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Amino acids (AAs) have broad nutritional, therapeutic, and medical significance and thus are one of the most common active ingredients of nutritional supplements. Analytical strategies for determining AAs are high-priced and often limited to methods that require modification of AA polarity or incorporation of an aromatic moiety. The aim of this work was to develop a new method for the determination of L-arginine, L-ornithine, and L-lysine on low-cost microchip electrophoresis instrumentation conjugated with capacitively coupled contactless conductivity detection. A solution consisting of 0.3 M acetic acid and 1 × 10−5 M iminodiacetic acid has been identified as the optimal background electrolyte, ensuring the shortest possible analysis time. The short migration times of amino acids (t ≤ 64 s) and method simplicity resulted in high analysis throughput with high precision and linearity (R2≥ 0.9971). The limit of detection values ranged from 0.15 to 0.19 × 10−6 M. The accuracy of the proposed method was confirmed by recovery measurements. The results were compared with CE-UV-VIS and HPLC-DAD methods and showed good agreement. This work represents the first successful demonstration of the ME-C4D analysis of L-arginine, L-ornithine, and L-lysine in real samples.

Dimeric forms of flavonoids, known as biflavonoids, are much less studied compared to monomeric forms. It is estimated that nearly 600 different natural biflavonoids have been described to date, containing various subtypes that can be subdivided according to the position of their combinations and the nature of the subunits. The group in which two monomers are linked by a 3′-8″-C atom includes the first isolated biflavonoid ginkgetin, derivatives of amentoflavone, and several other compounds. 3′-8″-biflavones recently attracted much attention as potential molecules with biological activity such as antiviral and antimicrobial activity and as effective molecules for the treatment of neurodegenerative and metabolic diseases and in cancer therapies. With the growing interest in them as pharmacologically active molecules, there is also increasing interest in finding new natural sources of 3′-8″-biflavones and optimizing methods for their extraction and identification. Herein, we have summarized the available data on the structural diversity, natural occurrence, role in plants, extraction, and identification of 3′-8″-biflavones.

The aim of this work was to investigate the influence of cold storage at 2 °C for 180 days on water content, total acids, reducing and total sugar, antioxidant activity, polyphenol oxidase activity, and polyphenol profile of three Croatian traditional apple cultivars (‘Kanadska Reneta’, ‘Ilzer Rosenapfel’, ‘Wagener’). A significant variation in polyphenol profile of apple cultivars under investigation was noted. The most represented polyphenols in all apple cultivars were procyanidin B2 followed by chlorogenic acid, epigallocatechin, epicatechin, and phloridzin. After storage, the content of procyanidin B2 and chlorogenic acid was higher, while content of epigallocatechin after storage was lower in all investigated apple cultivars. Water content before storage ranged between 79.17 and 80.76 %, and after storage between 75.62 and 78.56 %. Total acids, polyphenol oxidase activity, and flavonoids had decreased during storage, while reducing and total sugar content was higher after storage. Moreover, after storage, cultivar ‘Kanadska Reneta’ had the highest antioxidant activity measured by both ABTS and DPPH methods, 1.65 and 0.70 mmol trolox kg–1, respectively. A significant variation in polyphenol profile of apple cultivars under investigation was noted. The most represented polyphenols in all apple cultivars were procyanidin B2, followed by chlorogenic acid, epigallocatechin, epicatechin, and phloridzin. After storage, the content of procyanidin B2 and chlorogenic acid was higher, while content of epigallocatechin after storage was lower in all investigated apple cultivars. Overall, it can be concluded that investigated apple cultivars, after storage, preserved most of the antioxidants and functional properties.

The presence of mycotoxins and other toxic metabolites in hops (Humulus lupulus L.) was assessed for the first time. In total, 62 hop samples were sampled in craft breweries, and analyzed by a multi-toxin LS-MS/MS method. The study collected samples from craft breweries in all of the Croatian counties and statistically compared the results. Based on previous reports on Alternaria spp. and Fusarium spp. contamination of hops, the study confirmed the contamination of hops with these toxins. Alternaria toxins, particularly tenuazonic acid, were found in all tested samples, while Fusarium toxins, including deoxynivalenol, were present in 98% of samples. However, no Aspergillus or Penicillium metabolites were detected, indicating proper storage conditions. In addition to the Alternaria and Fusarium toxins, abscisic acid, a drought stress indicator in hops, was also detected, as well as several unspecific metabolites. The findings suggest the need for monitoring, risk assessment, and potential regulation of Alternaria and Fusarium toxins in hops to ensure the safety of hop usage in the brewing and pharmaceutical industries. Also, four local wild varieties were tested, with similar results to the commercial varieties for toxin contamination, but the statistically significant regional differences in toxin occurrence highlight the importance and need for targeted monitoring.

A lot of research has been conducted on economic and consumer aspects of ecological food products. However, we are witnessing the appearance of food products produced according to the principles of biodynamic growth which can be seen as a higher standard in ecological production process. Similar to ecological cultivation the biodynamic one also has proscribed methods and processes of production, processing, distribution and labelling as well as control and certification processes. However, such products are still not being recognized be the consumers in Croatia. This paper provides, based on authentic empirical research, some basic background information on the importance of labelling packages of food products as well as their quality and traceability. These also present the first results of research on the importance of labelling biodynamic products packages in Croatia.

