Nile red staining for rapid screening of plastic-suspect particles in edible seafood tissues.

Concerns regarding microplastic (MP) contamination in aquatic ecosystems and its impact on seafood require a better understanding of human dietary MP exposure including extensive monitoring. While conventional techniques for MP analysis like infrared or Raman microspectroscopy provide detailed particle information, they are limited by low sample throughput, particularly when dealing with high particle numbers in seafood due to matrix-related residues. Consequently, more rapid techniques need to be developed to meet the requirements of large-scale monitoring. This study focused on semi-automated fluorescence imaging analysis after Nile red staining for rapid MP screening in seafood. By implementing RGB-based fluorescence threshold values, the need for high operator expertise to prevent misclassification was addressed. Food-relevant MP was identified with over 95% probability and differentiated from natural polymers with a 1% error rate. Comparison with laser direct infrared imaging (LDIR), a state-of-the-art method for rapid MP analysis, showed similar particle counts, indicating plausible results. However, highly variable recovery rates attributed to inhomogeneous particle spiking experiments highlight the need for future development of certified reference material including sample preparation. The proposed method demonstrated suitability of high throughput analysis for seafood samples, requiring 0.02-0.06 h/cm2 filter surface compared to 4.5-14.7 h/cm with LDIR analysis. Overall, the method holds promise as a screening tool for more accurate yet resource-intensive MP analysis methods such as spectroscopic or thermoanalytical techniques.

Physicochemical and Sensory Characterization of Whey Protein-Enriched Semihard Cheese.

In view of potential future changes of German food legislation with regard to cheese product quality parameters, this study aimed to evaluate the quality of whey protein-enriched semihard cheese (WPEC). Model WPEC was produced in a pilot plant and on an industrial scale by adding defined amounts of high-heat (HH) milk to the cheese milk and comprehensively analyzed during cheese processing. The dry matter, total protein, pure protein, fat, and sodium chloride content of six-week ripened cheese samples were not significantly different (p < 0.05) when the technologically necessary heating of the curd was adapted to the amount of HH milk. However, the ripening, firmness, and melting behavior of WPEC was different compared to cheese without HH milk. During ripening, no formation of whey protein peptides was observed, but differences in the amount of some bitter peptides deriving from the casein fraction were found. Sensory data suggested a slightly more bitter taste perception by the panelists for the WPEC. Further technological adjustments are recommended to obtain marketable WPEC.

Fast and User-Friendly Detection of Flatfish Species (Pleuronectes platessa and Solea solea) via Loop-Mediated Isothermal Amplification (LAMP).

The detection of a Cytochrome b gene (cytb) for species differentiation in fish is intensively used. A fast alternative to expensive and time-consuming DNA barcoding is loop-mediated isothermal amplification (LAMP) in combination with efficient readout systems. For this reason, we developed LAMP assays for rapid species detection of Pleuronectes platessa and Solea solea, two economically important flatfish species in Europe that are prone to mislabeling. Species-specific primer sets targeting cytb were designed, and LAMP assays were optimized. With the optimized LAMP assays, we were able to detect up to 0.1 and 0.01 ng of target DNA of P. platessa and S. solea, respectively, and in each case up to 1% (w/w) of target species in mixtures with nontarget species. For future on-site detection, a lateral flow assay and a pocket-sized lab-on-phone assay were used as readout systems. The lab-on-phone assay with the S. solea specific primer set revealed cross-reactivity to Solea senegalensis. The assay targeting P. platessa proved to be highly specific. Both assays could be performed within 45 min and provided rapid and easy detection of fish species.

Identification of Marker Peptides for the Whey Protein Quantification in Edam-Type Cheese.

Several technologies are available for incorporating whey proteins into a cheese matrix. However, there is no valid analytical method available to determine the whey protein content in matured cheese, to date. Consequently, the aim of the present study was to develop a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the quantification of individual whey proteins based on specific marker peptides ('bottom-up' proteomic approach). Therefore, the whey protein-enriched model of the Edam-type cheese was produced in a pilot plant and on an industrial scale. Tryptic hydrolysis experiments were performed to evaluate the suitability of identified potential marker peptides (PMPs) for α-lactalbumin (α-LA) and ß-lactoglobulin (ß-LG). Based on the findings, α-LA and ß-LG appeared to be resistant to proteolytic degradation during six weeks of ripening and no influence on the PMP was observed. Good levels of linearity (R2 > 0.9714), repeatability (CVs < 5%), and recovery rate (80% to 120%) were determined for most PMPs. However, absolute quantification with external peptide and protein standards revealed differences in model cheese depending on the PMP, e.g., 0.50% ± 0.02% to 5.31% ± 0.25% for ß-LG. As protein spiking prior to hydrolysis revealed differing digestion behavior of whey proteins, further studies are required to enable valid quantification in various cheese types.

