|Year : 2018 | Volume
| Issue : 2 | Page : 94-99
Useful approaches for reducing aflatoxin M1 content in milk and dairy products
Fatemeh Naeimipour1, Jafar Aghajani2, Saba Amiri Kojuri3, Saman Ayoubi4
1 Department of Mycology, Faculty of Veterinary Medicine, Science and Research Branch, Islamic Azad University, Tehran, India
2 Young Researchers and Elite Club, Lahijan Branch, Islamic Azad University, Lahijan, Iran
3 Young Researchers and Elite club, Chalus branch, Islamic Azad University, Chalus, Iran
4 Young Researchers and Elite Club, Lahijan Branch, Islamic Azad University, Lahijan; Inflammatory Lung Disease Research Center, School of Medicine, Razi Hospital, Guilan University of Medical Sciences, Rasht, Iran
|Date of Web Publication||14-Jun-2018|
Dr. Saman Ayoubi
Young Researchers and Elite Club, Lahijan Branch, Islamic Azad University, Lahijan; lnflammatory Lung Disease Research Center, School of Medicine, Razi Hospital, Guilan University of Medical Sciences, Rasht
Source of Support: None, Conflict of Interest: None
The quality and safety of food are of major importance. Using contaminated animals' milk and meat may result in human disease. Among microorganisms, fungal toxins, especially aflatoxin B-1 (AFB1), are of special importance. Aflatoxin M-1 (AFM-1) is a metabolite that is produced by conversion and hydroxylation of AFB-1. Both toxins can cause acute and chronic mycotoxicosis mainly through ingestion of contaminated milk. Hence, it is critical to control and decrease these microorganisms. Despite cost-effective efforts, preventing foods contamination due to aflatoxins (AFs) is not only an expensive but also a difficult task. The best agricultural monitoring during preharvest and postharvest stages cannot eliminate the AFs, especially AFM-1 from milk and dairy products because of the high resistance of these toxins. There have been numerous studies investigating the methods of AF detoxification or reduction from infected milk. By focusing on advantages and disadvantages of preventative procedures using probiotics, antibodies, chemisorbents and even additives, one can choose one or several procedures to eliminate or reduce AFM-1 in milk and its byproducts efficiently.
Keywords: Aflatoxin B1, aflatoxin M1, milk, reduction methods
|How to cite this article:|
Naeimipour F, Aghajani J, Kojuri SA, Ayoubi S. Useful approaches for reducing aflatoxin M1 content in milk and dairy products. Biomed Biotechnol Res J 2018;2:94-9
|How to cite this URL:|
Naeimipour F, Aghajani J, Kojuri SA, Ayoubi S. Useful approaches for reducing aflatoxin M1 content in milk and dairy products. Biomed Biotechnol Res J [serial online] 2018 [cited 2019 Sep 22];2:94-9. Available from: http://www.bmbtrj.org/text.asp?2018/2/2/94/234464
| Introduction|| |
Aflatoxins (AFs) are secondary fungal metabolites that are naturally produced by some Aspergillus species especially Aspergillus flavus and Aspergillus parasiticus. Dairy products have favorable conditions to be contaminated by these molds at the stages of growth, processing, storage, and transportation., AFs are the most carcinogenic compounds. AF B1 (AFB-1) is an important fungal metabolite with a high potential to induce mutation and immunosuppression. In the liver, cytochrome P450 system converts AFB-1 into AF M-1 (AFM-1)., [Figure 1] shows pathways involved in metabolism and elimination of AFs in the liver [Figure 1].,,,,
|Figure 1: Pathways that are involved in metabolism and elimination of aflatoxins in the liver|
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The presence of AFM-1 in milk and dairy products is considered to pose certain hygienic risks for mammals especially human. Level of AFM-1 in infected milk or contaminated foodstuffs is directly associated with the levels of AFB-1. To decrease or detoxify AF from infected milk, some studies have suggested the usage of an absorbent such as soil-bentonite. It is shown that such components can reduce the levels of AFs efficiently. On the other hand, a majority of studies suggested using of probiotics., Probiotics are inexpensive and safe to decrease or detoxify AF in milk and advantages of using probiotics have been demonstrated in several researches. Considering to importance of preventive approaches to decontaminate food, the point is that how many and in which condition, the mentioned methods are allowed to be used.,,,
In this study, different methods to decrease or detoxify AFM-1 in milk and dairy products are briefly reviewed and the most effective approaches are highlighted.
