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Testing & AnalysisChemicalMethods

Using APGC to Improve Food Contaminant Monitoring

By Tania Portolés
December 14, 2018

Chemical contaminants may occur in our food from various sources. This is a result of various stages of production, processing, transport, or the environment. As these chemicals can be harmful to human health at higher concentrations, the use of many contaminants is limited by the European Union. Therefore, analyses of these contaminants in food samples are essential to ensure consumer safety and compliance with regulatory limits.

Environmental Contaminants
Contaminants are either man-made or naturally occurring substances that are present in the environment and bioaccumulate in the food chain. Examples of contaminants that enter the food chain include brominated flame retardants (BFRs), polychlorinated biphenyls (PCBs), and polycyclic aromatic hydrocarbons (PAHs) (see “A Closer Look at Food Contaminants”).

BFRs are man-made chemicals that are added to a wide variety of products, including for industrial use, to make them less flammable. These harmful compounds leach into the environment and pollute the air, soil, and water through waste residues or discharge from the factories that produce them.[1] As a result, the use of certain BFRs is banned or restricted in many countries.

BFRs enter the food chain when they reach the marine environment, as they are consumed by fish and shellfish, and cannot be excreted because they are lipophilic. This means that high levels can be present in seafood destined for human consumption.

PCBs also had widespread use in a range of industrial applications until they were banned in most countries in the 1980s, but they remain in the environment today as a result of their high stability. At high levels, they have been shown to cause health problems including carcinogenesis, endocrine disruption, and neurological problems.[2]

PAHs are generated mainly as a result of pyrolytic processes, especially the incomplete combustion of organic materials such as coal, oil, petroleum, and wood.[3] PAH exposure and its effects on human health have also been the focus of many studies, and some PAHs have been shown to be carcinogenic and mutagenic. The monitoring and regulation of PAHs are under constant change by advisory bodies such as the U.S. Occupational Safety and Health Administration and the National Institute for Occupational Safety and Health that stipulate exposure limits for PAH content.  

As a result of the changing regulatory landscape, there is need for more accurate identification and quantification of contaminants in environmental and food-related samples. Modern analytical techniques provide scope to remove contaminants from the food chain by obtaining unambiguous and highly accurate quantification of contaminants in complex food matrices at low concentrations. They can also be used to identify new or unexpected contaminants.

A technique for detection and quantification of contaminants in food samples of marine origin has been developed by researchers at the Research Institute for Pesticides and Water (IUPA), University Jaume I, in Spain. The aim was to develop advanced analytical methodology to improve the monitoring of compounds in food samples. Using atmospheric pressure gas chromatography (APGC), an innovative method was developed that is very effective for identifying and quantifying contaminants.

A Closer Look at Food Contaminants
Food contaminants are chemical substances that have been unintentionally added to food or feed, and can accumulate in the food chain. The major classes of contaminants include mycotoxins, heavy metals, dioxins and polychlorinated biphenyls, polycyclic aromatic hydrocarbons, 3-monochloropropanediol, melamine, erucic acid, and nitrates.

A number of analytical techniques are used in food safety testing. These include advanced instrumental techniques such as infrared spectroscopy, nuclear magnetic resonance, and mass spectrometry. Mass spectrometry is a versatile detection approach and is one of the most commonly employed tools for food safety testing, as it produces both qualitative and quantitative data, and allows detection of low concentrations of analytes in complex matrices.


Soft Ionization
Electron ionization (EI) has traditionally been used as an ionization technique to identify BFRs and other persistent organic pollutants. However, the technique’s limitations include extensive fragmentation and the absence or low intensity of the specific molecular ion. This lack of specificity makes the identification of these compounds difficult and can also reduce the technique’s sensitivity.

IUPA is utilizing a new chemical ionization source, APGC, that results in a “soft” ionization process to resolve the drawbacks of EI. The increased sensitivity enables quantification and confirmation of trace components at lower levels in the most complex samples.

The analysis of food samples by APGC enables improved selectivity when generating multiple reaction monitoring (MRM) transitions in comparison with the significant fragmentation experienced with an EI source. Operating the gas chromatography system at atmospheric pressure provides increased scope for ionization modes—namely charge and proton transfer.

Method Development and Validation
Using the APGC technique, IUPA collaborated with the Institute of Aquaculture Torre la Sal in Spain and the National Institute of Nutrition and Seafood Research in Norway to develop and test a method that would increase the number of contaminants detected, at much lower concentrations than previous methods reported, in a variety of food samples.

The method uses gas chromatography coupled to a tandem quadrupole tandem mass spectrometer with an atmospheric pressure chemical ionization source. The method is based on a modification of the unbuffered QuEChERS method (quick, easy, cheap, effective, rugged, and safe).

