Last Month's Most Accessed Feature: Mouldy mycotoxins: Imaging distributions in food

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  • Published: Feb 7, 2017
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Food mycotoxins

The distributions of mycotoxins in mould-infested food have been mapped by MALDI mass spectrometry imaging for the first time in a proof-of-principle study that could be extended to living plants.

Many food crops can be infected by fungi and moulds which produce toxic compounds that are dangerous to humans after consumption. These so-called mycotoxins come in different chemical forms with a range of effects, from poisoning the kidneys and liver to causing vomiting and gastrointestinal upsets. Some mycotoxins are also harmful to livestock, so their presence in foodstuffs is to be avoided at all costs.

When a plant is infected by a fungus and produces these toxins, they might be concentrated in one particular part of the plant, such as the fruit or skin.. Alternatively, they might infect the whole plant including the parts which are harvested for foodstuffs. Knowing where the mycotoxins are localised could help in food production by adapting the processing techniques such as surface treatment, milling and sorting, to reduce or eliminate their levels in the final food product.

To help achieve those goals, scientists in Münster, Germany, have applied MALDI mass spectrometry imaging to the analysis of mycotoxins in foodstuffs for the first time. This technique has been used successfully to plot the distributions of drugs and metabolites in human tissue as well as metabolites in plant tissue, but never mycotoxins. Hans-Ulrich Humpf, Sebastian Hickert, Benedikt Cramer and Matthias Letzel from Westfälische Wilhelms-University described their approach in Rapid Communications in Mass Spectrometry.

Mass spectrometry imaging

The researchers inoculated red and white grapes with Aspergillus carbonarius, a fungus known to infect various agricultural products. It is known to produce ochratoxin A, which has been detected in wine and is an especially dangerous mycotoxin as it is a potential carcinogen as well as a genotoxin and teratogen.

They also infected asparagus spears and maize kernels with Fusarium verticilloides which produces fumonisins, the most common mycotoxins found in maize. These toxins are hepatotoxic, nephrotoxic and immunosuppressive in humans in addition to causing birth defects and esophageal cancer.

After suitable incubation periods to allow the moulds to develop, the analysis process was initiated. Since sample preparation for the imaging experiments is somewhat laborious and the actual measurements can take several hours, the presence of mycotoxins in the foodstuffs was first confirmed by tandem mass spectrometry experiments on an LC/MS instrument using electrospray ionisation.

Then for the MALDI MSI experiments, the food samples were embedded in a cellulose medium and cut into very thin sections. These were covered with a suitable MALDI matrix and scanned with a laser to ionise the mycotoxins and produce 2D representations of their distributions across each slice of the plant tissue. These slice images can be studied individually or combined to reconstruct the distribution of the target compounds throughout the sample.

Mycotoxin distribution

The red grapes showed no traces of ochratoxin A after 12 days but fungal growth appeared on the white grapes within two days. The peak areas for the mycotoxin measured by LC/MS grew steadily to maximise after 7 days then remain constant. The imaging experiments showed that ochratoxin A was localised within the area of visible fungal growth and did not extend beyond those boundaries.

For maize, the fungus grew rapidly over a few hours and the fumonisin concentrations increased steadily thereafter. Several different fumonisins and partially hydrolysed fumonisins were identified from their tandem mass spectra. They could be found in different parts of the inner maize kernels and were also present in the outer parts of the kernels but at lower concentrations. They were absent from the germ buds.

The inoculated asparagus spears showed signs of fungal growth within 3 days but the fumonisins were only detectable after 14 days at very low concentrations, about 100-fold lower than in maize, so imaging experiments were dismissed.

This proof-of-principle study has shown that MALDI mass spectrometric imaging can produce maps of mycotoxin distribution within mould-infested food, which might prove useful to food producers who wish to remove the plant parts which have the highest concentrations. The method could also be extended to other types of food and to living plants and might be used to clarify the mechanisms of fungal infection in various plant species.

Related Links

Rapid Communications in Mass Spectrometry 2016, 30, 2508-2516: "Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry imaging of ochratoxin A and fumonisins in mold-infected food"

Article by Steve Down

The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.

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