Effects of digital processing on repeatability assessment of a multiple reaction monitoring liquid chromatography–tandem mass spectrometry system by ISO 11843-7

ISO 11843 part 7 (ISO 11843-7) can provide a standard deviation (SD) of area measurements of a target peak through the stochastic behaviors of instrumental noises. The purpose of this study is to demonstrate that ISO 11843-7 can be applied to assess repeatability in an isocratic liquid chromatography–tandem mass spectrometry (LC–MS/MS) system without repetitive measurements. The relative standard deviation (RSD) of the peak area of ergosterol picolinyl ester, which was used as an example, on a multiple reaction monitoring (MRM) chromatogram was determined by ISO 11843-7. The RSD by ISO 11843-7 (N = 1) was within a 95% confidence band of the RSD by repetitive measurements (N = 6). Moreover, the effects of digital smoothing, such as moving average, were also examined on the repeatability assessment in LC–MS/MS by ISO 11843-7. From the results of the comparisons of the RSDs obtained by ISO 11843-7 and the repetitive measurements, it was shown that suitable RSDs of the peak area were obtained from the smoothed MRM chromatograms by the moving average for narrow data point windows (e.g., one-sixth of the peak width). In conclusion, the utility of repeatability assessment based on ISO 11843-7 has been expanded for the validation of an LC–MS/MS system.     

Whole-cell luminescence-based flow-through biodetector for toxicity testing

A new type of biodetector was designed based on a bioluminescence test with the bacterium Vibrio fischeri performed in a liquid continuous flow-through system. Here we describe the modification of a commercial tube luminescence detector to work in the flow mode by building a new flow cell holder and a new case including “top cover” to connect the flow cell with the waste and the incubation capillary in a light-proof manner. As different samples were injected successively it was necessary to keep the individual peaks separated. This was done using an air-segmented flow in the reaction coil. To afford fast screening, the incubation time of the sample and the Vibrio fischeri, which equaled the dead time of the detection system, was set at 5.6 min. Rapid monitoring of toxic substances is achieved by using 20 μL of sample and flow-rates of 110–150 μL min⁻¹. As a proof-of-principle, we show results for the detection of five selected di-, tri- and tetrachlorophenols at different concentrations varying from 1 to 200 mg L⁻¹. Calculation of inhibition rates and EC₅₀ values were performed and compared with corresponding values from the DIN EN ISO 11348-2 microplate format. Compared with the latter, the inhibition rates obtained with our flow-through biodetector for the compounds tested were generally about twofold lower, but importantly, a much faster detection is possible. Figure Flow scheme of the biodetector setup     

Surface finishing of enamels by sol-gel coating

Sol-gel coatings composed of SiO₂, SiO₂-TiO₂, SiO₂-ZrO₂ and SiO₂-TiO₂-ZrO₂ were prepared on different technical enamels as substrates. Thin (from 80–300 nm) and thick (up to 2 μm) films were deposited by the dip coating method. The chemical durability of the coated and uncoated enamels was tested against acidic and basic attack in accordance with DIN ISO standards. The experimental results show that the thin SiO₂ coatings increase the chemical resistance of the coated enamels against oxalic acid attack by a factor of 4 to 22 compared to the uncoated ones. Especially efficient were SiO₂ films of 1 μm thickness, based on MTEOS-TEOS solutions. SiO₂ films do not protect the enamels against NaOH attack; however, an SiO₂-ZrO₂ coating on a non-resistant low-network former enamel significantly increases its stability against this basic attack. SNMS in-depth profiles show that in this case Na, Ca and Ba diffuse from the enamel substrate into the sol-gel layer, apparently stabilizing it.     

Detection capability of tetracyclines analysed by a fluorescence technique: comparison between bilinear and trilinear partial least squares models