IUPA used APGC for GC-tandem mass spectrometry (GC-APGC-MS/MS) analysis of PAHs, PCBs, and pesticides in 19 different matrices—including fish tissues, feeds, and feed ingredients.[4]

Sustainable plant-based feeds developed for marine fish farming have presented new challenges concerning contaminants. Unrefined plant oils obtained from oilseeds such as soybeans, rapeseeds, olive seeds, and sunflower seeds are known to contain elevated levels of PAHs.

The carcinogenic “heavy” PAHs (> 4–6 rings) have attracted extensive interest with regards to food safety. Studies related to plant oil PAH contamination, however, mainly focus on light (2–4 rings) PAHs, such as fluoranthene, naphthalene, anthracene, and phenanthrene, as they are most dominantly present in unrefined plant oils. These light PAHs are also on the U.S. Environmental Protection Agency list for environmentally relevant PAHs but are mostly neither carcinogenic nor genotoxic.

In addition to the 24 PAHs, researchers tested for 15 pesticides and seven PCB congeners to widen the scope of the method. The study was to determine trace levels (as low as 0.1 ng/L) of PAHs, PCBs, polybrominated diphenyl ethers, and some emerging flame retardants.

The team used a total of 76 samples from 19 different matrices. The list contains ingredients from different origins (plant, terrestrial animals, and marine) and feeds based on these ingredients, as well as fillets of Atlantic salmon and
gilthead sea bream reared on these feeds.

The high sensitivity of this technique allowed the simultaneous quantification of 19 different complex matrices from aquaculture using solvent calibration. The excellent sensitivity and selectivity provided by GC-APGC-MS/MS allowed the dilution of the sample extracts and quantification using calibration with standards in solvent for all 19 matrices tested.

Analysis of real-world samples revealed the presence of naphthalene, fluorene, phenanthrene, fluoranthene, and pyrene at concentration levels ranging from 4.8 to 187 ng/g. Studied PCBs, dichlorodiphenyltrichloroethane, and pesticides were not found in fillets from salmon and sea bream. The aim of this work was the elimination of matrix effects. Even so, limits of quantification of the developed method were 2 ng/g for most analytes in the same order, or better than those reported in previously published methods for similar matrices showing higher efficiency.

Increased Sensitivity and Selectivity
The research determined that APGC is a robust and sensitive technique to analyze a broad range of contaminants in several marine-based matrices.[5] The possibility of selecting the molecular ion or the protonated molecule as a precursor ion for MRM experiments provides greater sensitivity and selectivity. This allows for the dilution of the sample extracts and quantification using calibration with standards in solvent, so matrix-matched calibrations can be avoided in some cases.

Increased sensitivity enables quantification and confirmation of trace components at even lower levels in the most complex samples. The ability to eliminate the matrix effect thereby eliminates the need for time-consuming purification steps. The sensitivity and selectivity also reduce the cost of tests for contaminants.

The technique uses fewer solvents and materials in comparison with previous techniques. The ability to determine compounds at a lower concentration allows compliance with regulatory limits and the ability to inject less sample matrix, reducing effects of contamination on the GC-MS system and therefore increasing uptime.

Soft ionization is a key benefit because of the reduced fragmentation for many compounds when compared with techniques such as EI. Reduced fragmentation can give higher sensitivity and specificity, therefore simplifying precursor ion selection in MS/MS analyses.

The soft ionization that occurs in the APGC source generates spectral data typically rich in molecular or protonated molecule ion information. This notably facilitated the application of MS/MS methods and also the screening of contaminants with GC-MS, focusing the search to the highly diagnostic molecular ion.

The versatility of the technique is high because it is possible to have both GC and liquid chromatography (LC) coupled to the same mass spectrometer, and it is relatively quick and easy to change from LC to GC and vice versa.

Researchers are keen to understand more about the capabilities that APGC can provide. In particular, low-level detection is fundamental as researchers are often searching for the unknown in complex and challenging samples.  

The research undertaken using the innovative APGC technique provides great opportunities for food security in terms of improving monitoring pollutants and meeting stringent safety standards.    

Tania Portolés, Ph.D., is a researcher at the IUPA, University Jaume I, Spain.

References
1. www.efsa.europa.eu/en/topics/topic/brominated-flame-retardants.
2. www.efsa.europa.eu/en/topics/topic/dioxins-and-pcbs.
3. Abdel-Shafy, HI and MSM Mansour. 2016. “A Review on Polycyclic Aromatic Hydrocarbons: Source, Environmental Impact, Effect on Human Health and Remediation.” Egyptian J Petroleum 25(1):107–123.
4. Portolés, T, et al. 2017. “Multi-class Determination of Undesirables in Aquaculture Samples by Gas Chromatography/Tandem Mass Spectrometry with Atmospheric Pressure Chemical Ionization: Novel Approach for Polycyclic Aromatic Hydrocarbons.” Tatlanta 172:109–119.
5. www.waters.com/waters/library.htm?lid=134968887.


Author(s): Tania Portolés, Ph.D.

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Tania Portolés, Ph.D., is a researcher at the IUPA, University Jaume I, Spain.

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