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Analyzing Mycotoxins in Food with Liquid Chromatography

Mycotoxins are harmful secondary metabolites produced by certain fungi, which affect a broad range of crops and food goods. Hundreds of these toxic compounds have been discovered and characterized to date – although their toxic effects vary dramatically. 

Around a dozen distinct mycotoxins have been linked to severe long-term health effects where they have propagated through the food chain due to crop or feedstock contamination. Various analytical techniques, including liquid chromatography, have proven instrumental in characterizing these harmful mycotoxins and determining their presence and concentration in food goods.

Outlining Harmful Mycotoxins

Aflatoxins, Alternaria toxins, and zearalenone are three species of mycotoxin that have garnered enormous interest from the worldwide scientific and regulatory communities. This is primarily due to their severe adverse health effects for both humans and livestock; from acute poisoning shortly after exposure to long-term conditions like cancer and immune deficiency. 

Alongside these health concerns, mycotoxins also represent significant agro-economic challenges. Molds that produce mycotoxins can grow on food products at practically any point before and after harvest. Factors that influence mold growth and subsequent toxin development include extrinsic aspects like growth climate and storage conditions. However, complex intrinsic factors also contribute to mold growth, including fungal strain specificity, strain variation, and instability of toxigenic properties1 .

The optimal method for extracting and analyzing mycotoxins from food goods and crops involves solid-phase extraction (SPE) via high-performance liquid chromatography (HPLC) and fluorescence detection.

Detecting Mycotoxins with Liquid Chromatography

Liquid chromatography with fluorescence detection is diagnosed for the analysis of fungal metabolites like aflatoxins. These common mycotoxins are produced primarily by the Aspergillus species of fungi but have shown extreme resilience, contaminating food goods even after mold has been eliminated. The Food and Drug Administration (FDA) has established a series of action levels for aflatoxin levels in grains and cereals for various applications, with safe concentration levels for human consumption ranging from 0.5 – 20 micrograms per kilogram (μg/kg).
The underlying challenge in analyzing aflatoxins with fluorescence-enabled liquid chromatography is the complex photochemical behavior of the four distinct aflatoxins B1, B2, G1, and G2. This makes a high-throughput analysis of aflatoxins and other relevant mycotoxins via liquid chromatography intensely difficult. 

Similar challenges exist in the determination of Alternaria toxins via liquid chromatography. These are hazardous compounds produced by a widespread fungal species known as Alternaria alternata which primarily grows on cereals and seeds. It has been found on crops as varied as wheat, rice, and even tobacco. This group of mycotoxins exhibits both acute and chronic health effects. Tenuazonic acid (TeA) is a well-characterized Alternaria mycotoxin known to block the synthesis of proteins responsible for antibacterial, antitumor, and antiviral activity. Unlike aflatoxins, however, no regulatory standards currently exist for Alternaria concentration in food goods and feedstocks.

The unique complexities in mycotoxin analysis highlight the need for distinct SPE methodologies when using liquid chromatography and fluorescence detection. For example, UVE photochemical reactors are used to compensate for the low inherent fluorescence of aflatoxins B1 and G1. After the sample has eluted from the liquid chromatography column it is transferred to the photochemical reactor, where UV-light with approximate peak wavelengths of 254 nanometers (nm) derivatizes aflatoxins B1 and G1, causing them to undergo photo-induced hydroxylation. They can subsequently be analyzed based on fluorescence with comparative ease.

The complex taxonomy of Alternaria toxins can also be unraveled with the help of ultraviolet light. UV detection coupled with fluorescence detection in an isocratic liquid chromatography system has enabled the characterization of myriad mycotoxins in limited sample volumes and complex matrices.

Liquid Chromatography Solutions from Knauer

Knauer has a storied history of overcoming complex analytical challenges in the field of high-performance liquid chromatography. We can help you overcome the challenges in your food analysis processes. Simply contact a member of the team today to find out more.

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Sources:
[1] https://www.sciencedirect.com/science/article/pii/S1319610310000827

More about mycotoxins

Mycotoxin HPLC applications

AZURA HPLC system for sensitive aflatoxin determination

With post-column photochemical derivatization and fluorescence detection

The AZURA® Aflatoxin analysis system is specially designed to determine Aflatoxin B1, B2, G1, and G2 in food and feed products such as peanuts, corn and cottonseed. It utilizes efficient post-column photochemical derivatization to improve fluorescence sensitivity for aflatoxins B1 and G1 drastically.

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