T: Toxicology & Chemical Food Safety JFS T: Toxicology and Chemical Food Safety Quantification of Human IgE Immunoreactive Soybean Proteins in Commercial Soy Ingredients and Products Y.S. SONG, C. MARTINEZ-VILLALUENGA, AND E.G. DE MEJIA ABSTRACT: The objective of this study was the detection and quantification of human IgE immunoreactive soy- bean proteins in commercially available soy ingredients and products. Optimum dilutions of primary antibody and antigens as well as detection sensitivity were determined for the implementation of a sandwich ELISA method using plasma from soy sensitive subjects (IgE ranging from 0.35 to 98.7 IU/mL). Human IgE immunoreactivity of com- mercial soybean ingredients showed that the plasma of subjects with strong allergic reaction to soybean presented proportionally higher immunoreactive response. Soy protein isolate and soy protein concentrate contained less im- munoreactive proteins than soy flour and grits. As expected, a hypoallergenic soybean product presented the lowest IgE immunoreactivity. Hydrolyzed and fermented soy ingredients showed negligible human IgE immunoreactiv- ity when proteins and peptides were < 20 kDa. The IgE immunoreactivity of soymilk samples ranged from 3.4 to 68.9 ng IgE/mg extracted protein. Tofu contained about 20-fold higher IgE immunoreactivity than soymilk products (median 171 ng IgE/mg extracted protein). Furthermore, soy cheese products presented twice the IgE immunore- activity than tofu products (median 359 ng IgE/mg protein). Meat analogues presented considerably high extracted protein concentration (median 67.9 mg/g product). The findings of the current investigation demonstrate sandwich ELISA as a reliable immunochemical method with good repeatability, sensitivity, and low detection limit to quan- tify IgE immunoreactive proteins in soy ingredients and products. Quantitative measurement of specific IgE is likely to become an increasingly valuable tool for soybean industry to comply with food labeling for manufacturers, thus protecting soy-sensitive consumers. Keywords: allergen quantification, immunoreactivity, sandwich ELISA, soy ingredients, soybean products Introduction S oybean (Glycine max L.) is the world’s foremost source of pro- tein and oil for human food and animal feed purposes (Clarke and Wiseman 2000). In 2006, soybean production represented 39% in the United States, 23% in Brazil, 18% in Argentina, and 7% in China, which accounted for 88.5% of the global soybean produc- tion (FAO 2006). Yet, the United States is one of the largest soybean producers in the world (L’Hocine and Boye 2007). Soybean accep- tance has increased due to the low cost, nutritional quality, tech- nological properties, and health benefits of soybean protein prod- ucts (Friedman and Brandon 2001; L’Hocine and Boye 2007: Singh and others 2008). However, one of the difficult problems limiting the expanded use of soy protein products is the presence of aller- genic proteins. The Food Allergen Labeling and Consumer Protec- tion (FALCP) Act of 2004 includes soy in its definition of the “big 8” (FAO 2004), which comprises those foods that cause the 90% of all food allergies (Taylor and Hefle 2000; Kabourek and Taylor 2003; Cordle 2004). Food allergy is an immunological, usually immunoglobulin E (IgE)-related reaction resulting from the exposure (ingestion, skin contact, or inhalation) to exogenous food macromolecules (gen- erally proteins) known as allergens or antigens (Mills and others 2003; Beyer and Teuber 2004). Although most food allergies cause MS 20080140 Submitted 2/24/2008, Accepted 5/5/2008. Authors are with Dept. of Food Science and Human Nutrition, Univ. of Illinois at Urbana- Champaign, Urbana, IL 61801, U.S.A. Direct inquiries to author de Mejia (E-mail: edemejia@uiuc.edu). relatively mild and minor symptoms, some food allergies can cause severe reactions and may even be life-threatening (US FDA 2007). The estimated prevalence of soy allergies is about 0.5% of the total U.S. population (Stephan and others 2004; Sicherer and Sampson 2006). Clinical manifestations of soy allergies can provoke digestive disorders, respiratory (rhinitis, asthma), skin reactions (urticaria, atopic eczema), and IgE-mediated systemic multisystem reactions (Batista and others 2007). A recent clinical multicenter study on soy allergy in Europe determined the threshold levels for clinical reac- tivity to soy, predicting that 1% of subjects with soy allergy would react subjectively and objectively with 0.21 and 37.2 mg of soy pro- tein, respectively (Ballmer-Weber and others 2007). Soy allergy has become a public health problem that continues to challenge both the public community and the food industry, be- cause soybeans are widely used in processed foods and, therefore, represent a source of hidden allergens. Prophylactic treatments are still under investigation, but currently there is not a cure for food al- lergies caused by soy-based food products (Besler and others 2001). Currently, the only possible solution to prevent allergenic reactions is the strict avoidance of the offending allergen containing food (U.S. FDA 2007). In the United States, on March 2004, the Food Al- lergen Labeling and Consumer Protection Act (FALCPA) amended the Federal Food, Drug, and Cosmetic Act (FFDCA) and required that the label of a food product that is or contains an ingredient that bears or contains a “major food allergen” declare the presence of the allergens specified by FALCPA (Ben Rejeb and others 2004; US FDA 2007). This labeling law is effective since January 1, 2006. All ingredients derived or containing the 8 major food allergens require T90 JOURNAL OF FOOD SCIENCE—Vol. 73, Nr. 6, 2008 C  2008 Institute of Food Technologists R  doi: 10.1111/j.1750-3841.2008.00848.x Further reproduction without permission is prohibited