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This study aimed at the comparison of two different photobioreactors with focus on technology and sustainability. The mesh ultra-thin layer photobioreactor (MUTL-PBR) exhibited around 3-fold biomass based space–time-yield and an around 10-fold specific antioxidant capacity than the traditional reference photobioreactor. Life cycle assessment (LCA) was done under autotrophic conditions in both pilot scale reactors with focus on biomass production and on antioxidant capacity of the biomass, respectively. Biomass production within the reference reactor showed a lower environmental impact in most categories. A significantly higher energy demand for mixing and cooling of the cell suspension within the MUTL-PBR is the major reason for its environmental burden. This relates to high impacts in the categories “non-renewable energy” and “global warming potential“ per kg biomass. Comparing algal antioxidant capacity, environmental impact of the MUTL cultivation was 5–10 times lower. This clearly illustrates the potential of MUTL-PBR for sustainable production of bioactive substances.

As allergy towards apples is widespread, the evaluation of various cultivation and postharvest influences on the allergenic potential is of great importance. Therefore, the analysis of the Mal d 1 content was the focus of this study, originally dealing with investigating the influence of a selenium biofortification on apple quality. The Mal d 1 content of apples was in most cases reduced when the fruits were biofortified with selenium. Apple variety and climatic conditions were identified as further influencing factors for the Mal d 1 content of the fruits. The separate analysis of the peel and the fruit flesh showed that the content of Mal d 1 in the fruit flesh was significantly lower in the biofortified samples than in the controls. In conclusion, the results indicate that the selenium biofortification of apples and biochemical mechanism behind can reduce the allergenic potential regarding the content of Mal d 1.

High-performance thin-layer chromatography (HPTLC) is a suitable method for the analysis of peptides and proteins due to a wide selection of stationary and mobile phases and various detection options. Especially, two-dimensional HPTLC (2D-HPTLC) enables a higher resolution compared to one-dimensional HPTLC in the separation of complex peptide mixtures. Similar to 2D electrophoresis, characteristic peptide patterns can be obtained, allowing a differentiation of ingredients based on varying protein origins. The aim of this study was to evaluate 2D-HPTLC with regard to its suitability for the characterization of proteins/peptides and to verify whether it is possible to predict the retention behavior of peptides based on their properties. As models, the five most abundant milk proteins α-lactalbumin, β-lactoglobulin, α-, β-, and κ-Casein were used. In order to determine the repeatability of the peptide separation by 2D-HPTLC, each tryptic protein hydrolyzate was separated eight times. The standard deviations of the retardation factors for the separated peptides varied between 1.0 and 11.1 mm for the x-coordinate and 0.5–7.3 mm for the y-coordinate. It was also shown that after the chromatographic separation, peptides of the individual protein hydrolyzates were located in specific areas on the HPTLC plate, so that a clustering could be obtained for the whey proteins‘ as well as the caseins‘ hydrolyzates. For establishing correlations between the properties of the peptides and their retardation factors, 51 of 85 selected peptides were identified by matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry (MALDI-TOF-MS/MS). On this basis, statistically significant correlations (α = 0.05) between the retardation factors of the peptides and their isoelectric points, as well as the percentage of anionic and non-polar amino acids in the peptides were established. Finally, it was investigated, whether the retardation factors for peptides can be predicted on the basis of a linear regression of the percentage of non-polar amino acids in a peptide. For this purpose, a mixture of artifical (synthetic) peptides (n = 14) was separated by 2D-HPTLC and the measured retardation factors were compared with the corresponding retardation factors calculated. Absolute deviations of 0.3–17.9 mm were obtained. In addition, the universal applicability of the method to other protein sources other than milk proteins (animal protein) was tested using a mixture of pea peptides (plant protein, n = 3) resulting in absolute deviations of 0.7–8.6 mm.

Small plastic particles are found ubiquitously in marine and freshwater ecosystems and consequently, their inhabitants. When aquatic animals are consumed as seafood, human exposure to those plastic particles is possible. However, only a few studies, applying largely different analytical procedures, assessed microplastics content in the edible part of seafood. In this study, ten protocols for the extraction of microplastics from biota were chosen and tested for their suitability to digest the edible part of a broad range of seafood species. The following criteria were used for this assessment: 1) feasibility to filtrate the entire digested sample with one filter of approximately 1 μm pore size, 2) effect of reagents applied during the sample preparation on the polymer integrity as assessed by means of infrared and Raman spectroscopy as well as pyrolysis gas chromatography mass spectrometry, 3) total sample preparation time and the possibility to avoid the use of expensive reagents. The most suitable protocol was found to be an enzymatic-alkaline approach, consisting of a quick hydrolysis of proteins with pepsin for 2 h and a consecutive alkaline hydrolysis for 4 h, both at 37 °C for most of the tested seafood species. Digestion efficiency of the optimised protocol was tested with fish fillets and soft tissues of commercially relevant molluscs and crustacean species. Compared to most other protocols described in literature, an adequate digestion of the seafood matrix was achieved faster without being significantly more cost- or labour-intensive. Moreover, only negligible degradation of eleven commercially relevant polymers excluding polyacrylonitrile was observed. Polymer integrity was assessed by a change in particle weight or surface as well as spectroscopic and chromatographic data. The optimised sample preparation protocol aims to support future method standardisation efforts in order to assess the dietary uptake of microplastics in humans.

