Our working group deals with the identification and quantification of food ingredients and contaminants as well as their metabolites as biomarkers. Further focal points are the biological effects of food ingredients in vitro and in humans. For this purpose, our working group uses analytical coupling techniques such as HPLC-DAD, (U)HPLC-MS/MS, Tims ToF-MS or GC-MS. Different biomarkers of (oxidative) cell damage and cell response are used to characterize the preventive effects of food ingredients. The working group has established a broad range of methods, including comet assay, quantitative real-time PCR, photometric assays and western blot, which enable detailed clarification of cellular mechanisms of action, including those of food ingredients.
- Kirsch V., Stegmüller S, Richling E (2023) Metabolic activation of 2-methylfuran and reactivity towards biomolecules Naunyn-Schmiedeberg’s Archives of Pharmacology 396, pages 1–76 (no. 70)
- Bohlen D, Eichel L, Stegmüller S, Richling E (2023) Studies investigating the side-chain hydroxylation of 2,5-dimethylfuran Naunyn-Schmiedeberg’s Archives of Pharmacology 396, pages 1–76 (no. 68)
- Møller P, Azqueta A, Collia M, Bakuradze T, Richling E, Bankoglu EE, Stopper H, Bastos VC, Langie SAS, Jensen A, Ristori S, Scavone F, Giovannelli L, Wojewódzka M, Kruszewski M, Valdiglesias V, Laffon B, Costa C, Costa S, Teixeira JP, Marino M,Del Bo’ C, Riso P, Zheng C, Shaposhnikov S, Collins A (2023) Inter-laboratory variation in measurement of DNA damage by the alkaline comet assay in the hCOMET ring trial. Mutagenesis 38, 283–294. https://doi.org/10.1093/mutage/gead014
- Møller P, Azqueta A, Rodriguez-Garraus A, Bakuradze T, Richling E, Bankoglu EE, Stopper H, Bastos VC, Langie SAS, Jensen A, Ristori S, Scavone F, Giovannelli L, Wojewódzka M, Kruszewski M, Valdiglesias V, Laffon B, Costa C, Costa S, Teixeira JP, Marino M,Del Bo’ C, Riso P, Zheng C, Shaposhnikov S, Collins A (2023) DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trial. Mutagenesis 38, 273–282. doi.org/10.1093/mutage/gead019
Bioactive compounds in food, such as polyphenols, are found in many types of fruit and vegetables and are therefore part of our daily diet. The intake of these compounds is associated with a reduced risk of chronic diseases such as type 2 diabetes mellitus, Parkinson's disease, cardiovascular diseases or certain types of cancer. Various projects have already been carried out in the working group on the availability and biological effects of anthocyanins (a special subgroup of flavonoids). These projects were funded by the Funding for Industrial Collective Research (IGF) from the Federal Ministry for Economic Affairs and Energy (via AiF) via the Forschungsvereinigung Forschungskreis der Ernährungsindustrie e.V. (FEI) as part of the program for the promotion of joint industrial research (IGF). The aim of the project (AiF 20277N), which was completed in 2022, was to investigate the influence of fruit juice ingredients on human lipid metabolism. The focus was on the question of whether the secondary plant substances in fruit juices (especially colored juices) are able to regulate lipid metabolism. In vitro studies showed that chokeberry, cranberry and pomegranate extracts have a strong influence on lipid metabolism Niesen et al., 2022. The subsequent human study showed that fruit juice ingredients can regulate and positively influence lipid metabolism in humans Rahn et al., 2023. An ongoing project (AIF 22474 N) entitled "Identification and technological enrichment of bioactive ingredients in non-alcoholic wines and sparkling wines" aims to identify bioactive phenolic secondary metabolites in the course of activity-guided isolation from non-alcoholic wines and sparkling wines and to demonstrate the positive influences on carbohydrate and lipid metabolism in vitro and in vivo. Another ongoing project (AiF 23033N) entitled "Influence of extracts from fruit juices and their ingredients on inflammatory processes" deals with the identification of anti-inflammatory compounds from fruit juice extracts (in vitro). The extracts identified in the cell model will also be verified in a proof-of-concept study in humans.