According to the committee decision of 12 August 2002 (2002/657/EC) the capability of detection, CCβ, must be set in all analytical methods not only at concentration levels close to zero but also at the maximum permitted limit (PL). In this work we describe a methodology which evaluates the capability of detection of a fluorescence technique with soft calibration models (bilinear and trilinear PLS) to determine tetracyclines (group B1 substances from annex 1 of Directive 96/23/EC). Its estimation is based on the generalisation of the procedure described in International Union of Pure and Applied Chemistry and in the ISO standard 11843 for univariate signals which evaluates the probabilities of false positive (α) and false negative (β). The capability of detection, CCβ, estimated from the second-order signal and the trilinear PLS model is 9.93 μg l⁻¹ of tetracycline, 17.75 μg l⁻¹ of oxytetracycline and 26.31 μg l⁻¹ of chlortetracycline, setting α and β at 0.05. The capability of detection, CCβ, determined around the PL (100 μg kg⁻¹ in milk and muscle) with the second-order signal is 109.4 μg l⁻¹ of tetracycline, 117.0 μg l⁻¹ of oxytetracycline and 124.9 μg l⁻¹ of chlortetracycline, setting α and β at 0.05. The results were compared with those obtained with zero and first-order signals. The effect of the interferences on the capability of detection was also analysed as well as the number of standards used to build the models and their calibration range.When a tetracycline is quantified in presence of uncalibrated ones by means of the trilinear PLS model the errors oscillate between 14.70% for TC and 9.57% for OTC.     

Evaluation of the limit of performance of an analytical method based on a statistical calculation of its critical concentrations according to ISO standard 11843: Application to routine control of banned veterinary drug residues in food according to European Decision 657/2002/EC

Traceability of the measurement of analytical parameters capable of evaluating the performance of methods is an important concept for the assessment of quality for the routine control, especially for residue monitoring of non-authorized medicinal substances in food from animal origin. The European Decision no. 657/2002/EC recommends to calculate two statistical limits, CCα and CCβ, which allow to evaluate the critical concentrations above which the method reliably distinguish and quantify a substance taking into account the variability of the method and the statistical risk to take a wrong decision. The calculation, which can be derived from the ISO standard no. 11843 is applied on a routine basis. An example displays a very simple way for evaluating the performance of an LC-MSMS method which has been validated a few years ago and is qualified onto a Micromass Quattro LCZ tandem mass spectrometer to monitor and confirm the nitrofuran metabolite residues in food from animal origin. Community Reference Laboratory for Antimicrobial Veterinary Drug Residue Control in Food from Animal Origin     

An estimation of the measurement uncertainty for an optical emission spectrometric method

 A validation procedure based on the ISO/IEC 17025 standard was used to demonstrate the long-term stability of a calibration process and to assess the measurement uncertainty of a standard test method for optical emission vacuum spectrometric analysis of carbon and low-alloy steel (ASTM E 415–99a). The validation was used to provide documented evidence that the selected method fulfils the requirements and that the method is ”fit for purpose”. A test for drift was applied to determine statistically whether the analytical results vary systematically with time. The accuracy and traceability of the optimised method were tested by an analysis of closely matched matrix certified reference materials (CRMs). The measurement uncertainty estimations took account of the precision study, the bias and its uncertainty, and the qualification of uncertainties not considered in the overall performance studies. Received: 2 November 2002 Accepted: 2 January 2003 Acknowledgement The author expresses gratitude to Dr. Aleš Fajgelj for helpful discussions during the 3rd Central European Conference on Reference Materials and Measurements. Presented at CERMM-3, Central European Reference Materials and Measurements Conference: The function of reference materials in the measurement process, May 30–June 1, 2002, Rogaška Slatina, Slovenia Correspondence to T. Drglin     

Chlorophyll-a determination: results of an interlaboratory comparison

A chlorophyll-a interlaboratory comparison was carried out to compare three different analytical chlorophyll-a determination methods: a German standard DIN 38412-16, a method of the HELCOM-Combine-Manual and the different “in-house” methods of participating laboratories. Eleven laboratories took part. They had to determinate the chlorophyll-a and phaeopigment content in two seawater samples taken from the Baltic Sea. Furthermore, for the assessment three different statistical evaluation methods were applied: the German standard DIN 38402-42, the Q-method combined with an estimator according to Huber and the Cofino-method. All analytical methods were appropriate to determine the chlorophyll-a content. The relative standard deviations of reproducibility for chlorophyll-a varied between 12 and 31%. None of the analytical methods was appropriate to determine the phaeopigment content quantitatively. The relative standard deviations of reproducibility for phaeopigments ranged between 87 and 158%. The applied statistical evaluation methods resulted in different assessments. The Q-method combined with an estimator according to Huber proved to be the best qualified method.Presented at the Eurachem PT Workshop September 2005, Portorož, Slovenia     

Accreditation of reference material producers: Let's get it right!

