With the growth in bulk and container sea freight in the second half of last century, the modern food and feed distribution system became a global entity. This had significant positive implications on consumer choice and availability, but also had negative consequences for food borne toxins which manifested themselves during the extended phase of transportation and storage as well as growth and harvest.
The Codex Alimentarius Commission (CODEX) was first created between 1961 and 1963 by the Food and Agriculture Organisation (FAO) and the World Health Organization (WHO). The purpose of CODEX was to develop food standards and recommendations as the international benchmark for food safety and to protect consumer health. CODEX ensures food safety and quality standards by using risk management tools set by prudent scientific advice from a range of industrially and scientifically accredited organisations. World Trade Organisation members are encouraged to equate national feed regulations with international agreed standards. CODEX incorporates advice and feed management techniques that help to reduce the risk of mycotoxins, which are secondary metabolites of fungi. However it is impossible to completely remove such mycotoxins from the animal and human food chain given that feed and its precursors can be stored and transported throughout a range of time intervals, atmospheric humidity and temperatures. The modern feed distribution network needs to incorporate commercial methodologies to guarantee that feed contains minimum quantities of mycotoxins which can be detrimental to animal health and production. This paper reports on the development of a new mycotoxin binder which adsorbs mycotoxins into highly stable neutral complexes without compromising nutrient adsorption whilst at the same time de-activating and eliminating the causative mycotoxin producing fungi organisms.
The Problems Mycotoxins cause within animal production
Mycotoxins are a diverse family of toxins produced by certain fungi, especially by species of Aspergillus, Fusarium, Penicillium, Claviceps and Alternaria. Mycotoxins in food can cause huge problems for humans and animals. Consumption of a mycotoxin-contaminated diet may induce acute and long-term chronic effects resulting in teratogenic, carcinogenic, oestrogenic or immune suppressive effects. Direct consequences of consumption of mycotoxin-contaminated animal feed include: reduced feed intake, feed refusal, poor feed conversion, diminished body weight gain, increased disease incidence (due to immune-suppression), and reduced reproductive capacities (Fink-Gremmels and Malekinejad, 2007; Morgavi and Riley, 2007; Pestka, 2007; Voss and Haschek, 2007) which leads to economic losses (Huwig et al., 2001; Wu, 2004; Wu, 2006). The most common mycotoxins are aflatoxins, ochratoxin A, trichothecenes, zearalenone and fumonisins.
The only real solution
Globalisation of the agricultural commodities is now so embedded in the structure of our society that is has become an integral part of the stock exchange with its headquarters as the Chicago Board of Trade (CBOT). CBOT specialises in commodities trading and future sales of agricultural products. The diversity and scale of how the animal feed markets have become included within this global system has advantages in optimisation of production technologies but also its disadvantages. One significant disadvantage are the large distances between sites of production and the markets of consumption with the corresponding reliance on transportation links which go across different microclimates on their journey. Agricultural commodities do not hold enough commercial value to warrant the ultimate in packing technologies which would isolate their exposure to such changes in temperature and atmospheric humidity. Here lies the opportunity for mycotoxins to do their detrimental work, as well as during the cultivation and harvest stages of such crops. Therefore, if mycotoxins cannot be avoided, the only real practical solution is for a highly active mycotoxin adsorbent.
Broad spectrum mycotoxin adsorbents should include features such as safety, affordability, nutritionally beneficial to the animal but also that they can be easily included within animal feeds. Complete mycotoxin control may require a combination of different approaches with varying modes of action. Not all mycotoxin adsorbents have the same capacity to protect livestock against the detrimental effects of mycotoxins. Chung et al., (1990) has shown that a number of adsorbents may impair essential nutrient utilisation. It is fundamental that biological decontamination methods should incorporate the ability to decontaminate mild mycotoxin concentrations without losses of nutritional feed value or palatability.
Activated charcoal occurs as a result of pyrolysis of organic materials. It is a porous non-soluble powder with a high surface area to mass ratio (500 - 3500 m2/g). It adsorbs mycotoxins but different types of activated charcoals have a diverse affect against mycotoxins (A.Huwing et al., 2001).
