Comite Consultatif Pour La Quantite De Matiere Research Articles

PAWG pilot study on quantification of total haemoglobin in blood

Main textUnder the auspices of the Protein Analysis Working Group (PAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) a pilot study, CCQM-P201, was organised. Five National Metrology Institutes (NMIs) participated in this pilot study and provided a total of eleven measurement results using six different methods based on three different measurement principles. The purpose of this pilot study was to develop measurement capabilities for the accurate quantification of intact haemoglobin (Hb) whereby measurement results could be made directly traceable to the SI. The choice of protein resulted from its ability to be quantified via different measurement principles providing different routes of traceability. Hb also provided a model of an intact protein with a complex tetrameric structure which is often measured via quantification of a specific subunit or the incorporated haem group. To achieve appropriate results for the chosen measurand, it was necessary to distinguish between the whole protein and fragments of the tetrameric structure, such as dimers or monomers. The participants were encouraged to investigate several of the measurement methods available. This was considered particularly advantageous in this instance as various methods applied to the same analyte may assist in identifying possible previously unknown biases or interferences of the various methods. Hb is being used as a model system to build capacity in methods for the quantification of proteins with a molar mass ≤ 100 kDa in biological fluids in a concentration range > 1∙106 pmol/g.This was the first pilot study for the quantification of an inherent protein in blood with all the natural variations that protein shows in a clinical sample. The different methods applied by the participants included isotopic dilution (ID) organic mass spectrometry (MS) of specific peptides after tryptic digestion, species-specific ID inductively coupled plasma mass spectrometry (ICP-MS), post-column ID ICP-MS, quantification via total iron after chromatographic isolation of Hb and acid digestion as well as two optical methods. One of the latter was the reference method proposed by the International Committee for Standardisation in Haematology (ICSH) based on the conversion of Hb to hemiglobincyanide (HiCN), the other used the conversion of Hb with alkaline haematin detergent both followed by the measurement of spectroscopic absorbance.Considering the use of very different measurement techniques for the quantification of total Hb, the results are in good agreement with a reference value with corresponding expanded uncertainty of (120.6 ± 1.2) mg/g calculated as DerSimonian-Laird mean. However, the observed differences between the results are not completely reflected in the reported measurement uncertainties. The calculated interlaboratory standard deviation or Tau value (dark uncertainty) was approximately 3 % whilst the average reported measurement uncertainty was approximately half this (1.4 %). A possible reason for this large difference may lie in the underestimation of any necessary conversion factors and their uncertainty contributions, between the target analyte measured and the requested measurand.To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Subsequent bilateral key comparison study - organic solvent calibration solution - gravimetric preparation and value assignment of aflatoxin B1 (AfB1) in acetonitrile (ACN)

Main textThe original CCQM-K154.b comparison was coordinated by the Bureau International des Poids et Mesures (BIPM) and the Chinese National Institute of Metrology (NIM) on behalf of the Organic Analysis Working Group (OAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) for National Measurement Institutes (NMIs) and Designated Institutes (DIs) which provide measurement services in organic analysis under the 'Comité International des Poids et Mesures' Mutual Recognition Arrangement (CIPM MRA) and/or have participated in the BIPM's Mycotoxin Metrology Capacity Building and Knowledge Transfer (MMCBKT) project as part of its "Metrology for Safe Food and Feed in Developing Economies" Capacity Building Programme.Gravimetrically-prepared solutions having an assigned mass fraction of specified organic analytes are routinely used to calibrate measurement processes for the quantification of the same analytes in matrix samples. Appropriate assignments of the property value and associated uncertainty of calibration solutions thus underpin the traceability of routine analysis and are critical for accurate measurements. Evidence of successful participation in relevant international comparisons is needed to document calibration and measurement capability claims (CMCs) made by national metrology institutes and designated institutes. In total, eleven NMIs/DIs participated in the Track C, Model II, Key Comparison CCQM-K154.b [Gravimetric preparation and value assignment of aflatoxin B1 (AfB1) in acetonitrile (ACN)] for emerging areas of global interest and innovation. Participants were requested to gravimetrically prepare calibration solutions and value assign the mass fractions, expressed in mg/kg, of aflatoxin B1 (AfB1) in the acetonitrile (ACN) solution. Study samples, with assigned values and associated uncertainties were prepared by the comparison participants and sent to the coordinating laboratory for comparison. The Key Comparison Reference Values (KCRVs), calculated form values measured by the coordinating laboratory based on calibrations obtained from independent gravimetrically prepared calibrant solutions, agreed with participants reported values, within their stated uncertainties.AfB1 was selected to be representative of polar aflatoxins. Aflatoxins are a class of mycotoxins generally produced by fungi of the genus Aspergillus. It was anticipated to provide a challenge representative for the gravimetrical preparation and value assignment of calibration solutions in the mass fraction range of 2 mg/kg to 50 mg/kg of mycotoxins with broadly similar structural characteristics.In the original study CCQM-K154.b, nine participants of the MMCBKT programme were provided with a stock solution having a known AfB1 mass fraction and expanded uncertainty to use to gravimetrically prepare and value assign a calibration solution. Three NMIs/DIs also participated using their own calibration solutions. The use of in-house solutions required anadditional capacity to undertake a fit-for-purpose purity assessment. NIM was the only NMI participating using both the MMCBKT based and their own in-house assigned solutions in order to connect the two different groups.It was decided to propose separate KCRVs for each of the two ampoules provided by the participating NMIs/DIs based on the AfB1 mass fraction. This allowed participants to demonstrate the efficacy of their implementation of the approaches used to gravimetrically prepare calibrations solutions and to assess the AfB1 mass fraction.The majority of the AfB1 mass fraction KCRVs (wKCRV) for CCQM-K154.b spanned a mass fraction range of 2.02 mg/kg to 31.57 mg/kg. The relative expanded uncertainties U(wKCRV) ranged from 0.69 % to 2.93 %. Inspection of the degree of equivalence plots for the AfB1 mass fraction assignments in CCQM-K154.b indicated that there was an excellent agreement of results. Solely, the AfB1 mass fraction assignments of the INRAP (Institut National de Recherche et d'Analyse Physico-Chimique, Tunisia) did not agree with the KCRVs. It was found that the samples were altered as a result of an acid contamination.The INRAP has produced a new batch of AfB1 calibration solution using the AfB1 stock solution originally provided to MMCBKT participants to be able to demonstrate the efficacy of their implementation of the approaches used to gravimetrically prepare calibrations solutions and to assess the AfB1 mass fraction. The BIPM organized the present subsequent bilateral comparison CCQM-K154.b.1 to assess the new batch of AfB1 calibration solution of the INRAP. Inspection of the degree of equivalence plots for the AfB1 mass fraction assignments indicated that there is an excellent agreement of the INRAP results for CCQM-K154.b.1.To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Key Comparison Study - Organic Solvent Calibration Solution - Gravimetric preparation and value assignment of patulin (PAT) in acetonitrile (ACN) with 0.1 % formic acid (FA)

