Karen Bekaert

Recent advances in recognition elements of food and environmental biosensors: A review

A sensitive monitoring of contaminants in food and environment, such as chemical compounds, toxins and pathogens, is essential to assess and avoid risks for both, human and environmental health. To accomplish this, there is a high need for sensitive, robust and cost-effective biosensors that make real time and in situ monitoring possible. Due to their high sensitivity, selectivity and versatility, affinity-based biosensors are interesting for monitoring contaminants in food and environment. Antibodies have long been the most popular affinity-based recognition elements, however recently a lot of research effort has been dedicated to the development of novel recognition elements with improved characteristics, like specificity, stability and cost-efficiency. This review discusses three of these innovative affinity-based recognition elements, namely, phages, nucleic acids and molecular imprinted polymers and gives an overview of biosensors for food and environmental applications where these novel affinity-based recognition elements are applied.

Development and validation of an ultra-high performance liquid chromatography tandem mass spectrometry method for quantifying thyreostats in urine without derivatisation

Thyreostatic drugs, illegally administrated to livestock for fattening purposes, are banned in the European Union since 1981 (Council Directive 81/602/EC). For monitoring their illegal use, sensitive and specific analytical methods are required. In this study an UHPLC-MS/MS method was described for quantitative analysis of eight thyreostatic drugs in urine, this without a derivatisation step. The sample pretreatment involved a reduction step with dithiothreitol under denaturating conditions at 65 degrees C, followed by liquid-liquid extraction with ethyl acetate. This analytical procedure was subsequently validated according to the EU criteria (2002/657/EC Decision), resulting in decision limits and detection capabilities ranging between 1.1 and 5.5 microg L(-1) and 1.7 and 7.5 microg L(-1), respectively. The method obtained for all, xenobiotic thyreostats, a precision (relative standard deviation) lower than 15.5%, and the linearity ranged between 0.982 and 0.999. The performance characteristics fulfill not only the requirements of the EU regarding the provisional minimum required performance limit (100 microg L(-1)), but also the recommended concentration fixed at 10 microg L(-1) in urine set by the Community of Reference Laboratories. Future experiments applying this method should provide the answer to the alleged endogenous status of thiouracil.

Ultra-high performance liquid chromatography-tandem mass spectrometry in high-throughput confirmation and quantification of 34 anabolic steroids in bovine muscle.

An ultra-high performance liquid chromatography tandem mass spectrometry multi-residue method for the determination of 34 anabolic steroids (10 estrogens including stilbenes, 14 androgens and 10 gestagens) in meat of bovine origin is reported. The extraction and clean-up procedure involved homogenization with methanol, defatting with hexane, liquid/liquid extraction with diethylether and finally SPE clean-up with coupled Si and NH(2) cartridges. The analytes were separated on a 1.9 μm Hypersil Gold column (100×2.1 mm) and quantified on a triple quadrupole mass spectrometer (TSQ Vantage) operating simultaneously in both positive and negative atmospheric pressure chemical ionisation (APCI) modes. This analytical procedure was subsequently validated according to EU criteria (CD 2002/657/EC), resulting in decision limits and detection capabilities ranging between 0.04 and 0.88 μg kg(-1) and 0.12 and 1.9 μg kg(-1), respectively. The method obtained for all, natural and synthetic steroids, adequate precisions and intra-laboratory reproducibilities (relative standard deviation below 20%), and the linearity ranged between 0.991 and 0.999. The performance characteristics fulfill the recommended concentrations fixed by the Community Reference Laboratories. The developed analysis is sensitive, and robust and therefore useful for confirmation and quantification of anabolic steroids for research purposes and residue control programs.