Rhineland-Palatinate is Germany's largest wine-growing region with an annual production of 5.4 million hL of wine. Wine production generates large quantities of wine by-products such as pomace, rape, vine leaves and vine wood. The development of innovative, value-enhancing ways of utilizing these wine by-products is becoming increasingly important. Due to the high content of secondary plant substances, such as polyphenols, wine by-products are of interest from a nutritional point of view. As part of the project funded by the Rhineland-Palatinate Foundation for Innovation, extracts from wine by-products of the Vitis vinifera L. cv. Riesling grape variety, fractions of these as well as identified pure substances were investigated in vitro with regard to their biological effects on the human hepatocarcinoma cell line HepG2 Fuchs et al., 2019. In addition, wine press residues (pomace) are used as organic fertilizers due to their high content of macro- and micronutrients such as sugar, phosphorus, sodium and nitrogen. At the same time, depending on the cultivation and storage conditions, they may contain pesticide and mycotoxin residues. These in particular are capable of changing the microbiome of the soil. Little is currently known about the mobility of these substances and their long-term effects on the soil. The benefits and risks of using grape pomace as an organic fertilizer are currently being investigated in a soil model as part of a Carl-Zeiss-funded project (cooperation with AG Prof. Munoz, University of Kaiserslautern-Landuau, Landau campus).
Furan is a compound that has been the focus of the European Food Safety Authority for several years (EFSA, 2017). It is known that furan and methylfurans are particularly present in coffee and canned food. Metabolic activation by CYP2E1 produces a reactive diaaldehyde (but-2-endial), which can react with various cellular nucleophiles. Our research group has set itself the goal of identifying some of the metabolites and testing them for their suitability as biomarkers of exposure. Since June 2019, a DFG-funded project (German Research Foundation (DFG) - project number 423497382 (1176/12-1) has been investigating the metabolism of furan and alklyfurans (Kremer et al., 2023) and developing methods for analyzing the furan and alkylfuran content in food. A further focus is on the toxicological assessment (e.g. cytotoxicity and DNA damage) of other alkylfurans such as 2-methylfuran, 2,5-dimethylfuran and 2-ethylfuran in vitro (DFG Project number 490818715 (1176/13-1)).
Methylglyoxal (MG) is a highly reactive compound that is formed in food, e.g. in the course of the Maillard reaction, and can therefore be absorbed by humans. However, MG is also formed endogenously in humans as an unavoidable by-product of glycolysis. MG is associated with many different diseases such as neurodegenerative disorders or type 2 diabetes. The glyoxalase system is important for the detoxification of MG, but it is also found in humans and yeasts, for example. As part of the BioComp project (funded by the Rhineland-Palatinate research initiative), the working group established an HPLC-MS/MS method for quantifying MG in various matrices. Thus, the MG content in different yeast mutants could already be determined in cooperation with working group Herrmann, RPTU; Rödl et al., 2022. Furthermore, the focus is on investigations of the effects of glucose on the MG status in HepG2 cells, as well as on the analysis of DNA adducts of MG. A further aim of the project is to establish the UPLC-MS/MS method for the quantification of second messengers in bacteria. This will provide further insights into the significance of second messengers for the motility of various P. aeruginosa strains (cooperation with the working group of Prof. Dr. Frankenberg-Dinkel, RPTU).
Monoterpenes and monoterpenoids are a group of structurally diverse and volatile organic natural substances whose basic structure consists of two isoprene units. While most monoterpenes and monoterpenoids are classified as low to slightly toxic, most in vitro studies lack evidence of actual substance concentrations. Due to their lipophilic properties, monoterpenes can suffer substance losses during in vitro testing, e.g. through sorption, binding to serum components or volatility losses. As a result, large differences between the nominal and the actual concentration in in vitro tests can lead to an underestimation of the toxicological potential.
Accordingly, the aim of our research is to investigate the toxicological potential of different monoterpenes and monoterpenoids with regard to substance losses in in vitro testing and to establish different methods to enable constant exposure concentrations.
