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Aflatoxin B1 adsorption by yeast cell wall from commercial origin Kelly Moura Keller, Tatiana Xavier de Almeida, Rosane Nora Castro, Carina M. Pereyra, Ana Mara Dalcero, Lilia Rene Cavaglieri, Carlos Alberto da Rocha Rosa* (Argentina, Brazil).

Departamento de Microbiologia e Imunologa Veterinria. Universidade Federal Rural do Rio de Janeiro. Instituto de Veterinria. Rio de Janeiro. Brazil. 2Departamento de Qmica - Universidade Federal Rural do Rio de Janeiro. Instituto de Veterinria. Rio de Janeiro. Brazil. 3Departamento de Microbiologa e Inmunologa. Universidad Nacional de Ro Cuarto. Ruta 36 km. 601. (5800). Ro Cuarto, Crdoba. Argentina.

* Tel: ++5521 86048642 e-Mail: shalako1953@gmail.com Background. Among more than 300 mycotoxins described as yet, aflatoxin B1 (AFB1) and the group of fumonisins (FBs) are the toxins of major concern in tropical and sub-tropical regions. Aflatoxins are toxic secondary metabolites produced by species of Aspergillus genus, mainly A. flavus and A. parasiticus and its effects are carcinogenic, mutagenic, teratogenic and hepatoxic. Aflatoxin B1 is one of the most potent known hepatocarcinogens. Mycotoxin contamination of feed is a serious problem because they reduce feed consumption, decrease growth rate and reduce immunity. Besides the health aspects, which involves the presence of toxins or toxic metabolic products in food and / or by-products such as meat intended for human consumption, its presence in animal feed has also important economic connotations causing losses by increasing mortality and production (Rosa et al., 2001, 2006). One of the most effective methods to control risks of mycotoxins in animal husbandry is based on the use of specific materials that adsorb mycotoxins. These substances adsorb the toxins in the gastrointestinal tract to form insoluble complexes that are eliminated in the feces. Thus, the toxic effects are diminished by reducing the bioavailability of mycotoxins. In particular, the use yeast cell -glucans and mannans- walls (PL), mainly composed of oligosaccharides. They are usually introduced as a food additive in the animal production industry since the 90s. Objective. To evaluate the efficacy of a commercial yeast cell wall to adsorb AFB1. Materials and methods. Adsorbent: a yeast wall of commercial origin (Safmannan - Saf do Brasil - Agricultural Division) was resuspended in buffer at pH 2. pH 2 buffer solution: 50 mL of a solution of potassium chloride (KCl) 0.2 M was added 13 mL of a solution of hydrochloric acid (HCl) 0.2 M. Aflatoxin B1 Solution: 10 mg of AFB1 (Sigma) were resuspended in methanol (MeOH) to obtain a solution of 2 mg/mL. From this, the toxin was diluted to obtain the required concentrations for each test. Adsorption test: to determine the potential adsorption, saturation isotherms were previously made with different concentrations of the mycotoxin (250, 200, 100, 50, 10, 5, 2, 1 and 0, 1 mg/ml). Then, adsorption isotherms were made between the PL (2 mg/ml) with AFB1 (15,346, 10:33, 7:34, 2:08 and 4.83 mg/ml). Tests were assayed at pH 2 and 6, in triplicate. All mycotoxin concentrations were evaluated using high performance liquid chromatography (HPLC). Results and Discussion. Figures 1 and 2 show the adsorption isotherms at pH 2 and 6, respectively. Visual inspection shows a S-type isotherm, following a Hill model that explains the cooperative adsorption isotherm between the toxin and the adsorbent. The mathematical n n expression of the adj max [ZEA] /kD adsorbed ZEA by g of PL, n the number of sites and kD constant adsorption site.

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1,0 aa 0,8 0,6

0,8 0,6 0,4

ab

0,4 0,2 0,0 0 2 4 6 8

0,2 0,0 0 2 4 6 8

Figure 1. Adsorption isotherm of AFB1 by Safmannan, (a) pH 2, (b) pH 6. Among the most important parameters we consider a saturation value of 0.918 0.056 mg of adsorbed AFB1/g PL at pH 2 and 1.002 0.089 mg of adsorbed AFB1/g PL at pH 6. The association constant per site was 0.233 0.044 and 0.208 0.063 ng/mL-1 (Table 1). Table 1. Setting parameters obtained by Hill model to evaluate the adsorption isotherms from AFB1 and Safmannan at different pH.
Adsorbent Safmannan ppH D2,0 S6,0 kd (M) 4,2910,189 4,8060,303 0,2330,044 0,2080,063
max

AFB1eq/ M

AFB1eq/ M

(g/g)

n 3,8290,453 3,6190,459

N 5 5

RR2 00,996 00,996

0,9180,056 1,0020,089

kd = dissociation constant, = association constant, max = masimum capacity of adsorption, n = number of sites for coorperativism, N = number of curve points. Each point is the mean of triplicates.

Conclusion. Aflatoxin B1 was efficiently bound by PL through a cooperative attraction mechanism. This result shows the potential of this PL to prevent the toxic effects caused by the intake of ZEA. References. 1. Hooge, D.M. (2004), Meta-analysis of broilen chicken pen trials evaluating dietary mannan oligosaccharides 1993-2003, Int. J. Poult .Sci. 3, 163-174. 2. Newman, K. (1998). The biochemistry behind esterified glucomannans titrating mycotoxins out of the diet. En: Biotechnology in the feed Industry, Proceedings of Alltechs 14th Annual symposium. Nottingham University Press, UK, p. 369. 3. Oliveira, G.R.; Ribeiro, J.M.; Fraga, M.E.; Cavaglieri, L.R.; Direito, G.M.; Keller, K.M.; Dalcero, A.M.; Rosa, C.A.R. (2006) Mycobiota in poultry feeds and natural occurrence of aflatoxins, fumonisins and zearalenone in the Rio de Janeiro State, Brazil. Mycopathologia 162, 355-362. 4. Rosa, C.A.R.; Miazzo, R.; Magnoli, C.; Salvano, M.; Chiacchiera, S.M.; Ferrero, S.; Carvalho, E.Q.; Dalcero, A.M. (2001) Evaluation of the efficacy of bentonite from south of Argentina to ameliorate the toxic effects of aflatoxins in broilers. Poultry Sci. 80, 139-144. 5. Rosa, C.A.R.; Ribeiro, J.M.; Cavaglieri, L.R.; Fraga, M.E.; Gatti, M.J.; Magnoli, C.; Dalcero, A.M. (2006) Mycoflora of poultry feed and ochratoxin producing ability of isolated Aspergillus and Penicillium species. Veterinary Microbiology 113, 89-96.

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