Effects of different processing methods on digestibility of Scylla paramamosain allergen (tropomyosin)
Introduction
There is growing concern about food allergy because it can lead to autoimmune diseases. Food allergy is representative of type I allergies that are mediated by IgE antibodies. The main food allergens are mostly proteins or glycoproteins with molecular masses of 10–60 kDa (Shainti et al., 1993). A report of the Centers for Disease Control and Prevention has indicated an 18% increase in childhood food allergy from 1997 to 2007, with an estimated 3.9% of children currently affected (Branum and Lukacs, 2008). In addition, according to an epidemiological survey, adverse immune responses to foods affect approximately 5% of young children and 3–4% of adults in westernized countries, and appear to have increased in prevalence (Scott et al., 2010).
Among a number of allergenic foods, crustaceans are considered to be one of the most common causes of food allergy due to their widespread consumption, especially in coastal countries (Lehrer et al., 2003, Wild and Lehrer, 2005). Crustaceans are one of the eight major sources of food allergens proposed by the Food and Agriculture Organization (FAO) of the United Nations and World Health Organization (WHO) (FAO/WHO, 2001), and they are favored by people because of their delicacy and nutritious value. Recently, with the development of processed crab products, researchers around the world have paid much more attention to the study of crab allergens. It has been demonstrated that the major allergen of crab is tropomyosin (TM), a myofibrillar protein that is composed of two identical subunits with molecular masses of 35–38 kDa and an isoelectric point of 4.5. In contrast with other proteins, TM is stable and can tolerate heat, grinding and the normal processing methods (Leung et al., 1998, Motoyama et al., 2006, Motoyama et al., 2007).
According to the research of Lee and Park (2004), digestibility of common whelk allergens that were digested by simulated gastrointestinal fluid remained unchanged after thermal treatment, thus demonstrating that the main allergen of crustaceans is heat stable. Moreover, compare to the numerous food processing methods, ultrasound is an effective processing and preservation technology, on account of its good directivity and strong penetrability (Villamie and Jong, 2000). Currently, there have been few studies about the impact of ultrasound on food allergens. Li et al. (2006) have adopted ultrasonic treatment of shrimp protein extract and muscle. They have indicated that high-intensity ultrasound can clearly reduce the allergic properties of shrimp allergens, and this will be beneficial to allergic individuals. In addition, the effect of high pressure treatment (300 MPa) on the hydrolysis of dairy whey proteins in simulated gastrointestinal digestion has been analyzed. After treatment with high pressure, dairy whey proteins and β-lactoglobulin were hydrolyzed easily by protease, and the IgE-binding was much lower (Izquierdo et al., 2005, Peñas et al., 2006). Therefore, it is meaningful to study the effect of different processing methods on crab allergens, and to establish proper processing methods to reduce or remove the stability and reactivity of crab TM.
In the present study, in order to identify the method that is most effective in the degradation of TM and reduction of its IgE-binding reactivity, in vitro simulated gastrointestinal digestion was adopted to investigate the digestive stability of TM extracted from crab treated by boiling, combined ultrasound and boiling (CUB), and high pressure steaming (HPS). Meanwhile, western blotting and inhibition ELISA were used to analyze the IgG/IgE-binding of TM from the digested crab crude extract (CCE) from processed crabs.
Section snippets
Crab and chemicals
Crabs (Scylla paramamosain) were purchased alive at Jimei market, Xiamen. They were immediately used or stored by freezing at −80 °C.
Protein standards for SDS–PAGE were purchased from Fermentas (Lithuania). Prestained protein standards for western blotting were purchased from New England BioLabs (Beverly, MA, USA). Horseradish peroxidase (HRP)-labeled goat anti-human IgE was from Kirkegaard & Perry Laboratories (Gaithersburg, MD, USA). HRP-labeled goat anti-rabbit IgG and 3,3′-diaminobenzindin
Digestibility of processed CCE by SGF
In SGF, the digestibility of the CCE was analyzed by SDS–PAGE, and the original TM revealed a band with a molecular mass of 38 kDa (Fig. 1). Under digestion with pepsin, the original TM band and other proteins were gradually decreased with formation of digested fragments. On one hand, a main fragment of molecular mass 34 kDa was generated after 1 min, and the density of this fragment intensified during time extension and persisted until 1 h (Fig. 1A–C). On the other hand, compared with the
Discussion
Recently, studies on crab allergens have focused on their purification and characterization, and details about the effects of different processing methods on the digestive stability of TM are still not available. Crabs are always consumed after processing, followed by gastrointestinal digestion with proteases; the proteins are digested through the gastric and then the intestinal tracts. Since the human gastrointestinal digestive system was very complicated correlated to kinds of enzymes,
Conclusions
The digestive stability and reactivity of TM from S. paramamosain treated with different processing methods were studied by in vitro simulated gastrointestinal digestion and immunoassay. In conclusion, boiling was found to accelerate the digestion of TM in simulated gastric fluid. CUB could also increase the digestion of TM in SGF and SIF. Comparing the three processing methods, HPS was found the most effective method to promote the degradation of TM by digestive protease, and to decrease the
Conflict of Interest
The authors declare that there are no conflicts of interest.
Acknowledgements
This work was supported by grants from the National Natural Scientific Foundation of China (No. 20872049), key projects from the Ministry of Science and Technology of China (2008BAD94B09), Natural Scientific Foundation of Fujian Province (2010J06012, 2008J0067), the Fujian Finance Science and Technology Bureau (2010N0019), and the Foundation for Innovative Research Team of Jimei University (2010A005).
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