The accreditation of testing and calibration laboratories to ISO/IEC 17025 is increasingly calling for the accreditation of reference material (RM) producers. Two international guides, ISO Guide 34 (2000) " General requirements for the competence of reference material producers" issued by the ISO Committee on Reference Materials and ILAC-G 12 " Guidelines for the requirements for the competence of reference material producers" issued by the International Laboratory Accreditation Cooperation (ILAC), are already in use for this purpose. Recently however initiatives have been launched to accredit RM producers to ISO 17025 as calibration laboratories and it has been suggested that a combination of ISO/IEC 17025 " General requirements for the competence of testing and calibration laboratories" and ISO Guide 34 may be the best option. This publication is an expression of the position of the ISO Committee on Reference Materials (ISO/REMCO) on the standards and guides currently in use in the accreditation of RM producers. The paper discusses the advantages and disadvantages of these standards and guides from the perspective of benefit to RM producers and RM users. In conclusion, the use of ISO Guide 34 alone or in combination with ILAC-G 12 is the preferred system for the accreditation of RM producers. Therefore ISO/REMCO strongly encourages all accreditation bodies to adopt ISO Guide 34.     

Three-way models and detection capability of a gas chromatography-mass spectrometry method for the determination of clenbuterol in several biological matrices: the 2002/657/EC European Decision

Clenbuterol has been extracted by mixed solid-phase extraction from two biological matrices (bovine hair and urine) and detected by GC/MS (selected ion monitoring (SIM) and full-SCAN modes). The analytical signal has been modelled with univariate and three-way models, namely DTLD, PARAFAC, PARAFAC2, Tucker3 and trilinear PLS. Since clenbuterol is a banned substance a comparative study of the capability of detection (CCβ, X₀=0) has been performed as a function of the sample (hair, 74 μg kg⁻¹ and urine, 0.36 μg l⁻¹), the mode in which the signals are monitored (SCAN, 283 μg kg⁻¹ and SIM, 74 μg kg⁻¹) and the statistical model (univariate, 283 μg kg⁻¹ and trilinear PLS, 20.91 μg kg⁻¹). The capability of detection has been calculated as stated in ISO 11843 and Decision 2002/657/EC setting in all cases the probabilities of false positive and of false negative at 0.05.The identification of the mass spectra must be done to confirm the presence of clenbuterol and has been carried out through PARAFAC. The correlation coefficient between the spectra estimated by PARAFAC and the library spectra is 0.96 (hair, SCAN mode) and 1.00 (hair and urine, SIM mode).The Decision 2002/657/EC advocates the use of independent mass fragments to identify banned compounds. These recommendations together with the effect of the number of ions registered on the capability of detection have lead us to select five uncorrelated fragments (86, 243, 262, 264 and 277) from the data set of 210 ions by hierarchical clustering of variables.     

Towards more fair and effective doping tests based on chromatography with mass spectrometric detection

Van Eenoo and Delbeke in Accred Qual Assur (2009) have criticized Faber (in Accred Qual Assur, 2009) for not taking “all factors under consideration when making his claims”. Here, it is detailed that their criticism is based on a misunderstanding of examples that were merely intended to be illustrative. Motivated by this criticism, further discussion is provided that may help in the pursuit of more fair and effective doping tests, here exemplified by chromatography with mass spectrometric detection. Surely, any doping test can only be improved or even optimized if the risks of false positives and false negatives are well defined. This requirement is consistent with a basic principle concerning mathematical approximations (Parlett in “The symmetric eigenvalue problem”, Prentice-Hall, Englewood Cliffs, 1980): apart from just being good, they should be known to be good. Author’s reply to the response on “Regulations in the field of residue and doping analysis...” Papers published in this section do not necessarily reflect the opinion of the Editors, the Editorial Board and the Publisher.     

Comparison of models for estimation of measurement uncertainties. Determination of BOD<Subscript>5</Subscript> in wastewaters using a manometric method

Biochemical oxygen demand after 5 days (BOD₅) is a bioassay frequently used to determine the polluting strength of wastewaters. It quantifies the mass concentration of dissolved O₂ consumed during an incubation period of 5 days at 20 °C, in the dark, by biological oxidation of organic and/or inorganic matter in wastewater. In this work, the BOD₅ measurand is reported as γ(O₂). Reporting analytical results with estimation of uncertainty is an ISO/IEC 17025:2005 requirement and allows results to be compared and their fitness for purpose to be demonstrated. The γ(O₂) uncertainty was estimated using two models: the individual components’ model (model 1) and the quality-control-criteria model in which the trueness component is calculated using the equation for recovery from CRMs (model 2.A) and the standard deviation of all recoveries (model 2.B). Model 1 provides a realistic strategy for estimation of γ(O₂) uncertainty and for assessment of performance. Models 2.A and 2.B are expeditious and simple procedures for the 100–800 mg L⁻¹ range. When γ(O₂) < 100 mg L⁻¹ model 1 must be used because with both Models 2 uncertainty was underestimated, more drastically close to the limit of quantification. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

