Newsletter Food Allergen Community

AOAC Food Allergen Community

NEWSLETTER Volume 4 | Issue 2

IN THIS ISSUE

Editorial Comment

Editorial Comment

Risk perception: Dihydrogenmonoxide

Risk perception: Dihydrogenmonoxide1

News FDA defines ‘gluten-free’

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Health Canada’s Position on Highly Refined Oils Derived from Food Allergen Sources

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iFAAM – the world’s largest allergen study has started

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Guidance for the Validation of Gluten ELISA Methods coming to AOAC 4

Research Highlights Hide and Seek: The complex game of allergen detection in processed foods and the need for incurred reference standards 5 Special Issue on Food Allergens

Some years ago, a survey was conducted to evaluate the perception of risk in the general public. A simple question was asked: should dihydrogenmonoxide be banned or at least regulated in Europe. 75% of the people asked responded with “yes – banned”. But why would anybody regulate water? On the other hand, food allergens are a real and immediate risk to affected consumers, and producers and regulators have to ensure that people suffering from food allergies can make an informed choice about the products they are buying. The same applies to gluten levels in food for celiacs. In this issue, we report about two major advances towards risk reduction and consumer safety: The FDA has – dare I say it – finally issued the “Gluten-Free” Rule. The second advance is the start of the European iFAAM project, which for the first time combines clinical studies, reference materials and analytical techniques to get more reliable and consistent results, allowing significantly improved risk management. While these, in the grand scheme of things, are little steps, we are definitely on the right way and need to keep moving forward.

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Bert Popping | Editorial Team Member

Manufacturer’s Corner6 Upcoming Events6

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2013

Editorial Team

Editor in Chief: Carmen Diaz-Amigo

Editorial Members: Terry Koerner Jupiter Yeung Michael Abbott James Roberts Bert Popping

Graphic Design: Carmen Diaz-Amigo

AOAC Food Allergen Community Newsletter: [email protected]

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News FDA defines ‘gluten-free’ The U.S. FDA published a new regulation in the Federal Register defining the term “gluten-free” for voluntary food labeling on August 5, 2012 (http://www.gpo.gov/fdsys/pkg/FR-2013-0805/pdf/2013-18813.pdf ) and Questions and Answers on the final rule (http://www.fda.gov/Food/GuidanceRegulation/ GuidanceDocumentsRegulatoryInformation/Allergens/ ucm362880.htm.) The regulation is effective September 4, 2013, 30 days after publication, and compliance activities begin on August 5, 2014, one year after publication. This new definition standardizes the meaning of gluten-free claims across the food industry. It requires that, in order to use the term gluten-free” on its label, a food must meet all of the requirements of the definition, including that the food must contain less than 20 ppm gluten. This level is consistent with the regulations in EU and Canada, and aligns with the limit of detection of validated analytical methods. FDA’s enforcement approach for “gluten free” claims is built on validated analytical methods, even though their safety assessment led to lower thresholds. FDA also states:

available that can reliably detect gluten in a range of food matrices at levels below 20 ppm, we will reevaluate the < 20 ppm gluten level that we have included as part of the criteria for the definition of ‘gluten-free.’” (78 Fed. Reg. 47154, page 47161, at response 6)” FDA plans to issue a separate proposed rule to address its compliance approach for “gluten free” claims on fermented or hydrolyzed products, as the traditional analytical methods used to detect gluten are not appropriate for these foods. To the extent FDA does not issue such a proposed rule before August 5, 2014, the agency will issue guidance for “gluten free” claims on fermented or hydrolyzed foods. The agency also states its intent to exercise enforcement discretion with respect to the requirements for “gluten free” claims for FDA-regulated beers that currently make a “gluten free” claim and that are: (1) made from a non-gluten-containing grain; or (2) made from a gluten-containing grain, where the beer has been subject to processing that the manufacturer has determined will remove gluten below a 20 ppm threshold. Jupiter M. Yeung | Nestle Nutrition

“If future data indicate that the gluten content of < 20 ppm is not sufficiently protective of the health of individuals with celiac disease and analytical methods become

Join the Allergen Community at the AOAC Annual Meeting & Exposition Chicago, IL, August 25-28, 2013 Allergen and Gluten Scientific Sessions: Selection of Analytical Methods for Validation of Allergen Control Plans Co-Chairs: Steve Taylor (University of Nebraska) and George Dunaif (Grocery Manufacturers Association) Chasing Gluten” What Can you Detect? How to Deal with the Results? Co-Chairs: Carmen Diaz-Amigo (Eurofins) and Eric Marceau (Canadian Food Inspection Agency)

Subscribe to our FREE Newsletter HERE Send us an e-mail to [email protected] or complete the following electronic form and send it to us by clicking on the SUBMIT bottom: Name: Affiliation: E-mail: Comment:

What Comes Before the MS in LC-MS Food Allergen Analysis? Co-Chairs: Terry Koerner (Health Canada) and Mark Ross (U.S. Food and Drug Administration) More information »

SUBMIT

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Health Canada’s Position on Highly Refined Oils Derived from Food Allergen Sources Enhanced food allergen labelling regulations took effect in Canada on August 4, 2012. In part, these regulations require that whenever a priority food allergen is used as an ingredient or component of an ingredient in the manufacturing of prepackaged foods, the food allergen source must be declared on the label of the food, either in the list of ingredients or in an optional, separate “contains:” statement. “Food allergen” is defined in the regulations as any protein or modified protein, including any protein fraction that is derived from one of the priority allergens listed in the regulations. Since the regulatory definition of a food allergen is specific to protein or protein fractions derived from food allergens, ingredients that are derived from a food allergen but which have been processed in a way that removes the protein are not themselves considered to be food allergens. Edible oils can be derived from a number of priority food allergens, including soybean, peanuts, tree nuts (hazelnut, almond, Brazil nut, pecan, pistachio, pine nut, cashew, walnut, and macadamia nut), sesame seed and fish. In addition, specific grades of mustard seed are sometimes used to make canola oil. Refined oils are produced using a number of different steps. The crude oil is generally produced by mechanically pressing the source seed or bean in expellers after a preheating step in indirectly heated conditioners. The crude oil can then be treated using a number of different refining steps. Degumming, neutralizing, bleaching and deodorizing are all steps that are used to treat the crude oil and produce what is then considered to be a highly refined oil. Certain unrefined oils can contain significant quantities of protein, as high as 300 micrograms per gram (µg/g = parts per million). In contrast, highly refined oils contain very little protein, with published data showing low µg/g values or

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Quizz

lower. Each of the steps used in refining the oil will reduce the amount of protein in the final product. When highly refined oils are used as ingredients in other foods, the concentration of any residual protein would be further diluted in the final food. In Canada, sections B.01.009 (4) and B.01.010 of the Food and Drug Regulations require that whenever peanut oil is present as an ingredient, or component of an ingredient, in a food, the source of the oil, “peanut”, must always be identified. Highly refined oils may be derived from a priority food allergen source. However, because of the degree to which they have been refined, contain no protein or an amount of protein that is too small to pose a health risk. Health Canada’s position is that the enhanced labelling regulations would not apply to highly refined oils. This position was developed in consultation with, and agreement of, the Canadian Food Inspection Agency (CFIA). Companies must be prepared to provide evidence that allergen-derived ingredients (for example, allergen-derived oils) are highly refined and contain negligible levels of protein if they consider that the ingredient is not subject to the enhanced allergen labelling regulations. Highly refined oils would still have to be labelled according to all other existing labelling requirements, such as the requirement to declare the source for peanut oil (whether highly refined or not) and the requirement to declare the refined oil as an ingredient in the ingredient list of a prepackaged food to which it has been added, if applicable. For more information on Health Canada’s position on highly refined oils derived from food allergen sources please follow this link » Michael Abbott | Health Canada

See the response below (only available in the electronic pdf )

In the year 2012, which allergenic ingredient was reported to be the main cause of allergen-related official actions in Europe?

Egg

Milk

Soy

Wheat

Hazelnut

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2013

iFAAM – the world’s largest allergen study has started The European Commission funded under the 6th framework program two important projects related to allergy: Europrevall, which focused on clinical aspects of allergies, and MoniQA, which focused on the analytical aspects. One of the major issues with any of the allergen related project was, that none of them enabled the direct correlation of clinical response and analytical result. Professor Clare Mills submitted under the 7th framework program the iFAAM project proposal. iFAAM stands for Integrated Approaches for Food Allergen and Allergy Risk Assessment. The project combines for the first time these two aspects: clinical studies are conducted with well characterized lightly and highly processed, commutable reference material, looking for trigger levels of major allergens, including milk, egg, hazelnut and peanut. The very same material will be analysed by multi-allergen LC-MS/MS (mass spectrometry), following the recommendations driven through the MoniQA Project (published in the J. AOAC Int.) and an international expert group.