Mycorrhizal symbiosis is known to be the most prevalent form of fungal symbiosis with plants. Although some studies focus on the importance of mycorrhizal symbiosis for enhanced flavonoids in the host plants, a comprehensive understanding of the relationship still is lacking. Therefore, we studied the effects of mycorrhizal inoculation of onions (Allium cepa L.) regarding flavonol concentration and the genes involved in flavonol biosynthesis when different forms of nitrogen were supplied. We hypothesized that mycorrhizal inoculation can act as a biotic stress and might lead to an increase in flavonols and expression of related genes. The three main quercetin compounds [quercetin-3,4′-di-O-β-d-glucoside (QDG), quercetin-4′-O-β-d-glucoside (QMG), and isorhamnetin-4′-O-β-d-glucoside (IMG)] of onion bulbs were identified and analyzed after inoculating with increasing amounts of mycorrhizal inocula at two time points and supplying either predominantly NO3 − or NH4 + nitrogen. We also quantified plant dry mass, nutrient element uptake, chalcone synthase (CHS), flavonol synthase (FLS), and phenyl alanine lyase (PAL) gene expression as key enzymes for flavonol biosynthesis. Inoculation with arbuscular mycorrhizal fungi (highest amount) and colonization at late development stages (bulb growth) increased QDG and QMG concentrations if plants were additionally supplied with predominantly NH4 +. No differences were observed in the IMG content. RNA accumulation of CHS, FLS, and PAL was affected by the stage of the mycorrhizal symbiosis and the nitrogen form. Accumulation of flavonols was not correlated, however, with either the percentage of myorrhization or the abundance of transcripts of flavonoid biosynthesis genes. We found that in plants at late developmental stages, RNA accumulation as a reflection of a current physiological situation does not necessarily correspond with the content of metabolites that accumulate over a long period. Our findings suggest that nitrogen form can be an important factor determining mycorrhizal development and that both nitrogen form and mycorrhizas interact to influence flavonol biosynthesis.

Fishmeal and oil are regarded as byproducts resulting from fish processing. However, the procedures are hypothesized to be an ecological harm for maintaining the balance of the aquatic ecosystems. This study aimed at characterizing an innovative algae–yeast mix for a partial substitution of fishmeal and oil in fish feed. The microorganism mix consisted of the cyanobacterium Arthrospira sp., the microalgae Crypthecodinium cohnii, and the yeast Rhodotorula glutinis to create a balanced nutritional composition with an enrichment of bioactive compounds for inducing positive health effects in selected fish species. Such a modification can also be interpreted in terms of a positive (human) nutritional effect. From an ecological point of view, all these microorganisms can be cultivated without minimum effort. In fish feeding trials, pike-perch (Sander lucioperca) and rainbow trout (Oncorhynchus mykiss) were taken for evaluating the feed acceptance, fish health, and fish filet quality. The study revealed a successful feed development, which resulted in healthy organisms depending on the species and compared to conventionally fed organisms.

Besides its bioactivity, Opuntia dillenii cactus juice exhibits desirable physicochemical properties. In the present study, impact of innovative pasteurization techniques – pulsed electric fields (PEF) and high hydrostatic pressure (HHP) – on the microbial inactivation, selected physicochemical properties and rheological characteristics, ascorbic acid, flavonols, betacyanins, and the antioxidant activity was evaluated in comparison to the non-pasteurized fresh juice. Results showed that PEF and HHP, as well as thermal pasteurization (TP), reduced the microbial populations found in juice from 10 3 to less than 10 cfu/mL. The non-Newtonian pseudoplastic flow behaviour of juice was maintained by all three methods. PEF and HHP resulted in a better retention of ascorbic acid, while TP reduced 22% of the ascorbic acid content. In contrast, TP caused a slight transformation of isorhamnetin glycosides. PEF and HHP maintained antioxidant activity of juice comparatively better than TP.

A prototype gas chromatography-ion mobility spectrometry (GC-IMS) system, hyphenating temperature-ramped headspace GC to a modified drift time IMS cell, was evaluated and compared to a conventional, isothermal capillary column (CC)-IMS system on the example of the geographical differentiation of extra virgin olive oils (EVOO) from Spain and Italy. It allows orthogonal, 2D separation of complex samples and individual detection of compounds in robust and compact benchtop systems. The information from the high-resolution 3D fingerprints of volatile organic compound (VOC) fractions of EVOO samples were extracted by specifically developed chemometric MATLAB® routines to differentiate between the different olive oil provenances. A combination of unsupervised principal component analysis (PCA) with two supervised procedures, linear discriminant analysis (LDA) and k-nearest neighbors (kNN), was applied to the experimental data. The results showed very good discrimination between oils of different geographical origins, featuring 98 and 92% overall correct classification rate for PCA-LDA and kNN classifier, respectively. Furthermore, the results showed that the higher resolved 3D fingerprints obtained from the GC-IMS system provide superior resolving power for non-targeted profiling of VOC fractions from highly complex samples such as olive oil. Graphical abstract Principle of the determination of geographic origins of olive oils by chemometric analysis of three-dimensional HS-GC-IMS fingerprints.

Fish and fish oil are an important source of essential dietary components such as polyunsaturated long chain fatty acids like eicosapentaenoic acid and docosahexaenoic acid. Refining of crude fish oil is necessary to remove environmental contaminants e.g., dibenzo- p -dioxins, dibenzofurans, dioxin-like polychlorinated biphenyls, and non-dioxin-like polychlorinated biphenyls for reaching legal limits, ensuring a pleasant taste and for enhancing shelf-life. However, during the deodorization of crude fish oil, process contaminants such as 2- and 3-monochloropropane-diol esters and glycidyl esters may be formed. The aim of the present study was to provide a comprehensive overview on the content of selected environmental and process contaminants in raw and processed fish oils deriving from different fish species. In crude fish oils extracted from farmed fish, the contents of environmental contaminants were well below the legal limits for food grade fish oil. In contrast, fish oils extracted from captured fish did not meet the requirements for human consumption. When applying filtration over active carbon, a reduction in the content of dibenzo-p-dioxins, dibenzofurans and dioxin-like polychlorinated biphenyls was achieved. 2- and 3-monochloropropane-diol ester and glycidyl ester contents were considerably higher in the refined compared to the crude fish oils investigated. Additionally, the comparison of analyzed and labeled contents of polyunsaturated long chain fatty acids showed a satisfying compliance.