UPLC-MS/MS determination of doxazosine in human plasma

A sensitive, selective and rapid method for the analysis of doxazosine (DOX) in human plasma based on ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) is described. DOX and tamsulosin, used as an internal standard (IS), were extracted by liquid-liquid extraction, and the chromatography was performed on a C18 UPLC column packed with 1.7 μm particles. The total run time was 2 min. Detection was achieved by the multiple reaction monitoring of the following transitions: m/z 452→344 and m/z 409→228 for DOX and IS, respectively. Transitions of m/z 452→247 and m/z 409→271 were also collected for confirmation purposes. The calibration curve based on peak area ratio was linear up to at least 100 ng ml⁻¹, with a detection limit of 0.02 ng ml⁻¹ (a signal-to-noise ratio of 3). The method showed satisfactory reproducibility, and the short-term stability of the analyte was assessed. The method was successfully applied to the analysis of DOX in human plasma.     

Chemiluminescence system of Ce(IV)-RhB-bismerthiazol

A novel chemiluminescence system for the determination of bismerthiazol is first described in this paper. It is based on the chemiluminescence reaction of bismerthiazol and Ce(IV) in nitric acid solution. The emission intensity could be enhanced greatly by Rhodamine B. The chemiluminescence intensity was proportional to bismerthiazol concentration over the range 30–1000 μg/L. The detection limit was 12 μg/L (3σ) and the relative standard deviation is 2.4% for 500 μg/L of bismerthiazol (n = 11). The proposed method was successfully applied to the determination of bismerthiazol in water and in rice. The recovery was 96.4%–104.1%. __________ Translated from Chinese Journal of Applied Chemistry, 2007, 24(5): 593–595     

Feasibility studies for the preparation and certification of reference materials Part II: mineral oil contaminated waste materials

The 2-year international joint project HYCREF (Contract-No. G6RD-CT-2002-00854), funded by the European Commission in the 5th Framework programme, aimed to develop methods to prepare homogenous and stable water-, soil- and waste reference materials contaminated with mineral oil hydrocarbons and to test certify the mineral oil content by gas chromatographic methods. As mineral oil products are important sources for environmental contaminations, appropriate reference materials certified by using the new gas chromatographic methods (soil: ISO/FDIS 16703, waste: prEN 14039 and KW/04, water: ISO 9377-2) are highly needed. Additional to the HYCREF-results presented in part I–mineral contaminated soils–this second paper gives an overview of the feasibility study for the preparation and test certification of three waste reference materials (offshore marine sediment, building material and industrial waste). The following specifications, which had been defined in the project work plan, were reached successfully: uncertainty of the mineral oil content resulting from the certification exercise ≤5% and a between-bottle inhomogeneity of ≤3%. All three materials have been evaluated for long- and short term stability. They contain different levels and types of mineral oil and cover the full application range of ISO/FDIS 16703 and prEN 14039 (100–10,000 mg/kg). The expanded uncertainties U _cert of the three materials are about 6%. In this way, they are comparable to ERM^®-CC015a (U _cert=7.1%, mineral oil in a river sediment) and are suited to close the present gap of commercially available CRM for mineral oil determination in waste.     

Reliability of measurement results in food microbiology: the contribution of reference laboratories in the European Union and of international/European standardization

The contribution of reference laboratories in the European Union and of European/international standardization to the reliability of food microbiology measurement results is discussed. A set of European Union reference laboratories has been established. Each of them coordinates a network of national reference laboratories which, in turn, coordinate networks of laboratories in charge of official testing and sometimes own checks in each European Union country. Their contribution to the reliability of food microbiology measurement results is illustrated by three food safety cases: Listeria monocytogenes, coagulase positive staphylococci and milk/milk products. The contribution of European/international standardization focuses on two topics: method validation and measurement uncertainty. The standards covering these topics—EN ISO 16140 and ISO/TS 19036—are briefly discussed, and an update given on their ongoing revision.     

