Pyrrolizidine alkaloids (PAs) and their N-oxides (PANOs) are natural toxins, exclusively biosynthesized by a wide variety of plant species (>6000). They are secondary plant metabolites against herbivores and are believed to be one of the most widely spread natural toxins. PAs and PANOs can become a significant public human health problem from the intake of contaminated food of botanical or animal origin. Human poisoning cases have been documented; they are characterized by acute and chronic liver damage, pulmonary hypertension, cardiac or kidneys injuries, and can lead to death. Therefore, the development of efficient analytical methods is required to detect and quantify PAs/PANOs in a large range of food items.
The Belgian Scientific Institute of Public Health (WIV-ISP) has directed a project aiming to report occurrence levels of PAs and PANOs in targeted food items to perform a dietary exposure assessment to these natural contaminants and evaluate if they can pose a health problem, filling a data gap at Belgium’s level. Analytical methods based on UHPLC separation and MS/MS detection were developed for the analysis of 16 PAs and 14 PANOs in honey and honey based snacks, meat and meat products, milk and dairy commodities, prepared dishes, sauces, plant based products, dry teas and corresponding infusions, and food supplements. The sample preparation protocols were comprised of an acidic solid-liquid extraction of the analytes followed by SPE clean-up to avoid loss of sensitivity due to matrix effect. Several steps were carefully optimized for each targeted food matrices with a specific focus on very low limits of quantification (LOQs < 1 ng/g or ng/mL), taking into account the various interfering compounds to discard. The analytical methods were validated in-house and the method’s performances were within the criteria allowed by Directive 2002/657/CE and SANTE/11945/2015. A structured sampling plan has been established for more than 1,200 samples available on the Belgian market.
The analytical results showed that PAs/PANOs contamination in food is recurrent, with maximum levels ranging from 16 pg/g (ppt) in dairy products to several ppm in plant-based food supplements and dried culinary herbs. Surprisingly, high levels of heliotrine-type PAs/PANOs (>1000 ng/g) were detected in 2 Mediterranean spices/herbs mixes and certain salad mixes were also contaminated with significant levels of senecionine-type compounds. More than 85% of plant-based food supplements contained PAs/PANOs, with levels up to 557,000 ng/g. An original approach enabled us to demonstrate that only a part of the PAs/PANOs contamination is transferred during the brewing process of (herbal) teas (15-25%). This transfer rate contrasts with the conventional strategy used by the European Food Safety Authority (EFSA) consisting of applying a dilution factor to the PAs concentration found in the dry plant products to estimate the contamination level of the infusions.
The quantitative results for the different food groups were linked to the actual consumption data for different age groups from the Belgian population to assess their dietary intake to those natural contaminants. It appeared that milk and dairy products were by far the main contributors to the overall PAs/PANOs intake for each age category, even though the concentration levels detected in this food group were the lowest.