Biflavonoids are dimeric forms of flavonoids that have recently gained importance as an effective new scaffold for drug discovery. In particular, 3′-8″-biflavones exhibit antiviral and antimicrobial activity and are promising molecules for the treatment of neurodegenerative and metabolic diseases as well as cancer therapies. In the present study, we directly compared 3′-8″-biflavones (amentoflavone, bilobetin, ginkgetin, isoginkgetin, and sciadopitysin) and their monomeric subunits (apigenin, genkwanin, and acacetin) and evaluated their radical scavenging activity (with DPPH), antifungal activity against mycotoxigenic fungi (Alternaria alternata, Aspergillus flavus, Aspergillus ochraceus, Fusarium graminearum, and Fusarium verticillioides), and inhibitory activity on enzymes (acetylcholinesterase, tyrosinase, α-amylase, and α-glucosidase). All the tested compounds showed weak radical scavenging activity, while antifungal activity strongly depended on the tested concentration and fungal species. Biflavonoids, especially ginkgetin and isoginkgetin, proved to be potent acetylcholinesterase inhibitors, whereas monomeric flavonoids showed higher tyrosinase inhibitory activity than the tested 3′-8″-biflavones. Amentoflavone proved to be a potent α-amylase and α-glucosidase inhibitor, and in general, 3′-8″-biflavones showed a stronger inhibitory potential on these enzymes than their monomeric subunits. Thus, we can conclude that 3′-8″-dimerization enhanced acetylcholinesterase, α-amylase, and α-glucosidase activities, but the activity also depends on the number of hydroxyl and methoxy groups in the structure of the compound.

A multi-specimen, multi-mycotoxin approach involving ultra-sensitive LC-MS/MS analysis of breast milk, complementary food and urine was applied to examine mycotoxin co-exposure in 65 infants, aged 1–18 months, in Ogun state, Nigeria. Aflatoxin M1 was detected in breast milk (4/22 (18%)), while six other classes of mycotoxins were quantified; including dihydrocitrinone (6/22 (27%); range: 14.0–59.7 ng/L) and sterigmatocystin (1/22 (5%); 1.2 ng/L) detected for the first time. Seven distinct classes of mycotoxins including aflatoxins (9/42 (21%); range: 1.0–16.2 µg/kg) and fumonisins (12/42 (29%); range: 7.9–194 µg/kg) contaminated complementary food. Mycotoxins covering seven distinct classes with diverse structures and modes of action were detected in 64/65 (99%) of the urine samples, demonstrating ubiquitous exposure. Two aflatoxin metabolites (AFM1 and AFQ1) and FB1 were detected in 6/65 (9%), 44/65 (68%) and 17/65 (26%) of urine samples, respectively. Mixtures of mycotoxin classes were common, including 22/22 (100%), 14/42 (33%) and 56/65 (86%) samples having 2–6, 2–4, or 2–6 mycotoxins present, for breast milk, complementary food and urine, respectively. Aflatoxin and/or fumonisin was detected in 4/22 (18%), 12/42 (29%) and 46/65 (71%) for breast milk, complimentary foods and urine, respectively. Furthermore, the detection frequency, median concentrations and occurrence of mixtures were typically greater in urine of non-exclusively breastfed compared to exclusively breastfed infants. The study provides novel insights into mycotoxin co-exposures in early-life. Albeit a small sample set, it highlights transition to higher levels of infant mycotoxin exposure as complementary foods are introduced, providing impetus to mitigate during this critical early-life period and encourage breastfeeding.

As anionic surfactants are used as cleaning agents, they pose an environmental and health threat. A novel potentiometric sensor for anionic surfactants based on the 1,3-dioctadecyl-1H-imidazol-3-ium tetraphenylborate (DODI–TPB) ionophore is presented. The newly developed approach for DODI–TPB synthesis is faster and simpler than the currently used strategies and follows the green chemistry principles. The DODI–TPB ionophore was characterized by computational and instrumental techniques (NMR, LC–MS, FTIR, elemental analysis) and used to produce a PVC-based DODI–TPB sensor. The sensor showed linear response to dodecylbenzenesulfonate and dodecyl sulfate in concentration ranges of 6.3 × 10−7–3.2 × 10−4 M and 5.9 × 10−7–4.1 × 10−3 M, for DBS and SDS, respectively. The sensor exhibits a Nernstian slope (59.3 mV/decade and 58.3 mV/decade for DBS and SDS, respectively) and low detection limits (7.1 × 10−7 M and 6.8 × 10−7 M for DBS and SDS, respectively). The DODI–TPB sensor was successfully tested on real samples of commercial detergents and the results are in agreement with the referent methods. A computational analysis underlined the importance of long alkyl chains in DODI+ and their C–H∙∙∙π interactions with TPB− for the ionophore formation in solution, thereby providing guidelines for the future design of efficient potentiometric sensors.

Apples and apple-based products are particularly interesting due to being a good source of polyphenols in an everyday diet. Recently there has been increased interest in the preservation of traditional apple varieties due to studies that suggest that traditional apple varieties have a higher content of polyphenols and antioxidant activity compared to commercial varieties. This study shows that traditional apple varieties contain higher concentration of polyphenolic compounds than conventional ones, such as chlorogenic acid (1.29–456 mg/kg dw), catechin (0.70–312 mg/kg dw), epicatechin (1.75–244 mg/kg dw), procyanidin B1 + B2 (4.08–358 mg/kg dw) and quercetin-3-glucoside (0.96–231 mg/kg dw). This research underlies the great potential of traditional apple varieties as a source of natural antioxidants and polyphenolics.