Mineral oil content in sediments and soils: comparability, traceability and a certified reference material for quality assurance

The performance of twelve laboratories with previously established proficiency in the determination of the mineral oil content in a fresh water sediment is described. The summation parameter total petrol hydrocarbon (TPH) is defined according to ISO 16703:2004 with regard to the sample preparation to be applied, the flame ionisation detection (FID) and the boiling range of C₁₀–C₄₀ to be integrated. Comprehensive tests of homogeneity and stability have been carried out on the candidate material using appropriate models. The outcome of the study served as the basis for the certification of the candidate reference material as ERM-CC015a. The certified mass fraction is 1,820±130mgkg⁻¹and traceability was established by using an appropriate calibration standard certified for the mass fraction of C₁₀–C₄₀. The interlaboratory scatter of measurement results in this exercise can largely be explained by the variability of the individual calibrations based on this common calibration standard.     

A simple flow injection spectrophotometric determination method for iron(III) based on O-acetylsalicylhydroxamic acid complexation

A simple and rapid flow-injection spectrophotometric method for the determination of iron(III) and total iron is proposed. The method is based on the reaction between iron(III) and O-acetylsalicylhydroxamic acid (AcSHA) in a 2 % methanol solution resulting in an intense violet complex with strong absorption at 475 nm. Optimum conditions for the determination of iron(III) and the interfering ions were tested. The relative standard deviation for the determination of 5 μg L⁻¹ iron(III) was 0.85 % (n = 10), and the limit of detection (blank signal plus three times the standard deviation of the blank) was 0.5 μg L⁻¹, both based on the injection volumes of 20 μL. The method was successfully applied in the determination of iron(III) and total iron in water and ore samples. The method was verified by analysing a certified reference material Zn/Al/Cu 43XZ3F and also by the AAS method.     

Nonlinear modeling of equilibrium sorption of selected anionic adsorbates from aqueous solutions on cellulosic substrates. Part 1: model development

A new nonlinear isothermal sorption model, incorporating Donnan equilibrium and electrical neutrality in the classical sorption model of direct dyes onto cellulosic substrates, as model adsorbates, is proposed. The nonlinear isothermal model was used to simulate equilibrium sorption of adsorbates containing ionic charges (z) of −2 to −4 on cellulose adsorbents at various temperature (T) and sodium chloride concentrations ([NaCl]). A detailed analysis of simulation results demonstrates that results based on the nonlinear sorption model highly agree with those based on the log-linear sorption model when the deviation in the concentration of sodium ions in the aqueous solution ([Na⁺] _S ) relative to [NaCl] used in the sorption system is restricted to <5.0%. Compared to the log-linear model, the nonlinear model avoids using graphical techniques that are relatively insensitive for determining important sorption parameters such as the internal accessible volume (V) and the standard affinity associated with sorption (−Δμ°). The nonlinear sorption model was used to examine the correlation of fit for previously reported sorption data. The model parameters V and −Δμ° based on curve fits were used to estimate V for cellulose as well as −Δμ°. The values were found to match those based on the conventional log-linear model when deviations of [Na⁺] _S relative to [`([\textNa ⁺ ]_S )] \overline{{[{\text{Na}}^{ + } ]_{S} }} were below 5%. The nonlinear model therefore provides a convenient and accurate technique to interpret the sorption of a range of anionic adsorbates on cellulosic substrates.     

Absolute quantification of semicarbazide-sensitive amine oxidase in human umbilical artery by single-reaction monitoring with electrospray tandem mass spectrometry

Semicarbazide-sensitive amine oxidase (SSAO; E.C.1.4.3.6.) is widely distributed in different tissues, particularly in vascular smooth muscle and adipose tissue. Its physiological function remains unclear. Up to now, the common method to determine SSAO is based on enzymatic activity measurement. However, enzymatic activity could be easily influenced by the temperature, pH, and circumstance. In the present study, we have developed the single-reaction monitoring (SRM) approach for measuring the absolute amount of SSAO expression in human umbilical artery based on LC-ESI-MS/MS. The measurement of protein was converted to the measurement of a unique peptide of SSAO from Homo sapiens. The peptide (YQLAVTQR) was confirmed to be unique to the SSAO in human using the ExPasy blast tools, and thus the synthetic peptide was used as the standard which can produce abundant parent ion (m/z = 490.0) and daughter ion (m/z = 687.4) in the mass spectrometry. Trap drive and fragmentation energy of MS/MS of the unique peptide was 60 V and 0.6 V, respectively. The calibration curve was linear over the range of 1.99–127.8 fmol/μL, with 1.99 fmol/μl of the lower limit of quantification. The inter- and intra-day precisions and recoveries for all samples were satisfactory. The results demonstrated SSAO protein concentration was 7.75 fmol/g wet weight. It proved that the novel assay was sensitive and selective to measure the amount of SSAO protein originated from H. sapiens.