Progress 08/01/09 to 07/31/11
Outputs
OUTPUTS: The digestion stability of tree nut proteins may contribute to the intrinsic allergenicity of these foods. The digestion of soluble walnut and pecan proteins was carried out in simulated gastric fluid (SGF) and simulated intestinal fluid (SIF) using different ratios of protease to substrate. High molecular weight proteins of both tree nuts were found to be labile to digestion in SGF and SIF compared to low molecular weight proteins, especially the 2S albumins. These digestion-resistant proteins and peptides retain IgE binding despite extensive proteolytic digestion. Parvalbumin is recognized as an important pan-allergen in fish and frog that is capable of triggering IgE-mediated reactions in fish-allergic individuals. The reactivity of the polyclonal anti-cod parvalbumin antibody, and the commercially-available, monoclonal antifrog and anti-carp parvalbumin antibodies against raw muscle extracts of 25 fish species were determined using an indirect ELISA and IgG immunoblotting. The polyclonal anti-cod parvalbumin antibody showed reactivity to a wider range of fish species, whereas the monoclonal anti-frog parvalbumin antibody showed the least cross-reactivity. Additionally, the expression of parvalbumins in carp, catfish, and albacore tuna were evaluated using the indirect ELISA and SDS-PAGE. The parvalbumin content decreased from the anterior to the posterior positions of these fish species. These results suggested that the anti-cod parvalbumin antibody appeared to be more suitable for the detection of fish parvalbumin in foods, although limitations still exist regarding to the inconsistent binding to different species of fish. Various library databases and Current Contents are being screened continuously for publications on food allergens and clinical reports of food allergies to update the Food Allergy Research and Resource Program (FARRP) allergen database. This database has aided our bioinformatics expert in compiling a list of proteins for assessment and identification of key supporting documentation needed by the panel of allergy experts for inclusion of the allergen in the AllergenOnline database. The AllergenOnline database was updated twice during the granting period to its current version of 11.0 in February 2011. Version 11.0 contains a comprehensive list (1491 sequence entries from 265 species that are clustered into 553 protein groups) of unique proteins of known and putative allergenic proteins (food, airway, venom/salivary and contact) with sufficient evidence judged by the panel to be called allergens or putative allergens. The updates replace version 9.0 with 1386 sequences. Sequences and other reference information for version 11.0 were downloaded in May 2010 and compared with all sequences contained in version 9.0 and 10.0 including those listed as allergens and putative allergens and those that were not listed as they were judged to have insufficient evidence to be defined as allergens in version 10.0. Source and sequence information from NCBI as well as data from allergy studies were gleaned from AllergenOnline version 10.0, IUIS, Allergome, PubMed and individual publications. PARTICIPANTS: Jelena Spiric is a Masters student in the Department of Food Science and Technology at the University of Nebraska-Lincoln. The examination of the digestive stability of pecan and walnut allergens and characterization of intact protein and digestion-resistant fragments is her Masters research project. Poi-Wah Lee is a Ph.D. student in Department of Food Science and Technology at the University of Nebraska-Lincoln and the assessment of the allergenicity of fish and ingredients derived from fish is part of her Ph.D. dissertation research. Dr. Steve Taylor and Dr. Joseph Baumert are the principal investigators of this grant and supervise the graduate students previously mentioned. Jamie Kabourek (M.S., R.D.) is the Resource Manager with the Food Allergy Research and Resource Program (FARRP) at the University of Nebraska-Lincoln. Jamie manages the continuous update of the FARRP allergen database. John Wise is a bioinformatics specialist with FARRP at the University of Nebraska-Lincoln. John compiles the initial list of proteins for assessment and identifies key supporting documentation, develops the new version of AllergenOnline and makes the new version publically available on the AllergenOnline website. Dr. Rick Goodman is the director of AllergenOnline and supervises the work conducted by John Wise as well as leads the peer review team consisting of allergen experts from around the world. TARGET AUDIENCES: Three target audiences exist for the research being conducted to characterize the digestive stability of food allergens and assessment of the allergenicity of fish and ingredients derived from fish. Scientific researchers will benefit from the additional characterization of additional tree nut and fish allergens and evaluation of their relative stability under proteolytic and thermal processing conditions as this information may provide insight into additional research avenues that examine techniques that could be used to decrease the immunoreactivity of these allergenic proteins. Examination of the stability of food allergens may also aid in the improvement of immunoassays for detection of allergenic residues as the stable proteins identified in this research may be utilized in the immunoassays for improved detection in highly processed foods. Food processors may be interested in the characterization of fish allergens as this research could lead to the development of an immunoassay capable of detecting multiple fish species. Currently there are no immunoassays capable of detecting fish residue from multiple species so reliable detection is limited. Regulatory agencies may also be interested in these immunoassays to ensure that processed food products on the market are in compliance. Scientists in academia, government, and private industry will be interested in the new version of AllergenOnline for risk assessment of potential allergenicity of the genetically engineered or novel food proteins that they are researching. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
The prevalence of walnut and pecan allergies is common among allergic consumers with tree nut allergies. Allergic reactions to walnut and pecan can be severe and occasionally life threatening. There is limited information available on characterizing the stability of walnut and pecan allergens. The 2S albumins from walnut and pecan remain stable despite extensive in vitro gastrointestinal digestion. The digestion stability of the 2S albumins may contribute to allergic sensitization to walnut and pecan. Identification of digestion resistant peptides is an important factor contributing to assessment on food protein allergenicity. The presence of undeclared fish parvalbumin in foods can pose serious health risks among fish-allergic consumers and present liability issues for the food industry. However, current methods present shortcomings in consistently detecting the allergenic residues from all species of fish. Monoclonal antifrog and anti-carp parvalbumin antibodies are commercially available, however, the anti-cod parvalbumin antibody developed by the Food Allergy Research and Resource Program (FARRP) appeared to be more suitable for the detection of fish parvalbumin in foods, although limitations still exist regarding to the inconsistent binding to different species of fish. The variation of the parvalbumin content in muscle tissues located in different parts of the fish could affect the ability of these anti-parvalbumin antibodies to detect undeclared fish residues in foods, depending on which parts of fish muscles that are incorporated in the foods. The up-to-date literature in the FARRP allergen database has been used to develop scholarly review articles, website reviews, and other scholarly materials that benefit the broader research community on food allergens. The literature database has also been instrumental for the development of the new version of AllergenOnline which can be utilized by academic scientists, government regulators, and private industry for risk assessment of potential allergenicity of the genetically engineered or novel food proteins.