A low-cost and fast potentiometric surfactant sensor for cationic surfactants, based on the new ion-pair 1,3-dioctadecyl-1H-imidazol-3-ium-tetraphenylborate (DODI-TPB), is presented. The new cationic surfactant DODI-Br was synthesized and characterized by NMR, LC-MS, and elemental analysis, and was used for synthesis of the DODI-TPB ionophore. The DODI-TPB surfactant sensor was obtained by implementation of the ionophore in PVC. The sensor showed excellent response characteristics with near-Nernstian slopes to the cationic surfactants DMIC, CPC, CTAB, and Hyamine 1622. The highest voltage responses were obtained for DMIC and CPC (58.7 mV/decade of activity). DMIC had the lowest detection limit (0.9 × 10−6 M) and the broadest useful linear concentration range (1.8 × 10−6 to 1.0 × 10−4 M). An interference study showed remarkable stability. Potentiometric titration curves for the titration of cationic surfactants (DMIC, CPC, CTAB, and Hyamine 1622), with DDS and TPB used as titrants, showed sigmoidal curves with well-defined inflexion points and a broad signal change. The standard addition method was successfully applied with recovery rates from 98.9 to 101.2 at two concentrations. The amount of cationic surfactant found in disinfectants and antiseptics was in good agreement with the referent two-phase titration method and the surfactant sensor on the market. This new surfactant sensor represents a low-cost alternative to existing methods for cationic surfactant detection.

The botanical origin of starch is of importance in industrial applications and food processing because it may influence the properties of the final product. Current microscopic methods are time-consuming. Starch consists of an origin-dependent amylose/amylopectin ratio. Triiodide ions bind characteristically to the amylose and amylopectin depending on the botanical origin of the starch. The absorbance of the starch-triiodide complex was measured for wheat, potato, corn, rye, barley, rice, tapioca and unknown origin starch, and within the different cultivars. Each starch sample had specific parameters: starch-triiodide complex peak wavelength maximum (λmax/nm), maximum absorbance change at λmax (ΔA) and λmax shift towards the unknown origin starch sample values. The visible absorption spectra (500-800 nm) for each starch sample were used as a unique fingerprint, and then elaborated by cluster analysis. The cluster analysis managed to distinguish data of two clusters, a cereal type cluster and a potato/tapioca/rice starch cluster. The cereal subclusters extensively distinguished wheat/barley/rye starches from corn starches. Data for cultivars were mostly in good agreement within the same subclaster. The proposed method that combines cluster analysis and visible absorbance data for starch-triiodide complex was able to distinguish starch of different botanical origins and cultivars within the same species. This method is simpler and more convenient than standard time-consuming methods.

This study investigated the occurrence of 5 unregulated mycotoxins in a total of 250 traditional dry-cured meat products sampled in 2020 and 2021 in five Croatian regions (eastern, northern, central, western, and southern). Aflatoxin B1 (AFB1), ochratoxin A (OTA), sterigmatocystin (STC), citrinin (CIT), and cyclopiazonic acid (CPA) concentrations were related to the geographical region of the product’s origin and to local weather. The results revealed the contamination of 27% of samples, namely, STC in 4% of samples in concentrations of up to 3.93 µg/kg, OTA in 10% of samples in concentrations of up to 4.81 µg/kg, and CPA in 13% of samples in concentrations of up to 335.5 µg/kg. No AFB1 or CIT contamination was seen. Although no statistically significant differences in concentrations of individual mycotoxins across the production regions were found, differences in mycotoxin occurrence were revealed. The eastern and western regions, with moderate climate, delivered the largest number of contaminated samples, while the southern region, often compared with subtropics, delivered the smallest, so that the determined mycotoxins were probably mainly produced by the Penicillium rather than the Aspergillus species. Due to the interaction of various factors that may affect mycotoxin biosynthesis during production, the detected concentrations cannot be related solely to the weather.

Apples and apple-based products are among the most consumed fruits around the world. However, they are susceptible to infection with the fungi Penicilium expansum. In addition to the reduction of apple quality, secondary metabolism of this fungus produces a mycotoxin patulin that has a negative effect on human health. Currently, there is no available research in the literature on the resistance of Croatian traditional apple cultivars to contamination with P. expansum, and consequently, on the patulin content in apples and apple juice produced from those apples. Although the mechanism of apple resistance to fungal diseases has not yet been sufficiently investigated, some studies have shown that polyphenolic compounds have some impact on fungi growth. In order to contribute with new knowledge, this research deals with monitoring the growth of P. expansum on apples, patulin detection by LC/MS-MS, determination of polyphenol profile by validated HPLC method, and determining the effect of polyphenolic compounds on fungi growth and patulin production during apple storage. The results of this study have shown that Croatian traditional apple cultivars harvested from family farm Horvatić contain higher concentration of polyphenolic compounds and higher antioxidant activity. At the same time, they showed more resistance to infection by P. expansum than conventional ones. The higher content of dihydrochalcones and flavanols encouraged the biosynthesis of patulin in examined cultivars. However, the higher content of non-flavonoids such as 2-6 dimethoxybenzoic acid, 4-hydroxycinnamic acid and chlorogenic acid leads to decrease in content of patulin. In conclusion, it seems that content of polyphenols and patulin production are correlated.

