Regulatory landscape of dietary supplements and herbal medicines from a global perspective.

The number of Individuals that use dietary supplements and herbal medicine products are continuous to increase in many countries. The context of usage of a dietary supplement varies widely from country-to-country; in some countries supplement use is just limited to general health and well-being while others permit use for medicinal purposes. To date, there is little consensus from country to country on the scope, requirements, definition, or even the terminology in which dietary supplement and herbal medicines categories could be classified. Transparent science-based quality standards for the ingredients across these regulatory frameworks/definitions becomes even more important given the international supply chain. Meanwhile, there has been a rapid advancement in emerging technologies and data science applied to the field. This review was conceived at the Global Summit on Regulatory Sciences that took place in Beijing on September 2018 (GSRS2018) which is organized by Global Coalition for Regulatory Science Research (GCRSR) that consists of the global regulatory agencies from over ten countries including the European Union. This review summarizes a significant portion of discussions relating to a longitudinal comparison of the status for dietary supplements and herbal medicines among the different national jurisdictions and to the extent of how new tools and methodologies can improve the regulatory application.

Two-dimensional heart-cut LC-LC improves accuracy of exact-matching double isotope dilution mass spectrometry measurements of aflatoxin B1 in cereal-based baby food, maize, and maize-based feed.

Aflatoxins, mycotoxins of fungi of the Aspergillus sp., pose a risk to consumer health and are, therefore, regulated by more than 100 countries. To facilitate method development and validation as well as assessment of measurement capabilities, availability of certified reference materials and proficiency testing schemes is important. For these purposes, highly accurate determinations of the aflatoxin content in the materials used are necessary. We describe here the use of two-dimensional heart-cut LC-LC in combination with exact-matching double isotope dilution mass spectrometry to determine the content of aflatoxin B1 in three materials used in a proficiency testing scheme. The serious reduction in ionization suppression afforded by the two-dimensional heart-cut LC-LC had a positive effect on the precision of the measured isotope ratios of the exact-matching double isotope dilution mass spectrometry. This is evidenced by the expanded measurement uncertainty (k=2) of 0.017 µg/kg or 8.9 % relative to a mass fraction of aflatoxin B1 in a cereal-based baby food of 0.197 µg/kg. This value is in perfect agreement with the consensus value of this material from a proficiency test (PT) scheme for National Reference Laboratories executed by the European Reference Laboratory for Mycotoxins. The effort necessary to perform the described methodology precludes its frequent use but for specific applications we see it as a valuable tool.

Purity Assessment of Honey Based on Compound Specific Stable Carbon Isotope Ratios Obtained by LC-IRMS.

BACKGROUND: The use of stable carbon isotope ratios (δ13C) of sugar fractions of honey is a powerful tool to detect adulteration with sugar syrups. This is accomplished by calculating differences of the δ13C values between individual honey saccharides and comparing them to published purity criteria. A liquid chromatography-isotope ratio mass spectrometry (LC-IRMS) method for the determination of δ13C values of sugars in honey was previously validated by an interlaboratory comparison, but no further guidance was given how to include the obtained precision figures of the compound-specific δ13C values in the purity assessment of honey. OBJECTIVE: To use existing data to estimate the standard deviation of the repeatability (sr) and reproducibility (sR) of differences (Δ Î´13C) between the δ13C values of individual honey saccharides. METHODS: Previously published δ13C values were used to calculate differences (Δ Î´13C values) between δ13C fructose-δ13C glucose, δ13C glucose-δ13C disaccharides, etc. in a honey sample; sr and sR of Δ Î´13C values were calculated according to ISO 5725-2:2019. RESULTS: The Δ Î´13C sr and sR values were essentially of the same magnitude as the sr and sR values of δ13C values of the sugar fractions. The precision of the Δ Î´13C values was used to estimate the critical difference for comparing a test result with a reference value according to ISO 5725-6:1994. This varied between 0.26 and 1.10‰. CONCLUSION: The estimated critical differences can be used to determine whether a honey test result complies with published Δ Î´13C purity criteria. HIGHLIGHT: The proposed procedure will increase confidence in decisions based on compound-specific δ13C values regarding the conformity of honey with published purity criteria.