Main textThe CCQM-K154.d comparison was coordinated by the the Bureau International des Poids et Mesures (BIPM) and the Chinese National Institute of Metrology (NIM) on behalf of the Organic Analysis Working Group (OAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) for National Measurement Institutes (NMIs) and Designated Institutes (DIs) which provide measurement services in organic analysis under the 'Comité International des Poids et Mesures' Mutual Recognition Arrangement (CIPM MRA) and/or have participated in the BIPM's Mycotoxin Metrology Capacity Building and Knowledge Transfer (MMCBKT) project as part of its "Metrology for Safe Food and Feed in Developing Economies" Capacity Building Programme.Gravimetrically-prepared solutions having an assigned mass fraction of specified organic analytes are routinely used to calibrate measurement processes for the quantification of the same analytes in matrix samples. Appropriate assignments of the property value and associated uncertainty of calibration solutions thus underpin the traceability of routine analysis and are critical for accurate measurements. Evidence of successful participation in relevant international comparisons is needed to document calibration and measurement capability claims (CMCs) made by national metrology institutes and designated institutes. In total, twelve NMIs/DIs participated in the Track C, Model II, Key Comparison CCQM-K154.d [Gravimetric preparation and value assignment of patulin (PAT) in acetonitrile (ACN) with 0.1 % formic acid (FA)] for emerging areas of global interest and innovation. Participants were requested to gravimetrically prepare calibration solutions and value assign the mass fractions, expressed in mg/kg, of PAT in ACN with 0.1 % FA solution. Study samples, with assigned values and associated uncertainties were prepared by the comparison participants and sent to the coordinating laboratory for comparison. The Key Comparison Reference Values (KCRVs), calculated form values measured by the coordinating laboratory based on calibrations obtained from independent gravimetrically prepared calibrant solutions, agreed with participants reported values, within their stated uncertainties.PAT is a mycotoxin produced by a variety of moulds, particularly by fungi of the genus Aspergillus and Penicillium. Often found in rotting apples and apple products, PAT can also occur in various other moldy fruits, grains and foods. Major human dietary sources of PAT are apples and apple juice made from affected fruit. PAT is relatively stable when given a short-acting heat-treatment, especially in an acidic environment. It was anticipated to provide a challenge representative for the gravimetrical preparation and value assignment of PAT calibration solutions in the mass fraction range of 10 mg/kg to 100 mg/kg of mycotoxins with broadly similar structural characteristics.Eight participants of the MMCBKT programme were provided with a stock solution having a known PAT mass fraction and expanded uncertainty to use to gravimetrically prepare and value assign a calibration solution. Two NMIs/DIs used reference materials for preparing their solutions while five NMIs/DIs participated using their own calibration solutions. The use of in-house solutions required an additional capacity to undertake a fit-for-purpose purity assessment. NIM was the only NMI participating using both the MMCBKT based and their own in-house assigned solutions in order to connect the two different groups.It was decided to propose separate KCRVs for each of the two ampoules provided by the participating NMIs/DIs based on the PAT mass fraction. This allowed participants to demonstrate the efficacy of their implementation of the approaches used to gravimetrically prepare calibration solutions and to assess the PAT mass fraction.The majority of the PAT mass fraction KCRVs (wKCRV) for CCQM-K154.d spanned a mass fraction range of 9.72 mg/kg to 63.40 mg/kg. The relative expanded uncertainties U(wKCRV) ranged from 2.0 % to 5.4 %.Inspection of the degree of equivalence plots for the PAT mass fraction assignments in CCQM-K154.d indicated that there was an excellent agreement of results.To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

PAWG pilot study on quantification of SARS-CoV-2 monoclonal antibody - Part 2

Main textUnder the auspices of the Protein Analysis Working Group (PAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) a pilot study, CCQM-P216, was coordinated by the Chinese National Institute of Metrology (NIM), National Research Council of Canada (NRC) and the Bureau International des Poids et Mesures (BIPM).The global coronavirus pandemic has also led to increased focus on antibody quantitation methods. IgG are among the immunoglobulins produced by the immune system to provide protection against SARS-CoV-2. Anti-SARS-CoV-2 IgG can therefore be detected in samples from affected patients. Antibody tests can show whether a person has been exposed to SARS-CoV-2, and whether or not they potentially show lasting immunity to the disease. With the constant spread of the virus and the high pressure of re-opening economies, antibody testing played a critical role in the fight against COVID-19 by helping healthcare professionals to identify individuals who have developed an immune response, either via vaccination or exposure to the virus. Many countries have launched large-scale antibody testing for COVID-19. The development of measurement standards for the antibody detection of SARS-CoV-2 is critically important to deal with the challenges of the COVID-19 pandemic. In this study, the SARS-CoV-2 monoclonal antibody is being used as a model system to build capacity in methods that can be used in antibody quantification.The purpose of this pilot study was to develop measurement capabilities for larger proteins using a recombinant humanized IgG monoclonal antibody against Spike glycoprotein of SARS-CoV-2 (Anti-S IgG mAb) in solution. A Final Report on the first round of CCQM-P216 PAWG Pilot Study on Quantification of SARS-CoV-2 Monoclonal Antibody - Part 1 focusing on the assessment of both mass fraction determinations of different AAs in the material and mass fraction determinations of proteotypic peptides belonging to the constant region of the mAb has already been published in Metrologia. The present Final Report on the second round of CCQM-P216 - Part 2 was designed to investigate optional methods for the characterization of Anti-S IgG mAb in solution for the assessment of size heterogeneity purity determinations, mass fraction measurements of SARS-CoV-2 monoclonal antibody in the material by UV-VIS spectrophotometry, mass fraction measurements of proteotypic peptides belonging to the variable region of the mAb and mass fraction measurements of monomeric mAb in the material. Six Metrology Institutes or Designated Institutes and the BIPM participated in the second phase of the pilot study (Part 2).Acceptable agreement between all laboratories was for the assessment of size heterogeneity of the anti-S IgG mAb material. For example, a reference value with its corresponding expanded uncertainties of (99.48 ± 0.49) % has been obtained for the monomer. Good between laboratories for UV-VIS spectrophotometric analysis at a pre-agreed wavelength of 280 nm was obtained with individual relative expanded uncertainties from 2.8 % to 15.5 % and a mass concentration reference value and corresponding expanded uncertainty of (500.8 ± 15.4) mg/L.Mass fraction assignments of the target proteotypic peptide YSPSFQGQVTISADK in the variable region of the anti-S IgG mAb material by NIM and BIPM resulted in a reference value and corresponding expanded uncertainty of (9.8 ± 2.0) mg/kg.Both NIM and BIPM have provided estimates for the mass fraction of monomeric mAb in the material with individual relative expanded uncertainties of 4.3 % and 4.1 %, respectively. A mass fraction reference value and corresponding expanded uncertainty of (411 ± 88) mg/kg was calculated by applying an 'excess-variance' approach for the monomeric mAb mass fraction.In addition, a post-hoc assessment of the monomeric mAb mass fraction from the subset of that produced both amino acid and proteotypic peptide-based mass fraction results and SEC measurements during the 1st and present study round provided more detailed information. A reference value and its expanded uncertainty for the mass fraction of monomeric mAb in the anti-S IgG mAb material was calculated to be (434.3 ± 11.9) mg/kg applying an 'excess-variance' approach.The level of agreement was not significantly poorer than that achieved in previous studies with smaller or less complex proteins. The two-stage pilot study was a big step forward to build and improve capacity within NMIs/DIs to undertake future key comparisons in the field of mass fraction determinations of antibodies and other large proteins.To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Key comparison study - organic solvent calibration solution - gravimetric preparation and value assignment of deoxynivalenol (DON) in acetonitrile (ACN)

