The use of lecithin as an emulsifier in food supplements has increased in recent years. However, successful formation of liposomes or micelles requires an appropriate mixture of phospholipids in lecithin. To evaluate the emulsification properties of lecithin for food supplements, a reliable analytical procedure for characterizing phospholipids is necessary. A liquid chromatography–mass spectrometry method was developed to identify phospholipids in lecithin without standard reference materials. For efficient separation of phospholipids before mass spectrometric analysis, a reverse-phase high-performance liquid chromatography method was optimized using a Waters XBridge Protein BEH C4 column. The optimized chromatographic method demonstrated good linearity and precision. Molecular ions were detected in full scan mode to determine accurate mass-to-charge ratios for individual peaks in the chromatogram. A custom Python program was then used to generate a list of possible phospholipid species for each peak based on the measured mass-to-charge ratios. Tandem mass spectrometry was performed to confirm the identity of specific phospholipids by comparing experimental fragmentation patterns with theoretical predictions. Identification of the phospholipids was also confirmed with four commercially available standard reference compounds, demonstrating the reliability of the proposed approach. The developed method offers a practical and cost-effective strategy for identifying phospholipids in complex matrices, especially when standard reference compounds are unavailable. Additionally, it enables targeted selection of standard compounds for future quantitative analyses, making it a valuable tool for comprehensive lipid profiling.

Monitoring peptide and protein self-association is essential for understanding biological function, formulation stability, and aggregation mechanisms. While size-exclusion chromatography (SEC) is routinely used to quantify protein-size variants under native conditions, its hyphenation to high-resolution mass spectrometry (HRMS) for simultaneous structural characterization remains limited. Here, we report the development and validation of a robust SEC-UV/HRMS method optimized for native-like analysis of bovine serum albumin (BSA) monomers and higher-order oligomers using standard-flow electrospray ionization. Systematic evaluation of source parameters, mobile-phase composition, and chromatographic conditions enabled retention of native BSA structure, minimized in-source unfolding, and enhanced MS sensitivity, allowing detection of oligomers up to the heptamer. A short, narrow-bore 200 Å UHPLC SEC separation column was used. Low-flow separations (~0.05 mL/min) enabled efficient ionization and 10 min run times. An accelerated 60 °C stress-testing protocol demonstrated that SEC-MS can semi-quantitatively monitor oligomerization dynamics, complementing UV-based quantification and revealing transient species not resolved by UV alone. The method showed acceptable linearity, precision, and sample stability, and comparison with SEC-RALS/LALS confirmed molecular-weight trends across aggregation states. Overall, the developed SEC-UV/HRMS workflow provides a rapid, sensitive, and widely accessible approach for UV-based quantification of monomer- and HRMS-based characterizing protein aggregation in research and quality control in pharmaceutical laboratories.

The glycosylation process plays a crucial role in the structural integrity and biological activity of glycoproteins, where glycans are attached to a protein backbone. There are many kinds of glycans, the most common being N-glycans, which can be arranged into three classes, that is, complex, hybrid, and high mannoses, forming a structurally very diverse set of polar compounds that are difficult to detect and separate. Most commonly, N-glycans are labeled before separation by charged or fluorescence tags for better MS or fluorescence detection, respectively. This study examines the influence of ionic strength and organic modifier selection on the separation of fluorescently labeled dextran ladders in Hydrophilic Interaction Liquid Chromatography (HILIC). Using a Glycan BEH Amide column and varying the ammonium formate buffer concentration along with acetonitrile and methanol ratios, we investigated analyte retention, separation efficiency, and post-column conductivity changes. Our findings reveal that changes in the ionic strength of the mobile phase do not contribute to changes in selectivity, neither when acetonitrile nor methanol were used as organic modifiers to the mobile phase. However, the addition of methanol significantly changes the separation mechanism where two different prevailing separations mechanisms can be identified. It was assumed that the addition of methanol influences the folding pattern of dextrans around the permanent positive charge on the added tag, which influences the changes of separation selectivity. This work presents a systematic approach to altering mobile phase composition (buffer concentration, organic modifier type) to control retention and selectivity in complex glycan analysis. The discovery that methanol significantly alters separation behavior provides a potential new method for refining HILIC separations of polar compounds.

