Mycotoxins

Mycotoxins are chemical substances naturally produced by certain molds in foods and animal feed. These compounds can be found particularly in cereals, dried fruits, nuts, and spices. The formation of mycotoxins requires suitable temperature and humidity conditions. Mycotoxins are regarded as undesirable contaminants in food and are monitored for food safety purposes.

Main Types of Mycotoxins

The most common and significant types of mycotoxins include:

Aflatoxins: Toxic compounds produced by the species Aspergillus flavus and Aspergillus parasiticus. They can be found in peanuts, hazelnuts, dried fruits, spices, and cereals. Aflatoxin B1 is the most toxic and exhibits genotoxic properties.

Ochratoxin A: Produced by species of Aspergillus and Penicillium. It is detected in cereals, raisins, coffee, cocoa, wine, and spices.

Fusarium Toxins: Mycotoxins produced by fungi of the genus Fusarium. Deoxynivalenol (DON), T-2 and HT-2 toxins, fumonisins, and zearalenone belong to this group. They are commonly found in cereals grown in temperate climates.

Patulin: A mycotoxin primarily formed in apples and apple products, as well as in moldy fruits and cereals.

These mycotoxin types are monitored for food safety and controlled using various analytical methods.

Formation Conditions

Mycotoxins arise as a result of the metabolic activity of specific mold species under favorable environmental conditions. Temperature, humidity, and water activity play critical roles in this process. The optimal temperature range for the growth of Aspergillus species is generally between 25–40 °C, while mycotoxin production requires high humidity and a water activity level above 0.65. Mycotoxin formation in agricultural products increases under unsuitable temperature and humidity conditions during pre-harvest or post-harvest stages, particularly during storage. For example, contamination with mycotoxins is more intense in dried figs during the overripening phase. Controlling humidity, temperature, and packaging conditions during storage is essential to reduce mycotoxin risk.

Analytical Methods

Various analytical methods are used to detect mycotoxins in food and feed. These methods ensure reliable results and compliance with national and international standards. One of the most commonly used techniques is High-Performance Liquid Chromatography (HPLC), which provides high sensitivity for the quantitative determination of mycotoxins. For more advanced analyses, Liquid Chromatography–Tandem Mass Spectrometry (LC-MS/MS) is preferred, as it can accurately detect mycotoxins even at low concentrations. Enzyme-Linked Immunosorbent Assay (ELISA) is a rapid and widely used method, especially for screening tests. In products such as dried figs, the Ultraviolet Fluorescence Test (BGYF) is also applied. In this method, figs are examined under UV light; products exhibiting bright greenish-yellow fluorescence are considered contaminated and removed from the batch. In all analyses, recovery rates and measurement uncertainties are evaluated according to international standards such as AOAC and TS EN ISO/IEC 17043, ensuring the reliability and traceability of results.

Main Analytical Techniques

Analytical techniques used for the detection of mycotoxins in food and feed have been developed based on national and international standards to ensure reliable results. These techniques enable accurate determination of mycotoxins even at low levels.

Chromatographic Techniques

Chromatography-based techniques are among the most widely used methods in mycotoxin analysis. High-Performance Liquid Chromatography (HPLC) and Liquid Chromatography–Tandem Mass Spectrometry (LC-MS/MS) are employed for the separation, identification, and quantitative measurement of mycotoxins. These techniques provide reliability and sensitivity, especially in multi-mycotoxin analyses.

Immunological Techniques

Among immunological techniques, Enzyme-Linked Immunosorbent Assay (ELISA) stands out. This method is preferred for screening analyses due to its practicality and rapid results. ELISA tests can be effectively used for the analysis of large numbers of samples.

Fluorescence-Based Techniques

The Ultraviolet Fluorescence Test (BGYF) is a screening method used specifically for detecting aflatoxins in dried figs. In this technique, fig fruits are examined under UV light at a wavelength of 360 nm, and those exhibiting bright greenish-yellow fluorescence are identified as contaminated.

Competence and Standards

These analytical techniques are evaluated according to international standards AOAC and TS EN ISO/IEC 17043 with regard to recovery rates, measurement uncertainties, and accuracy criteria. This ensures the reliability and traceability of mycotoxin analyses.

Reliability and Laboratory Standards

In mycotoxin analysis, reliability is of great importance and depends on the accuracy, reproducibility, and conformity of results with internationally accepted standards. Therefore, laboratories operate according to various quality management systems and standards.

International Standards

Mycotoxin analyses conducted in Türkiye are based on internationally recognized standards. In this context, proficiency testing programs compliant with the TS EN ISO/IEC 17043 standard are implemented. This standard defines criteria for the planning, execution, and evaluation of proficiency tests, ensuring that laboratory results are comparable and reliable.

Proficiency Testing

Since 2013, the National Food Reference Laboratory (NFRL) has organized proficiency testing cycles for mycotoxin analyses. These tests involve participation from both public and private laboratories. Proficiency testing helps identify and address potential challenges encountered during analysis and ensures the traceability of laboratory performance at national and international levels.

Sampling and Homogenization

Due to the uneven distribution of mycotoxins in food and feed, appropriate sampling and homogenization procedures are essential for reliable analytical results. In accordance with relevant notifications of the Turkish Food Codex, numerous small samples are collected to represent the batch, combined into a composite sample, ground, homogenized, and prepared for analysis. Reference samples are also separated using the same method.