The availability of reliable sensitive multi-analyte methods for unambiguous determination of mycotoxins is crucial for ensuring food and feed safety, considering their adverse health effects and (co-)occurrence in various foods. Accordingly, a multi-mycotoxin confirmatory method for simultaneous determination of 11 mycotoxins regulated in cereals within the European Union (EU) using ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS) was developed and in-house validated to fit the EU legislation requirements for analytical methods. A simple sample preparation was based on a solid–liquid extraction using a solvent mixture acetonitrile/water/formic acid (79/20/1, v/v/v) and a dilution of raw extract using water/acetonitrile/formic acid (79/20/1, v/v/v) before instrumental analysis. Average recoveries in all three validated cereal crop types (maize, wheat, and barley), spiked at multiple levels, were found acceptable for all analytes when matrix-matched calibration was used, ranging from 63.2% to 111.2% and also showing very good repeatability, with relative standard deviations below 20%. Matrix effect (SSE) evaluation revealed maize as the most complex of the three analyzed cereal matrices, with strong SSE (150%) recorded for all 11 analyzed mycotoxins. An additional method verification was performed through successful participation in proficiency testing schemes, with the achieved z-scores generally in the acceptable range of −2 ≤ z ≤ 2. The obtained validation results demonstrated the suitability of the developed confirmatory multi-mycotoxin UHPLC-MS/MS method based on a dilute-and-shoot principle for the simultaneous determination of low concentrations of 11 EU-regulated mycotoxins in cereals, including aflatoxins B1, B2, G1 and G2, deoxynivalenol, fumonisins B1 and B2, zearalenone, T-2 and HT-2 toxins, and ochratoxin A.

A total of 209 samples of various cereal crops (maize, wheat, barley, rye and oats) grown in Croatian fields during 2016 and 2017 were collected to analyze and determine the occurrence and co-occurrence of EU regulated mycotoxins in cereals (AFB1, AFB2, AFG1, AFG2, DON, FB1, FB2, ZEA, T-2, HT-2 and OTA). The analysis, performed by a validated confirmatory LC-MS/MS method based on a dilute and shoot principle, highlighted Fusarium mycotoxins as the main contaminants, often co-occurring in samples from both years (50.0% in 2016 and 33.7% in 2017). DON was found to be the most frequent mycotoxin, present in 72.5% of the 2016 samples and 32.6% of the 2017 samples, while maize proved to be the most contaminated cereal type of both years with FUM as the most abundant mycotoxins, with an average concentration of 1180 µg/kg. Moderate temperatures with periods of high humidity favored the accumulation of DON in wheat samples instead of other Fusarium mycotoxins, while similar conditions favored maize contamination with FUM. A total of 8.3% of all the 2016 harvest samples and 7.9% of the 2017 harvest samples were assessed as non-compliant, containing mycotoxins in concentrations higher than the levels set by the EU legislation for food.

Abstract Increased interest in fullerene C60 and derivatives in recent years implies an intensification of their environmental spread. Yet, the potential risks for living organisms are largely unknown, including the interaction of C60 with fungal organisms. This may be especially relevant for mycotoxigenic fungi since C60 may both scavenge and produce reactive oxygen species (ROS), and oxidative stress induces mycotoxin production in fungi. Therefore, this study examined effects of environmentally plausible concentrations of C60 (0, 10, 50, and 100 ng/mL) on Aspergillus flavus growth and aflatoxin production in culture media. In addition, ROS-dependent oxidative stress biomarkers—thiobarbituric acid reactive substances (TBARS), reduced and oxidised glutathione ratio, superoxide dismutase isoenzymes, catalase, glutathione peroxidase, and glutathione reductase were determined in mycelia. Nanoparticles of fullerene C60 (nC60) did not exhibit strong antifungal activity against A. flavus. At the same time, nC60 caused an antiaflatoxigenic effect at 10–100 ng/mL, and 50 ng/mL unexpectedly enhanced aflatoxin production. The TBARS content, reduced and oxidised glutathione ratio, and copper, zinc superoxide dismutase activity suggest that 10 ng/mL nC60 exerted antioxidative action and reduced aflatoxin B1 production within fungal cells. Detected prooxidative effects of 50 ng/mL fullerene exceeded cellular defenses and consequently enhanced aflatoxin B1 production. Finally, the results obtained with 100 ng/mL nC60 point to prooxidative effects, but the absence of increase in aflatoxin output may indicate additional, presumably cytotoxic effects of nC60. Thus, a range of rather low levels of nC60 in the environment has a potential to modify aflatoxin production in A. flavus. Due to possible implications, further studies should test these results in environmental conditions.