| Methods for Reducing the Aflatoxin M-1 Content in Milk|| |
The levels of AFM-1 in milk and dairy products of different countries are variable. It can be related to climate condition of each geographical area and differences in dairy cattle feeding system., Therefore, to decrease the level of AFM-1 in milk, AFs in both animal feed and milk should be evaluated continuously., In the last decades, by understanding the serious effects of mycotoxins on human and animals health and to protect human health, food companies in different countries attempted to adjust content of mycotoxins in food and feed., [Table 1] shows the accepted level of the toxin in some countries description of [Table 1].
|Table 1: Maximum acceptable levels of aflatoxin M-1 in milk and milk products in some countries|
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Several methods have been introduced to reduce AFs in milk and dairy products. Some studies showed that preventing the crops from contamination in preharvest and postharvest stages is a useful strategy to eliminate fungal toxins. Other studies proposed direct methods for reducing AFM-1 in milk.
Approaches to prevent milk contamination
It is believed that prevention and control of fungal growth are an important technique to reduce AFM-1 content in milk. There is a linear correlation between AFB-1 in animals feed and quantity of AFB-1 in their milk. To prevent contamination of milk, dairy animals should be vaccinated against AFs especially AFB-1. Some studies showed that vaccination might reduce the risk of contamination of animal/dairy products by AFs. The levels of anti-AFB-1 antibodies in vaccinated heifers decrease during pregnancy and after calving and at the beginning of the milk production cycle, they return to the previous range. Such changes in levels of anti-AFB-1 antibodies after vaccination may demonstrate the effectiveness of such strategy., Another valuable method to control AFB-1 and consequently AFM-1 is to inhibit the initial contamination of feed consumed by Dairy Cattles. In a research, it was emphasized that the usage of damaged portion of crops should be prohibited and such crops must be removed during the harvesting process. Moreover, place, temperature, and moisture of storages should be controlled since they can induce the fungal growth in storehouses.
It is reported that the inhibition of contamination through livestock feed control is very important. The quality of livestock feed has an impact on the AFs content in the produced milk. During production of crop in the farm and storage of products in traditional or industrial tanks, the monitoring instruments and health considerations need to be controlled.
An in vitro investigation proved that specific anti-AFB-1 antisera could reduce the amount of AFM-1 in milk. Another research showed that using antifungal agents should be done before the formation of the toxin. Organic acids are the main inhibitors and include acetic, benzoic, formic and lactic acids. To prevent the formation of AFs, performing biological control and using absorbent clays are very useful. These methods are been reviewed in the next paragraphs of the current study.
Overall, considering above-mentioned evidences, the preventative approaches to inhibit entrance of AFM-1 into milk are essential. It should be noted that controlling can reduce toxin but cannot remove it completely. In addition, not all of the prophylaxis methods can be used in farm or factories. Therefore, some other methods for mitigation of AFM-1 in milk should be utilized.
Reducing of aflatoxin M-1 content during processing of milk
Thermal treatment during processing stages is not considered as a magnificent way to reduce AFM-1 in milk. Although pasteurization and sterilization may eliminate many microorganisms from milk, they are not effective for mitigating AFs. Some researchers believe that heat does not cause significant changes in amount of AFM-1, while others have reported different levels of AFs detoxification through these techniques. For instance, in a study, it was shown that pasteurization at 62°C for 30 min could reduce the AFM-1 content in milk by 32%. Another study showed that heating might decrease AFs-content by 12%–35% (depending on the conditions). It is also reported that AFM-1 may be relatively stable during milk pasteurization process. Processing stages of milk may remove other dangerous contaminants from milk, but it cannot effectively remove AFs from infected milk.
Role of probiotics to reduce aflatoxin M-1
The meaning of probiotic is “for life,” but in biology, the term is used to call microorganisms that are cost-effective for human and animals. Probiotics have been considered to have several advantages as follows: they can potentially decrease populations of gastrointestinal pathogens, lessen the gastrointestinal pain, support the immune system, affect and improve the skin's function, regulate the intestine and reduce flatulence and bloating, induce the resistance to pollen allergens, protect DNA, proteins and lipids from oxidative damages, and maintain the normal flora of intestine while receiving antibiotics treatment.