Uridine as a non-toxic actinometer for UV-C treatment: influence of temperature and concentration.

UV-C treatment is an effective method to inactivate microorganisms and therefore gets increasingly more attention in food industry, especially for liquid products. To test and monitor different UV-C reactor designs, a photochemical actinometer is required that gives reliable UV-C dose values and is non-toxic allowing frequent control of the production chain. Here, a variable concentrated aqueous uridine solution is tested as a photochemical actinometer. Uridine reacts at 262 nm by photohydration to a single photoproduct not absorbing any light. A concentration dependent quantum yield (Ф) was quantified in the range of 0.2-3.0 mM uridine. Results show that uridine is as accurate as the commonly accepted iodide/iodate actinometry, but not as precise. Especially at higher concentrations a higher number of measurements becomes necessary. Further, a temperature correction is presented for 10 °C > ϑ > 30 °C. Taking these results into account, uridine can certainly be considered as a non-toxic dosimeter for UV-C systems.

Objectively measured adherence to physical activity among patients with coronary artery disease: Comparison of the 2010 and 2020 World Health Organization guidelines and daily steps.

Background: Tailored recommendations for patients after percutaneous coronary interventions (PCI) need physical activity (PA) to be objectively measured and assessed for adherence to guidelines. The recent WHO guidelines removed the daily recommended bout duration, while the potential impact of this change on patients after PCI remains unclear. Aim: We evaluated prevalence estimates of adherence to PA recommendations among patients after PCI across the 2010 [≥30 min moderate- to vigorous-intensity PA (MVPA) at ≥ 10-min bout duration] and 2020 WHO guidelines (≥30 min of MVPA of any bout duration), as well as 7,500 and 10,000 steps. Methods: We conducted an observational longitudinal single-center study with patients after PCI for chronic or acute coronary syndrome (ACS); maximal age 80 years. Wrist-worn accelerometers recorded participants' PA data from the evening of hospital discharge over the next 18 days. Results: We analyzed data from 282 participants with sufficient minimum wear time (7 days of ≥12 h), including 45 (16%) women; and 249 (88%) with ACS. Median wear time was 18 (17, 18) days. Median participant age was 62 (55, 69) years. Fifty-two participants (18.4%) fulfilled 2010 WHO guidelines and 226 (80.1%) fulfilled the 2020 WHO guidelines. Further, 209 (74.1%) participants achieved ≥7,500 steps/day and 155 (55.0%) performed ≥10,000 steps/day. Conclusion: Among participants after PCI, most MVPA was accumulated in bouts <10 min, leading to a fourfold discrepancy between participants fulfilling the 2010 and 2020 WHO PA recommendations. The number of steps/day may be a valid proxy to recent WHO PA recommendations as it is not dependent on the bout-length definition. Clinical trial registration: [ClinicalTrials.gov], identifier [NCT04663373].

Influence of processing and storage on the iodine content of meat and fish products using iodized salt.

Table salt fortified with KIO3 is commonly used to prevent iodine deficiency disorders. However, there is a lack of reliable data about the stability of KIO3 during food processing. In this study several meat and fish products were prepared with iodized salt and the iodine stability was determined through the whole production process. Applied processes included heating, fermenting, freezing, hot smoking, ripening by enzymes and storing. In all products an increase in iodine content was observed after addition of iodized salt. The iodine content remained constant during most of the applied processes. The only iodine loss was observed in ham after heating and can be explained by loss of iodine containing brine. During subsequent storage no iodine loss was observed in any of the products. The use of KIO3 fortified salt in the investigated products might therefore be beneficial for the iodine supply.

High-Performance Thin-Layer Chromatography-Immunostaining as a Technique for the Characterization of Whey Protein Enrichment in Edam Cheese.