There is a growing interest in alternative diets such as vegan or lacto-ovo vegetarian diet. The choice is made mainly for ethical, health reasons or both. Creatinine is a waste product of creatine phosphate metabolism in muscles, and the excretion rate is relatively constant. Forty-nine participants were tested in total: 5 lacto-ovo vegeterians, 14 vegans, and 30 omnivores. To compare the groups, all participants had a similar diet consisting of bread, bran flakes and wheat bran. The creatinine level was measured before the initiation of the diet in the first morning void, and in the sample of 24-h urine. The results confirmed slightly higher mean creatinine levels in males (1139 ± 517 mg/L), compared to females (901 ± 539 mg/L). Regular consumption of different diets did not show statistically significant differences in datasets on either first void or 24-h samples. The highest median result after 24-h was determined in lacto-ovo vegetarians (2175 mg/L), followed by omnivores (1328 mg/L), while vegans had the lowest median (1235 mg/L). Based on these results, there is no evidence that the vegetarian or vegan lifestyle influences the creatinine excretion rate.

The methods for fungal genomic DNA isolation for PCR amplification, including commercially available kits, must often be adapted in order to produce sufficient amounts of high-quality DNA from specific fungal species. The aim of this study was to select an optimal method for the isolation of DNA from Aspergillus flavus suitable for PCR reaction. Four different methods were compared according to their efficiency in isolating pure DNA, their price and time consumption. DNA quantification and purity estimation were performed using the NanoDropTM 1000 UV/VIS spectrophotometer and DNA integrity and PCR products were determined by gel-electrophoresis. DNA quantity ranged from 92.77 ± 11.52 to 5477.4 ± 22.03 ng/µL, with A260/280 from 1.14 ± 0.10 to 1.94 ± 0.16, and A260/230 0.37 ± 0.05 to 1.91 ± 0.17. There were also great differences in time consumption per sample, ranging from 1 hr 15 min to 7 hr 5 min. The determined costs per sample were ranging from 0.12 € to 2.29 € per sample. All tested methods were suitable for the isolation of A. flavus genomic DNA and subsequently for PCR reaction.

Penicillium expansum is a necrotrophic plant pathogen among the most ubiquitous fungi disseminated worldwide. It causes blue mould rot in apples during storage, transport and sale, threatening human health by secreting patulin, a toxic secondary metabolite that contaminates apples and apple-derived products. Nevertheless, there is still a lack of sufficient data regarding the resistance of different apple cultivars to P. expansum, especially ancient ones, which showed to possess certain resistance to plant diseases. In this work, we investigated the polyphenol profile of 12 traditional and 8 conventional apple cultivar and their resistance to P. expansum CBS 325.48. Eight polyphenolic compounds were detected; the most prominent were catechin, epicatechin and gallic acid. The highest content of catechin was detected in ‘Apistar’—91.26 mg/100 g of fresh weight (FW), epicatechin in ‘Bobovac’—67.00 mg/100 g of FW, and gallic acid in ‘Bobovac’ and ‘Kraljevčica’—8.35 and 7.40 mg/100 g of FW, respectively. The highest content of patulin was detected in ‘Kraljevčica’ followed by ‘Apistar’—1687 and 1435 µg/kg, respectively. In apple cultivars ‘Brčko’, ‘Adamčica’ and ‘Idared’, patulin was not detected. Furthermore, the patulin content was positively correlated with gallic acid (r = 0.4226; p = 0.002), catechin (r = 0.3717; p = 0.008) and epicatechin (r = 0.3305; p = 0.019). This fact indicates that higher contents of gallic acid, catechin and epicatechin negatively affected and boost patulin concentration in examined apple cultivars. This can be related to the prooxidant activity of polyphenolic compounds and sensitivity of P. expansum to the disturbance of oxidative status.

A 1,3-dihexadecyl-1H-benzo[d]imidazol-3-ium-tetraphenylborate (DHBI-TPB) ion-pair implemented in DHBI-TPB surfactant sensor was used for the potentiometric quantification of anionic surfactants in detergents and commercial household care products. The DHBI-TPB ion-pair was characterized by FTIR spectroscopy and computational analysis which revealed a crucial contribution of the C–H∙∙∙π contacts for the optimal complex formation. The DHBI-TPB sensor potentiometric response showed excellent analytical properties and Nernstian slope for SDS (60.1 mV/decade) with LOD 3.2 × 10−7 M; and DBS (58.4 mV/decade) with LOD 6.1 × 10−7 M was obtained. The sensor possesses exceptional resistance to different organic and inorganic interferences in broad pH (2–10) range. DMIC used as a titrant demonstrated superior analytical performances for potentiometric titrations of SDS, compared to other tested cationic surfactants (DMIC > CTAB > CPC > Hyamine 1622). The combination of DHBI-TPB sensor and DMIC was successfully employed to perform titrations of the highly soluble alkane sulfonate homologues. Nonionic surfactants (increased concentration and number of EO groups) had a negative impact on anionic surfactant titration curves and a signal change. The DHBI-TPB sensor was effectively employed for the determination of technical grade anionic surfactants presenting the recoveries from 99.5 to 101.3%. The sensor was applied on twelve powered samples as well as liquid-gel and handwashing home care detergents containing anionic surfactants. The obtained results showed good agreement compared to the outcomes measured by ISE surfactant sensor and a two-phase titration method. The developed DHBI-TPB surfactant sensor could be used for quality control in industry and has great potential in environmental monitoring.