Encapsulated formulations have emerged as a promising tool for increasing nutrient absorption in the food supplement and cosmetic industries. Although the theoretical amplification factors for improving the bioavailability of encapsulated formulations are very high for poorly soluble active compounds, it has long been known that encapsulation can also enhance the absorption of water-soluble ingredients. These findings have led to the development of new technologies for encapsulating nutrients for use in the food industry. However, accurate quantification of nutrients in encapsulated formulations in the food supplement industry remains a challenge. This study presents the development and validation of novel analytical procedures for determining artemisinin in various food supplement formulations. Three formulations were prepared using different emulsifying procedures for artemisinin encapsulation. High-performance liquid chromatography with UV/Vis detection (HPLC-UV/Vis) was used for analysis. Separation was performed using a Waters ACQUITY Premier BEH C18 column. Specialized sample preparation procedures were designed to efficiently disrupt encapsulation and extract artemisinin for precise quantification. Three different sample preparation procedures were required to accurately determine the artemisinin content in the tested formulations. All methods were validated. The precision, linearity expressed as R2, LOD, and LOQ of the chromatographic method were 0.39%, 0.9995, 18 µg/mL, and 26 µg/mL, respectively. Recoveries of the sample preparation methods were above 94%. The developed procedures enable accurate determination of artemisinin in encapsulated formulations, ensuring product quality and safety. These findings suggest that, for quality control of encapsulated food products, specialized analytical procedures for individual formulations may need to be developed and validated.

This study investigates the effect of taxifolin and arabinogalactan premixed with dihydroquercetin (Lavitol V) at different concentrations on the quality changes of trout soup at 4 +/- 1 degrees C for 31 days. Shelf life studies included physiochemical, sensory, and microbiological quality measures. The shelf lives of the fish soups were extended for 2-4 and 7-10 days with taxifolin and Lavitol V, respectively, with significant variations (p < 0.05), the longest shelf life corresponding to Lavitol V for 31 days. The highest TBA and TVB-N were obtained for the control as 3.72 mg MA/kg and 11.21 mg/100 g, respectively. Therefore, Lavitol V has a good potential for future applications in seafood preservation.

Hydrophilic interaction liquid chromatography (HILIC) is widely used for the analysis of glycans and oligosaccharides, yet the molecular basis of retention remains incompletely understood. In this study, we investigated dextran ladders labelled with 2-aminobenzamide (2-AB) and Rapifluor-MS™ (Waters, Milford, MA, USA) across a wide range of degrees of polymerization (DP 2–15), temperature conditions (10 °C to 70 °C), and gradient programs using a Acquity™ Premier Glycan BEH Amide column (Bridged Ethylene Hybrid, Waters, Milford, MA, USA). Van’t Hoff analysis revealed distinct enthalpic and entropic contributions to retention, allowing identification of a mechanistic transition from enthalpy-dominated docking interactions at low DP to entropy-driven dynamic adsorption at higher DP. This transition occurred reproducibly between DP 4–6, depending on the fluorescent label, while gradient steepness primarily influenced the location of the minimum enthalpy. Molecular dynamics simulations provided additional evidence, showing increased conformational flexibility and end-to-end distance variability for longer oligomers. This finding is consistent with entropy-dominated adsorption accompanied by displacement of structured interfacial water. Together, these results establish a molecular-level framework linking retention thermodynamics, conformational behavior, and solvation effects, thereby advancing our mechanistic understanding of glycan separation in HILIC.

Kynurenic acid (KYNA) has been attributed many beneficial properties, such as antioxidant, antiproliferative, anti-inflammatory, and anti-obesogenic, as it is believed to affect metabolism and weight gain. A rapid and simple HPLC-MS/MS method for the determination of kynurenic acid (KYNA) in honey has been developed. HPLC-MS/MS system allowed us to perform the analyzes without any special extraction or treatment of the samples. The study was carried out on different honeys: Chestnut (C), Linden (L), Acacia (A), Spruce (S), Silver Fir (SF), Forest (Fo) and Flower (F). The highest mean concentration, 682 μg/g, was determined for chestnut honey, making it one of the foods with the highest KYNA content.