High-protein diet is a promising strategy for diabetes treatment supporting body weight control, improving glycaemic status, cardiovascular risk factors and reducing liver fat. Here, we investigated effects of diets high in animal (AP) or plant (PP) protein on oxidative stress and antioxidant status in individuals with type 2 diabetes (T2DM). 37 obese individuals (age 64.3 ± 1.0 years) with T2DM were randomized to an isocaloric diet (30 energy(E)% protein, 30 E% fat and 40 E% carbohydrates) rich in AP or PP for 6 weeks. Markers of oxidative and nitrosative stress and antioxidant status in plasma and nitrate/nitrite levels in urine were assessed. Gene expression in subcutaneous adipose tissue (SAT) was analysed by RNA-Seq and real-time PCR. Both AP and PP diets similarly reduced plasma levels of malondialdehyde (PAP = 0.003, PPP = 1.6 × 10−4) and protein carbonyls (PAP = 1.2 × 10−4, PPP = 3.0 × 10−5) over 6 weeks. Nitrotyrosine (NT) increased upon both AP and PP diets (PAP = 0.005, PPP = 0.004). SAT expression of genes involved in nitric oxide (NO) and oxidative stress metabolism and urine NO metabolite (nitrate/nitrite) levels were not changed upon both diets. Plasma levels of carotenoids increased upon PP diet, whereas retinol, alpha- and gamma-tocopherol slightly decreased upon both diets. AP and PP diets similarly improve oxidative stress but increase nitrosative stress markers in individuals with T2DM. Mechanisms of the NT regulation upon high-protein diets need further investigation., Graphical abstract Image 1, Highlights • Effects of 6-week isocaloric diets high in animal (AP) or plant (PP) protein in subjects with type 2 diabetes were studied. • AP and PP diets reduced plasma levels of malondialdehyde and protein carbonyls and increased nitrotyrosine. • Expression of genes of nitric oxide (NO) and oxidative stress metabolism and urine NO metabolite levels were not changed. • Plasma retinol and tocopherol decreased upon both diets, and carotenoids increased upon PP diet.

Two field trials were conducted to investigate the influence of fungicide and fertilization management on the potato tubers' metabolome (

Metabolomics-based approaches are still receiving growing attention with regard to food authenticity testing. Such studies require enormous sample numbers with negligible experimental or analytical variations to obtain statistically reliable results. In this context, an extraction protocol in line with optimized ionization parameters was developed in consideration of potential starch-derived matrix effects focusing on the polar lipids of potatoes. Therefore, well-known extractions (Bligh and Dyer, Folch, Matyash, and a n-hexane-based procedure) were compared in a non-targeted and a targeted approach regarding the extractability of their lipids such as phosphatidylcholines, phosphatidylethanolamines, galacto- and glucocerebrosides, di- and triglycerides, and acylated steryl glucosides. The selected Folch method was also scrutinized in view of its ability to remove the matrix&rsquo, s starch and consequently improved by substituting trichlormethane with ethyl acetate as a &ldquo, greener&rdquo, Folch approach. Moreover, the challenge of starch-derived contamination and imminent ion suppression in the electrospray ionization source (ESI) was addressed by an optimization of ionization parameters varying desolvation settings, removing injection peaks, and increasing the angles and distances of the ESI-device. Long-term stability tests over five days were performed successfully with a combination of appropriate extraction and decreased desolvation settings during ionization. In conclusion, the present methodology provided the basis for on-going large-scale metabolomic studies with respect to the botanical origin of potatoes using UPLC-IMS-QToF (ultra-high performance liquid chromatography ion mobility spectroscopy quadrupole-time of flight mass spectrometer).

The electrochemical behavior of the vitamers cholecalciferol and ergocalciferol was investigated in order to determine whether it is possible to evaluate phase-I and phase-II metabolism of these steroids and yield metabolites that can serve as reference material. The vitamers were electrochemically-oxidized using an electrochemical system (ROXY&trade, EC system). The influence of pH value, solvent, and potential was evaluated. When using methanol or ethanol, the formation of artificial methoxy or ethoxy groups, respectively, was observed, while the use of acetonitrile did not show any formation of further functional groups. A neutral pH value and use of a constant potential led to the highest number of oxidation products with intensive signals. Additionally, a binding study between vitamin D and glucuronic acid as an example for phase-II conjugation was carried out. It was possible to detect adduct formation. Coupling mass spectrometry directly to electrochemistry (EC-MS) is a promising approach for generating vitamin D metabolites and/or yielding a number of metabolites without in vivo or in vitro test systems. It can support or even replace animal studies in the long-term and might be promising for yielding reference compounds.

One hundred eighty-two authentic potato samples ( Solanum tuberosum) of known variety were collected from various German regions in 2016 and 2017. Samples were extracted with a liquid-liquid-extraction protocol that included isopropanol, methanol, and water in order to focus on lipophilic metabolites. The analysis of nonpolar extracts was performed using an UPLC-IMS-QToF-MS system; data sets obtained were evaluated via multivariate data analysis. A selection of 14 key metabolites with a significant difference in their abundance profiles was identified. This set of markers contained four hydroxylated glucocerebrosides, two phosphoinositols, one phosphocholine, and seven acylated sterol glucosides based on stigmasterol and β-sitosterol, which primarily enable the varietal discrimination. Fragments and neutral losses commonly appearing within one class or subclass of lipids were summarized within a new database that included ion mobility data. The performance of the approach was verified with twenty-nine commercial potato samples.

For centuries, Amaranthus sp. were used as food, ornamentals, and medication. Molecular mechanisms, explaining the health beneficial properties of amaranth, are not yet understood, but have been attributed to secondary metabolites, such as phenolic compounds. One of the most abundant phenolic compounds in amaranth leaves is 2-caffeoylisocitric acid (C-IA) and regarding food occurrence, C-IA is exclusively found in various amaranth species. In the present study, the anti-inflammatory activity of C-IA, chlorogenic acid, and caffeic acid in LPS-challenged macrophages (RAW 264.7) has been investigated and cellular contents of the caffeic acid derivatives (CADs) were quantified in the cells and media. The CADs were quantified in the cell lysates in nanomolar concentrations, indicating a cellular uptake. Treatment of LPS-challenged RAW 264.7 cells with 10 &micro, M of CADs counteracted the LPS effects and led to significantly lower mRNA and protein levels of inducible nitric oxide synthase, tumor necrosis factor alpha, and interleukin 6, by directly decreasing the translocation of the nuclear factor &kappa, B/Rel-like containing protein 65 into the nucleus. This work provides new insights into the molecular mechanisms that attribute to amaranth&rsquo, s anti-inflammatory properties and highlights C-IA&rsquo, s potential as a health-beneficial compound for future research.