Effect of surgical castration, immunocastration and chicory-diet on the meat quality and palatability of boars

This study evaluates 1) carcass quality, meat quality and palatability for barrows, immunocastrates and boars and 2) the effect of chicory supplemented feed during 10 days before slaughter on boar meat quality. At comparable carcass weights, estimated carcass lean meat percentage was higher in immunocastrates and boars than in barrows. Muscle thickness was higher for immunocastrates and barrows compared to boars, while fat thickness was lowest for immunocastrates and boars. Barrows, immunocastrates and boars differed in water holding capacity and boar taint. Home consumer panels were conducted to evaluate palatability. The consumers did detect differences in tenderness and juiciness, but not for boar taint. The chicory feed supplemented in boar feed decreased skatole concentration in backfat, without largely influencing meat quality or palatability. Not only boar taint, but also carcass and meat quality should be considered when evaluating alternatives for surgical castration.

Influence of breed and slaughter weight on boar taint prevalence in entire male pigs.

Piétrain (P), Large White (LW) and Belgian Landrace stress negative (BN) boars were slaughtered at 50, 70, 90 or 110 kg live weight to investigate breed differences and the effect of slaughter weight on boar taint prevalence. Boar taint was quantified by four different methodologies: sensory evaluation of neckfat heated with a hot iron in the slaughterhouse, sensory evaluation of meat by consumer panels, sensory evaluation of fat and meat by expert panels and laboratory analysis of indole, skatole and androstenone in backfat. Skatole levels in backfat were significantly higher for LW and BN than for P boars. The androstenone levels and the hot iron method revealed a significant interaction between breed and slaughter weight. On the other hand, experts detected an effect of weight on the androstenone odour perception, which was significantly higher in fat from boars slaughtered at 90 kg compared with 50 kg, and significantly higher in meat from boars slaughtered at 110 kg compared with 50 kg. Consumers did not detect differences in the sensory characteristics among breeds or slaughter weight. These results indicate opportunities to minimise the risk of boar taint in entire male pigs by carefully selecting a combination of breed and slaughter weight. Along with the optimal slaughter weight, the effectiveness of reducing boar taint by lowering slaughter weight appeared to be breed dependent.

A validated ultra-high performance liquid chromatography coupled to high resolution mass spectrometry analysis for the simultaneous quantification of the three known boar taint compounds.

Boar taint is an off-odour that can occur when meat or fat from entire male pigs is heated. Most of the currently available analytical methods are not capable of detecting the three known boar taint compounds (indole, skatole and androstenone) simultaneously, which renders their analysis often labour-intensive and time-consuming as separate analyses are required. In this study a validated U-HPLC-HR-Orbitrap-MS analysis method is described for the quantitative determination of the three boar taint compounds in fat. The sample pre-treatment involves a melting step followed by extraction with methanol and clean-up consisting of a freezing step and solid phase extraction (HLB cartridges). The analytes are then chromatographically separated and detected with an Exactive high-resolution mass spectrometer. Due to the absence of guidelines for the analysis of boar taint in fat, the Commission Decision 2002/657/EC [18] and ISO 17025 [19] guidelines were used as guideline for validation of the developed detection method. This resulted in limits of detection and limits of quantification between 2.5 and 7 μg kg(-1) and between 5 and 10 μg kg(-1) for the three compounds, respectively, which is far below the threshold values set at 100 μg L(-1) for indole, 200 μg L(-1) for skatole and 1000 μg L(-1) for androstenone in pig fat samples. The method obtained for the three compounds a repeatability (RSD) lower then 12.7% and a within-laboratory reproducibility (RSD) lower than 16.9%. The recovery of the three compounds ranged between 99 and 112 and an excellent linearity (R(2) ≥ 0.99) was found. In the future, this method may be extended with other compounds that turn out to be correlated with boar taint.

Effect of information provisioning on attitude toward surgical castration of male piglets and alternative strategies for avoiding boar taint.