Whey protein-enriched cheese can be produced by means of a high-temperature treatment of a part of the cheese milk. In this way, the nutritional quality of the resulting cheeses can be increased while resources are conserved. High-performance thin-layer chromatography-immunostaining (HPTLC-IS) using specific ß-lactoglobulin (ß-LG) antibodies was applied to study the implementation and stability of ß-LG in two different sample sets of whey protein-enriched Edam model cheeses, including industrial-scale ones. Two methods were compared for the extraction of the proteins/peptides from the cheese samples. By applying tryptic hydrolysis directly from a suspended cheese sample instead of a supernatant of a centrifuged suspension, a better yield was obtained for the extraction of ß-LG. When applying this method, it was found that selected epitopes in the tryptic ß-LG peptides remain stable over the ripening period of the cheese. For four of the tryptic ß-LG peptides detected by immunostaining, the amino acid sequence was identified using MALDI-TOF-MS/MS. One of the peptides identified was the semi-tryptic peptide VYVEELKPTP. A linear relationship was found between the content of this peptide in cheese and the proportion of high-heated milk in the cheese milk. ß-LG enrichment factors of 1.72 (n = 3, sample set I) and 1.33 ± 0.19 (n = 1, sample set II) were determined for the cheese samples containing 30% high-heated milk compared to the non-enriched samples. The relative ß-LG contents in the cheese samples with 30% high-heated milk were calculated to be 4.35% ± 0.39% (sample set I) and 9.11% ± 0.29% (sample set II) using a one-point calibration. It can be concluded that the HPTLC-IS method used is a suitable tool for the analysis of whey protein accumulation in cheese, being therefore potentially directly applicable on an industrial scale. For more accurate quantification of the whey protein content in cheese, an enhanced calibration curve needs to be applied.

Two-dimensional high-performance thin-layer chromatography for the characterization of milk peptide properties and a prediction of the retention behavior - a proof-of-principle study.

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, ß-lactoglobulin, α-, ß-, 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.

A DNA microarray for the authentication of giant tiger prawn (Penaeus monodon) and whiteleg shrimp (Penaeus (Litopenaeus) vannamei): a proof-of-principle.

This proof-of-principle study describes the development of a rapid and easy-to-use DNA microarray assay for the authentication of giant tiger prawns and whiteleg shrimp. Following DNA extraction and conventional end-point PCR of a 16S rDNA segment, the PCR products are hybridised to species-specific oligonucleotide probes on DNA microarrays located at the bottom of centrifuge tubes (ArrayTubes) and the resulting signal patterns are compared to those of reference specimens. A total of 21 species-specific probes were designed and signal patterns were recorded for 47 crustacean specimens belonging to 16 species of seven families. A hierarchical clustering of the signal patterns demonstrated the specificity of the DNA microarray for the two target species. The DNA microarray can easily be expanded to other important crustaceans. As the complete assay can be performed within half a day and does not require taxonomic expertise, it represents a rapid and simple alternative to tedious DNA barcoding and could be used by crustacean trading companies as well as food control authorities for authentication of crustacean commodities.

Effect of reduction of sodium content on the microbial ecology of Edam cheese samples.

Sodium intake is a major risk factor for non-communicable diseases. Consequently, reformulation of cheeses such as Edam to contain less sodium may contribute to lowering disease risk. However, sodium is essential for cheese manufacture, influencing starter culture bacteria activity and abundance during fermentation. This study aimed to assess the microbial diversity of reformulated Edam cheese samples with a reduced sodium content using culture-independent technique. The microbial diversity of samples produced using simple sodium reduction, as well as by substituting salt with a mineral salt compound containing potassium, were analysed in comparison to regular control Edam samples during manufacture and the subsequent 6-week ripening period using 16S rDNA metagenomics. In addition, a challenge test using Listeria (List.) innocua as a surrogate species for List. monocytogenes was performed. Reducing sodium content did not influence the microbiological composition of reformulated samples in comparison to that of regular samples. The starter culture bacteria dominated the microbial diversity and no increase in spoilage or potentially pathogenic bacterial growth was detected, including that of List. innocua. From a microbiological perspective, it can be concluded that lowering sodium content in Edam samples without affecting the microbial composition is achievable through simple sodium reduction and through implementation of a mineral salt replacement approach.

Developing an Automatic Color Determination Procedure for the Quality Assessment of Mangos (Mangifera indica) Using a CCD Camera and Color Standards.