Probiotics can be used to lessen or detoxify toxins in milk, dairy products, and other foods. Such useful microorganisms can be added to cattle feed or may be utilized at the different stages of milk processing.
Probiotics for biological control in the field
Growth of yeasts and molds can cause decay in crops and animal food (feed). Contamination of crops may lead to considerable economic damages. Probiotics as biocontrol agents may absorb toxins, compete with fungi for an ecological niche or nutrient, or destroy AF. Therefore, they can prevent fungal growth and toxin secretion in feed.,
The most important bacteria used as probiotics belong to bacterial genera including Lactobacillus, Streptococcus, Pseudomonas, Bifidobacteria and Burkholderia. The commercial strains can reduce AFM-1 contents in phosphate-buffered saline, milk, and yogurt. Some yeast such as, Saccharomyces species and nontoxigenic Aspergillus strains, have been used as competitive biocontrol agents.
Probiotics as biocontrol agents during storage of food
Probiotics have an important role in biocontrol of the storage houses. They have the ability to impair the growth of fungi and reduce AFs content through the above mentioned mechanisms. Below, some studies with emphasizing the effectiveness of probiotics are reviewed.
In a research, Saccharomyces Kefir and Lactobacillus casei were used to reduce AFM-1 in milk. The results revealed the positive significant role of these microorganisms to eliminate high level of the toxin in milk. In 2011, the role of lactic bacteria such as Lactobacillus bulgaricus and Streptococcus thermophilus for removal of AFM-1 in dairy products during manufacturing was investigated. It is found that L. bulgaricus has more binding ability than S. thermophilus. Therefore, L. bulgaricus can reduce the amount of toxin. In another research, the beneficial effects of specific dairy strains of lactobacilli to remove AFM-1 from milk were highlighted. The role of probiotics to improve liver functions as well as their positive effects on human health particularly children and the high-risk population were also emphasized.
It is shown that the attachment of AFM-1 to cell wall components of probiotic microorganisms may results in the reduction of the toxin in milk. The enzymatic degradation of AFM-1 by Lactobacillus, Pseudomonas, and Burkholderia species can also inhibit fungal growth and production of Afs. The Saccharomyces cerevisiae is one of the yeasts that can bind to AFM-1 effectively. Among bacteria, Lactobacillus rhamnosus L60 and Lactobacillus fermentum L23 have a high ability to inhibit mycelium growth of aflatoxigenic Aspergillus strains and reduce the AFB-1 production.
All of these researches show the significant roles of probiotics. The culture of lactic bacteria is an easy task. Consequently, as these microorganisms are safe, inexpensive, and have high potential ability for detoxification, they are valuable candidates for reducing AFM-1 in milk and dairy products.
Role of adsorption in reducing the aflatoxin M-1 content in milk
Toxicants enter into an organ through an adsorption process. For reducing AFs in the gastrointestinal tract and therefore diminish the rate of the toxin distribution in the tissues, adsorbing agents can be added to animal feed or use separately during their meal times. The absorbents can reduce AFs contents in milk because of their high variety and ability to bind to AFM-1 in a stable mode. Such compounds may be affected by pH and temperature. The bentonite, vermiculite, hydrated sodium calcium aluminum silicate (HSCAS), and activated carbon are the well-known absorbents.
Bentonite clay works such as a magnet that bonds to toxins and helps to the excretion from the body. Sodium-bentonite and calcium-bentonite are important types of bentonite clay. When activated by water, sodium-bentonite has an electromagnetic properties and can swell up six times in the size. This makes it very absorbent and helpful for drawing out toxins. Calcium-bentonite has smaller than sodium-bentonite in size. Their small size makes them to be more effective for remineralizing. These small particles can pass through the colon wall into the bloodstream where they get rid of toxins and leaves minerals behind.