This study demonstrates the effects of the freezing and frozen storage of anchovies prior to brining and dry-salting on quality changes and food safety during refrigerated storage (4 ± 1 °C). Fresh anchovies were divided into two groups, one of which was used as a control representing fresh raw material; the other was the experimental group and consisted of frozen and thawed anchovies stored at −18 °C for a year. Five different salt concentrations were used for brining (10, 15, 20, 25 and 30%) in addition to the dry-salting method. Microbiological, chemical, physical and sensory analyses were carried out during storage. Salt concentration and salting method had significant effects on the shelf-life of salted anchovy products, with the highest shelf-life corresponding to dry-salted anchovies (p < 0.05). The effect of using frozen and thawed raw materials for salting on the shelf-life depended on the processing method since the experimental dry-salted group had a shelf-life one month longer than that of the control group, while the opposite situation occurred for the brined samples, with one exception. Strong correlations were usually found between sensory values and chemical quality parameters (R2: 0.83–0.99 for the control group and 0.63–0.99 for the experimental group). The results demonstrated that the experimental group, with some exceptions, had better values for most quality and food safety parameters in comparison to the control group, indicating the advantage of using frozen and thawed raw materials before salting to prevent spoilage and enhance food safety. Considering that the experimental group was produced from one-year-stored raw material compared to the control group, the advantage of the freezing and frozen storage of anchovies can be accepted as much higher versus when freezing is not implemented. The positive effect is due to the fact that frozen anchovies absorb salt faster after thawing, especially in groups with high salt concentrations. Therefore, it can be concluded that frozen salted anchovies can be utilized for longer as they have a longer shelf-life, particularly when using either the 30% brining or dry-salting method.

Studies of protein adsorption on reversed-phase and ion exchange stationary phases demonstrated an increase in retention with increasing pressure, which is interpreted as a standard partial molar volume decrease during the transition of the protein from a mobile to a stationary phase. Investigation of the pressure effect on the retention of lysozyme and IgG on a cation exchange column surprisingly revealed a negative retention trend with the increase of pressure. Further investigation of this phenomenon was performed with β-lactoglobulin, which enabled adsorption to be studied on both cation and anion exchange columns using the same mobile phase with a pH of 5.2. The same surface charge and standard partial molar volume in the mobile phase allowed us to examine only the effect of adsorption. Interestingly, a negative retention trend with a pressure increase occurred on an anion exchange column while a positive trend was present on a cation exchange column. This indicates that the interaction type governs the change in the standard partial molar volume during adsorption, which is independent of the applied pressure. Increasing the protein charge by decreasing the pH of the mobile phase to 4 reversed the retention trend (into a negative) with a pressure increase on the cation exchange column. A further decrease of the pH value resulted in an even more pronounced negative trend. This counterintuitive behavior indicates an increase in the standard partial molar volume during adsorption with the protein charge, possibly due to intermolecular repulsion of adsorbed protein molecules. While a detailed mechanism remains to be elucidated, presented results demonstrate the complexity of ion exchange interactions that can be investigated simply by changing the column pressure.

This study represents new information on the nutritional value of soup prepared from cultured Rainbow trout (Oncorhyncus mykiss). The proximate contents of soup were represented by 87.56% moisture, 7.22% protein, 1.59% crude fat, 0.07% carbohydrate, 1.17% dietary fibre and 2.44% ash. The energy value was calculated as 48.0 kcal/100g. Amino acid values were in the range of 183.5-1128.5mg/100g while mineral contents were varied as 0.6µg/g-19.5mg/g. The value of polyunsaturated fatty acids (PUFA) was higher than saturated fatty acids (SFA). Total PUFA and eicosapentaenoic acid + docosahexaenoic acid (EPA+DHA) were observed as 37.2 and 7.2%, respectively. These values corresponded to the values of 673.3 and 123.2 mg/100g of the edible portion of the soup within the same respect. The results of this study demonstrated that a portion of trout soup would almost cover daily recommended n-3 PUFA intake, however, higher amounts are required for the necessary levels of EPA+DHA. The vitamin B12 content was well above the recommended levels. The values of α and β tocopherols were observed as 272.0 and 1131.0 µg/100g (wwb), respectively. This study indicates that trout soup has a good nutritional value for human consumption for a healthy diet.