Aluminosilicates (zeolites, Hydrated sodium calcium aluminosilicates HSCAS, clays) Aluminosilicates for the adsorption of mycotoxins has been studied for more than 20 years (Masimango et al., 1978; Mumpton and Fishman, 1977). Zeolites are crystalline aluminosilicate minerals (Harben, 1999). There are about fifty different natural zeolites and another hundred and fifty which have been synthesised for specific applications such as industrial catalysts. Clinoptilolite, which in Greek means 'oblique feather stone', is a member of the zeolite group of minerals (Harben, 1999). Clinoptilolite is a naturally occurring zeolite, formed by the devitrification of volcanic ash in lake and marine waters millions of years ago. It is the most researched of all zeolites and is widely regarded as the most useful. Clinoptilolite is able to bind a range of mycotoxins, forming highly stable complexes (Tomasevic-Canovic et al, 2001; Huwig et al, 2001). Studies have shown that clinoptilolite absorbs toxins created by moulds in animal feeds, as well as enhancing nutrient absorption by cattle, pigs, lambs and other animals. Clinoptilolite binds a range of mycotoxins, which is a benefit to the health of animals into whose feed clinoptilolite has been added (Huwig, 2001; Phillips, 1990).
Yeast and probiotics. Yeast cell walls can be used as adsorbents for mycotoxins (Grunkemeier, 1990; Bauer, 1994). In recent years it has been shown, that probiotics have the ability to bind and biotransform mycotoxins (Biagi, 2009).
Polymers - Polyvinylpyrrolidone (PVP) is a highly polar amphoteric polymer adsorbent with capacity to adsorb zearalenone in vitro (Alegakis et al, 1999).
Anti-fungal Antifungal agents (essential oils) do not get rid of mycotoxins themselves, but kill the causative organisms which produce them. This ensures that mycotoxin concentrations will not increase, and this perfectly complements the mode of action of mycotoxin binders.
Efficacy of different adsorbents for the binding of mycotoxins
Benefits of Aluminosilicates and Zeolites as mycotoxin binders in experimental studies
It is suggested, that activated charcoal, which is a porous non-soluble powder (mass ratio 500-3500 m2/g) might inactivate mycotoxins. Activated charcoal may potentially absorb essential nutrients if the concentrations in feed are higher than mycotoxins.
Aflatoxins are some of the most extensively researched mycotoxins. Clinoptilolite is able to adsorb aflatoxins resulting in measurable improvements in the health of swine, sheep and chickens (Mumpton, 1999; Parlat et al, 1999; Kyriakis et al, 2002). The addition of clinoptilolite to the diets of swine, poultry, ruminants and other animals resulted in improved weight gain and feed conversion ratios with improved milk yields in dairy herds (Kyriakis et al, 2002; Mumpton, 1985). The incidence of scours, enteritis and other intestinal diseases were found to be substantially decreased (Mumpton, 1985). An abundance of published data indicates that the dietary use of natural zeolites reduces the frequency, severity and duration of diarrhoea in calves and pigs. (Mumpton et al., 1977). Katsoulos et al., (2006) showed that clinoptilolite was effective to prevent milk fever at an inclusion level of 2.5% of the feed concentrate resulted in a lower incidence of ketosis.
Hempken et al showed that cows diets containing urea which had been supplemented with clinoptilolite significantly reduced rumen NH3 concentration. The same trend was observed by the dietary addition of clinoptilolite in steers and lambs.
It has been discovered that Hydrated Sodium Calcium Aluminosilicates (HSCAS) and/or Activated Charcoal included in diets that contained 102 ppb aflatoxins reduced or essentially eliminated histopathologic lesions in the livers (Bonna et al., 1991). Kubena et al. (1998) suggested that HSCAS provides a level of protection against Aflotoxin, but not T-2 toxin, in young broiler chicks. Types of clay products including calcium bentonite clay were similar in effectiveness to HSCAS in restoring performance to pigs after the consumption of aflatoxins (Schell et al., 1993a; Schell et al., 1993b). Different clays showed adsorption rates for ochratoxin A and zearalenone in the range of 0-100%.
Zeolites have been used effectively for the prevention of heavy metal toxicity in animals due to their high ion-exchange capacity (Pond et al., 1983).
Zeolites also have the capacity to bind radioactive elements as well as heavy metals and have therefore been suggested as a means of altering toxin uptake and excretion from the body (Papaioannou, et al., 2005). Ivan et al., (1992) observed a significantly reduced Cu concentration in sheep's liver and suggested that zeolites can be used to prevent copper poisoning. Wells and McHugh (1983) fed 10% clinoptilolite to rat's diet. They found that this removed parasites from the intestinal lumen when infected with the nematode Nippostrongylus brasiliensis.
The use of different mycotoxins binders for adsorption was first investigated by in vitro experiments and demonstrated that most mycotoxins were sufficiently bound by at least one adsorbent (Phillips et al., 1988, 1990; Bauer, 1994; Galvano et al., 1997, 1998; Huebner et al., 1999). There is now a broad base of scientific data to verify that the dietary use of zeolites contributes to the improvement of animal health status. Such studies additionally show a secondary effect of improvement in final meat and diary product quality.