Main textThe CCQM-K154.c comparison was coordinated by the the Bureau International des Poids et Mesures (BIPM) and the Chinese National Institute of Metrology (NIM) on behalf of the Organic Analysis Working Group (OAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) for National Measurement Institutes (NMIs) and Designated Institutes (DIs) which provide measurement services in organic analysis under the 'Comité International des Poids et Mesures' Mutual Recognition Arrangement (CIPM MRA) and/or have participated in the BIPM's Mycotoxin Metrology Capacity Building and Knowledge Transfer (MMCBKT) project as part of its "Metrology for Safe Food and Feed in Developing Economies" Capacity Building Programme.Gravimetrically-prepared solutions having an assigned mass fraction of specified organic analytes are routinely used to calibrate measurement processes for the quantification of the same analytes in matrix samples. Appropriate assignments of the property value and associated uncertainty of calibration solutions thus underpin the traceability of routine analysis and are critical for accurate measurements. Evidence of successful participation in relevant international comparisons is needed to document calibration and measurement capability claims (CMCs) made by national metrology institutes and designated institutes. In total, nine NMIs/DIs participated in the Track C, Model II, Key Comparison CCQM-K154.c [Gravimetric preparation and value assignment of deoxynivalenol (DON) in acetonitrile (ACN)] for emerging areas of global interest and innovation. Participants were requested to gravimetrically prepare calibration solutions and value assign the mass fractions, expressed in mg/kg, of deoxynivalenol (DON) in the acetonitrile (ACN) solution. Study samples, with assigned values and associated uncertainties were prepared by the comparison participants and sent to the coordinating laboratory for comparison. The Key Comparison Reference Values (KCRVs), calculated from values measured by the coordinating laboratory based on calibrations obtained from independent gravimetrically prepared calibrant solutions, agreed with participants reported values, within their stated uncertainties.DON belongs to the large group of trichothecene mycotoxins. It is produced by certain fungi of the genus Fusarium that predominantly infect wheat, corn, oats, barley, rice, and other grains in the field or during storage. It was anticipated to provide a challenge representative for the gravimetrical preparation and value assignment of calibration solutions in the mass fraction range of 10 mg/kg to 100 mg/kg of mycotoxins with broadly similar structural characteristics.Ten participants of the MMCBKT programme were provided with a stock solution having a known DON mass fraction and expanded uncertainty to use to gravimetrically prepare and value assign a calibration solution. Three NMIs/DIs also participated using their own calibration solutions. The use of in-house solutions required an additional capacity to undertake a fit-for-purpose purity assessment. NIM was the only NMI participating using both the MMCBKT based and their own in-house assigned solutions in order to connect the two different groups.It was decided to propose separate KCRVs for each of the two ampoules provided by the participating NMIs/DIs based on the DON mass fraction. This allowed participants to demonstrate the efficacy of their implementation of the approaches used to gravimetrically prepare calibration solutions and to assess the DON mass fraction.The majority of the DON mass fraction KCRVs (w KCRV ) for CCQM-K154.c spanned a mass fraction range of 9.88 mg/kg to 123.45 mg/kg. The relative expanded uncertainties U(w KCRV ) ranged from 2.8 % to 6.8 %.Inspection of the degree of equivalence plots for the DON mass fraction assignments in CCQM-K154.c indicated that there was an excellent agreement of results.To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Key comparison study on peptide purity - hexapeptide of HbA0

Main textUnder the auspices of the Protein Analysis Working Group (PAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) a key comparison, CCQM-K115.2018, was coordinated by the Bureau International des Poids et Mesures (BIPM), the Health Sciences Authority (HSA) of Singapore and the Chinese National Institute of Metrology (NIM). Ten Metrology Institutes or Designated Institutes and the BIPM participated. Participants were required to assign the mass fraction of the hexapeptide of HbA0 (VE) present as the main component in the comparison sample for CCQM-K115.2018. The comparison samples were prepared by HSA/BIPM from synthetic VE purchased from a commercial supplier and used as provided without further treatment or purification.VE was selected to be representative of the performance of a laboratory's measurement capability for the purity assignment of chemically synthesized peptides of known sequence, without cross-links, up to 5 kDa and without modification. It was anticipated to provide an analytical measurement challenge representative for the value-assignment of compounds of broadly similar structural characteristics.The majority of participants used amino acid analysis (PICAA) with a correction for structurally-related peptide impurities approach as the amount of material that had been provided to each participant (25 mg) was insufficient to perform a full mass balance based characterization of the material by a participating laboratory. The coordinators, the BIPM, the HSA and the NIM, were the laboratories to use the mass balance approach as they had more material available.It was decided to propose KCRVs for both the VE mass fraction and the mass fraction of the peptide related impurities as indispensable contributor regardless of the use of PICAA, mass balance or any other approach to determine the VE purity. This allowed participants to demonstrate the efficacy of their implementation of the approaches used to determine the VE mass fraction. In particular, it allowed participants to demonstrate the efficacy of their implementation of peptide related impurity identification and quantification.More detailed studies on the identification/quantification of peptide related impurities revealed that the integrity of the impurity profile of the related peptide impurities obtained by the participant is crucial for the impact on accuracy of the VE mass fraction assignment.The assessment of the mass fraction of peptide impurities is based on the assumption that only the most consistent set of results is taken for the calculation of the KCRVPepImp. The sum of the combined cis/trans VE depsipeptide impurities (only identified/quantified by NRC and confirmed by BIPM and HSA) and mass fractions of peptide related impurities that have been identified by at least two participants have been used to establish the KCRVPepImp. The KCRVPepImp of 53.0 mg/g is associated with a corresponding expanded uncertainty of 17.3 mg/g (k = 2) providing a more realistic basis of evaluation for the capabilities of the participants to identify/quantify peptide related impurities. Inspection of the degree of equivalence plots for the mass fraction of peptide impurities and additional information obtained from the peptide related impurity profile indicates that in all cases the major related peptide impurity, VE depsipeptide, has not been identified. The VE depsipeptide impurity was initially and uniquely identified and quantified by the NRC by the use of 1H-NMR. The related peptide impurity mass fraction results of only four participants (NRC, LGC, LNE and HSA) are in agreement with the KCRVPepImp. The NRC has identified and quantified the VE depsipeptide whereas the LGC, LNE and HSA have not identified the VE depsipeptide but accounted for that contribution.The approach selected to obtain a KCRVVE for the mass fraction of VE is based on a mass balance calculation that takes into account the most consistent set of results for the peptide related impurities KCRVPepImp, TFA mass fraction and the water mass fraction. The KCRVVE for CCQM-K115.2018 is 613 mg/g with a corresponding expanded uncertainty of the KCRVVE of 20 mg/g (k = 2).To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

High polarity analyte(s) in aqueous media: determination of L-PFOA and L-PFOS in ground water

Main textThe CCQM-K156 comparison was coordinated by TUBITAK UME on behalf of the Organic Analysis Working Group (OAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) for National Measurement Institutes (NMIs) and Designated Institutes (DIs) which provide measurement services in organic analysis under the 'Comité International des Poids et Mesures' Mutual Recognition Arrangement (CIPM MRA).Perfluoro alkyl substances (PFAS) such as PFOS and PFOA have been used in numerous industrial applications and products. Because of their high stability and resistance to biodegradation, atmospheric photooxidation, direct photolysis and hydrolysis, they are extremely persistent in the environment. The European Union (EU) Water Framework Directive lists PFOS as a priority hazardous substance that poses a significant risk to the aquatic environment. The use of PFOS-containing Aqueous Film-Forming Foams (AFFFs) has been banned since June 2011 in the EU. As relatively water-soluble, effectively non-degradable compounds, PFOS and PFOA migrate to ground water. They are not removed in the conventional drinking water treatment, and therefore cause health risks in polluted areas. The EU Drinking Water Directive and the European Commission has proposed a limit value of 100 ng/L for the sum of 20 PFAS, including PFOS and PFOA.This study provides the means for assessing measurement capabilities for determination of high polarity measurands in a procedure that requires extraction, clean-up, analytical separation and detection. Successful participation in CCQM-K156 demonstrates measurement capabilities in determining mass fraction of organic compounds, with a molecular mass of 200 g/mol to 700 g/mol, having high polarity pKow > -2, in a mass fraction range from 0.5 ng/kg to 500 ng/kg in aqueous media.Nine NMIs and DIs participated in the CCQM-K156 key comparison. Seven institutes reported their results. SPE was applied in the sample pre-treatment and LC-MS was applied for detection. All participating laboratories applied isotope dilution mass spectrometry (IDMS) techniques for quantification. Participants established the metrological traceability of their results using certified reference materials (CRMs) from NMIs with stated traceability; where commercially available high purity materials were used the purity was determined in-house. The CCQM-K156 results for L-PFOA and L-PFOS range from 2.75 ng/kg to 5.50 ng/kg with a % RSD of 19.5 % for L-PFOA and from 2.04 ng/kg to 4.45 ng/kg with a % RSD of 21.3 % for L-PFOS. The KCRV was assigned using a Hierarchical Bayesian Random Effects Model (HB REM) estimator from the values reported by six of the participants. One participant result of L-PFOS and one result of L-PFOA were excluded from the KCRV for technical reasons. The KCRV was 4.9 ng/kg ± 0.4 ng/kg for L-PFOA and 3.8 ng/kg ± 0.4 ng/kg for L-PFOS. The six institutes that were included in the assignment of consensus KCRV all agreed within their standard uncertainties.To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Key comparison study on peptide purity - glycated hexapeptide of HbA1c