In recent years, anthocyanins, a special subgroup of flavonoids, have become the focus of our research work. Bioavailability and numerous biological effects have been investigated as part of various projects. For example, in vitro studies showed that the encapsulation of an anthocyanin-containing blueberry extract did not affect antioxidant efficacy Schantz et al., 2014. The results obtained in a human study showed that the encapsulation of anthocyanins was able to modulate their bioavailability compared to non-encapsulated anthocyanins Müller et al., 2017; Müller et al., 2018. It was also observed that after ingestion of a blueberry extract Kropat et al., 2013 and an anthocyanin-rich mixed fruit juice FEI final report AiF 18068N, short version DNA- protective and antioxidant effects occurred in healthy volunteers Bakuradze et al., 2019. The project completed in 2018 (AiF 18506N) entitled "Activity-guided fractionation of fruit juice extracts to identify active substances for influencing glucose absorption in humans" dealt with the influence of anthocyanins on glucose metabolism. By inhibiting the activity of various key enzymes of carbohydrate digestion (α-amylase, α-glucosidase), it can indirectly lead to a reduction in postprandial blood glucose levels (Ostberg-Potthoff et al., 2019, Berger et al., 2020).
As part of the ERDF (European Regional Development Fund) -funded Interreg project BIOVAL, a cross-border value chain was to be developed based on the brewery residue brewer's spent grains, which are produced worldwide at around 39 million tons per year. In addition to three TUK working groups, the universities of Liège, Lorraine and Saarland as well as the Luxembourg Institute of Health and the Belgian company Celabor were involved. Brewer's grains, a main by-product after the mashing process in beer production, are currently mainly used as animal feed due to their high cellulose content. However, other ingredients such as proteins, hemicelluloses and minor components such as polyphenols offer the potential for further applications. Prof. Dr. Elke Richling's working group conducted research into the effects of substances and extracts from spent grains on human health. The focus was on the biological effects of the ingredients with regard to their influence on glucose metabolism and the toxicological evaluation of potential components using in silico and in vitro methods (Kirsch et al., 2020, Becker et al., 2021).
The project was also presented at ACHEMA 2018 https://idw-online.de/en/news696124
Coffee is one of the most widely consumed beverages in the world. It is a complex mixture of over a thousand different substances, some of which have biological activity. In addition to carbohydrates, lipids, proteins and free amino acids, coffee contains secondary plant substances such as alkaloids (especially caffeine and trigonelline), phenolcarboxylic acids (e.g. chlorogenic acids), roasting products, volatile aromatic substances (e.g. pyrazines) as well as diterpenes (cafestol, kahweol), vitamin B3 (niacin) and minerals. In our working group, twelve different alkyl pyrazines in coffee were quantified using SIVA-GC-MS Pickard et al., 2013. In the past, several intervention studies were carried out with test subjects to investigate the biological effects of coffee drinks on the one hand and the metabolism of selected coffee ingredients on the other. The results of the intervention studies show that regular coffee consumption is associated with DNA-protective and antioxidant effects (Bakuradze et al., 2014; Bakuradze et al., 2015; Bakuradze et al., 2016). In addition, our studies found an increase in high density lipoprotein (HDL) in the blood and an inhibition of phosphodiesterase activity in platelets after several weeks of coffee consumption (Riedel et al., 2014; Montoya et al., 2014). The metabolism of coffee ingredients was investigated in a human intervention study. The excretion of niacin, a semi-essential vitamin, and its metabolites were recorded. Using stable isotope dilution analysis (SIVA), detailed kinetics of renally excreted niacin metabolites following coffee consumption could be established. The results confirm that coffee consumption contributes to the niacin supply Kremer et al., 2018.
Acrylamide is an α,β-unsaturated carbonyl compound that can be formed from amino acids such as asparagine and reducing sugars when food is heated (T > 120 °C). Starchy foods in particular, such as French fries, potato chips and crispbread, but also roasted coffee, contain high amounts of acrylamide. After absorption, acrylamide is partially metabolically activated to the genotoxic epoxide glycidamide. In human studies, selected metabolites of acrylamide were detected as biomarkers in urine using stable isotope dilution analysis (SIVA). The studies of our working group showed that these compounds are largely excreted renally as mercapturic acids in a dose-dependent manner from acrylamide intake via the diet Watzek et al.,2012. Thus, these could be validated under controlled conditions as short-term biomarkers of acrylamide exposure Rünz et al., 2016. Under uncontrolled conditions, it was shown that other exogenous factors, e.g. (tobacco) smoke, influence the excretion of mercapturic acid Goerke et al., 2019. In addition, our studies showed a disproportionate excretion of mercapturic acid compared to a minimized dietary acrylamide intake. Oral administration of stable isotope-labeled acrylamide ruled out storage and delayed release in the organism Goempel et al., 2017. The working group is now investigating the hypothesis of a possible endogenous formation of acrylamide in the body.