were calculated for the three species using the optimized system parameters. Detection limits are the following: for monobutyltin (MBT) 5.6 ng Sn kg −1, for dibutyltin (DBT) 6.6 ng Sn kg −1 and for tributyltin (TBT) 3.4 ng Sn kg −1 obtained. Determination of the butyltin compounds were carried out by means of species-specific isotope dilution analysis. The spike solution contained all species investigated but with altered isotopic composition. Each species were enriched in their 119Sn isotope. Concentrations found in Hungarian freshwater origin mussel and sediment samples ranged between 19 and 39 ng g −1for MBT, between 1.2 and 6.3 ng g −1 for DBT and between 1.2 and 3.2 ng g −1 for TBT indicated as Sn in dry weight. Validation of the method was done by means of certified reference materials (BCR CRM 646 and 477). Good agreement was found between certified and experimental values. Normalized deviation ( E n) was also computed in order to validate the method used. E n values obtained ranged between 0.07 and 0.11 for mussel samples and between 0.26 and 0.72 for sediment samples. These values show that isotope dilution–GC–ICPMS methodology is valid for the determination of MBT, DBT and TBT from both types of matrices.

Stable isotope dilution analysis of N‐acetylaspartic acid in urine by liquid chromatography electrospray ionization tandem mass spectrometry

N-acetylaspartic acid (NAA) is a specific urinary marker for Canavan disease, an autosomal recessive leukodystrophy. We developed a 'dilute and shoot' stable isotope dilution liquid chromatography tandem mass spectrometry (LC-MS/MS) method for determination of NAA in urine. Deuterated internal standard d(3)-NAA was added to untreated urine and the mixture was injected into the LC-MS/MS system operated in the negative ion mode. Chromatography was carried out on a C(8) minibore column using 50% acetonitrile solution containing 0.05% formic acid at a flow rate of 0.25 mL/min. The retention time was 1.6 min and the turnaround time was 2.2 min. NAA and d(3)-NAA were analyzed in multiple reaction monitoring mode. Calibrators and quality control samples were prepared in pooled control urine. The assay was linear up to 2000 micromol/L with limit of quantification at 1 micromol/L (S/N = 12). Interassay and intraassay coefficients of variation were less than 7% and recovery at three different concentrations was 98.9-102.5%. The LC-MS/MS method for NAA as described involves no extraction and no derivatization, showed no interference and gave excellent recovery with low variability and short analytical time. The method was successfully applied for the retrospective analysis of urine from 21 Canavan disease cases.

New method for caffeine quantification by planar chromatography coupled with electropray ionization mass spectrometry using stable isotope dilution analysis

A new high-performance thin-layer chromatography/electrospray ionization mass spectrometry (HPTLC/ESI-MS) method for the quantification of caffeine in pharmaceutical and energy drink samples was developed using stable isotope dilution analysis (SIDA). After sample preparation, samples and caffeine standard were applied on silica gel 60 F254 HPTLC plates and over-spotted with caffeine-d3 used for correction of the plunger positioning. After chromatography, densitometric detection was performed by UV absorption at 274 nm. The bands were then eluted by means of a plunger-based extractor into the ESI interface of a single-quadrupole mass spectrometer. For quantification by MS the [M+H]+ ions of caffeine and caffeine-d3 were recorded in the positive ion single ion monitoring (SIM) mode at m/z 195 and 198, respectively. The calibration showed a linear regression with a determination coefficient (R2) of 0.9998. The repeatability (RSD, n=6) in matrix was<or=+/-3.75%. The intermediate precisions (RSD, n=2) for two samples of different brand names were determined three times and ranged between RSD+/-0.68% and+/-2.64% (sample 1) and between+/-3.44% and+/-8.60% (sample 2). The method accuracy was evaluated by comparing the results obtained by HPTLC/SIDA-ESI-MS with those from the validated HPTLC/UV method. The results for pharmaceutical and energy drink samples were (ng/band) 99.82+/-3.75 and 338.09+/-4.87 by HPTLC/SIDA-ESI-MS and 104.74+/-1.51 and 334.86+/-5.63 by HPTLC/UV. According to the F-test (homogeneity of variances) and the t-test (comparison of means) the two methods show no significant difference. The detection and quantification limits were 75 and 250 microg L-1 (0.75 and 2.5 ng/band), respectively, which were a factor of 13 lower than those established for HPTLC/UV. The positioning error (RSD+/-6%) was calculated by comparing HPTLC/SIDA-ESI-MS with HPTLC/ESI-MS. However, using SIDA the positioning error was nullified. HPTLC/SIDA-ESI-MS was demonstrated to be a highly reliable method for the quantification of compounds by planar chromatography coupled online with mass spectrometry.

Uranium-lead dating method at the Pará-Iso isotope geology laboratory, UFPA, Belém - Brazil

Analytical procedures for U-Pb isotope dilution analyses at the Pará-Iso isotope geology laboratory of the Federal University of Pará (UFPA) are described in detail. The procedures are applied to zircon, titanite, rutile, apatite, columbite-tantalite and whole rock. Reagent preparation and chemical processing are done in clean-room conditions. Samples are dissolved using Teflon microcapsules in steel jacket Teflon Parr Instrument bomb or Teflon screw cap containers. U and Pb are separated using anion exchange AG 1 x 8 resin columns. Typical blanks for mineral sample amounts of 0.01 to 1.0 mg are less than 1 pg U and 20-30 pg Pb. Isotope analysis of the U and Pb from the same filament are carried out using a Finnigan MAT 262 mass-spectrometer in static and dynamic modes. The current analytical level is demonstrated on analyses of international standard zircon 91500 with three different 235U-205Pb and 235U-208Pb isotope tracers and whole rock standards. Results of analyses of two zircon samples are also presented.

Substoichiometric determination of mercury with cetyltrimethylammonium bromide

A radiometric method based on substoichiometric isotope dilution analysis has been developed for the determination of mercury using cetyltrimethylammonium bromide (CTAB) as the substoichiometric reagent. The method is based on the extraction of mercury as an ionassociation complex of tetraiodomercurate(II) ion with a substoichiometric amount of CTAB in benzene. The calibration graph was found to be linear in the range 0–10 μg Hg (II) in the aqueous phase of 10 ml volume. A detection limit of 0.06 μg·ml−1 could be achieved by the proposed method. The relative standard deviation of the method was found to be 3.0% and the method has been successfully applied to study the determination of mercury in various synthetic mixtures.