The potential benefit of data fusion based on different complementary analytical techniques was investigated for two different classification tasks in the field of foodstuff authentication. Sixty-four honey samples from three different botanical origins and 53 extra virgin olive oil samples from three different geographical areas were analyzed by attenuated total reflection IR spectroscopy (ATR/FT-IR) and headspace gas chromatography-ion mobility spectrometry (HS-GC-IMS). The obtained datasets were combined in a low-level data fusion approach with a subsequent multivariate classification by principal component analysis-linear discriminant analysis (PCA-LDA) or partial least squares-discriminant analysis (PLS-DA). Performing a back projection of PCA loadings, the influence of variables in the FT-IR spectra (one-dimensional) and the GC-IMS profiles (two-dimensional) on the discrimination was visualized within the original axis of the two data sources. Validation results of the classification models were compared to the results that could be obtained by using the individual data blocks separately. For both the honey and olive oil samples, a decreased cross-validation error rate and more robust model was obtained due to the low-level data fusion. The results show that data fusion is an effective strategy for improving the classification performance, particularly for challenging classification tasks such as the discrimination of olive oils with different geographical origin. Graphical abstract.

In this study, a fully automated two-dimensional liquid chromatography system was evaluated for pesticide analysis. The first dimension was a hydrophilic interaction liquid chromatography column for matrix separation. The analytes were collected on a trap column and transferred to the second dimension with a reversed phase column. This separation system was coupled to a quadrupole time-of-flight mass spectrometer. Investigations were started to elucidate, whether this system is useful for screening purposes. Therefore, analytes with different masses, pKa and log Kow values were chosen. The aim was to test, if broadening the scope of this method, without a time-intensive adjustment of the valve switching times is possible. All in all, the limits of the system were determined. It is important that the analytes elute from the hydrophilic interaction liquid chromatography column within a small window. It could be deduced that the window within which the analytes elute expanded too far with the chosen analytes. Therefore, optimization with different buffers, columns, column temperatures, and flow rates was started to minimise the window. Furthermore, differences in the analytes’ elution behavior during hydrophilic interaction liquid chromatography separation were elucidated.

Consumption of Brassica vegetables is linked to health benefits, as they contain high concentrations of the following secondary plant metabolites (SPMs): glucosinolate breakdown products, carotenoids, chlorophylls, and phenolic compounds. Especially Brassica vegetables are consumed as microgreens (developed cotyledons). It was investigated how different ontogenetic stages (microgreens or leaves) of pak choi (Brassica rapa subsp. chinensis) and kale (Brassica oleracea var. sabellica) differ in their SPM concentration. The impact of breadmaking on SPMs in microgreens (7 days) and leaves (14 days) in pak choi and kale as a supplement in mixed wheat bread was assessed. In leaves, carotenoids, chlorophylls, and phenolic compounds were higher compared to those of microgreens. Breadmaking caused a decrease of SPMs. Chlorophyll degradation was observed, leading to pheophytin and pyropheophytin formation. In kale, sinapoylgentiobiose, a hydroxycinnamic acid derivative, concentration increased. Thus, leaves of Brassica species are suitable as natural ingredients for enhancing bioactive SPM concentrations in bread.

There are a lot of health-beneficial properties that are described for plant phenolic compounds. However, many of them have not been sufficiently proven to date. While there is an increase in knowledge about the bioavailability of phenolic compounds, the physiological modes of action are still discussed controversially. One possible explanation for the high number of varying results might be the interactions between phenolic compounds and other compounds (which are still not considered sufficiently). From the several possibilities of interactions, covalent reactions seem to be the most important ones, as they irreversibly affect the properties of both reaction partners. The phenolic compounds are highly reactive and many of them are able to be oxidized to their corresponding semiquinones and quinones, which are further capable of undergoing covalent reactions with an enormous number of nucleophiles, especially proteins that seem to be a nonneglectable reaction partner, on different occasions and reaction sites and due to their concentration. As a result, properties of both reaction partners are significantly affected. Consequently, analysis of such reaction products is quite complex and therefore challenging. Although there are already many possibilities to use methods related to proteomics, the development of innovative methods seems to be necessary to decipher reaction mechanisms and physiological consequences of such protein derivatives.

This document is the Accepted Manuscript version of a Published Work that appeared in final form in Journal of Agricultural and Food Chemistry, copyright © American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see https://doi.org/10.1021/acs.jafc.9b06119., The aroma properties of cocoa nibs obtained by applying a novel postharvest treatment were investigated using methods of the molecular sensory science approach, i.e., solvent extraction and solvent-assisted flavor evaporation, aroma extract dilution analysis (AEDA), stable isotope dilution analysis, calculation of odor activity values (OAVs), and orthonasal sensory evaluation; those properties were then compared to the unfermented and dried raw material and a traditionally fermented sample of the same harvest. For the treatment, unfermented and dried cocoa nibs were, first, rehydrated with lactic acid and ethanol solution to adjust the pH value to 5.1 and, second, incubated under aerobic conditions for 72 h at 45 °C and subsequently dried. This treatment was used to induce enzymatic reactions within the cotyledon matrix, which also occur inside the bean during microbial fermentation of the surrounding fruit pulp. The results of the AEDA showed that many of the key aroma compounds found in fermented and dried cocoa increased during the incubation treatment. Especially some “fruity” esters were found with an equal or even higher flavor dilution (FD) factor in the incubated sample compared to the fermented sample, whereas the fermented sample showed high FD factors for “pungent, sour” and “sweaty” acids, such as acetic acid and 2- and 3-methylbutanoic acids. The quantitative data and calculated OAVs for the samples supported the findings of the AEDA, underlining the potential of this approach as a controllable and reproducible alternative postharvest treatment.