Color is one of the key sensory characteristics in the evaluation of the quality of mangos (Mangifera indica) especially with regard to determining the optimal level of ripeness. However, an objective color determination of entire fruits can be a challenging task. Conventional evaluation methods such as colorimetric or spectrophotometric procedures are primarily limited to a homogenous distribution of the color. Accordingly, a direct assessment of the mango quality with regard to color requires more pronounced color determination procedures. In this study, the color of the peel and the pulp of the mango cultivars "Nam Dokmai", "Mahachanok", and "Kent" was evaluated and categorized into various levels of ripeness using a charge-coupled device (CCD) camera in combination with a computer vision system and color standards. The color evaluation process is based on a transformation of the RGB (red, green, and blue) color space values into the HSI (hue, saturation, and intensity) color system and the Natural Color Standard (NCS). The results showed that for pulp color codes, 0560-Y20R and 0560-Y40R can be used as appropriate indicators for the ripeness of the cultivars "Nam Dokmai" and "Mahachanok". The peels of these two mango cultivars present two distinct colors (1050-Y40R and 1060-Y40R), which can be used to determine the fruit maturity during the post-ripening process. However, in the case of the cultivar "Kent", peel color detection was not an applicable approach for determining ripeness; thus, the determination of the pulp color with the color code 0550-Y20R gave promising results.

Microbial composition of sweetness-enhanced yoghurt during fermentation and storage.

The reformulation of dairy products to contain less added sugar can contribute to reducing sugar consumption, thereby reducing the risk of non-communicable diseases. The objective of this study was to investigate the microbial ecology of reformulated yoghurt, which was produced using bi-enzymatic modification of lactose to increase its sweetness by a factor of 2-3. Ultimately, this reformulation strategy could reduce the amount of added sugar needed for equal sweetness of the end product. The bi-enzymatic modification relied on utilisation of a ß-galactosidase enzyme to convert the milk sugar lactose to galactose and glucose, followed by the enzymatic conversion of the glucose moiety to fructose using a glucose isomerase. The microbial ecology of reformulated yoghurt produced with two mixed starter culture preparations containing either Streptococcus (S.) thermophilus and Lactobacillus (Lb.) delbrueckii or S. thermophilus, Lb. acidophilus and Bifidobacterium sp. strains, was analysed during fermentation and cool storage using 16S rRNA based metagenomics. None of the yoghurt samples showed a significant difference in microbial composition between sweetness-enhanced and regular milk at all sampling time points during manufacture and storage of yoghurt. However, a significant difference between the microbiota of inoculated milk before and after fermentation was observed. In both types of yoghurt, the starter culture genera dominated the microbial ecology at the end of fermentation as expected, reducing the possibility of growth of potentially pathogenic or spoilage bacteria possibly resulting from a changed carbohydrate spectrum.

[Fix the Supply Side! Demand Stimulus Will Not Cure What Ails]. / Corona-Krise: (Wirtschafts-)politische Perspektiven: Die Reflexe aus der Finanzkrise sind nicht genug!

The explosion of Covid-19 cases is looming in Germany. The German Society for Epidemiology has warned that the number of cases could soon overshoot the capacity of the healthcare system. This may be true even if Germany follows the 'flatten-the-curve'- approach to reduce infection rates. A suppression of the virus remains the best solution for the crisis. Supply will suffer as long the virus persists. Until then, demand side measures will not cure the epidemic. Coordinated measures for business that ensure compliance and European debt instruments may be part of a strategy to solve the crisis.

Design of a user-friendly and rapid DNA microarray assay for the authentication of ten important food fish species.

Seafood is particularly susceptible to the substitution of species. In order to guarantee authentic seafood products, seafood processors and traders must perform self-checks on the authenticity of imported and purchased goods. However, the conventional Sanger sequencing of PCR products for the authentication of seafood species is time-consuming and requires advanced infrastructure. DNA microarrays (DNA chips) with species-specific oligonucleotide probes represent a rapid alternative to sequencing-based species authentication. So far, though, only DNA microarrays for the authentication of land vertebrate species have achieved market success. In this work, a user-friendly DNA microarray assay was developed for the authentication of ten important food fish species that can be performed in four to five hours from start to end. The assay was tested with authenticated specimens from 67 different fish species, and by comparing the probe signal patterns all target species and even closely related non-target species could be distinguished.

Correction to: Chemometric tools for the authentication of cod liveroil based on nuclear magnetic resonance and infrared spectroscopy data.

The article Chemometric tools for the authentication of cod liver oil based on nuclear magnetic resonance and infrared spectroscopy data, written by Editha Giese, Sascha Rohn and Jan Fritsche.

Chemometric tools for the authentication of cod liver oil based on nuclear magnetic resonance and infrared spectroscopy data.