Pork consumers know little about boar taint and the methods used to avoid it. As such, relevant information is necessary to assist consumers to judge the acceptability of different strategies to avoid boar taint. The effect of basic (T1) or extensive (T2) written information or T2 with supplementary audio-visual information (AV) on the opinion concerning immunocastration (IC), raising entire male pigs (EM) and surgical castration with anaesthesia (SA) as compared to castration without anaesthesia (SC) was investigated in a student population. Overall, IC was significantly preferred over SC. The information condition influenced the preference for IC and EM as compared to SC. Participants exposed to AV were more positive to IC than participants exposed to T1 and T2, and more positive to EM than participants exposed to T2. The impact of information condition was not affected by gender, farming experience, knowledge about the boar taint issue or personal relevance of pig welfare. Potential effects of providing background information and media campaigns on public surveys ought to be considered. Supplementary audio-visual information increased the impact of information provisioning.

Ultra-high performance liquid chromatography coupled to triple quadrupole mass spectrometry detection of naturally occurring thiouracil in urine of untreated livestock, domesticated animals and humans

Thiouracil belongs to the xenobiotic thyreostats, which are growth-promoting agents illegally used in animal production. Recently it has been reported that thiouracil is suspected to have a natural origin. The European Union of Reference Laboratory guidance paper of 2007 acknowledged this by stating that thiouracil concentrations below 10 µg l−1 might have a natural origin derived from the consumption of Brassicaceae. The present research aimed at endorsing this possible natural occurrence. Urine samples of animals (livestock and domesticated) with known and unknown clinical backgrounds were analysed for thiouracil with a newly developed ultra-high performance liquid chromatography coupled to a triple quadrupole mass spectrometric analysis method without derivatisation. In addition, a small-scale 9-day human experiment with Brassicaceae vegetables was performed to investigate if this natural prevalence could be extrapolated to the human population. The untreated animals had thiouracil concentrations below 10 µg l−1 acknowledging the alleged natural occurrence of thiouracil. As for the humans, in 66.7% of the urine samples thiouracil was found above the CCα of 2.2 µg l−1. However, the correlation with the Brassicaceae diet proved to be non-significant (p = 0.095). Nevertheless, these results clearly demonstrate the natural occurrence of thiouracil in urine of animals and humans. The exact origin of this natural thiouracil trace still needs to be identified.

Evaluation of different heating methods for the detection of boar taint by means of the human nose

No automated detection system for boar taint detection is currently available, thus boar taint at the slaughterline can currently only be assessed using the singeing method (olfactory scoring). This study compares several heating methods (microwave, soldering iron and pyropen) and evaluates the effect of habituation, cleaning the soldering iron, singeing the fat twice in the same place, and variations in the technical procedures. All methods seem to be suitable for detecting boar taint but the choice of heating method for sensory scoring of boar taint depends on habituation of the trained assessor and specific conditions applied. The pyropen seems to be most suitable because it does not contact the fat and is easy to handle (wireless). Finally, the intensity score may also be influenced by: contamination from not cleaning the soldering iron, singeing the fat twice in the same place, and the effect of habituation.

Influence of soiling on boar taint in boars

It has been suggested that skatole, one of the main compounds responsible for boar taint, can be lowered by keeping pigs clean, as skatole can be absorbed through skin and/or lungs (Hansen, Larsen, Jensen, HansenMoller & Bartongade, 1994). With this experiment, we further investigated this hypothesis by comparing extremely clean with extremely dirty animals with regard to the occurrence of boar taint. One group of boars was washed daily and pens were mucked on and littered down daily (CLEAN), a second group of boars was rubbed with faeces daily (DIRTY) and a third group of boars was kept in control conditions (CONTROL). The treatment was performed during the last four weeks before slaughter. According to the standardised consumer panel evaluations, boars subjected to extra soiling had a higher concentration of boar taint than boars that were kept extra clean. In contrast, expert panels judged general meat flavour to be inferior in CLEAN than CONTROL pigs. The home consumer panel, the hot iron method, and laboratory analyses, i.e., the presence of indole, skatole and androstenone in fat and serum, all showed no significant differences. So no clear indications towards skatole reduction by improving cleanliness of pigs were found.