In this study, comparative analyses were carried out with ion chromatography mass-spectrometry (IC-MS/MS) which has no derivatization step, high-performance liquid chromatography (HPLC) technique, as well as two quantitative and two semi-quantitative immunoassays. The results demonstrated that HPLC and quantitative immunoassay methods were well-correlated with IC-MS/MS in determining histamine in various types of fish products. The best correlation was observed with the HistaSure ELISA Fast Track kit (R2 = 0.9903). More than half of the values (68%) obtained by two methods were also statistically similar. The results of semi-quantitative test kits also supported histamine values estimated by quantitative methods, with some exceptions. The best results were found for HistaSure Lateral Flow in supporting the quantitative techniques. Therefore, these methods are found suitable for monitoring histamine in fish products in terms of food safety. Good correlations were also observed HPLC and IC-MS/MS in determining cadaverine, putrescine, and tyramine with the highest value observed for tyramine as R2 = 0.9785. However, no correlation was observed for other biogenic amines, and the majority of the results were significantly different from each other for these amines (p < 0.05). The differences may be caused by the drawbacks reported previously for HPLC. However, further studies are required to confirm the possible effects. This study provides a comparative evaluation of several methods in terms of their suitability in determining biogenic amines in fish products for both monitoring and regulatory purposes.

A new method was proposed for the determination of underivatized biogenic amines based on ion-exchange chromatography coupled with mass spectrometry detection. The method was applied to the analysis of 10 biogenic amines in fresh and processed fish products. The amines were extracted from muscle tissue with water without any additional derivative step or sample clean-up. Separation of biogenic amines was done by the IonPac (4 × 50 mm) column, applying a gradient eluent by mixing formic acid (2 mol L−1) and Milli-Q water (formic acid concentration from 400 mM to 2 M). The results demonstrated a linear response in the range of 0.01 to 10 mg L−1. The detection limits for the fish products ranged from 20 ng/g up to around 400 ng/g for histamine and putrescine, respectively. Spermidine and spermine showed significantly higher detection limits. This current method can be used for the determination of biogenic amines in both fresh and processed fish products for regulatory purposes and monitoring food-safety issues relating to these amines, particularly histamine. It is also a useful method for evaluation of other commercial analytical test kits and commonly used methods that are possibly affected by the food matrix due to processing or other drawbacks arising from the derivatization process.

Human insulin and six most used therapeutic analogues are very similar in terms of retention on a reversed-phase column. Thus, the LC methods prescribed in the European Pharmacopoeia monographs for insulin and insulin analogues include many similar separation methods, which tend to be time consuming when separating individual products of insulins or are inadequate when handling a mixture. In this study, we present a simple, robust, versatile and accessible HPLC-UV separation method for identification and quantification of human insulin and its analogues in one run. The simultaneous separation and detection is possible by fine-tuning the mobile phase properties that affect the separation mechanism on a mixed mode column combining anion exchange and reversed-phase characteristics. Also developed was a simple and effective SPE sample cleaning procedure with insulin recoveries ranging from 80 to 100% for all analogues. On the other hand, the concentration of major excipients such as phenol and m-cresol fall below 1%. The two developed and validated separation methods differ in their compatibility with the use of a quaternary or binary pump, thus enabling sample characterisation independent of the HPLC solvent delivery system. The methods are compatible with the use of a mass spectrometric detector for an indisputable identification.

Introduction. The aim of this study is to estimate the nutritional value of fish soup from cultured brook trout in terms of a healthy diet for human consumption. Materials and methods. Fish soup was prepared from 23.65% previously cooked trout mince and 18.76% vegetables, and cooked for 35 mins. The final product was analyzed for the proximate composition, fatty acids and, mineral contents as well as carotenoids and vitamins B1, B2 and B6. Inductively coupled plasma Mass Spectrometer (ICPMS) was used for mineral content after decomposition of lyophilized samples. Fatty acids methyl esters were separated by gas chromatography by flame ionization detector (FID). High performance liquid chromatography (HPLC) was used for the estimation the contents of vitamins and carotenoids. Results and discussion. The proximate contents of soup were represented by 87.79% moisture, 8.18% protein, 2.89% crude fat, 1.17% dietary fibre, 0.62% ash and 0.03% carbohydrate. The energy value was calculated as 58.82 kcal/100g. The value of total polyunsaturated fatty acids (Sigma PUFA) was higher than the values of total monounsaturated (Sigma MUFA) and saturated fatty acids (Sigma SFA) and accounted as 43.89, 34.93 and 19.83%, respectively. The main PUFA corresponded to linoleic acid as 27.14% followed by docosahexaenoic acid (DHA) as 7.92%. Total eicosapentaenoic acid+docosahexaenoic acid (Sigma EPA+DHA) was observed as 9.21% which was accounted as 239.04 mg/100g soup. The results of this study demonstrated that a portion of trout soup (about 200g) would well cover daily recommended n-3 PUFA intake while slightly higher amount is required for daily EPA+DHA intake. Mineral contents were varied in the range of 1.77-31.52 mg/g (dwb), while the results obtained for vitamin B1, B2 and B6 and for carotenoids were comparable with the data given for different types of soups in literature. Conclusion. This study indicates that a nutritious fish soup can be produced from brook trout for human consumption suitable for a healthy diet.

