Serum-Derived Bovine Immunoglobulin/Protein Isolate Bind Aflatoxins B2 and G1
Authors: Rose Alexis , Okerlund Adam , Warner Christopher 
Published: May 2024

Aflatoxins are highly toxic biproducts of mold growth which commonly contaminate agricultural commodities, including food and feed. The United States Centers for Disease Control and Prevention estimates approximately half of people worldwide are chronically exposed to aflatoxins through their regular food consumption. The gut serves as both the initial target of toxicity and the location of absorption, leading to systemic exposure. Serum-derived bovine immunoglobulin/protein isolate (SBI) is a purified protein powder enriched in immunoglobulins IgG (55%), IgM (5%), and IgA (1%) used to help manage gut enteropathy. SBI binds a variety of pro-inflammatory antigens and decreases inflammatory markers in the gut by neutralizing or preventing translocation of antigen across the intestinal barrier. The objective of this study is to determine if the immunoglobulins in SBI bind to aflatoxins. Due to the protective effects of SBI, we hypothesize SBI and a highly purified immunoglobulin fraction (>95%), b ovine gamma globulin (BGG), can bind to aflatoxin B2 (AFB2) and aflatoxin G1 (AFG1). Fluorescence spectroscopy was used to quantify the binding of aflatoxins to SBI. Previous work has shown the binding of aflatoxins to tryptophan containing proteins, such as bovine serum albumin, reduces the fluorescence of the protein’s tryptophan residue. The same principle works for SBI, upon excitation at 280 nm, the emission spectra of SBI exhibits a peak at 370 nm which is decreased upon aflatoxin binding. Either SBI (5 μM) or BGG (5 μM) was titrated with an increasing concentration of either AFB2 or AFG1 (0-40 μM). The protein solution was excited at 280 nm and the emission spectra was recorded at 350-450 nm with quenching quantified at 370 nm. The fluorescence from SBI or BGG was reduced in a concentration-dependent manner as the concentration of AFB2 or AFG1 increased. The quenching constant (Ksv) was calculated using the Stern-Volmer equation, while a derivative from the equation was used to calculate the binding constant (Kb). All protein-aflatoxin pairs tested showed binding with Ksv values ranging from 0.9–1.2 x 104 M−1 (R2=0.99) and Kb values ranging from 2.5 x 104 to 4.21 x 105, with SBI-AFB2 exhibiting the tightest binding. Further, the Gibbs free energy for each pair was -6.0 to -7.7 kcal/mol, indicating favorable binding. The quenching of both protein solutions indicates structural and conformational changes in the native structures of both SBI and BGG, likely caused by the interaction of the immunoglobulins with the aflatoxin. The quenching of BGG suggests immunoglobulins in SBI play a significant role for the observed phenomenon, though other serum proteins within SBI could contribute to the quenched fluorescence. Future work will examine if the binding of the aflatoxins by SBI can prevent or mitigate aflatoxin toxicity in the gut. Proliant Health and Biologicals. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.