Electrospray mass spectrometry-profiling guided the metabolome investigation of a C18 reversed phase adsorbate of Opuntia stricta var. dillenii fruits following analytical, and semi-preparative high-performance countercurrent chromatography (HPCCC) fractionation, and visualization of molecular weight elution profiles based on selected single ion-traces. Experimental partition-ratio values KD, and peak widths for detected metabolites were determined. Structural characterization of metabolites, and co-elution effects were monitored in the scan range m/z 150-2200. The polar ion-pair activated solvent system tert.-butylmethylether - n-butanol - acetonitrile - water (0.7% trifluoroacetic acid) [2:2:1:5]) was used for partition-ratio KD improvement of ionic betalains. HPCCC operations were in the head-to-tail mode using the elution-extrusion approach. Selected ESI-MS ion traces visualized the elution of fourty-three metabolites, whereas twenty-one were identified as known betalain pigments, and their classic degradation products, and chlorinated betacyanin artefacts. Potentially, novel fruit metabolites of Opuntia were recognized by unknown molecular weights, and MS/MS fragmentations. Off-line ESI-MS fraction monitoring determined peak elution windows, and resulted in KD-based chromatographic scales. Detectable metabolites were compared by separation- α, and resolution-factors RS revealing a better performance of the analytical HPCCC experiment. Experimental metabolite KD-values from analytical and semi-preparative HPCCC runs were widely consistent, and confirmed the reproducibility of the technique based on the used sample material.

Due to the adaption to environmental stress, biomasses of microalgae and cyanobacteria contain high-value pigments and phenolic compounds with antioxidant capacity to be used as ingredients for food, cosmetics, and pharmaceuticals. To expand the current state of knowledge about microalgae and cyanobacteria as sources of high-value compounds biomasses and extracts of 13 microalgal and cyanobacterial species of the four divisions, namely Chlorophyta, Rhodophyta, Cyanophyta and Diatoms, with regards to their antioxidant capacity, were characterized. Moreover, cell disruption prior to extraction of the slurry of the alga Acutodesmus obliquus was investigated for increasing yields of compounds with antioxidant capacity. The total chlorophyll and carotenoids contents of the tested green algae was with up to 60 mg g−1 and 12 mg g−1, respectively, substantially higher compared to the other algae. Results revealed that aqueous extraction is effective for extracting phenolic compounds from biomass. The quantity of phenolic compounds in the extracts revealed no superior species. Nevertheless, Neochloris oleobundans, Phormidium sp., and Wilmottia murravi can be recommended as species with high contents in phenolic compounds (>20 mg gallic acid eq. g−1). Phormidium ambiquum and Phormidium sp. were the strains with highest contents in water-soluble compounds (12–14 μmol ascorbic acid eq. g−1), while N. oleobundans and A. obliquus were the strains with highest contents in lipid-soluble compounds (60–80 μmol trolox eq. g−1). Cell disruption prior to extraction doubled the yields of compounds with antioxidant activity, which underlines the necessity of an effective pre-treatment. Results of this study contribute to the proceeding towards a utilization of microalgal and cyanobacterial biomasses for the formation of high-value compounds with antioxidant properties and potential for food, cosmetic, and pharmaceutical industries.

Extracts of Opuntia stricta var. dillenii fruits were fractionated by semi-preparative high-performance countercurrent chromatography (HPCCC) to study the secondary metabolite formation, whereby HPCCC showed a superior separation capacity to fractionate minor metabolites compared to HPLC. A family of new peptides was detected in semi-polar fractions when monitoring the HPCCC separation by off-line injections of fractions to ESI-MS/MS. Planar structures of the major compounds, two 14-ring-membered cyclopeptide alkaloids, which were named opuntisines A and B, were elucidated by 1D- and 2D-NMR spectroscopy and HR-ESI-MS/MS spectrometry, while a combination of chemical derivatisation and degradation revealed the stereo-configurations. Specifically, the methods of Marfey and Mosher indicated l -Glu, l -Ile, l -Phe and 1S-configurations, respectively; ROESY correlations revealed 8S, 9S. The novel opuntisine A showed moderate activity against the Gram-negative bacterium Escherichia coli, but no further antibacterial, antifungal nor cytotoxic effects. This bioactive natural product class is reported for the first time in the plant family Cactaceae.

Schisandra chinensis is a widely-consumed nutraceutical providing advantageous nutritional and bioactive properties. This study aimed at characterizing the chemical composition and antioxidant capacity of S. chinensis fruits as well as the underestimated leaves and stems. A special aim of this study was to characterize the contribution of single constituents (lignans and further phenolic compounds) to the overall antioxidant activity using an HPLC-online TEAC approach. By using LC-DAD/ESI-ToF-MS, twenty-eight lignans and twenty phenolic compounds were identified. Among the 48 characterized components, twelve phenolic compounds were identified for the first time in Schisandra. Gomisin D was the only lignan being able to scavenge ABTS+ radicals, suggesting that not its amount but rather its chemical structure is responsible. Overall, clorogenic acid isomers and quercetin glycosides contribute to over 80% of the total antioxidant activity. This study indicates that leaves and stems of S. chinensis may be used for obtaining bioactive formulations and functional foods.