Fullerol C60(OH)24 nanoparticles (FNP)-wheat-A. flavus interaction outcome is more complicated in the presence of drought. This study sheds light on how the presence of FNP affects food and feed safety from the perspective of mycotoxin contamination. The study aims to determine the influence of FNP at environmentally plausible concentrations on wheat growth under drought stress and on the aggressiveness of A. flavus during wheat germination, as well as the influence of FNP on the secondary metabolite profile during the inappropriate wheat storage. The co-occurrence of drought and FNP inhibited germination and shoot growth, while an application of FNP alone had no negative effect on plant growth. Wheat pre-treated with FNP showed a concentration dependent resistance pattern to A. flavus aggressiveness. Nevertheless, using a LC-MS/MS based multi-mycotoxin method, six secondary fungal metabolites: 3-nitropropionic acid (−1), aflatoxin B1 (−1) and B2 (−1), aflatoxicol (−1), kojic acid (−1), and O-methylsterigmatocystin (−1), were detected. FNP affected secondary metabolism of A. flavus during inappropriate wheat storage and increased the concentration of secondary metabolites in a concentration-dependent pattern (3-nitropropionic acid and kojic acid). In addition, aflatoxicol production was provoked in FNP treated samples.

A novel, simple, low-cost, and user-friendly potentiometric surfactant sensor based on the new 1,3-dihexadecyl−1H-benzo[d]imidazol−3-ium-tetraphenylborate (DHBI–TPB) ion-pair for the detection of cationic surfactants in personal care products and disinfectants is presented here. The new cationic surfactant DHBI-Br was successfully synthesized and characterized by nuclear magnetic resonance (NMR), Fourier transform infrared (FTIR) spectrometry, liquid chromatography–mass spectrometry (LC–MS) and elemental analysis and was further employed for DHBI–TPB ion-pair preparation. The sensor gave excellent response characteristics for CTAB, CPC and Hyamine with a Nernstian slope (57.1 to 59.1 mV/decade) whereas the lowest limit of detection (LOD) value was measured for CTAB (0.3 × 10−6 M). The sensor exhibited a fast dynamic response to dodecyl sulfate (DDS) and TPB. High sensor performances stayed intact regardless of the employment of inorganic and organic cations and in a broad pH range (2−11). Titration of cationic and etoxylated (EO)-nonionic surfactant (NSs) (in Ba2+) mixtures with TPB revealed the first inflexion point for a cationic surfactant and the second for an EO-nonionic surfactant. The increased concentration of EO-nonionic surfactants and the number of EO groups had a negative influence on titration curves and signal change. The sensor was successfully applied for the quantification of technical-grade cationic surfactants and in 12 personal care products and disinfectants. The results showed good agreement with the measurements obtained by a commercial surfactant sensor and by a two-phase titration. A good recovery for the standard addition method (98–102%) was observed.

To investigate into the T-2 and HT-2 toxin occurrence, 240 samples of unprocessed cereals (maize, wheat, barley, and oats) were sampled from different fields located in three Croatian regions during 2017–2018. In all samples, sum concentrations of T-2/HT-2 toxin were determined using the ELISA method, while the LC-MS/MS was used as a confirmatory method for both mycotoxins in positive samples (>LOD) and the establishment of T-2 over HT-2 toxin ratios. The results showed oats to be the most contaminated cereal, with T-2/HT-2 toxins detected in 70.0% of samples, followed by barley (40.9%), maize (26.8%) and wheat (19.2%), with the mean T-2/HT-2 ratio ranging from 1:2.7 in maize to 1:4.4 in oats. Sum T-2/HT-2 concentrations in two maize samples were higher than the indicative level recommended by the European Commission, necessitating subsequent investigations into the conditions under which these poorly investigated mycotoxins are produced. Statistically significantly (p < 0.05) higher concentrations of T-2/HT-2 toxin were determined in oats throughout study regions as compared to those found in wheat, but not maize and barley, while the concentrations of these mycotoxins were related to the regional weather in Croatia.

Cereals are still one of the most important food and feed sources, thus determining cereal’s safety, i.e., compliance with legislation, is extremely important. As systematic investigations of nowadays unavoidable secondary fungal metabolites and other common legally regulated contaminants occurrence in Croatian cereals are still lacking, this research aims to monitor the contamination levels of nation-wide crops by mycotoxins, pesticide residues, and heavy metals by employing UHPLC-MS/MS, GC-MS/MS, and atomic absorption spectrometer (AAS) validated analytical methods. The most common secondary fungal metabolites were found to be Fusarium mycotoxins, with DON being the most occurring present in 73.7% of the samples. At least one pesticide residue was found in 331.8% of the samples, and Hg and Cd were the most occurring heavy metals. A total of 8.5% of the samples was non-compliant to the European Union (EU) legislation for food regarding the found mycotoxins concentrations, 4.5% regarding pesticide residues and none regarding heavy metals. The unusual presence of certain pesticide residue and heavy metal indicates the importance of systematic control of the contaminant presence, in order to gather enough occurrence data for proper risk assessment that these contaminants represent for the consumer’s health.