Cod liver oil is a popular dietary supplement marketed as a rich source of omega-3 fatty acids as well as vitamins A and D. Due to its high market price, cod liver oil is vulnerable to adulteration with lower priced vegetable oils. In this study, 1H and 13C nuclear magnetic resonance spectroscopy, Fourier transform infrared spectroscopy, and gas chromatography (coupled to a flame ionization detector) were used in combination with multivariate statistics to determine cod liver oil adulteration with common vegetable oils (sunflower and canola oils). Artificial neural networks (ANN) were able to differentiate adulteration levels based on infrared spectra with a detection limit of 0.22% and a root mean square error of prediction (RMSEP) of 0.86%. ANN models using 1H NMR and 13C NMR data yielded detection limits of 3.0% and 1.8% and RMSEPs of 2.7% and 1.1%, respectively. In comparison, the ANN model based on fatty acid profiles determined by gas chromatography achieved a detection limit of 0.81% and an RMSEP of 1.1%. The approach of using spectroscopic techniques in combination with multivariate statistics can be regarded as a promising tool for the authentication of cod liver oil and may pave the way for a holistic quality assessment of fish oils. Graphical abstract.

Recent Developments and Digital Perspectives in Food Safety and Authenticity.

Food safety is of fundamental importance for the food processing industry, food retailers and distributors, and competent authorities because of its potentially direct impact on the health of consumers. Next to the prevention of microbiological, chemical, and physical hazards, increasing efforts are currently made to combat risks associated with food fraud or food authenticity. Food safety management systems nowadays comprise food safety, food defense, and food fraud prevention measures, trying to cope with the increasing complexity and globalization of the food supply chains. Future digital opportunities include the prediction of food safety and food authenticity issues by handling structured and unstructured data retrieved from various sources and origins to ensure the health of consumers and to minimize economical losses.

Determining quality parameters of fish oils by means of 1H nuclear magnetic resonance, mid-infrared, and near-infrared spectroscopy in combination with multivariate statistics.

Fish oil is becoming increasingly popular as a dietary supplement as well as for its use in animal feed, which is mainly due to its high contents of the health promoting omega-3 fatty acids. However, these polyunsaturated fatty acids are highly susceptible to oxidation, which results in a decrease of the fish oil quality. This study investigated the potential of 1H NMR, FT-MIR, and FT-NIR spectroscopy in the quality assessment of fish oils. A total of 84 different fish oils, of which 22 were subjected to accelerated storage with varying temperature and light exposure, were used to develop models for predicting the peroxide value (PV), the anisidine value (AnV), and the acid value (AV). Predictions were based on comprehensive spectroscopic data in combination with Artificial Neural Networks (ANN) as well as Partial Least Squares Regression (PLSR). The best ANN model for PV was obtained from NMR data, with a predictive coefficient of determination (Q2) of 0.961 and a Root Mean Square Error of Prediction (RMSEP) of 1.5meqO2kg-1. The combined MIR/NIR data provided the most reliable ANN model for AnV (Q2=0.993; RMSEP=0.74). For AV, the ANN model based on the MIR data yielded a Q2 of 0.988 and an RMSEP of 0.43mgNaOHg-1. In most cases, the accuracy of the ANN models was superior to the respective PLSR models. Variable selection and data dimensionality reduction turned out to improve the performance of the ANN models in some cases. The application of 1H NMR, FT-MIR, and FT-NIR spectroscopy in combination with ANN can be considered very promising for a rapid, reliable, and sustainable assessment of fish oil quality.

Mitigation strategies for ester bound 2-/3-MCPD and esterified glycidol in pre-fried breaded and frozen fish products.

Pre-frying of chloride-containing raw materials (e.g., breaded frozen fish products) can lead to the formation of fatty acid esters of 2-monochloropropane-1,3-diol, 3-monochloropropane-1,2-diol (MCPD-E), and glycidol (G-E). The aim of the present study was to identify relevant parameters for the formation of these process contaminants during the pre-frying. Secondly, several mitigation approaches have been investigated. The major proportion of the MCPD-E and G-E in the fish products resulted from the pre-frying oil absorbed, while the temperature and the heating period of the pre-frying oil showed the strongest impact. A significant reduction of the MCPD-E content in the pre-frying oil was achieved by filtering-off solid breading particles. Additionally, the G-E content decreased resulting from the use of adsorbent materials. Moreover, the analyses of total polar material and the color intensity of the pre-frying oil are suggested as screening methods for estimating the MCPD-E and G-E contents in the fish products.