SCOPE Benzyl isothiocyanate (BITC), which occurs in Brassicales, has demonstrated chemopreventive potency and cancer treatment properties in cell and animal studies. However, fate of BITC in human body is not comprehensively studied. Therefore, the present human intervention study investigates the metabolism of the glucosinolate (GSL) glucotropaeolin and its corresponding BITC metabolites. Analyzing BITC metabolites in plasma and urine should reveal insights about resorption, metabolism, and excretion. METHODS AND RESULTS Fifteen healthy men were randomly recruited for a cross-over study and consumed 10 g freeze-dried Indian cress as a liquid preparation containing 1000 μmol glucotropaeolin. Blood and urine samples were taken at several time points and investigated by LC-ESI-MS/MS after sample preparation using SPE. Plasma contained high levels of BITC-glutathione (BITC-GSH), BITC-cysteinylglycine (BITC-CysGly), and BITC-N-acetyl-L-cysteine (BITC-NAC) 1-5 h after ingestion, with BITC-CysGly appearing as the main metabolite. Compared to human plasma, the main urinary metabolites were BITC-NAC and BITC-Cys, determined 4-6 h after ingestion. CONCLUSION This study confirms that consumption of Indian cress increases the concentration of BITC metabolites in human plasma and urine. The outcome of this human intervention study supports clinical research dealing with GSL-containing innovative food products or pharmaceutical preparations.

In previous studies, it was hypothesized that isorhamnetin glycosides are the dominant flavonol derivatives occurring in the peels and cladodes of cactus Opuntia ficus-indica fruits and might be used as markers for analyzing the authenticity of food products containing (whole) cactus fruits or cladodes. In the present work, use of the whole yellow-orange fruit from cactus O. ficus-indica as an ingredient of rice- and corn-based snacks was evaluated. The impact of extrusion cooking on the flavonol profile was characterized in rice- and corn-based extruded products fortified with a freeze-dried preparation from the whole cactus fruit. HPLC analysis showed that the flavonol profile of cactus O. ficus-indica consisted of five prominent isorhamnetin glycosides with a concentration of 148.9 μg/100 mg dw. In contrast, the rice or corn grits used in the present study did not contain any flavonol at all. The freeze-dried fruit was added in different ratios (0, 2, 6, and 10%) to the rice or corn grits prior to the extrusion process. Analysis of the final extrusion products showed that flavonol profiles from the fortified rice or corn products, even at the lowest level of addition (2%), were similar to the original profile of the fruit before processing. Furthermore, the total flavonol content in extruded products was not significantly affected by extrusion cooking. This study showed that the unique flavonol profile of fruits from cactus O. ficus-indica might serve as a biochemical marker for the evaluation of the authenticity of products made from whole cactus fruit or the fruit's peel.

Amaranth is a plant genus of global importance comprising more than 60 species that are dually used for human consumption. While the grains are used as pseudo-cereals mainly in America and Asia, leaves are also consumed as leafy vegetable in African countries. Besides further secondary plant metabolites, saponins are described as major bioactive constituents in amaranth species. These triterpenoid saponins belonging to the oleanane-type are assumed to be part of the plant defense system and are often also associated with potential health risks for the consumer, mainly due to hemolytic properties. However, data concerning amaranth saponins are limited to the grains of single cultivars of only a few species investigated. The aim of the present work was to determine the saponin profile in leaves of various amaranth cultivars grown under identical conditions. Out of 15 cultivars, six did not show any indications for the presence of saponins in HPLC–MS analysis. Two saponin-rich cultivars (one of Amaraanthus hybridus and one of Amaranthus hypochondriacus) as well as commercially available amaranth grains were selected for an in-depth analysis using a combined approach of high resolution and multi stage mass spectrometry. Three previously undescribed monodesmosidic and four bidesmosidic saponins could be assigned according to the MS data. Four novel saponins were also found in commercial grain amaranth analyzed for comparison. The investigation of the hemolytic effects revealed that only one saponin exerts significant activity whilst the further saponins did not lyse erythrocytes in vitro. The results show that the saponin profile of amaranth cultivars is more diverse than reported so far. However, the biological activity seems to be different for the single structures. Thus, a more comprehensive case-by-case investigation of amaranth saponins is required to evaluate the impact of these secondary metabolites on humans and plants.

In this study, a clean-up approach using a two-dimensional liquid chromatography (2D-LC) consisting of a hydrophilic interaction liquid chromatography column and a reversed phase column was investigated. A fully automated 2D-LC system was used and compared with a traditional quick, easy, cheap, effective, rugged, and safe (QuEChERS) liquid chromatography (QuE-LC) method. The comparison was based on the results of a validation of selected analytes. It was investigated whether the detectability of analytes could be improved by the use of the 2D-LC. On the basis of these results, the relative detection rates were determined for every matrix. By means of those detection rates, the matrices were categorized regarding their complexity. Furthermore, the applicability of the 2D-LC was tested by participation in the European Proficiency Test in Fruits and Vegetables Screening Methods. In order to evaluate the separation and the elution profile of matrix components, multivariate data analysis was applied. For this purpose, ten matrices were processed in accordance to a QuEChERS protocol and the protocol for 2D-LC analysis. Moreover, the reagent blanks of the corresponding matrix were processed and analyzed by QuE-LC and 2D-LC. The results allowed evaluating the number of detected compounds for both methods. Additionally, the influence of compounds originating from reagent blanks can be estimated. In general, less compounds could be detected when 2D-LC was used. Especially, these were very polar compounds and compounds with m/z values >1500. These compounds seem to originate primarily from the used reagents especially from the citrate salts. However, the most of these compounds could be separated and were not detectable any more when 2D-LC was used. The results of the comparison based on validation and participation in the European Proficiency Test also show a better detectability for the most analytes with 2D-LC.

So far, only a few cases of immunoglobulin E (IgE)-mediated coconut allergies have been described in the literature. Due to a growing consumption of coconut-containing foods in occidental countries, the number of coconut allergies may also increase. As there is no causative immunotherapy in clinical routine, appropriate food labelling is particularly important, also with regard to cross-contamination, to prevent serious health consequences. The purpose of this study was to develop a DNA-based detection method for coconut (Cocos nucifera). Initially, three sets of coconut-specific primers were designed and tested. A TaqMan&trade, probe was then developed to identify and quantify coconut by real-time PCR assay. With 27 other plant and animal species, the specificity of the primer/probe system was tested and cross reactivity was excluded. In a dilution series, a limit of detection of 1 pg/&micro, L was determined. Thus, the developed real-time PCR assay is a suitable method to detect coconut in food.