Main textUnder the auspices of the Protein Analysis Working Group (PAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) a key comparison, CCQM-K115.c, was coordinated by the Bureau International des Poids et Mesures (BIPM), the Health Sciences Authority (HSA) of Singapore and the Chinese National Institute of Metrology (NIM). Nine Metrology Institutes or Designated Institutes and the BIPM participated. Participants were required to assign the mass fraction of the glycated hexapeptide of HbA1c (GE) present as the main component in the comparison sample for CCQM-K115.c. The comparison samples were prepared by HSA/BIPM from synthetic GE purchased from a commercial supplier and used as provided without further treatment or purification.GE was selected to be representative of the performance of a laboratory's measurement capability for the purity assignment of chemically synthesized peptides of known sequence, without cross-links, up to 5 kDa and modification (mono-glycation). It was anticipated to provide an analytical measurement challenge representative for the value-assignment of compounds of broadly similar structural characteristics.The majority of participants used amino acid analysis (PICAA) or quantitative nuclear magnetic resonance (PICqNMR) spectroscopy with a correction for structurally-related peptide impurities approach as the amount of material that has been provided to each participant (25 mg) is insufficient to perform a full mass balance based characterization of the material by a participating laboratory. The coordinators, the BIPM, the HSA and the NIM, were the laboratories to use the mass balance approach as they had more material available.It was decided to propose KCRVs for both the GE mass fraction and the mass fraction of the peptide-related impurities as indispensable contributor regardless of the use of PICAA, PICqNMR or mass balance to determine the GE purity. This allowed participants to demonstrate the efficacy of their implementation of the approaches used to determine the GE mass fraction. In particular, it allows participants to demonstrate the efficacy of their implementation of peptide-related impurity identification and quantification.More detailed studies on the identification/quantification of peptide-related impurities revealed that the integrity of the impurity profile of the related peptide impurities obtained by the participant is crucial for the impact on accuracy of the GE mass fraction assignment.The assessment of the mass fraction of peptide impurities is based on the assumption that that all results are directly taken for the calculation of the KCRVPepImp by use of random-effects meta-analysis (DerSimonian-Laird (DSL) variance-weighted mean). The KCRVPepImp of 45.4 mg/g is associated with a corresponding expanded uncertainty of 9.5 mg/g (k =2.26) providing a more realistic basis of evaluation for the capabilities of the participants to identify/quantify peptide-related impurities. Inspection of the degree of equivalence plots for the mass fraction of peptide impurities and additional information obtained from the peptide-related impurity profile indicates that in most cases the major related peptide impurities have been identified and quantified.The approach selected to obtain a KCRVGE for the mass fraction of GE is based on the DerSimonian-Laird (DSL) variance-weighted mean. The DSL mean takes into account the uncertainties of the results while introducing sufficient excess variance to allow for their observed dispersion resulting in a larger expanded uncertainty U(KCRVGE). The KCRVGE for CCQM-K115.c is 628 mg/g with a corresponding expanded uncertainty of the KCRVGE of 27 mg/g (k =2.26). All GE mass fraction results are in agreement with the KCRVGE.To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Pilot study on peptide purity - glycated hexapeptide of HbA1c

Main textUnder the auspices of the Protein Analysis Working Group (PAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) a pilot study, CCQM-P55.2.c, was coordinated by the Bureau International des Poids et Mesures (BIPM), the Health Sciences Authority (HSA) of Singapore and the Chinese National Institute of Metrology (NIM). Three Metrology Institutes or Designated Institutes and the BIPM participated. Participants were required to assign the mass fraction of glycated hexapeptide of HbA1c (GE) present as the main component in the comparison sample for CCQM-P55.2.c. The comparison samples were prepared by HSA/BIPM from synthetic GE purchased from a commercial supplier and used as provided without further treatment or purification.GE was selected to be representative of the performance of a laboratory's measurement capability for the purity assignment of chemically synthesized peptides of known sequence, without cross-links, up to 5 kDa and modification (mono-glycation). It was anticipated to provide an analytical measurement challenge representative for the value-assignment of compounds of broadly similar structural characteristics.The majority of participants used amino acid analysis (PICAA) or quantitative nuclear magnetic resonance (PICqNMR) spectroscopy with a correction for structurally-related peptide impurities. It was decided to assign reference values (RVs) based on the KCRVs of CCQM-K115.c for both the GE mass fraction and the mass fraction of the peptide related impurities as indispensable contributor regardless of the use of PICAA, mass balance or any other approach to determine the GE purity. This allowed participants to demonstrate the efficacy of their implementation of the approaches used to determine the GE mass fraction. In particular, it allows participants to demonstrate the efficacy of their implementation of peptide related impurity identification and quantification.More detailed studies on the identification/quantification of peptide related impurities revealed that the integrity of the impurity profile of the related peptide impurities obtained by the participant is crucial for the impact on accuracy of the GE mass fraction assignment.The assessment of the mass fraction of peptide impurities is based on the assumption that all results are directly taken for the calculation of the RVPepImp by use of random-effects meta-analysis (DerSimonian-Laird (DSL) variance-weighted mean). The RVPepImp of 45.4 mg/g is associated with a corresponding expanded uncertainty of 9.5 mg/g (k =2.26) providing a more realistic basis of evaluation for the capabilities of the participants to identify/quantify peptide related impurities. Inspection of the degree of equivalence plots for the mass fraction of peptide impurities and additional information obtained from the peptide related impurity profile indicates that in all cases the major related peptide impurities have been identified and quantified.The approach selected to obtain a RVGE for the mass fraction of GE is based on the DerSimonian-Laird (DSL) variance-weighted mean. The DSL mean takes into account the uncertainties of the results while introducing sufficient excess variance to allow for their observed dispersion resulting in a larger expanded uncertainty U(RVGE). The RVGE based on CCQM-K115.c is 628 mg/g with a corresponding expanded uncertainty of the RVGE of 27 mg/g (k =2.26). All GE mass fraction results are in agreement with the RVGE.To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Key Comparison Study - Organic Solvent Calibration Solution - Gravimetric preparation and value assignment of aflatoxin B1 (AfB1) in acetonitrile (ACN)