Measurement Uncertainty and Recovery

The reliability of analytical results is ensured by evaluating measurement uncertainty and recovery rates in accordance with established standards. In this context, analytical results are assessed for compliance with regulatory requirements according to AOAC and European Union guidelines.

Audits and Scope Expansion

Mycotoxin analyses are subject to regular audits to ensure food safety and maintain compliance with international trade standards. These audits aim to evaluate the accuracy of methods applied by laboratories and their conformity with international standards.

The Mycotoxin Section within the National Food Reference Laboratory (NFRL) has been organizing proficiency testing cycles in accordance with the TS EN ISO/IEC 17043 standard since 2013. These tests are conducted to measure the performance of both public and private laboratories and to ensure the reliability of results. Additionally, laboratories are subject to detailed evaluations when requesting scope expansion.

In the scope expansion process, the essential criterion is the assessment of laboratories’ technical and scientific competence to implement new analytical methods or updates to existing ones. This process is supported by reviewing submitted scope expansion documentation and conducting on-site inspections. Thus, laboratories are enabled to provide reliable results not only for their current analyses but also for new methods and parameters.

During audits, compliance of sampling and homogenization procedures with regulations is also reviewed. Due to the uneven distribution of mycotoxins in food and feed, homogenization of composite samples prepared by appropriate methods is of critical importance. These steps are carried out in accordance with the “Notification on Criteria for Sampling, Sample Preparation and Analytical Methods for Official Control of Mycotoxin Levels in Food.”

Effects on Public Health

The effects of mycotoxins on human health are characterized by carcinogenic, mutagenic, nephrotoxic, and immunosuppressive properties.

Carcinogenic Effect: Aflatoxin B1, one of the most potent mycotoxins, forms adducts with DNA molecules. These adducts cause damage to genetic material and increase the risk of liver cancer.

Mutagenic Effect: Mycotoxins can cause permanent changes in genetic material, leading to DNA damage and mutations during cell division.

Nephrotoxic Effect: Ochratoxin A can adversely affect kidney function and lead to chronic health problems.

Immunosuppressive Effect: Some mycotoxins suppress the immune system, making humans and animals more susceptible to infections.

These effects demonstrate that mycotoxins represent a significant food safety issue for public health at the international level.

Economic Impacts

Mycotoxin contamination is of major importance not only from a health perspective but also economically. Türkiye is among the leading global producers of dried figs, hazelnuts, and raisins; therefore, mycotoxin contamination in these products is a critical issue for international trade. The European Union has set a maximum total aflatoxin limit of 10 ppb in dried figs. Exceeding this limit results in the prohibition of exports, causing serious economic losses for producers and exporters.

Under official inspections, contaminated products are identified and prevented from entering the food chain. For example, aflatoxin contamination in dried figs is detected using the UV Fluorescence Test (BGYF), and contaminated products are destroyed in accordance with regulations of the Ministry of Environment and Urbanization. These practices aim to protect public health and maintain the reliability of export markets.

Mechanism of Aflatoxin B1 Formation of DNA Adducts

Aflatoxin B1 is metabolized in the human body into a reactive epoxide form. This epoxide forms covalent bonds with guanine bases in DNA, creating DNA adducts. These adducts cause structural damage to genetic material and induce mutations. This process creates the foundation for uncontrolled cell proliferation and the initiation of carcinogenic mechanisms.

Epidemiological Data

Epidemiological studies have shown that high exposure to aflatoxin B1 significantly increases the risk of hepatocellular carcinoma (liver cancer). These studies reveal that chronic exposure to aflatoxin B1 in regions where mycotoxins are prevalent elevates cancer-related mortality rates.

Prevention Strategies

Preventing mycotoxin contamination is a fundamental requirement for ensuring food safety and protecting public health. Prevention strategies encompass many stages, from agricultural production to storage and marketing.

First, appropriate harvesting techniques and timely harvest are crucial. Since mycotoxin contamination intensifies during overripeness (e.g., the overripening phase in dried figs), timely collection of products is necessary. Controlling moisture content during post-harvest drying reduces mycotoxin formation.

During storage, humidity and temperature levels play a critical role. To prevent mycotoxin formation, humidity should not exceed 24% and water activity should remain at or below 0.65. Additionally, safe storage conditions can be maintained for up to one year by storing products in cold storage at +4 °C. Low moisture permeability of packaging materials is also an important preventive factor.

Among control methods, the UV Fluorescence Test (BGYF) is prominent. In this method, samples are taken from incoming batches of dried figs and examined under a 360 nm UV lamp. Fruits exhibiting bright greenish-yellow fluorescence are considered contaminated and removed from the batch. Contaminated products are destroyed in accordance with regulations to prevent them from entering the food chain. This process is carried out by incineration in cement plants, in compliance with regulations of the Ministry of Environment and Urbanization.

Additionally, under notifications issued by the Ministry of Agriculture and Forestry, sampling, preparation, and analytical methods have been standardized. In this context, proficiency testing programs compliant with TS EN ISO/IEC 17043 are implemented, and food safety is systematically monitored.

Effective implementation of prevention strategies contributes to protecting public health and preventing potential economic losses in international trade.