A broad range of compounds/ingredients for the substitution of fishmeal and fish oil in feeds for aquaculture was suggested over the last decades. This study aimed at characterizing changes in antioxidant activity from the raw materials over the fish feed production to the accumulation of potential health-beneficial antioxidants in the fish fillets of Oncorhynchus mykiss, when being fed with an innovative microorganism-based fish feed. The feed used consisted of a mix of the cyanobacterium Arthrospira sp., the microalgae Crypthecodinium cohnii, and the yeast Rhodotorula glutinis. These ingredients formed a balanced feed to demonstrate no limitations in fishmeal and oil substitution with regard to lipids, proteins, and antioxidants. The enrichment with bioactive compounds stabilizing the feed as well as enhancing bioactivity in the fish was successful. Also, radical scavenging activity was documented from the raw materials over the extrusion process to the accumulation in the fillets of rainbow trout (Oncorhynchus mykiss) over the whole fish production period of 16 weeks. As an additional quality parameter, the accumulation of pigments in the fish fillets was determined. The study revealed changes in the antioxidant profile over the whole value chain, which resulted in healthy organisms of higher quality compared to conventionally fed organisms.

Biofortified apples seem to be a suitable produce. In this study, different selenium forms and application levels were applied to the two apple varieties ‘Golden Delicious’ and ‘Jonagold’, grown in the years 2017 and 2018 in order to increase the selenium uptake within a typical Western diet. It was shown that the biofortification, which was performed as a foliar application implemented in usual calcium fertilization, led to significantly increased selenium contents in the fruits. Furthermore, biofortification affected the total phenolic content (TPC), the polyphenol oxidase activity (PPO), as well as the antioxidant activity (AOA), the latter measured with the two well-known assays Trolox Equivalent Antioxidant Capacity Assay (TEAC) and Oxygen Radical Absorbance Capacity Assays (ORAC). The varying selenium forms and application levels showed a differing influence on the parameters mentioned before. Higher fertilizer levels resulted in higher selenium accumulation. It was found that PPO activity fluctuates less in biofortified apples. With regard to TPC, selenate led to higher amounts when compared to the untreated controls and selenite resulted in lower TPC. AOA analysis showed no clear tendencies as a result of the selenium biofortification. In the case of ‘Jonagold’, a higher AOA was generally measured when being biofortified, whereas, in the case of ‘Golden Delicious’, only one form of application led to higher AOA. Additionally, differences in the amount of major phenolic compounds, measured with High Performance Liquid Chromatography Mass Spectrometry (HPLC-DAD-ESI-MSn), were observed, depending on the conditions of the biofortification and the variety.

The present human intervention trial investigated the health-promoting potential of B. carinata, with a focus on effects of thermal processing on bioactivity. Twenty-two healthy subjects consumed a B. carinata preparation from raw (allyl isothiocyanate-containing) or cooked (no allyl isothiocyanate) leaves for five days in a randomized crossover design. Peripheral blood mononuclear cells were exposed to aflatoxin B1 (AFB1), with or without metabolic activation using human S9 mix, and subsequently analyzed for DNA damage using the comet assay. Plasma was analyzed for total antioxidant capacity and prostaglandin E2 (PGE2) levels. Cooked B. carinata significantly reduced DNA damage induced by AFB1 as compared to baseline levels (+S9 mix: 35%, −S9 mix: 33%, p ≤ 0.01, respectively). Raw B. carinata only reduced DNA damage by S9-activated AFB1 by 21% (p = 0.08). PGE2 plasma levels were significantly reduced in subjects after consuming raw B. carinata. No changes in plasma antioxidant capacity were detectable. A balanced diet, including raw and cooked Brassica vegetables, might be suited to fully exploit the health-promoting potential. These results also advocate the promotion of B. carinata cultivation in Eastern Africa as a measure to combat effects of unavoidable aflatoxin exposure.

Background As the gut microbiota contributes to metabolic health, it is important to determine specific diet-microbiota interactions that influence host metabolism. Bile acids and dietary fat source can alter phenotypes of diet-induced obesity, but the interplay with intestinal microorganisms is unclear. Here, we investigated metabolic consequences of diets enriched in primary bile acids with or without addition of lard or palm oil, and studied gut microbiota structure and functions in mice. Results In combination with bile acids, dietary lard fed to male C57BL/6N mice for a period of 8 weeks enhanced fat mass accumulation in colonized, but not in germ-free mice when compared to palm oil. This was associated with impaired glucose tolerance, lower fasting insulin levels, lower counts of enteroendocrine cells, fatty liver, and elevated amounts of hepatic triglycerides, cholesteryl esters, and monounsaturated fatty acids. Lard- and bile acid-fed mice were characterized by shifts in dominant gut bacterial communities, including decreased relative abundances of Lachnospiraceae and increased occurrence of Desulfovibrionaceae and the species Clostridium lactatifermentans and Flintibacter butyricus. Metatranscriptomic analysis revealed shifts in microbial functions, including lipid and amino acid metabolism. Conclusions Caution is required when interpreting data from diet-induced obesity models due to varying effects of dietary fat source. Detrimental metabolic consequences of a diet enriched with lard and primary bile acids were dependent on microbial colonization of the host and were linked to hepatic lipid rearrangements and to alterations of dominant bacterial communities in the cecum. Electronic supplementary material The online version of this article (10.1186/s40168-018-0510-8) contains supplementary material, which is available to authorized users.