Validation procedures for quantitative gluten ELISA methods: AOAC allergen community guidance and best practices.

The food allergen analytical community is endeavoring to create harmonized guidelines for the validation of food allergen ELISA methodologies to help protect food-sensitive individuals and promote consumer confidence. This document provides additional guidance to existing method validation publications for quantitative food allergen ELISA methods. The gluten-specific criterion provided in this document is divided into sections for information required by the method developer about the assay and information for the implementation of the multilaboratory validation study. Many of these recommendations and guidance are built upon the widely accepted Codex Alimentarius definitions and recommendations for gluten-free foods. The information in this document can be used as the basis of a harmonized validation protocol for any ELISA method for gluten, whether proprietary or nonproprietary, that will be submitted to AOAC andlor regulatory authorities or other bodies for status recognition. Future work is planned for the implementation of this guidance document for the validation of gluten methods and the creation of gluten reference materials.

Detection and Quantification of Botanical Impurities in Commercial Oregano (Origanum vulgare) Using Metabarcoding and Digital PCR.

DNA technology for food authentication is already well established, and with the advent of Next Generation Sequencing (NGS) and, more specifically, metabarcoding, compositional analysis of food at the molecular level has rapidly gained popularity. This has led to several reports in the media about the presence of foreign, non-declared species in several food commodities. As herbs and spices are attractive targets for fraudulent manipulation, a combination of digital PCR and metabarcoding by NGS was employed to check the purity of 285 oregano samples taken from the European market. By using novel primers and analytical approaches, it was possible to detect and quantify both adulterants and contaminants in these samples. The results highlight the high potential of NGS for compositional analysis, although its quantitative information (read count percentages) is unreliable, and other techniques are therefore needed to complement the sequencing information for assessing authenticity ('true to the name') of food ingredients.

Essential terminology and considerations for validation of non-targeted methods.

Through their suggestive name, non-targeted methods (NTMs) do not aim at a predefined "needle in the haystack." Instead, they exploit all the constituents of the haystack. This new type of analytical method is increasingly finding applications in food and feed testing. However, the concepts, terms, and considerations related to this burgeoning field of analytical testing need to be propagated for the benefit of those associated with academic research, commercial development, or official control. This paper addresses frequently asked questions regarding terminology in connection with NTMs. The widespread development and adoption of these methods also necessitate the need to develop innovative approaches for NTM validation, i.e., evaluating the performance characteristics of a method to determine if it is fit-for-purpose. This work aims to provide a roadmap for approaching NTM validation. In doing so, the paper deliberates on the different considerations that influence the approach to validation and provides suggestions therefor.

Evaluation of the quality of postharvest rapeseed by means of an electronic nose.

BACKGROUND: Rapeseed is a valuable source of edible oil. The presence of even a small amount of mouldy or burnt rapeseed in a particular production batch deteriorates the quality of the edible oil obtained from it. Since the traditional method of using a panel of experts is time-consuming, there is a need for fast and easy methods for rapeseed quality evaluation by intelligent devices to replace human labour. RESULTS: For rapeseed quality evaluation, an electronic nose equipped with an array of eight quartz microbalance sensors and four metal oxide semiconductor sensors was used. Signals generated by the sensors were analysed by principal component analysis and discriminant function analysis. Identification of samples that contained small proportions of mouldy or burnt rapeseed was possible despite the differences between the particular varieties studied. CONCLUSION: Electronic nose technology has shown the possibility of detecting samples of faulty rapeseed at very low contamination levels and distinguishing them with high probability from sound rapeseed.

Analytical methods for detection of gluten in food--method developments in support of food labeling legislation.

The current essential therapy of celiac disease is a strict adherence to a gluten-free diet. Besides food products that are naturally gluten-free, "very low gluten" and "gluten-free" bakery products have become available. The availability of immunochemical and other analytical methods to determine gluten markers in foods is of utmost importance to ensure the well being of gluten-sensitive individuals. The aim of this review was to evaluate if currently available methodologies are suitable to meet the requirements of food labeling standards for individual gluten source declaration, in order to achieve policy objectives. Codex Alimentarius and European Union (EU) legislation and gluten detection methodologies applicable at present have been summarized and compared. In 2009, the European Commission issued Regulation No. 41/2009 concerning the composition and labeling of foodstuffs suitable for people intolerant to gluten. This review constitutes a basis to investigate the possibility to develop a proteomic-based method for the specific detection of gluten-containing cereals in food products, especially at or around the limits specified in EU legislation.