Publications
- Lee P, Nordlee JA, Koppelman SJ, Baumert JL, Taylor SL. 2011. Evaluation and comparison of the species-specificity of 3anti-parvalbumin IgG antibodies. J Agri Food Chem. (Accepted for publication).
- Lee P, Nordlee JA, Koppelman SJ, Baumert JL, Taylor SL. 2011. Characterization of IgG and IgE binding to parvalbumin derived from commercially important fish species. J Allergy Clinical Immunol. 127: AB32.
- Spiric J, Knulst A, Nordlee JA, Koppelman SJ, Taylor SL, Baumert JL. 2011. Identification and characterization of digestion-resistant allergens in walnut and pecan species. Allergy. 66(Suppl. 94):532.
- Lee P, Nordlee JA, Koppelman SJ, Baumert JL, Taylor SL. 2010. Binding characteristics of anti-parvalbumin antibodies to the muscle extracts derived from various fish species and distribution with whole fish. Allergy. 65(Suppl. 92):324.
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Progress 08/01/09 to 07/31/10
Outputs
OUTPUTS: This project was recently funded, starting in August of 2009. Good progress has already been made toward the four main goals and objectives outlined in this project. Objective 1: Pepsin digestion experiments are underway to examine the stability of 2S albumins from peanut and tree nuts under reducing and non-reducing conditions. Identification and characterization of digestion-resistant proteins and peptide fragments will follow. Objective 2: The immunoreactivity of parvalbumin from 15 different species of fish has been evaluated using 3 anti-parvalbumin antibodies (anti-frog parvalbumin, anti-cod parvalbumin, and anti-carp parvalbumin). Differences in parvalbumin content across the longitudinal position of carp and catfish using indirect ELISA and SDS-PAGE techniques are currently being evaluated to determine if sub-sampling from different areas of the fish will affect detectable parvalbumin content. Objectives 3 and 4: Various library databases and Current Contents are being screened continuously for publications on food allergens and clinical reports of food allergies to update the Food Allergy Research and Resource Program (FARRP) allergen database. This database has aided our bioinformatics expert in compiling a list of proteins for assessment and identification of key supporting documentation needed by the panel of allergy experts for inclusion of the allergen in the AllergenOnline database. The expert panel is currently reviewing the list of potential allergenic proteins and the version 10 of AllergenOnline will be developed in the coming months with a public release anticipated in January 2010. PARTICIPANTS: Jelena Spiric is a Masters student in Food Science and Technology at the University of Nebraska-Lincoln. The examination of the digestive stability of food allergens and characterization of intact protein and digestion-resistant fragments is her Masters research project. Poi-Wah Lee is a Ph.D. student in Food Science & Technology at the University of Nebraska-Lincoln and the assessment of the allergenicity of fish and ingredients derived from fish is part of her Ph.D. dissertation research. Dr. Steve Taylor and Dr. Joe Baumert are the principle investigators of this grant and supervise of the graduate students previously mentioned. Jamie Kabourek (M.S., R.D.) is the Resource Manager with the Food Allergy Research and Resource Program (FARRP) at the University of Nebraska-Lincoln. Jamie manages the continuous update of the FARRP allergen database. John Wise is a bioinformatics specialist with FARRP at the University of Nebraska-Lincoln. John compiles the initial list of proteins for assessment and identifies key supporting documentation, develops the new version of AllergenOnline and makes the new version publically available on the AllergenOnline website. Dr. Rick Goodman is the director of AllergenOnline and leads the peer review team consisting of allergen experts from around the world. TARGET AUDIENCES: Three target audiences exist for the research being conducted to characterize the digestive stability of food allergens and assessment of the allergenicity of fish and ingredients derived from fish. Scientific researchers will benefit from the additional characterization of peanut, tree nut, and fish allergens and evaluation of their relative stability under proteolytic and thermal processing conditions as this information may provide insight into additional research avenues that examine techniques that could be used to decrease the immunoreactivity of these allergenic proteins. Examination of the stability of food allergens may also aid in the improvement of immunoassays for detection of allergenic residues as the stable proteins identified in this research may be utilized in the immunoassays for improved detection in highly processed foods. Food processors may be interested in the characterization of fish allergens as this research could lead to the development of an immunoassay capable of detecting multiple fish species. Currently there are no immunoassays capable of detecting fish residue from multiple species so reliable detection is limited. Regulatory agencies may also be interested in these immunoassays to ensure that processed food products on the market are in compliance. Scientists in academia, government, and private industry will be interested in the new version of AllergenOnline for risk assessment of potential allergenicity of the genetically engineered or novel food proteins that they are researching. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
The outcomes/impacts of this research cannot be accurately measured at this time as the research data is just beginning to be analyzed. The up-to-date literature in the FARRP allergen database has been used to develop scholarly review articles, website reviews, and other scholarly materials that benefit the broader research community on food allergens. The literature database has also been instrumental for the development of the new version of AllergenOnline.
Publications
- No publications reported this period
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