Benefits of Yeast, Probiotics and Polymers as mycotoxin binders in experimental studies
Biernasiak et al., (2006) reported that the use of microorganisms as adsorbents is a successful strategy for the management of mycotoxins in animal feeds. He concluded that Saccharomyces cerevisiae and lactic acid bacteria had unique aflatoxin (B1, B2, G1, G2) and ochratoxin A decontaminati0on properties. Saccharomyces cerevisiae has a great ability to bind different forms of mycotoxins that have detrimental effects on animal's health and performance. Based on a number of different studies, it has been suggested that mycotoxin decontamination by Saccharomyces cerevisiae functions as a result of adhesion to the cell wall surface. It has also been reported that dead yeast cell walls have a good functional ability to bind mycotoxins. Mannan components of yeast cell walls bind significant quantities of aflatoxins, ochratoxin A and T-2 toxins (Biernasiak et al., 2006). In addition to the mycotoxin binding capacity of Saccharomyces cerevisiae there is also a nutritional value with essential amino acids, vitamins and minerals all of which are vital for animal health and performance.
Biagi (2009) has stated that researches are becoming more interested in specific strains of lactic acid forming bacteria that are able to bind and biotransform mycotoxins. Niderkorn et al., (2007) identified Lactobacillus and Leuconostoc bacteria that biotransformed zearalenone. It was also found that Streptococcus and Enterococcus strains have the ability to bind deoxynivalenol, zearalenone, and fumonisins. It has been shown, that the strain Lactobacillus rhamnosus can bind aflatoxin B1 in vitro (Haskard et al., 2001; Lahtinen et al., 2004) and in vivo (Gratz et al., 2006). El-Nezami et al., (2002) identified that some strains of Lactobacillus and Propionibacterium bind trichothecenes in vitro. Schatzmayr et al., (2006) demonstrated that a species of Eubacterium has the ability to deactivate trichothecenes.
Synthetic polymers can also be used for the purposes of Mycotoxin management. The Investigation of a synthetic water soluble polymer polyvinlypyrrolidone (PVP) has found that it can eliminate zearalenone and aflatoxin concentrations in vitro (Alegakis et al., 1999). Ramos et al (1996b) suggested that PVP can adsorb zearalenone in vitro at aconcentration of 0.3 mg/g. However Alegakis et al (1999) discovered that an improved cryogel of cross-linked polyvinylpyrrolidone may increase elimination values of zearalenone up to 2.1 mg/g (Alegakis et al. 1999).
Benefits of Antifungal agents in experimental studies
The ability of oregano essential oil to inhibit growth was studied using three mycotoxin causing species of Aspergillus and a species of Penicillium at the Faculty of Veterinary Medicine, Univesity Putra Malaysia. Standard fungi culture concentrations at 104CFU/ml were homogenously inoculated onto SDA agar. Standard sterile discs were impregnated with oregano essential oil and incubated for 5 days at room temperature. The range of zones of inhibition shown below demonstrate that oregano essential oil inhibits the growth of these two fungi species.
Carvacrol and thymol induced >80% inhibition of the biofilm when used at concentrations of 0.06% and showed the strongest anti-biofilm activities against the C. albicans ATCC 3153 strain (Dalleau et al. 2008). (Adam 1998) found that oregano essential oil inhibited growth of three species of human skin fungi Malassezia furfur, Trichophyton rubrum and Trichosporon beigelii. (Dulger 2005) also found that oregano essential oil had antifungal properties tested by inhibition of inoculated agar against the fungi Rhodotorula rubra. Carvacrol and Eugenol which are the main phenolic components of some aromatic plants showed antifungal properties on rat's tongues when challenged with Candida albicans (Chami et al. 2004). Antifungal mycotoxin activity of Oregano Essential Oil against Fusarium verticillioides mycotoxins in vitro was the best of all plant essential oils and in vivo tests on the use of oregano oil on the grains of Zea mays highly inhibited the production of fumonisin B1 (Lopez et al. 2004). A similar experiment was performed on Zea mays by Velluti 2002, who found that antifungal and antimycotoxigenic activity of the essential oils was shown to depend on the environmental conditions of temperature and moisture content.
Mycotoxin binders can work in three ways. They can physically bind toxins and metal ions by adhesion. They can bind toxins by electrostatic charge or cation exchange capacity. They can eliminate the source of the toxins by increasing cell membrane permeability of the fungi, which are the cause of mycotoxin production. There are a range of commercially available mycotoxin binders and antifungal agents that have been shown to have varying potency effects at reducing the presence or eliminating the toxicity of mycotoxins and fungi in animal feed diets, as well as plant animal feed ingredients. The development of a successful commercial mycotoxin binder incorporates the best of each of these active ingredients at the required concentration to ensure that the overall function acts to reduce the harmful effects of mycotoxins in animal nutrition.
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