Current perspectives and recommendations for the development of mass spectrometry methods for the determination of allergens in foods.

Allergen detection and quantification is an essential part of allergen management as practiced by food manufacturers. Recently, protein MS methods (in particular, multiple reaction monitoring experiments) have begun to be adopted by the allergen detection community to provide an alternative technique to ELISA and PCR methods. MS analysis of proteins in foods provides additional challenges to the analyst, both in terms of experimental design and methodology: (1) choice of analyte, including multiplexing to simultaneously detect several biologically relevant molecules able to trigger allergic reactions; (2) choice of processing stable peptide markers for different target analytes that should be placed in publicly available databases; (3) markers allowing quantification (e.g., through standard addition or isotopically labeled peptide standards); (4) optimization of protease digestion protocols to ensure reproducible and robust method development; and (5) effective validation of methods and harmonization of results through the use of naturally incurred reference materials spanning several types of food matrix.

Results of an Interlaboratory Comparison of a Liquid Chromatography-Isotope Ratio Mass Spectrometry Method for the Determination of 13C/12C Ratios of Saccharides in Honey.

BACKGROUND: Stable carbon isotope analysis of sugars in honey by LC-isotope ratio mass spectrometry (IRMS) is a useful tool for detecting adulteration of honey with extraneous sugar. Purity criteria based on 13C/12C ratios of saccharides in honey, determined by LC-IRMS of a large number of authentic honey samples, have been elaborated. However, no interlaboratory comparison (ILC) has yet been performed to estimate the precision of the method under reproducibility conditions. OBJECTIVE: To address this knowledge gap an ILC involving 14 laboratories and using six honey samples was conducted. METHODS: The participants were allowed to use their LC-IRMS-based method of choice for sample preparation and compound separation. RESULTS: The precision figures were estimated according to ISO 5725:1994. The repeatability relative standard deviation (RSDr) for the determination of δ13C values of fructose and glucose varied between 0.3 and 0.5%, with 0.3 and 1.0% for disaccharides, and 0.7 and 2.8% for trisaccharides. The RSDR varied between 0.8 and 1.8% for the monosaccharides, 1.0 and 1.5% for disaccharides, and 1.4 and 2.8% for trisaccharides. CONCLUSION: Based on the obtained precision data the LC-IRMS method for the determination of 13C/12C ratios of saccharides in honey was considered fit for the conformity assessment of honey with established purity criteria. HIGHLIGHTS: Precision estimates for a LC-IRMS method to determine 13C/12C ratios of saccharides in honey were obtained through an ILC. The data created can form the basis for the standardization of the method by interested standards-developing organizations for use in official control.

DNA Accounting: Tallying Genomes to Detect Adulterated Saffron.

The EU General Food Law not only aims at ensuring food safety but also to 'prevent fraudulent or deceptive practices; the adulteration of food; and any other practices which may mislead the consumer'. Especially the partial or complete, deliberate, and intentional substitution of valuable ingredients (e.g., Saffron) for less valuable ones is of concern. Due to the variety of products on the market an approach to detect food adulteration that works well for one species may not be easily applicable to another. Here we present a broadly applicable approach for the detection of substitution of biological materials based on digital PCR. By simultaneously measuring and forecasting the number of genome copies in a sample, fraud is detectable as a discrepancy between these two values. Apart from the choice of target gene, the procedure is identical across all species. It is scalable, rapid, and has a high dynamic range. We provide proof of concept by presenting the analysis of 141 samples of Saffron (Crocus sativus) from across the European market by DNA accounting and the verification of these results by NGS analysis.

Are the elemental fingerprints of organic and conventional food different? ED-XRF as screening technique.