This establishes a direct correlation between allergic response and analytical result using LC-MS/MS. This is a truly new development and will allow manufacturers to perform better risk management-based on sound results, which can be correlated back to clinical responses. However, this European project will not only establish a direct correlation between clinical response and analytical results, it will also produce reference materials which can be used to calibrate suitable conventional methods. More information: Link 1 » and Link 2 » This 4-year project is funded with 9M Euros by the European Commission and started in March 2013. Bert Popping | Eurofins

Guidance for the Validation of Gluten ELISA Methods coming to AOAC Celiac disease occurs in 1% of the population resulting in an intolerance to the storage proteins (gluten) of wheat, rye, barley and their cross bred species. With the only prescription being avoidance of gluten in the diet, well validated methods are needed to accurately measure the inadvertent gluten contamination of products in the diet of people with celiac disease. This information is vital for the food industry to accurately label products as gluten-free and for consumers to be confident in the food they purchase for themselves and their families. The international Codex Alimentarius revised Standard 118 -1979 for gluten-free foods in 2008, which defines the term gluten and the requirements for products to be considered gluten-free. Many jurisdictions have, or are considering, adopting this standard that would require gluten-free foods ready for consumption not to exceed a level of 20 mg of gluten per kg of food.

these validation requirements in Ottawa, Canada. A second meeting in the spring of 2012 introduced the draft guidance to a larger group during the 7th Food Allergen Methodologies Workshop in Montreal, Canada. With all input considered, the guidance document was submitted to AOAC in late 2012 and was accepted for publication in May 2013. This is a consensus document with contributors from international institutions including government, industry, and academia. The document is divided into two broad sections for the information that would be required by a method developer and guidance around the implementation of a multi-laboratory validation study. The guidance document provides specific information requirements for cross reactivity towards both celiac sensitive cereals and non-celiac cereals. There is also specific guidance around gluten reference materials, spiking methods and food matrices important in the diet of people with celiac disease.

ELISA methods are the current choice for measuring the gluten content in both raw ingredients and processed foods. Gluten is a complex mixture of proteins with diverse structural and chemical properties and the ability of the ELISA to detect these proteins is further complicated by a wide variety of matrices and processing conditions. In view of these factors, there is a need to develop a harmonized approach to the validation of gluten ELISA methods. Important steps towards harmonization will require (i) harmonized validation protocols, (ii) a commonly accepted gluten reference material, and (iii) at least one independent reference method to verify the routine methods.

This guidance document represents one piece of a complete approach to validating ELISA methods to be used for the measurement of the unintended gluten contamination in foods for people with celiac disease. Members of the AOAC allergen community in collaboration with other international organizations are working towards the others including gluten reference materials and an independent method of analysis. These will be discussed in more detail in further issues of this newsletter.

In August of 2011 members of the AOAC allergen community assembled a gluten working group to address the first of

The protocol will be published in the upcoming issue (September/October) of the Journal of AOAC International. Terry Koerner | Health Canada

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Research Highlights Hide and Seek: The complex game of allergen detection in processed foods and the need for incurred reference standards Accurate detection of allergens when present in foods is an important component of allergen risk management. For some food allergy sufferers, the inadvertent presence of even small quantities of allergenic proteins can result in severe health impacts. ELISA methods commonly used for allergen detection require the use of calibrated reference standards. To verify the performance of these kits, standards are spiked into food matrices for testing and validation. Spiked samples are generally not subjected to food processing treatments and as such are very different from incurred reference materials where target allergens are incorporated into the food matrix prior to processing (e.g., baking). Incurred samples are therefore more representative of foods that allergic sufferers are likely to consume. The recovery of allergens in incurred reference materials vs spiked materials can vary significantly which raises questions about the accurate detection and quantification of allergens in processed foods when spiked references are used. In recent studies we determined that gluten and casein recoveries in a cookie dough decreased from 93-72% to 3121% and from 88-75% to 35 -19%, respectively, after baking. Recoveries for samples incurred with egg and soy were similarly decreased to the point where false negatives were detected. We also observed that increasing baking temperature and time had negative impacts on allergen recovery. Additionally, physical characteristics such as the size of the cookie had an impact on recovery with larger cookie sizes yielding higher recoveries than smaller cookies, likely due to greater thermal transfer to the smaller cookies. Similar studies on the effect of thermal processing on the antigenicity of ovalbumin using ovalbumin, egg white and whole egg powders and incurred pasta preparations made with egg protein, using both LCMS and ELISA, yielded negative results for all incurred pasta samples after cooking. (A)

(B)

These studies and others reported in the literature demonstrate that allergen recovery and detection is affected by processing. Foods are processed in a variety of ways and with a multitude of ingredients. The composition of the food as well as the processing technique used determine its final form and structure and can impact protein extractability. Thermal processing, as an example, may result in protein denaturation, association, aggregation and complexation. These changes affect protein conformation which can modify allergenic epitopes. Proteins also interact with other components in foods (e.g., carbohydrates, phenolics) to create complex networks (e.g., through the Maillard reaction) that can render them difficult to extract (e.g., in cookie dough, pasta, chocolate). The use of appropriate buffers and extraction conditions that increase protein recovery in processed foods without modifying allergenic epitopes is therefore important as this will enhance allergen detection. Furthermore, using incurred reference standards for allergen detection may be preferable to the use of spiked standards. A major difference between spiked and incurred samples is that target allergens in incurred materials encounter processing conditions that can impact their extractability and recovery in a similar manner as for the food of interest. Use of incurred standards in allergen detection should be increasingly considered as they may provide more accurate information about the presence or absence of allergens in foods. Gomaa, A. & Boye, J.I. (2013) Impact of thermal processing time and cookie size on the detection of casein, egg, gluten and soy allergens in food. Food Research International, 52(2), pp. 483-489. Abstract » Joyce Irene Boye | Agriculture and Agri-Food Canada

