Chemistry:Aflatoxin M1

Aflatoxin M₁ is a chemical compound of the aflatoxin class, a group of mycotoxins produced by three species of Aspergillus - Aspergillus flavus, Aspergillus parasiticus, and the rare Aspergillus nomius - which contaminate plant and plant products.

Aspergillus flavus produces only B-type aflatoxins. Aflatoxin M₁ is the hydroxylated metabolite of aflatoxin B₁ and can be found in milk or milk products obtained from livestock that have ingested contaminated feed. The carcinogenic potency of aflatoxin M₁ in sensitive species is about one order of magnitude less than that of aflatoxin B₁. Aflatoxin M₁ is usually considered to be a detoxication by-product of aflatoxin B₁. The main sources of aflatoxins in feeds are peanut, meal, maize and cottonseed meal.

Chemical structure

The chemical structure of aflatoxin M₁. Aflatoxin M₁ is the 4-hydroxy derivative of aflatoxin B₁ and is secreted in the milk of mammals that consume aflatoxin B₁. Aflatoxin M₁ has a relative molecular mass of 328 Da and has the molecular formula C₁₇H₁₂O₇.

Schematic presentation summarizing the major contamination and exposure routes and adverse health effects to human for the aflatoxins AFB1 and AFM1

Sources of exposure

Aflatoxin M₁ is found in milk, like human milk. Cows, sheep, goats and buffaloes that have consumed feeds contaminated with aflatoxin B₁, aflatoxin M₁ will be formed as a result of the metabolic process in the livers of ruminants and excreted in milk. In other words, human beings can be exposed to the toxins through consumption of contaminated milk and other foods.

Pathology

The effect of aflatoxin M₁ was much weaker than aflatoxin B₁ in producing liver cancer. The limited animal studies carried out to determine toxicity of aflatoxin M₁. Aflatoxin M₁ has toxic and carcinogenic properties. The toxicity of aflatoxin M₁ in duckling and rats seems to be slightly less than that of aflatoxin B₁. The carcinogenicity is probably one to two orders of magnitude less than that of the highly carcinogenic aflatoxin B₁.

Bioconversion pathway

Aflatoxin M₁ (AFM1), the principal hydroxylated metabolite of AFB1, found in milk (hence the designation M) of mammals fed upon contaminated feedstuff. Carry-over of aflatoxin B₁ (AFB1) as AFM1 in the milk of dairy cows has been established to range from 0.3% to 6.2%. However, AFM1 was also found in lactating mother’s milk . Several studies reported carcinogenic and immunosuppressive effects similar to that of AFB1, on both humans and other animals, even if with a less potent effect. However, AFM1 is the only mycotoxin for which maximum residue limits (MRLs) in milk were established.

Toxicological study

Several studies have been undertaken of the toxic effects of aflatoxin M₁ in laboratory animals. However, in comparison to aflatoxin B₁, relatively little is known about the toxicity of aflatoxin M₁, primarily because of the difficulty in obtaining sufficient quantities of the pure compound necessary for extensive toxicity testing.

Species	Sex	Dose	Result
Trout (fry)up to 12 month	Male	8μg/kg 4-64μg/kg(120 treated/group) 800μg/kg	no tumours(0/110) Liver tumour response:0.086% 50% tumours at 9 months
Fischer rats	Male	0μg/kg 0.5μg/kg 5μg/kg 50μg/kg	Liver cancer 0% neoplastic nodules 0% Liver cancer 0% neoplastic nodules 0% Liver cancer 0% neoplastic nodules 0% Liver cancer 5% neoplastic nodules 16%
Fischer rats	Male	1 mg/rat	Liver cancer 1 of 29

Genotoxicity

The potency of aflatoxin B₁ and aflatoxin M₁ in inducing DNA damage and genotoxicity was tested in Drosophila melanogaster. Aflatoxin M₁ was found to be a DNA-damaging agent, with an activity about one-third that of aflatoxin B₁.

Analytical methods

Many methods have become available for the determination of aflatoxin M₁ in milk. In particular, solid-phase correction and immunoaffinity chromatography cartridges offer good possibilities for efficient clean up. Both thin-layer chromatography (TLC) and high performance liquid chromatography (HPLC) are adequate techniques to separate and determine aflatoxin M₁ in extracts of milk. Enzyme-linked immune sorbent assay (ELISA) is more popular, due to ease of use and properties which are conducive to rapid screening and semi-quantitative determination. Aflatoxins determinations are usually expensive and employ environmentally unfriendly procedures, thus, the search for new materials and technologies, that are both ecologically safe and inexpensive its rice husk as an adsorbent method.

Regulation

According to a recent review conducted by the Dutch National Institute for Public Health and the Environment (RIVM) on behalf of the Food and Agriculture Organization (FAO) approximately 60 countries have set specific limits for aflatoxin M₁. The European Union countries generally apply a maximum level of 0.05 μg/kg milk. Some countries in Africa, Asia and Latin America also propose this level. In contrast, the USA as well as some European and several Asian countries accept a maximum level of 0.5 μg/kg aflatoxin M₁ in milk, which is also the harmonized MERCOSUR limit applied in Latin America. The 0.5 μg/kg limit for aflatoxin M₁. Thus, the maximum permitted level of aflatoxin M₁ in milk in the EU is among the lowest in the world, and is based on the ALARA (As Low As Reasonably Achievable) principle. Considering the carry-over into milk and the established adverse effects on animal health, approximately 45 countries have set specific levels for aflatoxin B₁ in feed for dairy animals. To support compliance with the maximum levels in milk intended for human consumption, stringent maximum levels were also set in the EU for feed stuffs which might be consumed by dairy cows. A limit of 0.005 mg/kg feed for dairy cattle is applied in the EU countries and in the new member states as well as in Europe Union countries, but only in few countries outside Europe. This level is below the no-effect level in target animals.