Main textThe CCQM-K154.b comparison was coordinated by the Bureau International des Poids et Mesures (BIPM) and the Chinese National Institute of Metrology (NIM) on behalf of the Organic Analysis Working Group (OAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) for National Measurement Institutes (NMIs) and Designated Institutes (DIs) which provide measurement services in organic analysis under the 'Comité International des Poids et Mesures' Mutual Recognition Arrangement (CIPM MRA) and/or have participated in the BIPM's Mycotoxin Metrology Capacity Building and Knowledge Transfer (MMCBKT) project as part of its "Metrology for Safe Food and Feed in Developing Economies" Capacity Building Programme.Gravimetrically-prepared solutions having an assigned mass fraction of specified organic analytes are routinely used to calibrate measurement processes for the quantification of the same analytes in matrix samples. Appropriate assignments of the property value and associated uncertainty of calibration solutions thus underpin the traceability of routine analysis and are critical for accurate measurements. Evidence of successful participation in relevant international comparisons is needed to document calibration and measurement capability claims (CMCs) made by national metrology institutes and designated institutes. In total, eleven NMIs/DIs participated in the Track C, Model II, Key Comparison CCQM-K154.b [Gravimetric preparation and value assignment of aflatoxin B1 (AfB1) in acetonitrile (ACN)] for emerging areas of global interest and innovation. Participants were requested to gravimetrically prepare calibration solutions and value assign the mass fractions, expressed in mg/kg, of aflatoxin B1 (AfB1) in the acetonitrile (ACN) solution. Study samples, with assigned values and associated uncertainties were prepared by the comparison participants and sent to the coordinating laboratory for comparison. The Key Comparison Reference Values (KCRVs), calculated form values measured by the coordinating laboratory based on calibrations obtained from independent gravimetrically prepared calibrant solutions, agreed with participants reported values, within their stated uncertainties.AfB1 was selected to be representative of polar aflatoxins. Aflatoxins are a class of mycotoxins generally produced by fungi of the genus Aspergillus. It was anticipated to provide a challenge representative for the gravimetrical preparation and value assignment of calibration solutions in the mass fraction range of 2 mg/kg to 50 mg/kg of mycotoxins with broadly similar structural characteristics.Nine participants of the MMCBKT programme were provided with a stock solution having a known AfB1 mass fraction and expanded uncertainty to use to gravimetrically prepare and value assign a calibration solution. Three NMIs/DIs also participated using their own calibration solutions. The use of in-house solutions required an additional capacity to undertake a fit-for-purpose purity assessment. NIM was the only NMI participating using both the MMCBKT based and their own in-house assigned solutions in order to connect the two different groups.It was decided to propose separate KCRVs for each of the two ampoules provided by the participating NMIs/DIs based on the AfB1 mass fraction. This allowed participants to demonstrate the efficacy of their implementation of the approaches used to gravimetrically prepare calibration solutions and to assess the AfB1 mass fraction.The majority of the AfB1 mass fraction KCRVs (wKCRV) for CCQM-K154.b spanned a mass fraction range of 2.02 mg/kg to 31.57 mg/kg. The relative expanded uncertainties U(wKCRV) ranged from 0.69 % to 2.93 %.Inspection of the degree of equivalence plots for the AfB1 mass fraction assignments in CCQM-K154.b indicated that there was an excellent agreement of results. Solely, the AfB1 mass fraction assignments of INRAP did not agree with the KCRVs. It was found that the samples were altered as a result of an acid contamination.To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Pilot study on peptide purity - hexapeptide of HbA0

Main textUnder the auspices of the Protein Analysis Working Group (PAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) a pilot study, CCQM-P55.2.2018, was coordinated by the Bureau International des Poids et Mesures (BIPM), the Health Sciences Authority (HSA) of Singapore and the Chinese National Institute of Metrology (NIM). Three Metrology Institutes or Designated Institutes and the BIPM participated. Participants were required to assign the mass fraction of hexapeptide of HbA0 (VE) present as the main component in the comparison sample for CCQM-P55.2.2018. The comparison samples were prepared by HSA/BIPM from synthetic VE purchased from a commercial supplier and used as provided without further treatment or purification.VE was selected to be representative of the performance of a laboratory's measurement capability for the purity assignment of chemically synthesized peptides of known sequence, without cross-links, up to 5 kDa and without modification. It was anticipated to provide an analytical measurement challenge representative for the value-assignment of compounds of broadly similar structural characteristics.The majority of participants used amino acid analysis (PICAA) or quantitative nuclear magnetic resonance (PICqNMR) spectroscopy with a correction for structurally related peptide impurities. It was decided to assign reference values (RVs) based on the KCRVs of CCQM-K115.2018 for both the VE mass fraction and the mass fraction of the peptide-related impurities as indispensable contributor regardless of the use of PICAA, mass balance or any other approach to determine the VE purity. This allowed participants to demonstrate the efficacy of their implementation of the approaches used to determine the VE mass fraction. In particular, it allows participants to demonstrate the efficacy of their implementation of peptide-related impurity identification and quantification.More detailed studies on the identification/quantification of peptide related impurities revealed that the integrity of the impurity profile of the related peptide impurities obtained by the participant is crucial for the impact on accuracy of the VE mass fraction assignment.The assessment of the mass fraction of peptide impurities is based on the assumption that only the most consistent set of results is taken for the calculation of the RVPepImp. The sum of the combined cis/trans VE depsipeptide impurities (only identified/quantified by NRC and confirmed by BIPM and HSA) and mass fractions of peptide-related impurities that have been identified by at least two participants have been used to establish the RVPepImp. The RVPepImp of 53.0 mg/g is associated with a corresponding expanded uncertainty of 17.3 mg/g (k = 2) providing a more realistic basis of evaluation for the capabilities of the participants to identify/quantify peptide related impurities. Inspection of the degree of equivalence plots for the mass fraction of peptide impurities and additional information obtained from the peptide-related impurity profile indicates that in all cases the major related peptide impurity, VE depsipeptide, has not been identified. The VE depsipeptide impurity was initially and uniquely identified and quantified by the NRC by the use of 1H-NMR in the parallel CCQM-K115.2018. The related peptide impurity mass fraction results of only four participants (NRC, LGC, LNE and HSA) are in agreement with the RVPepImp. The NRC has identified and quantified the VE depsipeptide whereas the LGC, LNE and HSA have not identified the VE depsipeptide but accounted for that contribution.The approach selected to obtain a RVVE for the mass fraction of VE is based on a mass balance calculation that takes into account the most consistent set of results for the peptide-related impurities RVPepImp, TFA mass fraction and the water mass fraction. The RVVE based on CCQM-K115.2018 is 613 mg/g with a corresponding expanded uncertainty of the RVVE of 20 mg/g (k = 2). Inspection of the degree of equivalence plots for the mass fraction of VE shows that all results obtained by PICAA agree with the RVVE while the result obtained by the BIPM with PICqNMR disagrees with the RVVE.To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Supplementary comparison study - measurement capabilities for the quantification of ethanol in water

Main textThe accurate quantification of ethanol in water is essential for forensic applications such as blood and breath alcohol testing and for commercial applications such as the assessment of alcoholic beverages.The intercomparison EURAMET.QM-S14 is part of a capacity building project named ALCOREF "Certified forensic alcohol reference materials" [1] that is running within the European Metrology Programme for Innovation and Research (EMPIR) [2]. The intercomparison should allow project partners and other interested National Metrology Institutes (NMIs) and Designated Institutes (DIs) to benchmark their analytical methods for the quantification of ethanol in water. The study plan was agreed by the European Association of National Metrology Institutes (EURAMET) Subcommittee Bio- and Organic Analysis (SCBOA) and the Organic Analysis Working Group (OAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) in February and April 2019, respectively. The intercomparison was coordinated by BAM. Two concentration levels relevant for the calibration and verification of evidential breath alcohol analysers were distributed to study participants. Fifteen institutes from 15 countries registered for the intercomparison and returned results. Participants mostly applied gas chromatography with flame ionisation detection (GC-FID) or mass spectroscopy (GC-MS), one participant used titrimetry and one participant employed a test bench for breath analyser calibration ("bubble train"). Participants did either in-house purity assessment of their commercial ethanol calibrants by Karl-Fischer titration, chromatographic methods, quantitative nuclear magnetic resonance spectroscopy (qNMR) and/or density measurements; or they used ethanol/water Certified Reference Materials (CRMs) from NMIs/DIs for calibration.CCQM OAWG agreed to use a consensus value from participants' results that utilizes the reported uncertainties as Key Comparison Reference Value (KCRV). The Gaussian Random effects model with Hierarchical Bayesian solution (HB-REM) is a reasonable approach in this case. The KCRVs and Degrees of Equivalence (DoEs) were calculated with the NIST consensus builder version 1.2 Hierarchical Bayes procedure [3].Successful participation in the interlaboratory comparison has demonstrated the capabilities in determining the mass fraction of ethanol in aqueous matrices in the range 0.1 mg/g to 8 mg/g. Fourteen out of 15 participants have successfully quantified both samples, one participant successfully quantified only the lower-level (0.6 mg/g) sample.To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Supplementary comparison study - fipronil-sulfone in chicken egg powder (APMP.QM-S16)