In this study a new method for evaluating the pressure effect on separations of oligonucleotides and proteins on an anion exchange column was developed. The pressure rise of up to 500 bar was attained by coupling restriction capillaries to the column outlet to minimize differences in pressure over the column. Using pH transient measurements it was demonstrated that no shift in ion exchange equilibria occurs due to a pressure increase. Results from isocratic and gradient separations of oligonucleotides (model compounds) were evaluated by stoichiometric displacement and linear gradient elution model, respectively. Both elution modes demonstrated that for smaller oligonucleotides the number of binding sites remained unchanged with pressure rise while an increase for large oligonucleotides was observed, indicating their alignment over the stationary phase. From the obtained model parameters and their pressure dependencies, a thermodynamic description was made and compared between the elution modes. A complementary pattern of a linear increase of partial molar volume change with a pressure rise was established. Furthermore, estimation of the pressure effect was performed for bovine serum albumin and thyroglobulin that required gradient separations. Again, a raise in binding site number was found with pressure increase. The partial molar volume changes of BSA and Tg at the maximal investigated pressure and minimal salt concentration were -31.6 and -34.4 cm3/mol, respectively, indicating a higher rigidity of Tg. The proposed approach provides an insight into the molecule deformation over a surface at high pressures under nondenaturing conditions. The information enables a more comprehensive UHPLC method development.

A new method for determination of underivatized biogenic amines in cheese based on ion exchange chromatography coupled with tandem mass spectrometric detection was proposed. The method was applied to the analysis of 10 biogenic amines (trimethylamine, putrescine, cadaverine, histamine, 2-phenylethylamine, spermine, spermidine, tryptamine, agmatine, and tyramine) in different types of cheese. The amines were extracted only with water without any additional derivatization step or sample cleanup. This is a great advantage in terms of simplicity of sample pretreatment procedure compared with other currently existing methods in the literature. Biogenic amines were separated using cation exchange column, under gradient elution conditions by mixing formic acid (1.00 M) and deionized water. Detection was achieved using tandem MS/MS, with the instrument set into multiple reaction monitoring mode to ensure high specificity. The detection and quantification limits were in the ranges of 12-46 μg/L and 40-153 μg/L, respectively. The exceptions were spermidine and spermine, with detection limits of 0.8 and 5.4 mg/L, respectively. The linearity for most of the biogenic amines was from 10 μg/L up to 10 mg/L. The best recoveries were observed for trimethylamine, tyramine, and cadaverine, and were 89, 94, and 102%, respectively. The results showed that this method can be used for routine determination of biogenic amines in different types of cheeses as well other food matrices. It must be stressed that the proposed method is capable of determining 10 biogenic amines, including tyramine, which is reported to cause food intoxication commonly associated with cheeses.