In this study, the occurrence of multiple fungal metabolites including mycotoxins was determined in four different winter wheat varieties in a field experiment in Croatia. One group was naturally infected, while the second group was inoculated with a Fusarium graminearum and F. culmorum mixture to simulate a worst-case infection scenario. Data on the multiple fungal metabolites including mycotoxins were acquired with liquid chromatography with mass spectrometry (LC-MS/MS) multi-(myco)toxin method. In total, 36 different fungal metabolites were quantified in this study: the Fusarium mycotoxins deoxynivalenol (DON), DON-3-glucoside (D3G), 3-acetyldeoxynivalenol (3-ADON), culmorin (CULM), 15-hydroxyculmorin, 5-hydroxyculmorin, aurofusarin, rubrofusarin, enniatin (Enn) A, Enn A1, Enn B, Enn B1, Enn B2, Enn B3, fumonisin B1, fumonisin B2, chrysogin, zearalenone (ZEN), moniliformin (MON), nivalenol (NIV), siccanol, equisetin, beauvericin (BEA), and antibiotic Y; the Alternaria mycotoxins alternariol, alternariolmethylether, altersetin, infectopyron, tentoxin, tenuazonic acid; the Aspergillus mycotoxin kojic acid; unspecific metabolites butenolid, brevianamid F, cyclo(L-Pro-L-Tyr), cyclo(L-Pro-L-Val), and tryptophol. The most abundant mycotoxins in the inoculated and naturally contaminated samples, respectively, were found to occur at the following average concentrations: DON (19,122/1504 µg/kg), CULM (6109/1010 µg/kg), 15-hydroxyculmorin (56,022/1301 µg/kg), 5-hydroxyculmorin (21,219/863 µg/kg), aurofusarin (43,496/1266 µg/kg). Compared to naturally-infected samples, Fusarium inoculations at the flowering stage increased the concentrations of all Fusarium mycotoxins, except enniatins and siccanol in Ficko, the Aspergillus metabolite kojic acid, the Alternaria mycotoxin altersetin, and unspecific metabolites brevianamid F, butenolid, cyclo(L-Pro-L-Tyr), and cyclo(L-Pro-L-Val). In contrast to these findings, because of possible antagonistic actions, Fusarium inoculation decreased the concentrations of the Alternaria toxins alternariol, alternariolmethylether, infectopyron, tentoxin, tenuazonic acid, as well as the concentration of the nonspecific metabolite tryptophol.

Despite the efforts to control mycotoxin contamination worldwide, extensive contamination has been reported to occur in food and feed. The contamination is even more intense due to climate changes and different stressors. This study examined the impact of fullerol C60(OH)24 nanoparticles (FNP) (at 0, 1, 10, 100, and 1000 ng mL−1) on the secondary metabolite profile of the most relevant foodborne mycotoxigenic fungi from genera Aspergillus, Fusarium, Alternaria and Penicillium, during growth in vitro. Fungi were grown in liquid RPMI 1640 media for 72 h at 29 °C, and metabolites were investigated by the LC-MS/MS dilute and shoot multimycotoxin method. Exposure to FNP showed great potential in decreasing the concentrations of 35 secondary metabolites; the decreases were dependent on FNP concentration and fungal genus. These results are a relevant guide for future examination of fungi-FNP interactions in environmental conditions. The aim is to establish the exact mechanism of FNP action and determine the impact such interactions have on food and feed safety.

Micro matrix solid phase dispersion (micro-MSPD) was optimized by response surface methodology for the extraction of polyphenols from the peel of twelve traditional and eight commercial apple varieties grown in Croatia. The optimized micro-MSPD procedure includes the use of 0.2 g of sample, 0.8 g of dispersant, a 57% solution of methanol in water as the solvent and 5 mL of extract volume. The total polyphenolic index (TPI) and antioxidant activity (AA) were measured by spectrophotometric assays. Eighteen polyphenolic compounds were identified in all investigated apples by HPLC-DAD and LC-(ESI)-MS. The peel of traditional apple varieties had higher contents of all investigated polyphenols. Calculated relative contribution of polyphenol groups indicated non-flavonoids (28.6%) and flavanols (46.2%) as the major contributors to the total polyphenolic content in traditional and commercial apple varieties, respectively. The most abundant polyphenol in traditional apple peel was chlorogenic acid, procyanidin B2 and epicatechin (1143 ± 755 µg/g dw, 954 ± 343 µg/g dw and 560 ± 362 µg/g dw, respectively). The peel of varieties ‘Apistar’, ‘Bobovac’ and ‘Božićnica’ could be highlighted as an important source of polyphenols.

Objective. The aims of this study were to determine the HCV-RNA viral load, genotype distribution, risk factors and symptoms of HCVRNA positive viral load in HCV antibody-positive patients from north-eastern Croatia. Materials and methods. From January 2009 to December 2011, 203 HCV antibody- positive patients (130 men and 73 women; median age 44.5 years) were analyzed for HCV-RNA by the COBAS TaqMan HCV test and genotyped by the Linear Array HCV Genotyping test (both from Roche). All patients completed a structured questionnaire about risk factors and symptoms. Results. The HCV-RNA percentage was 61.1% and was similar for men and women. The HCV-RNA viral load increased with age: while 55% of 20-50 year old patients were HCV-RNA positive, 73% of patients >50 years were positive (p=0.021). Genotype 1 was the most prevalent genotype (79.8%), followed by 3 (12.9%), 4 (6.5%), and 2 (0.8%); genotypes 5 and 6 were not determined. Patients with genotype 1 (median, 50 years) were older than patients with 3 (median, 33.5 years) or 4 (median, 38 years). The blood transfusions performed in Croatian hospitals before 1993 was significantly associated with HCV-RNA positive viral load (p

Climatic changes influence considerably the distribution and occurrence of different secondary metabolites in cereals. The aim of this investigation was to assess the changes in metabolite prevalence observed in six different winter barley varieties over a statistically significant period of three years by linking agro-climatic conditions with metabolite concentrations in chosen samples. The results showed that temperatures and precipitation levels varied during the observed timeframe and that the multi-toxin concentrations followed the trend of changing climatic conditions depending on the variety. All quantified (fungal) metabolites showed significant variations throughout the years and, for some (tryptophol and the cyclic dipeptides cyclo(L-Pro-L-Tyr) and cyclo(L-Pro-L-Val)), an unexpected, but clear connection can be made with temperature changes and precipitation levels during the growing season.