Main textThe APMP.QM-S16 comparison was coordinated by the National Institute of Metrology, China (NIM) on behalf of the Technical Committee on Amount of Substance (TCQM) of the Asia Pacific Metrology Programme (APMP) and the Organic Analysis Working Group (OAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) for National Measurement Institutes (NMIs) and Designated Institutes (DIs) which provide measurement services in organic analysis under the 'Comité International des Poids et Mesures' Mutual Recognition Arrangement (CIPM MRA).Fipronil is a broad-spectrum insecticide attributed to the phenylpyrazole family. Due to its high toxicity to invertebrates and long persistence, it has been one of the most widely used insecticides to control household and agricultural pests since its appearance in the market. Fipronil has also shown genotoxicity and cytotoxicity to many vertebrate animals, like mammals, birds, fish, reptiles and so on. Moreover, fipronil could have negative effects on the liver, kidneys and thyroid glands of humans. Its oxidation metabolite is fipronil-sulfone that could induce even higher toxicity than fipronil. In 2017, a fipronil-in-egg scandal occurred in the European Union (EU) that involved more than 15 states. The detected concentration of the sum of fipronil and fipronil-sulfone was many times higher than the EU maximum residue limit in bird egg (5 μg/kg). Besides European countries, fipronil-contaminated eggs were also found in Asian areas like China, Hong Kong and South Korea. Systematic monitoring and control of pesticide residues are necessary to safeguard public health.Fipronil-sulfone was selected to be representative of pesticide residues in this comparison. This study provides the means for assessing measurement capabilities for determination of low-polarity measurands in a procedure that requires extraction, clean-up, analytical separation, and selective detection in a food matrix. Generally, it provides a demonstration of a laboratory's capabilities in determining the medium molecular weight analytes (mass range 300-500) with low polarity (pKow < -2) at mass fraction levels of 4 to 400 μg/kg in a high fat, high protein and low carbohydrate food matrix that falls within Sector 4 of the AOAC Food-matrix Triangle.Seven NMIs/DIs participated in the APMP.QM-S16 comparison. Different methods such as liquid-liquid extraction, QuEChERS and SPE were applied in the sample pre-treatment and HPLC-MS/MS, HPLC-HRMS, and GC-MS/MS were applied for detection by the participants. Most participating laboratories applied isotope dilution mass spectrometry (IDMS) techniques with fipronil-sulfone-13C4,15N2 as internal standard, except EC-JRC-Geel which used matrix matched external calibration method. In the aspect of the traceability for the measurement results, most participants either used their own CRMs, other NMIs' CRMs or the pure materials assessed in-house to guarantee traceability to SI unit. One lab was excluded from the SCRV evaluation, as they did not meet the CIPM metrological traceability requirements. As a result, six sets of valid results were used for the estimation of the SCRV. Finally Hierarchical Bayes was considered to be the appropriate estimator to calculate SCRV, which was determined as 121.82 μg/kg with a standard uncertainty of 1.77 μg/kg. Inspection of the degree of equivalence plots for the fipronil-sulfone mass fraction assignments in APMP.QM-S16 indicated that there was excellent agreement of results.To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

PAWG Pilot Study on Quantification of SARS-CoV-2 Monoclonal Antibody - Part 1

Main textUnder the auspices of the Protein Analysis Working Group (PAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) a pilot study, CCQM-P216, was coordinated by the Chinese National Institute of Metrology (NIM), National Research Council of Canada (NRC) and the Bureau International des Poids et Mesures (BIPM). Eleven Metrology Institutes or Designated Institutes and the BIPM participated in the first phase of the pilot study (Part 1). The purpose of this pilot study was to develop measurement capabilities for larger proteins using a recombinant humanized IgG monoclonal antibody against Spike glycoprotein of SARS-CoV-2 (Anti-S IgG mAb) in solution. The first phase of the study was designed to employ established methods that had been previously studies by the CCQM Protein Analysis Working Group, involving the digestion of protein down to the peptide or amino acid level.The global coronavirus pandemic has also led to increased focus on antibody quantitation methods. IgG are among the immunoglobulins produced by the immune system to provide protection against SARS-CoV-2. Anti-SARS-CoV-2 IgG can therefore be detected in samples from affected patients. Antibody tests can show whether a person has been exposed to the SARS-CoV-2, and whether or not they potentially show lasting immunity to the disease. With the constant spread of the virus and the high pressure of re-opening economies, antibody testing plays a critical role in the fight against COVID-19 by helping healthcare professionals to identify individuals who have developed an immune response, either via vaccination or exposure to the virus. Many countries have launched large-scale antibody testing for COVID-19. The development of measurement standards for the antibody detection of SARS-CoV-2 is critically important to deal with the challenges of the COVID-19 pandemic. In this study, the SARS-CoV-2 monoclonal antibody is being used as a model system to build capacity in methods that can be used in antibody quantification. Amino acid reference values with corresponding expanded uncertainty of 36.10 ± 1.55 mg/kg, 38.75 ± 1.45 mg/kg, 18.46 ± 0.78 mg/kg, 16.20 ± 0.67 mg/kg and 30.61 ± 1.30 mg/kg have been established for leucine, valine, phenylalanine, isoleucine and proline, respectively. Agreement between nearly all laboratories was achieved for the amino acid analysis within 2 to 2.5 %, with one participant achieving markedly higher results due to a technical issue found in their procedure; this result was thus excluded from the reference value calculations. The relatively good agreement within a laboratory between different amino acids was not dissimilar to previous results for peptides or small proteins, indicating that factors such as hydrolysis conditions and calibration procedures could be the largest sources of variability.Peptide reference values with corresponding expanded uncertainty of 4.99 ± 0.28 mg/kg and 6.83 ± 0.65 mg/kg have been established for ALPAPIEK and GPSVFPLAPSSK, respectively. Not surprisingly due to prior knowledge from previous studies on peptide quantitation, agreement between laboratories for the peptide-based analysis was slightly poorer at 3 to 5 %, with one laboratory's result excluded for the peptide GPSVFPLAPSSK. Again, this level of agreement was not significantly poorer than that achieved in previous studies with smaller or less complex proteins.To reach the main text of this paper, click on Final Report.