Bentonite is an aluminum phyllosilicate clay. The negative charges of the molecule enhance its adsorptive properties. The absorbent is able to pass through the intestinal tract and attract the positively charged toxins. The molecule is bound to the toxin by ions exchange resulting in the internalizing and absorbance of the toxic molecule to the clay molecule. Subsequently, the host body can excrete the toxin-filled clay through normal gut movements.,,
Bentonite is capable for maximum detoxification and has a low impact on the nutritional properties of milk. While producing pelleted feeds, the clay is used to bind and flush AFM-1 and bring the level of AFM-1 below the European standard limits for the toxin. Enrichment the feed by sodium bentonite and activated carbon can reduce AFM-1 content in dairy products of early lactating goats. In a research conducted in Iran, it was shown that the bentonite can reduce AFM-1 in milk and adding the clay entrosorbent in the diet of lactating dairy cattle and goats can reduce the concentration of AFM-1 significantly.
The hydrated HSCAS is a phyllosilicate clay with high effectiveness to prevent aflatoxicosis in chickens, goats, cows, and pigs. It has high affinity and capacity to absorb AFB-1 in contaminated milk and oils.
Using the calcium hydroxide and mono methylamine simultaneously can result in 94%–100% destruction of AFB-1. These substances can open the lactone ring. In a study, the calcium hydroxide was added to the dry diet at a concentration of 2%. Then, mono methylamine was added at a concentration of 0.5% and treatment was performed at 25°C and 10% of moisture in 4 h. As a result, the mycotoxins content was reduced by 50%.
Activated carbon is manufactured by treatment of carbonaceous source materials such as coal, coconuts, nutshells, peat, and wood and can be applied to purify liquids and gases. Activated carbons and esterified glucomannan have been shown to reduce the AFM-1 content in infected milk. It is reported that supplementation of early lactating goats by sodium bentonite and activated carbon (by 1%) can results in significant reduction of AFM-1 in milk without changing the milk composition. In a report, a mixture of activated carbon and HSCAS was added to AFB-1 contaminated feed of dairy cows (with an inclusion rate of 2%). Activated carbon and HSCAS reduced AFB-1 converting to AFM-1 in milk by 50% and 36%, respectively.
Chemical approaches to reduce aflatoxin M-1
Chemical processes can be used to reduce, destruct, or inactivate AF in milk. They include ammoniating, acidic treatment, oxidizing, and reducing techniques. There are several issues and risks associated with these methods. First, it is difficult to detoxify the AF without reducing nutritive value and palatability. Moreover, a number of parameters such as reaction time, temperature, and moisture must be monitored. Some necessary additional cleaning treatments are expensive and time-consuming and toxic byproducts may be produced. [Figure 2] compares effectiveness of some compounds to reduce AF content in milk [Figure 2].
|Figure 2: Effectiveness of some compounds to reduce aflatoxin content in milk|
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Ammonia and hydrogen peroxide
A high number of chemicals, such as acids, bases and bisulfite oxidizing agents, have been tested for their ability to degrade or inactive AFs. Ammonia has been used to destroy AF. Ammoniation is a reaction in which ammonia is added in stoichiometric proportions of an interested compound. Ammoniation can mitigate the AFM-1 content in contaminated foods by 79%–90%. Ammonia reacts with AFs molecule by breaking oxygen bond. It can also open lactone ring of the toxin to produce compounds with lower toxicity.
Hydrogen peroxide (H2O2) is a colorless, unstable, and oily liquid and is usually used in the aqueous solution. It is a powerful oxidizing agent used as bleach for textiles in rocket fuels. Using of H2O2 at different temperatures has been suggested to detoxify AFs in milk. An interesting report showed that using H2O2 followed by a heat treatment at 36°C for 30 min and boiling for 5 min could inactive AFs by 27.8%. Utilizing H2O2 followed by heat treatment at 75°C for 15 s and boiling for 5 min inactivated AFs by 28.8%. Another study showed that the addition of H2O2 or sodium hypochlorite in diet and storage at room temperature for 7 days can destruct 62% and 82% of AFB-1 content, respectively. Combined treatments such as ultra violet radiation followed by ultra-filtration can degrade AFM-1.