The hygroscopicity of organic aerosol in the atmosphere can be represented by a semi-empirical single parameter, κ. In this work we test possibilities for developing quantitative structure-property relationship (QSPR) models for κ based on chemical similarity. Models were developed in two ways: by manually assessing the suitability of several plausible physico-chemical descriptors; and by systematically evaluating hundreds of constitutional (e.g. number of particular atoms, bond types, molecular weight,...), topological, electrostatic, geometrical and quantum-chemical descriptors with the QSPR modelling software CODESSA (COmprehensive DEscriptors for Structural and Statistical Analysis). A set of 74 compounds with measured κ values was taken from the literature and prediction capabilities of the developed models were evaluated by leave-one-out cross-validation procedure. A 5-parameter linear regression model obtained with CODESSA was found to be the most suitable. Among the five descriptors, the two providing the highest contributions to the total variance were found to be (i) the final heat of formation divided by the number of atoms (69 %) and (ii) the ratio of molecular weight and molecular volume (16 %), although other topological and electrostatic descriptors were also of non-negligible importance for prediction of κ. The squared correlation coefficient and the root mean square error of a leave-one-out cross-validation procedure were 0.80 and 0.037, respectively. The results show that quantitative structure-property relationship approaches are useful for modeling κ.

A new variable distance weighted zero order connectivity index was used for development of structure activity relationship for modeling reactivity of OH radical with alkanes and non-conjugated alkenes in the atmosphere. The proposed model is based on the assumptions that the total reaction rate can be obtained by summing all partial reaction rates and that all reaction sites are interrelated by influencing each other. The results suggest that these assumptions are justified. The model was compared with the EPA implemented model in the studied application domain and showed superior prediction capabilities. Further, optimized values of the weights that were used in our model permit some insight into mechanisms that govern the reaction OH + alkane/alkene. The most important conclusion is that the branching degree of the forming radical seems to play a major role in site specific reaction rates. Relative qualitative structural interpretation is possible, e.g. allylic site is suggested to be much more reactive than even tertiary spa carbon.Novel modeling software MACI, which was developed in our lab and is now available for research purposes, was used for calculations. Various variable topological indices that are again starting to be recognized because of their great potentials in simplicity, fast calculations, very good correlations and structural information, were implemented in the program. (C) 2016 The Authors. Published by Elsevier Ltd.

In order to perform the screening of new potential pollution points and to estimate their impact on the environment, a quantitative structure-activity research (QSAR) modeling procedure was used to estimate possible toxicological effects of substances that were introduced into the environment. We have focused our study initially to known persistent toxic compounds with defined toxicological mechanisms. Such groups of studied chemicals are polychlorinated aromatic compounds. Three multiple linear regression (MLR) models were developed for the calculation of aryl hydrocarbon receptor (AhR) binding affinity of polychlorinated biphenyls (PCBs), dibenzofurans (PCDFs) and dibenzodioxins (PCDDs). In all cases models were able to explain more than 70% of the total variance. Additionally, the correlating capability of the newly developed variable distance-based topological index was tested. It showed reasonable modeling abilities and superior interpretation abilities compared to classical MLR QSAR models.

Tetramantanes, and all diamondoid hydrocarbons, possess carbon frameworks that are superimposable upon the cubic diamond lattice. This characteristic is invaluable in assigning their H-1 and C-13 NMR spectra because it translates into repeating structural features, such as diamond-cage isobutylmoieties with distinctively complex methine to methylene signatures in COSY and HMBC data, connected to variable, but systematic linkages of methine and quaternary carbons. In all tetramantane C22H28 isomers, diamond-lattice structures result in long-range (4)J(HH), W-coupling in COSY data, except where negated by symmetry; there are two highly symmetrical and one chiral tetramantane (showing seven (4)J(HH)). Isobutyl-cage methines of lower diamondoids and tetramantanes are the most shielded resonances in their C-13 spectra (<29.5ppm). The isobutyl methylenes are bonded to additional methines and at least one quaternary carbon in the tetramantanes. W-couplings between these methines and methylenes clarify spin-network interconnections and detailed surface hydrogen stereochemistry. Vicinal couplings of the isobutyl methylenes reveal positions of the quaternary carbons: HMBC data then tie the more remote spin systems together. Diamondoid C-13 NMR chemical shifts are largely determined by alpha and beta effects, however.-shielding effects are important in [123]tetramantane. H-1 NMR chemical shifts generally correlate with numbers of 1,3-diaxial H-H interactions. Tight van der Waals contacts within [123]tetramantane's molecular groove, however, form improper hydrogen bonds, deshielding hydrogen nuclei inside the groove, while shielding those outside, indicated by Delta delta of 1.47 ppm for geminal hydrogens bonded to C-3,21. These findings should be valuable in future NMR studies of diamondoids/nanodiamonds of increasing size. Copyright (C) 2015 John Wiley & Sons, Ltd.