Key comparison study—organic solvent calibration solution—gravimetric preparation and value assignment of trans-zearalenone (trans-ZEN) in acetonitrile (ACN)

The CCQM-K154.a and subsequent CCQM-K154.a.1 comparisons were coordinated by the the Bureau International des Poids et Mesures (BIPM) and the Chinese National Institute of Metrology (NIM) on behalf of the Organic Analysis Working Group (OAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) for National Measurement Institutes (NMIs) and Designated Institutes (DIs) which provide measurement services in organic analysis under the 'Comité International des Poids et Mesures' Mutual Recognition Arrangement (CIPM MRA) and/or have participated in the BIPM's Mycotoxin Metrology Capacity Building and Knowledge Transfer (MMCBKT) project as part of its "Metrology for Safe Food and Feed in Developing Economies" Capacity Building Programme.Gravimetrically-prepared solutions having an assigned mass fraction of specified organic analytes are routinely used to calibrate measurement processes for the quantification of the same analytes in matrix samples. Appropriate assignments of the property value and associated uncertainty of calibration solutions thus underpin the traceability of routine analysis and are critical for accurate measurements. Evidence of successful participation in relevant international comparisons is needed to document calibration and measurement capability claims (CMCs) made by national metrology institutes and designated institutes. In total, eleven NMIs/DIs participated in the Track C, Model II, Key Comparison CCQM-K154.a and the Subsequent Comparison CCQM-K154.a.1 [Gravimetric preparation and value assignment of trans-zearalenone (trans-ZEN) in acetonitrile (ACN)] for emerging areas of global interest and innovation. Participants were requested to gravimetrically prepare calibration solutions and value assign the mass fractions, expressed in mg/kg, of trans-zearalenone (trans-ZEN) in the acetonitrile (ACN) solution. Study samples, with assigned values and associated uncertainties were prepared by the comparison participants and sent to the coordinating laboratory for comparison. The Key Comparison Reference Values (KCRVs), calculated form values measured by the coordinating laboratory based on calibrations obtained from independent gravimetrically prepared calibrant solutions, agreed with participants reported values, within their stated uncertainties.trans-ZEN was selected to be representative of non-polar Fusarium mycotoxins. It was anticipated to provide a challenge representative for the gravimetrical preparation and value assignment of calibration solutions in the mass fraction range of 10 mg/kg to 100 mg/kg of mycotoxins with broadly similar structural characteristics.Six participants of the MMCBKT programme were provided with a stock solution having a known trans-ZEN mass fraction and expanded uncertainty to use to gravimetrically prepare and value assign a calibration solution. Four NMIs/DIs also participated using their own calibration solutions. The use of in-house solutions required an additional capacity to undertake a fit-for-purpose purity assessment. NIM was the only NMI participating using both the MMCBKT based and their own in-house assigned solutions in order to connect the two different groups.It was decided to propose separate KCRVs for each of the two ampoules provided by the participating NMIs/DIs based on the trans-ZEN mass fraction. This allowed participants to demonstrate the efficacy of their implementation of the approaches used to gravimetrically prepare calibration solutions and to assess the trans-ZEN mass fraction.The majority of the trans-ZEN mass fraction KCRVs (wKCRV) for CCQM-K154.a and CCQM-K154.a.1 spanned a mass fraction range of 12.18 mg/kg to 15.29 mg/kg. Solely the solutions of NRC exhibited higher trans-ZEN mass fraction KCRVs of 66.68 mg/kg and 66.90 mg/kg. The relative expanded uncertainties U(wKCRV) ranged from 0.90 % to 4.54 %.Inspection of the degree of equivalence plots for the trans-ZEN mass fraction assignments in CCQM-K154.a indicated that there was an excellent agreement of results in general.The CCQM-K154.a.1 subsequent comparison was organized for INTI to repeat measurements with new ampoules. The trans-ZEN mass fraction assignments of INTI within CCQM K154.a.1 were in agreement with the KCRVs.Main textTo reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database kcdb.bipm.org/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Key comparison study on peptide purity - synthetic oxytocin

Under the auspices of the Protein Analysis Working Group (PAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) a key comparison, CCQM-K115.b, was coordinated by the Bureau International des Poids et Mesures (BIPM) and the Chinese National Institute of Metrology (NIM). Seven Metrology Institutes or Designated Institutes and the BIPM participated. Participants were required to assign the mass fraction of oxytocin (OXT) present as the main component in the comparison sample for CCQM-K115.b. The comparison samples were prepared from synthetic OXT purchased from a commercial supplier and used as provided without further treatment or purification.OXT was selected to be representative of the performance of a laboratory's measurement capability for the purity assignment of chemically synthesized peptides of known sequence, with one cross-link and up to 5 kDa. It was anticipated to provide an analytical measurement challenge representative for the value-assignment of compounds of broadly similar structural characteristics.The majority of participants used amino acid analysis (PICAA) or quantitative nuclear magnetic resonance (PICqNMR) spectroscopy with a correction for structurally-related peptide impurities approach as the amount of material that has been provided to each participant (25 mg) is insufficient to perform a full mass balance based characterization of the material by a participating laboratory. The coordinators, both the BIPM and the NIM, were the laboratories to use the mass balance approach as they had more material available.It was decided to propose KCRVs for both the OXT mass fraction and the mass fraction of the peptide related impurities as indispensable contributor regardless of the use of PICAA, PICqNMR or mass balance to determine the OXT purity. This allowed participants to demonstrate the efficacy of their implementation of the approaches used to determine the OXT mass fraction. In particular, it allows participants to demonstrate the efficacy of their implementation of peptide related impurity identification and quantification.More detailed studies on the identification/quantification of peptide related impurities and the hydrolysis efficiency revealed that the integrity of the impurity profile of the related peptide impurities obtained by the participant is crucial for the impact on accuracy of the OXT mass fraction assignment.The assessment of the mass fraction of peptide impurities is based on the assumption that only the Largest Consistent Subset (LCS) of results is taken for the calculation of the KCRVPepImp by use of the weighted mean. The KCRVPepImp of 31.6 mg/g is associated with a small corresponding expanded uncertainty of ±1.4 mg/g (k =2) providing a more realistic basis of evaluation for the capabilities of the participants to identify/quantify peptide related impurities. Inspection of the degree of equivalence plots for the mass fraction of peptide impurities and additional information obtained from the peptide related impurity profile indicates that in many cases the major related peptide impurities have been identified and quantified.The approach selected to obtain a KCRVOXT for the mass fraction of OXT is based on random-effects meta-analysis (DerSimonian-Laird (DSL) variance-weighted mean). The DSLmean takes into account the uncertainties of the results while introducing sufficient excess variance to allow for their observed dispersion resulting in a larger expanded uncertainty U(KCRVOXT). The KCRVOXT for CCQM-K115.b is 787.2 mg/g with a corresponding expanded uncertainty of the KCRVOXT of ±12.9 mg/g. All OXT mass fraction results except the result of NMIM are in agreement with the KCRVOXT. It should be pointed out that the mass balance approaches show smaller uncertainties than PICAA or PICqNMR approaches. Mass balance approaches seem to produce slightly higher OXT mass fractions while PICAA approaches deliver slightly lower OXT mass fractions.Main textTo reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database kcdb.bipm.org/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Pilot study on peptide purity—synthetic oxytocin