Pu&rsquo, er tea produced from Camellia sinensis var. assamica is a widely appreciated and consumed beverage that can be divided into two kinds of tea depending on the different fermentation processed used, the special sensory characteristics, and their chemical composition. However, authentication seems to be very important for such teas, as they are traded to comparatively high prices, especially in Europe. The results for selected biochemical markers showed that aged raw pu&rsquo, er tea contained 210.2 mg GAE/g polyphenols, of which 2.2 mg/g were gallic acid, 16.1 mg/g theogallin, 35.1 mg/g (&minus, )-epigallocatechin gallate, and 40.1 mg/g (&minus, )-epicatechin gallate, on average. Young ripened pu&rsquo, er tea contained about 104.6 mg GAE/g polyphenols, of which 5.5 mg/g gallic acid, 0.9 mg/g theogallin, 0.7 mg/g (&minus, )-epigallocatechin gallate, and 1.8 mg/g (&minus, )-epicatechin gallate, on average. An additional objective of the present study was to unravel the best brewing conditions for optimal extraction of the bioactive compounds. Infusions of nineteen commercial teas (from pu&rsquo, er cakes) were obtained at different time-temperature ratios for studying the content of bioactive compounds (flavan-3-ols, flavonols, caffeoylquinic acids, methylxanthines). Brewing at 90 &deg, C for 5 min was the best condition to obtain a high content of total polyphenols in ripened pu&rsquo, er tea. Principal component analysis and hierarchical cluster analysis showed, that young ripened and aged raw pu&rsquo, er tea can be successfully differentiated by the analyzed chemical compounds. Principal component analysis results indicated that young ripened pu&rsquo, er tea has higher contents of gallic acid, quercetin, and kaempferol than aged raw pu&rsquo, er tea.

For the first time, this study describes a HS-GC-IMS strategy for analyzing non-targeted volatile organic compounds (VOCs) profiles to distinguish between virgin olive oils of different classification. Correlations among measured flavor characteristics and sensory attributes evaluated by a test panel were determined by applying unsupervised (PCA, HCA) and supervised (LDA, kNN and SVM) chemometric techniques. PCA and HCA were applied for natural clustering of the samples and LDA, kNN, and SVM methods were used to create predictive models for olive oil classification. Identification of 26 target compounds revealed which compounds are responsible for discrimination, and how their distribution correlates with the sensory evaluation. In the investigated samples, LDA, kNN, and SVM models correctly classified 83.3%, 73.8%, and 88.1% of the samples, respectively. This suggests that mathematical correlations of HS-GC-IMS 3D fingerprints with the sensory analysis may be appropriate for calculating a good predictive value to classify virgin olive oils.

Glucosinolates and their breakdown products, especially isothiocyanates (ITCs), are hypothesized to exert a broad range of bioactivities. However, physiological mechanisms are not yet completely understood. In this study, formation of protein conjugates after incubation with benzyl isothiocyanate (BITC) was investigated in vitro. A survey of protein conjugates was done by determining BITC cysteine and lysine amino acid conjugates after protein digestion. Therefore, a liquid chromatography-tandem mass spectrometry (LC-ESI-MS/MS) method was developed and validated. Stability studies showed that cysteine conjugates are not stable under alkaline conditions, and lysine conjugates did not show any correlation to pH values, although stability increased at low temperatures. Lysine conjugates were the preferred form of protein conjugates, and longer BITC exposure times led to higher amounts. Knowledge about the reaction sites of ITCs in eukaryotic cells may help to understand the mode of action of ITCs leading to health promoting as well as toxicological effects in humans.

Microalgae bear a great potential to produce lipids for biodiesel, feed, or even food applications. To understand the still not well-known single-cell dynamics during lipid production in microalgae, a novel single-cell analytical technology was applied to study a well-established model experiment. Multidimensional single-cell dynamics were investigated with a non-supervised image analysis technique that utilizes data from epi-fluorescence microscopy. Reliability of this technique was successfully proven via reference analysis. The technique developed was used to determine cell size, chlorophyll amount, neutral lipid amount, and deriving properties on a single-cellular level in cultures of the biotechnologically promising alga Acutodesmus obliquus. The results illustrated a high correlation between cell size and chlorophyll amount, but a very low and dynamic correlation between cell size, lipid amount, and lipid density. During growth conditions under nitrogen starvation, cells with low chlorophyll content tend to start the lipid production first and the cell suspension differentiated in two subpopulations with significantly different lipid contents. Such quantitative characterization of single-cell dynamics of lipid synthesizing algae was done for the first time and the potential of such simple technology is highly relevant to other biotechnological applications and to deeper investigate the process of microalgal lipid accumulation.

Zearalenone (ZEN) and its phase II sulfate and glucoside metabolites have been detected in food and feed commodities. After consumption, the conjugates can be hydrolyzed by the human intestinal microbiota leading to liberation of ZEN that implies an underestimation of the true ZEN exposure. To include ZEN conjugates in routine analysis, reliable standards are needed, which are currently not available. Thus, the aim of the present study was to develop a facilitated biosynthesis of ZEN-14-sulfate, ZEN-14-glucoside and ZEN-16-glucoside. A metabolite screening was conducted by adding ZEN to liquid fungi cultures of known ZEN conjugating Aspergillus and Rhizopus strains. Cultivation conditions and ZEN incubation time were varied. All media samples were analyzed for metabolite formation by HPLC-MS/MS. In addition, a consecutive biosynthesis was developed by using Fusarium graminearum for ZEN biosynthesis with subsequent conjugation of the toxin by utilizing Aspergillus and Rhizopus species. ZEN-14-sulfate (yield: 49%) is exclusively formed by Aspergillus oryzae. ZEN-14-glucoside (yield: 67%) and ZEN-16-glucoside (yield: 39%) are formed by Rhizopus oryzae and Rhizopus oligosporus, respectively. Purities of ≥73% ZEN-14-sulfate, ≥82% ZEN-14-glucoside and ≥50% ZEN-16-glucoside were obtained by 1H-NMR. In total, under optimized cultivation conditions, fungi can be easily utilized for a targeted and regioselective synthesis of ZEN conjugates.

This work describes a simple approach for the untargeted profiling of volatile compounds for the authentication of the botanical origins of honey based on resolution-optimized HS-GC-IMS combined with optimized chemometric techniques, namely PCA, LDA, and kNN. A direct comparison of the PCA-LDA models between the HS-GC-IMS and