Wageningen University research scientists at RIKILT Institute of Food Safety are looking for new and innovative approaches to detect banned and registered chemical substances in foods and animal feeds. They typically target residues such as growth promoters, veterinary drugs, natural toxins, and pesticides in a variety of matrices (meat, milk, eggs, seafood, honey, urine, hair, and other forensic samples). The goal of the residue testing team is to identify residues in our food supply and ensure that residue limits meet national and international regulations, while also scoping for forbidden residues to assess the illegal use of banned substances in food farming and manufacturing practices.
The use of steroidal hormones in the food chain
Worldwide, farmers raise an extensive quantity of food-producing animals to meet global food demands. In some cases, feedlots are given steroidal hormones to promote faster growth, with the goal of improving productivity. Testosterone surrogates, estrogens, or progestins are administered to livestock to enhance growth and fertility. Ultimately, the use of these substances is designed to make the animals more productive – either through gaining leaner muscle more rapidly or becoming more fertile to grow the brood. Unfortunately, steroidal hormone use results in the potential accumulation of these residues in the edible tissues of the animal, increasing risks when consumed by humans. Hormone consumption can cause disruption in the natural hormonal balance in the human body, and consuming animal products laced with hormones can result in adverse health effects, particularly in children whose bodies are still developing and are more sensitive to hormone consumption. Suggested side effects of excessive hormone consumption include premature sexual development, development of ovarian cysts in young girls, and autoimmune problems, among others. Due to these negative effects, many global food regulators monitor animal tissue, urine, or hair to identify the presence of steroidal hormones to help to enforce and regulate the use of these substances in food-producing animals.
Analysis of steroidal hormones in hair
Steroidal hormones are delivered to animals as synthetic esters (to improve the half-life of the compound in the animal and enhance its efficacy), and the routine testing of these synthetic steroidal esters is often a big challenge. The esters are rapidly hydrolyzed into substances that are endogenously present in urine; therefore, scoping urine for the presence of these steroidal compounds is not always an effective approach to determine the illegal use of these residues. The team at RIKILT is addressing this by proposing an interesting alternative – to test for the intact steroidal esters in animal hair samples. Not only is hair easy to collect and transport, but there is also along traceability, and testing can be done long after slaughter. By analyzing for steroidal residues in hair, the steroids are directly detected, giving concrete evidence of the use of the steroids in the animal. Unfortunately, hair can be a difficult matrix to test, with many matrix interferences that can cause challenges with the accurate analysis and quantitation of the residues of interest. Scientists at RIKILT are using their SCIEX QTRAP® 5500 and QTRAP® 6500 Systems with MRM3 workflows to overcome the matrix interferences produced by the hair to more accurately and selectively quantify low-level steroidal esters in hair samples.
MRM vs. MRM3
MRM3 takes routine MRM detection to the next level. With MRM, the precursor ion is selected in Q1, fragmented in Q2, and the product ions are selected in Q3 and detected. In MRM3, rather than selecting the product ion in Q3, those product ions are accumulated in Q3 and fragmented further by collisional-induced dissociation. This results in fragmentation of the product ion, giving a third-generation fragment that can be detected by the mass spectrometer. So, rather than having a unique precursor/product ion pair, you detect the precursor/product/sub-product ion triad, adding an additional dimension of selectivity that can improve signal-to-noise by reducing the impact of matrix interferences on the results. And, the MRM3 results can be used for quantitation, also resulting in improved accuracy and precision of the reported results. Shown in Figure 1 is an example of the detection of testosterone acetate in hair using MRM (left side) and MRM3 (right side). It is clear that there is a huge matrix interference at the retention time of the compound of interest (5.23 mins) in MRM mode, but that interference is significantly reduced when using MRM3 mode. This is just one way that RIKILT is using MRM3 on their QTRAP® Systems to overcome matrix interferences and get better quantitative results for low-level steroidal esters in hair samples.
Food testing scientists are always looking for new and creative approaches to accurately detect residues that might make their way into our food supply. RIKILT is paving the way in food testing innovations, using QTRAP® Technology and advanced scan functions to improve selectivity for their residue analyses, reduce the interferences that occur from the matrix, and enable low-level quantitation of banned and registered chemical substances used in food production. Learn more about the unique work at RIKILT by viewing their webcast presentation, which also includes an overview of the QTRAP® 6500 LC/MS/MS System to enable improved sensitivity and selectivity for residue testing in food and food-related matrices.