Under the auspices of the Protein Analysis Working Group (PAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) a pilot study, CCQM-P55.2.b, was coordinated by the Bureau International des Poids et Mesures (BIPM) and the Chinese National Institute of Metrology (NIM). Three Metrology Institutes or Designated Institutes and the BIPM participated. Participants were required to assign the mass fraction of oxytocin (OXT) present as the main component in the comparison sample for CCQM-P55.2.b. The comparison samples were prepared from synthetic OXT purchased from a commercial supplier and used as provided without further treatment or purification.OXT was selected to be representative of the performance of a laboratory's measurement capability for the purity assignment of chemically synthesized peptides of known sequence, with one cross-link and up to 5 kDa. It was anticipated to provide an analytical measurement challenge representative for the value-assignment of compounds of broadly similar structural characteristics.The majority of participants used quantitative nuclear magnetic resonance (PICqNMR) spectroscopy with a correction for structurally-related peptide impurities. Other participants provided results obtained by peptide impurity corrected amino acid analysis (PICAA) or Inductively coupled plasma mass spectrometry (PICICP). It was decided to assign reference values based on the KCRVs of CCQM-K115.b for both the OXT mass fraction and the mass fraction of the peptide related impurities as indispensable contributor regardless of the use of PICAA, mass balance or any other approach to determine the OXT purity. This allowed participants to demonstrate the efficacy of their implementation of the approaches used to determine the OXT mass fraction. In particular, it allows participants to demonstrate the efficacy of their implementation of peptide related impurity identification and quantification.More detailed studies on the identification/quantification of peptide related impurities and the hydrolysis efficiency revealed that the integrity of the impurity profile of the related peptide impurities obtained by the participant is crucial for the impact on accuracy of the OXT mass fraction assignment.The assessment of the mass fraction of peptide impurities is based on the assumption that only the Largest Consistent Subset (LCS) of results is taken for the calculation of the KCRVPepImp by use of the weighted mean. The KCRVPepImp of 31.6 mg/g is associated with a small corresponding expanded uncertainty of ±1.4 mg/g (k = 2) providing a more realistic basis of evaluation for the capabilities of the participants to identify/quantify peptide related impurities. Inspection of the degree of equivalence plots for the mass fraction of peptide impurities and additional information obtained from the peptide related impurity profile indicates that in many cases the major related peptide impurities have been identified and quantified.The approach selected to obtain a KCRVOXT for the mass fraction of OXT is based on random-effects meta-analysis (DerSimonian-Laird (DSL) variance-weighted mean). The DSLmean takes into account the uncertainties of the results while introducing sufficient excess variance to allow for their observed dispersion resulting in a larger expanded uncertainty U(KCRVOXT). The KCRVOXT for CCQM-K115.b is 787.2 mg/g with a corresponding expanded uncertainty of the KCRVOXT of ±12.9 mg/g. It should be pointed out that the mass balance approaches show smaller uncertainties than PICAA, PICqNMR or PICICP approaches. Inspection of the degree of equivalence plots for CCQM-P55.2.b for the mass fraction of OXT shows that all results, except the UME qNMR result, agree with the reference value.Main textTo reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database kcdb.bipm.org/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Final report on APMP.QM-S10: elements in food supplement

APMP.QM-S10 was coordinated by Government Laboratory, Hong Kong SAR (GLHK) under the auspices of the Inorganic Analysis Working Group (IAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM). Fourteen national metrology institutes (NMIs) or designated institutes (DIs) registered for the examination of zinc, manganese, calcium and magnesium. All institutes submitted the results for zinc and manganese, and twelve institutes submitted the results for calcium and magnesium. The supplementary comparison was designed to enable participating institutes to demonstrate their measurement capabilities in measuring the mass fractions of the analytes at mg/kg levels in a test sample of food supplement by various analytical techniques.For examination of zinc, manganese, calcium and magnesium, most of the participants used microwave-assisted acid digestion methods for sample dissolution. A variety of instrumental techniques including inductively coupled plasmas mass spectrometry (ICP-MS), isotope dilution inductively coupled plasmas mass spectrometry (ID-ICP-MS), inductively coupled plasmas optical emission spectrometry (ICP-OES), flame atomic absorption spectrometry (FAAS), and instrumental neutron action analysis (INAA) were employed by the participants for determination. For this supplementary comparison, inorganic core capabilities have been demonstrated by the concerned participants with respect to methods including ICP-MS (without isotope dilution), ID-ICP-MS, ICP-OES, FAAS and INAA on the determination of elements (zinc, manganese, calcium and magnesium) in a food matrix of food supplement.Generally, the participants' results of APMP.QM-S10 were found to be consistent for all measurands according to their equivalence statements. Except with some extreme values, most of the participants obtained the values of d i/U(d i) within ± 1 for the measurands.Main textTo reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database kcdb.bipm.org/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

CCQM-K104 key comparison (avermectin B1a) on the characterization of organic substances for chemical purity

Under the Comité Consultatif pour la Quantité de Matière (CCQM), a key comparison, CCQM-K104, was coordinated by the National Institute of Metrology (NIM). The comparison was designed to demonstrate a laboratory's performance in determining the mass fraction of the main component in a complex high purity organic material. Nine NMIs or DIs participated in the comparison. Eight participants reported their results. An additional impurity was resolved from the avermectin B1a peak and was tentatively identified as an unknown impurity by NMIA (National Measurement Institute (Australia)). It was subsequently identified by NIM as a diastereoisomer of avermectin B1a at the C-26 position. Final reference value (KCRV) = 924.63 mg/g, with uncertainty (k=1) = 3.89 mg/g, and expanded uncertainty = 8.97 mg/g. The degrees of equivalence with the avermectin B1a KCRV for each participant were reported. The measurement results and degrees of equivalence should be indicative of the performance of a laboratory's measurement capability for the purity assignment of organic compounds of high structural complexity (relative molecular mass range of 500 Da -1000 Da and low polarity (-log KOW ⩽ -2).Main textTo reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database kcdb.bipm.org/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Key comparison study on peptide purity—synthetic human C-peptide

Under the auspices of the Protein Analysis Working Group (PAWG) of the Comité Consultatif pour la Quantité de Matière (CCQM) a key comparison, CCQM-K115, was coordinated by the Bureau International des Poids et Mesures (BIPM) and the Chinese National Institute of Metrology (NIM). Eight Metrology Institutes or Designated Institutes and the BIPM participated. Participants were required to assign the mass fraction of human C-peptide (hCP) present as the main component in the comparison sample for CCQM-K115. The comparison samples were prepared from synthetic human hCP purchased from a commercial supplier and used as provided without further treatment or purification.hCP was selected to be representative of the performance of a laboratory's measurement capability for the purity assignment of short (up to 5 kDa), non-cross-linked synthetic peptides/proteins. It was anticipated to provide an analytical measurement challenge representative for the value-assignment of compounds of broadly similar structural characteristics.The majority of participants used a peptide impurity corrected amino acid analysis (PICAA) approach as the amount of material that has been provided to each participant (25 mg) is insufficient to perform a full mass balance based characterization of the material by a participating laboratory. The coordinators, both the BIPM and the NIM, were the laboratories to use the mass balance approach as they had more material available.It was decided to propose KCRVs for both the hCP mass fraction and the mass fraction of the peptide related impurities as indispensable contributor regardless of the use of PICAA, mass balance or any other approach to determine the hCP purity. This allowed participants to demonstrate the efficacy of their implementation of the approaches used to determine the hCP mass fraction. In particular it allows participants to demonstrate the efficacy of their implementation of peptide related impurity identification and quantification. More detailed studies on the identification/quantification of peptide related impurities and the hydrolysis efficiency revealed that the integrity of the impurity profile of the related peptide impurities obtained by the participant is crucial for the impact on accuracy of the hCP mass fraction assignment.The assessment of the mass fraction of peptide impurities is based on the assumption that only the most exhaustive and elaborate set of results is taken for the calculation of the KCRVPepImp. The KCRVPepImp for the peptide related impurity mass fractions of the material was 83.3 mg/g with a combined standard uncertainty of 1.5 mg/g. Inspection of the degree of equivalence plots for the mass fraction of peptide impurities and additional information obtained from the peptide related impurity profile indicates that in many cases only a very small number of impurities have been identified and quantified resulting in an underestimation of the peptide related impurity mass fractions.The approach to obtain a KCRVhCP for the mass fraction of hCP is based on a mass balance calculation that takes into account the most exhaustive and elaborate set of results for the peptide related impurities KCRVPepImp, the TFA mass fraction value, water and other minor counter ions obtained by the coordinating laboratories. Differences in the quality of the results obtained for both peptides related impurity mass fractions and hCP mass fractions are better weighted and reflected in smaller uncertainties. The KCRVhCP for CCQM-K115 is 801.8 mg/g with a corresponding combined standard uncertainty of 3.1 mg/g.In general, mass balance approaches show smaller uncertainties than PICAA approaches and the majority of results obtained by the PICAA approach are in agreement because of larger corresponding uncertainties.Main textTo reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database kcdb.bipm.org/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).