Using plant extracts to reduce aflatoxin M-1 in milk
One interesting technic to diminish toxins, especially AFM-1 in foods, is using the plant extracts. Some plants such as broccoli, garlic, black cumin, and curcumin have been reported to be efficient for reduction of AFs. The mentioned extracts may also have inhibitory effect on AFs. The extracts are also known as antioxidant, antifungal, and anti-inflammatory agents because their ability to inhibit infection or inflammation inducing molecules.
Broccoli, garlic, and black cumin for reducing aflatoxin M-1
Broccoli extract is an attractive novel additive with indirect antioxidant properties. In a study, it was shown that the broccoli extract has the ability to reduce the AFM-1 content in milk. This is important since milk is consumed by humans especially children.
Water extracts of garlic, black cumin, and carrot were attempted to use for detoxification of AFM-1. Ayoub et al. showed that the level of AFM-1 was significantly reduced by adding the mentioned extracts to milk in ratio of 1:10. There are limited data about ability of such extracts to reduce the toxin in milk, but above mentioned finding may reflect the advantages of plant extracts to detoxify AFs in dairy products during fermentation stages.
In a review, the effect of plant extracts including curcumin and Indian turmeric on AFM-1 in milk was reviewed. The results showed that such extracts can have anti-inflammatory, anticarcinogenic, antimicrobial, and gastrointestinal effects. Among the extracts, curcumin was able to enhance immunity. Considering the role of herbs against aflatoxicosis, plants like coumarins, flavonoid and curcuminoids have been reported to inhibit biotransformation of AFs to their active epoxide derivatives. Using 0.05%, 1.0%, and 2.0% of turmeric concentration, the level of AF were decreased significantly. In addition, turmeric could inhibit spore count and AFs production by A. flavus.
Antioxidants and aflatoxins reduction
Antioxidants can aid human body to detoxify toxins in liver and other cells, and consequently decrease the appearance of mycotoxicosis. Antioxidants, such as Vitamin C and Vitamin E, can be added in food to prevent chemical reactions in which oxygen is combined with other substances. Such reactions are harmful for human cells. Vitamin E, Vitamin C and selenium are involved in the formation of glutathione peroxidase that is vital for detoxification mechanisms in cells. using glutathione, glutathione peroxidase reduces lipid hydroperoxides to alcohols and free H2O2 to water. Glutathione is a very simple molecule that is produced naturally in human body. The molecule can attach to free radicals and heavy metals through its sulfur group.
| Conclusion|| |
Mycotoxins are secondary metabolite of fungi such as A. flavus, Aspergillus fumigatus and Penicillium spp and can cause mycotoxicosis. AF is a carcinogenic mycotoxin. Milk contamination with AFs is a big problem since milk is an essential nutriment to keep humans healthy and strong. Milk is a rich source of calcium, phosphorus, essential amino acids and vitamins.
Some technologies are available to prevent fungal growth during pre- and post-harvest times and in storing of crops. However, these methods are not accessible in undeveloped or under developing countries. There are some approaches to detoxify AF in milk (in both preharvest and postharvest times) including using probiotics, absorbent chemicals, and plant extracts. Yet, these methods have their pose and cons. Therefore, while using one or a combination of these approaches, their ability to diminish or degrade the toxin as well as their effects on the nutritional properties of milk should be considered.
Prevention of food contamination is very importan; however, it could not eliminate the toxin completely. After performing prevention controls such as prevent fungi growth in storage, vaccination of lactating mammals, and combination of dairy animals food with absorbents, some toxin may remain in row dairy products. Absorbents such as bentonite and activated carbon can reduce the high level of toxin, but they are expensive and may reduce the proteins of milk. Hence, some other treatments such as using processing stages to prepare pasteurized milk are performed. Several studies have shown that pasteurization could not efficiently reduce AFs content in milk since a high number of mycotoxins are resistant to thermal inactivation.
Probiotics play an important role in reduction of AFM-1 in milk. Besides the ability to reduce AFM-1, they are inexpensive and can be used in fermentation stages. It is known that these little microorganisms are very useful for the gastrointestinal tract, but it should be noted that some strains may produce AFs when they have been used as probiotics. In the case of using plant extracts as powerful detoxification agents, more studies are needed. In sum, each of the above-mentioned methods can be used based on their ability to reduce the toxin, their benefits for health and financial conditions.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2]