(C)

Percentage recoveries of (A) casein, (B) egg and (C) soya in incurred cookie as detected by ELISA and flow cytometry. (Large, mid-size, and small refers to cookie sizes of 10 mm thickness and 38, 58 and 76 mm diameters, respectively; 10, 15, 25 min refers to cookie baked for these different times in an oven at 177 °C). Source: ScienceDirect »

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Special Issue on Food Allergens The Journal of Agricultural and Food Chemistry Vol 61(24) 2013 includes a special issue on Food Allergens. This issue is co-edited by Mark M. Ross and Lauren Jackson (U.S. FDA) and compiles a series of studies presented at the symposium “Advances in Food Allergen Detection” held at the 243rd National Meeting of the American Chemical Society (ACS) in March 2012 in San Diego, CA. Ross M.M. & Jackson L. Summary of the ACS Symposium on Advances in Food Allergen Detection. pp 5621–5623. Abstract » Cucu, T.; Jacxsens, L. & De Meulenaer, B. Analysis To Support Allergen Risk Management: Which Way To Go? pp 5624–5633. Abstract » Gendel, S.M. The Regulatory Challenge of Food Allergens. pp 5634–5637. Abstract » Hebling, C.M.; McFarland, M.A.; Callahan, J.H. & Ross, M.M. Global Proteomic Screening of Protein Allergens and Advanced Glycation Endproducts in Thermally Processed Peanuts. pp 5638–5648. Abstract » Fu, T.J. & Maks, N. Impact of Thermal Processing on ELISA Detection of Peanut Allergens. pp 5649–5658. Abstract »

Stay Tuned! Can’t wait to hear from us? Do you want to share good news with the community?

Newsome, G.A. & Scholl, P.F. Quantification of Allergenic Bovine Milk αS1-Casein in Baked Goods Using an Intact 15N-Labeled Protein Internal Standard. pp 5659–5668. Abstract » Eischeid, A.C.; Kim, B.H. & Kasko, S.M. Two Quantitative Real-Time PCR Assays for the Detection of Penaeid Shrimp and Blue Crab, Crustacean Shellfish Allergens. pp 5669–5674. Abstract » Sakai, S.; Adachi, R.; Akiyama, H. & Teshima, R. Validation of Quantitative and Qualitative Methods for Detecting Allergenic Ingredients in Processed Foods in Japan. pp 5675-5680. Abstract » Diaz-Amigo, C. & Popping, B. Accuracy of ELISA Detection Methods for Gluten and Reference Materials: A Realistic Assessment. pp 5681–5688. Abstract »

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AOAC Food Allergen Community Newsletter You can contribute with articles, news items or suggestions Submission deadline for the 3rd Issue of 2013: Nov 22, 2013

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Mark your calendar !!!

Manufacturer’s Corner Biomedal® launched AlerTox® Sticks Casein, its latest rapid, specific and user-friendly kit. The test detects 2.5 ppm of casein and does not cross react with other milk proteins such as betalactoglobulin. Designed for solid and liquid samples and surfaces. Results obtained in less than 20 minutes. More information »

Upcoming Events Eighth Workshop on Food Allergen Methodologies Vancouver (Canada), May 5-8, 2014 Health Canada is pleased to announce the Eighth Workshop on Food Allergen Methodologies. This workshop will be taking place at the Sheraton Vancouver Wall Center Hotel in Vancouver, British Columbia (Canada) on May 5-8, 2014. These workshops are always very informative and provide opportunities for discussion and networking with others who share an interest in food allergens and particularly allergen detection methods. More information will be soon available. More information »

99 Regulatory Updates 99 Food Industry Initiatives 99 Regional developments 99 Upcoming events 99 Questions for our Experts 99 Interested in a topic?

The AOAC Food Allergen Community is a forum serving the scientific community working on Food Allergens: The community is aimed to help AOAC INTERNATIONAL in its consensus-based scientific and advisory capacity on methods of analysis for allergens in foods and other commodities. It is also meant to serve the broader Stakeholder Community whose objectives it is to enhance the protection of food allergic consumers worldwide. Contact us at [email protected]

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