Research has been conducted the last years to assess whether organically grown food is chemically different from produce of conventional agriculture and which markers are appropriate to discriminate between them. Most articles focus on one single food commodity, produced under strict controlled organic farming conditions, leaving open the question whether the difference would be seen when applied to the same commodity under different growing conditions. In this work 118 organic and 151 conventional samples of commercially available paprika powder, cinnamon, coffee, tea, chocolate, rice, wheat flour, cane sugar, coconut water, honey and bovine milk were characterised for their elemental composition using energy dispersive X-ray fluorescence. Resulting profiles were analysed using univariate and multivariate statistical techniques. Organic samples of a given commodity clustered together and were separated from their conventional counterparts. Differences in the elemental composition of food, could be used to develop statistical models for verifying the agronomical production system.

Capabilities of laboratories to determine melamine in food-results of an international proficiency test.

A proficiency test to assess the capabilities of laboratories to determine melamine in a milk powder and a baking mix, representing starch-containing foods like bread and biscuits, was carried out in January 2009. The need for such an interlaboratory comparison arose from a health scare in China about melamine-tainted powdered milk in the second half of 2008. Laboratories in 31 countries, including Australia, China, India, Japan, New Zealand and the USA, and 21 of the 27 Member States of the European Union participated and reported back 114 results for the milk powder and 112 for the baking mix test materials. The reported results were compared to reference values determined by exact-matching double isotope dilution mass spectrometry. The so-determined assigned values were 10.0 +/- 0.6 mg/kg melamine in the milk powder and 3.18 +/- 0.17 mg/kg melamine in the baking mix. A coverage factor k of 2 was applied to calculate the expanded uncertainties. Three quarters of all reported results for both materials had associated z scores which were satisfactory (z

Validation procedures for quantitative food allergen ELISA methods: community guidance and best practices.

This document provides supplemental guidance on specifications for the development and implementation of studies to validate the performance characteristics of quantitative ELISA methods for the determination of food allergens. It is intended as a companion document to other existing publications on method validation. The guidance is divided into two sections: information to be provided by the method developer on various characteristics of the method, and implementation of a multilaboratory validation study. Certain criteria included in the guidance are allergen-specific. Two food allergens, egg and milk, are used to demonstrate the criteria guidance. These recommendations will be the basis of the harmonized validation protocol for any food allergen ELISA method, whether proprietary or nonproprietary, that will be submitted to AOAC and/or regulatory authorities or other bodies for status recognition. Regulatory authorities may have their own particular requirements for data packages in addition to the guidance in this document. Future work planned for the implementation and validation of this guidance will include guidance specific to other priority allergens.

Tools to combat food fraud - A gap analysis.

A complex legal and institutional framework exists in the EU to ensure the safety of the feed-food chain, while such an integrated system for combating food fraud is under development. The European Commission (EC) Knowledge Centre for Food Fraud and Quality is charged with the provision of scientific insight for the policy making of EC services dealing with food fraud, and the creation of expert networks with the competent authorities of the EU Member States. To flag gaps in the existing infrastructure needed for effectively and efficiently fighting food fraud, the Centre together with the competent authorities and several EC services undertook a stocktaking exercise of what works well and which areas will need improvement. Out of several focus areas, (i) the development of early warning systems, (ii) the availability of compositional databases of vulnerable foods, and (iii) the creation of centres of competence were prioritised for further action.

Evaluation of performance characteristics of multistep analytical methods from collaborative study of linked samples.

Following unexpectedly variable results from an international comparison study of the determination of selected polychlorinated biphenyl (PCB) congeners in shellfish tissue, a group of national metrology institutes collaboratively explored the analytical characteristics of their measurement systems using a designed study with four sample materials. This "Uncertainty Suite" consisted of a 10-congener mixture of PCBs in relatively nonvolatile isooctane, a 5-congener mixture in relatively volatile methylene chloride, a methylene chloride extract of freeze-dried mussel (Mytilus edulis) tissue, and the (homogenized) mussel tissue itself. These related-but-different samples presented the participants' measurement processes with a linked series of analytical challenges. Data evaluation tools were developed to combine and visualize measurement results for the different congeners of interest for each material and, exploiting the linkages among the samples, to help identify causes for observed changes in performance. In addition to characterizing individual measurement processes, (1) the limiting sources of measurement uncertainty were found to be chromatographic separation and signal quantification in a natural matrix, (2) the achievable among-participant total measurement uncertainty for PCB calibration solutions is approximately 1.9% over the mass fraction range from 40 to 500 ng/g, and (3) the achievable among-participant measurement precision for the determination of PCB congeners in mussel tissue at levels above 0.5 ng/g mass fraction is approximately 5.4%.

Optimisation of the GC-MS conditions for the determination of the 15 EU foodstuff priority polycyclic aromatic hydrocarbons.

European legislation has recently established a list of 15 priority polycyclic aromatic hydrocarbons (PAHs) to be monitored in foodstuff. Thus, the accurate determination of these compounds has become a highly relevant issue. The fact that some of these European Union (EU) PAHs differ from those typically analysed, requires the re-evaluation of the instrumental conditions for the proper determination of the new target compounds. In this study, the influence of the stationary phase and dimensions of the GC capillary column on the chromatographic resolution of the 15 EU PAHs has been investigated. Apolar (DB-5 type) and medium polar (DB-17 type) stationary phases with different lengths and film thickness have been evaluated for the separation of the target compounds, with special emphasis on those coelutions involving isomers such as the three benzofluoranthenes included in the EU PAHs. In addition, the influence of the injection technique and the column dimensions on the recovery of the high molecular mass PAHs has been studied. A programmable temperature vaporising (PTV) injector has been used in three different operational modes and the results were compared to those obtained using on-column injection. The experimental parameters involved in the injection step were optimised by using experimental design.

Comparison of food colour regulations in the EU and the US: a review of current provisions.

This review describes the European Union and the US regulations applicable to food colours. Despite the different regulatory frameworks, the overall approach is similar, based on well-established risk-assessment procedures and risk-management measures. However, differences impacting free movement of goods can be found in the details and implementation of regulations. Using additives approved only in the US or in the EU implies that producers aiming to export need to adjust their product composition to the export market. Failure to comply may give rise to claims of adulteration, misbranding or non-compliance and rejection at the border or recall from the market. A careful comparison of the level of protection provided by the two sets of regulations, the criteria of good manufacturing practice (GMP) inspections and the certification requirements could be key to aligning the rules and to negotiating mutual recognition agreements. This review provides an extensive overview of the similarities and differences in regulating food colours in the EU and the US.

The feasibility of harmonizing gluten ELISA measurements.

Many publications have highlighted that routine ELISA methods do not give rise to equivalent gluten content measurement results. In this study, we assess this variation between results and its likely impact on the enforcement of the EU gluten-free legislation. This study systematically examines the feasibility of harmonizing gluten ELISA assays by the introduction of: a common extraction procedure; a common calibrator, such as a pure gluten extract and an incurred matrix material. The comparability of measurements is limited by a weak correlation between kit results caused by differences in the selectivity of the methods. This lack of correlation produces bias that cannot be corrected by using reference materials alone. The use of a common calibrator reduced the between-assay variability to some extent, but variation due to differences in selectivity of the assays was unaffected. Consensus on robust markers and their conversion to "gluten content" are required.

Comparative evaluation of methods for the detection of 2-alkylcyclobutanones as indicators for irradiation treatment of cashew nuts and nutmeg.

Irradiation of food products and ingredients must be indicated by proper labeling. This study evaluated the appropriateness of the European Standard EN 1785:2003 for the detection of 2-alkylcyclobutanones, which are radiolysis products of fatty acids, in cashew nuts and nutmeg and confirmed its suitability to detect irradiation of cashew nut samples at average absorbed doses of 1 kGy and above. An alternative method was developed, which is based on matrix solid phase dispersion and subsequent separation and detection of oxime derivatives of 2-alkylcyclobutanones by high performance-high resolution mass spectrometry. It is more rapid, less resource consuming, and more sensitive than EN 1785:2003. This method allowed detection of 2-alkylcyclobutanones in cashew nuts irradiated at 100 Gray and in nutmeg irradiated at 400 Gray. None of the 26 cashew nut and 14 nutmeg samples purchased in different EU Member States contained traces of 2-alkylcyclobutanones.