The aim of this study was to assess the impact of steeping water change and Fusarium graminearum contamination level on different multi-toxin types and concentrations in barley malt. Malt samples were subjected to two micromalting regimes—steeping water change and the other with no steeping water change. Malt was contaminated with different F. graminearum contamination levels (0%, 10%, and 20%). The results indicate that malt with higher F. graminearum contamination levels ensured higher concentrations of toxins. Higher fungal metabolite concentrations were determined in samples exposed to freshly-changed steeping water, especially zearalenone and its derivates whose values were three to four times higher than in samples with no water change. Zearalenone-4-sulfate showed four (in 10% contamination) and even thirty times (in 20% contamination) higher concentrations than in samples with no water change. Water change during malting resulted in higher levels of multi-toxins in the final product.

The aim of this study was to develop a liquid chromatography/tandem mass spectrometry LC-MS/MS method for the simultaneous determination of acrylamide and hydroxymethylfurfural (HMF) in corn snack products enriched with food industry by-products: brewer’s spent grain (BSG), sugar beet pulp (SBP) and apple pomace (AP). Development of the method included the study of different sources for ionization, different mobile phases, different extraction conditions as well as different methods of sample preparation. Finally, the single LC-MS/MS method was developed for the analysis of both analytes in one step with a duration of 20 min using a simple single-step extraction. The method with apparent recoveries of 91.4 and 90.4 for acrylamide and HMF, respectively, was applied for the analysis of non-extruded and extruded samples. The obtained results shown that the acrylamide content was

The focus of present study is on Codium bursa collected from the Adriatic Sea. C. bursa volatiles were identified by gas chromatography and mass spectrometry (GC-FID; GC-MS) after headspace solid-phase microextraction (HS-SPME), hydrodistillation (HD), and supercritical CO2 extraction (SC-CO2). The headspace composition of dried (HS-D) and fresh (HS-F) C. bursa was remarkably different. Dimethyl sulfide, the major HS-F compound was present in HS-D only as a minor constituent and heptadecane percentage was raised in HS-D. The distillate of fresh C. bursa contained heptadecane and docosane among the major compounds. After air-drying, a significantly different composition of the volatile oil was obtained with (E)-phytol as the predominant compound. It was also found in SC-CO2 extract of freeze-dried C. bursa (FD-CB) as the major constituent. Loliolide (3.51%) was only identified in SC-CO2 extract. Fatty acids were determined from FD-CB after derivatisation as methyl esters by GC-FID. The most dominant acids were palmitic (25.4%), oleic (36.5%), linoleic (11.6%), and stearic (9.0%). FD-CB H2O extract exhibited better antifungal effects against Fusarium spp., while dimethyl sulfoxide (DMSO) extract was better for the inhibition of Penicillium expansum, Aspergillus flavus, and Rhizophus spp. The extracts showed relatively good antifungal activity, especially against P. expansum (for DMSO extract MIC50 was at 50 µg/mL).

A series of newly disubstituted (compounds 4a,b) and trisubstituted 1,3,4-thiadiazines 5a–l with various substituents was prepared utilizing different thiosemicarbazides and 3-α-bromoacetylcoumarins as starting compounds. The structures of the synthesized 1,3,4-thiadiazines are elucidated and confirmed utilizing the corresponding analytical and spectroscopic data. All of the new thiadiazine derivatives were tested for their antioxidant activity, employing different antioxidant assays (DPPH scavenging activity, iron chelating activity, power reducing activity). Compounds 5b, 5f, 5j and 4b were proven to be the best DPPH radical scavengers, while compounds 5h and 5j have shown the best iron chelating activity. Thiadiazine derivatives were also tested on their antifungal activity against four mycotoxicogenic fungi, Aspergillus flavus, A. ochraceus, Fusarium graminearum and F. verticillioides. The best antifungal against A. flavus was proven to be compound 5e, while compounds 4a and 5c were the best antifungals on A. ochraceus, and compound 5g showed the best antifungal activity on F. verticillioides.

Contamination with phytopathogenic forms of Fusarium, besides field crops, may also occur in stored products. Addition of antifungal substances to stored livestock feed is therefore common. This paper examined the effectiveness of a mixture of synthetic and natural antioxidants against the growth of Fusarium graminearum and F. verticillioides in a concentrate mixture. The most effective inhibition of growth was achieved with a mixture of butylated hydroxyanisole, propyl paraben and thymol.

Fungi, yeasts, and bacteria are common microorganisms on cereals used in malting and brewing industries. These microorganisms are mostly associated with the safety and quality of malt and beer, but also with the health safety of by-products used in animal nutrition. The real problem is their harmful metabolites—toxins that, due to their thermostable properties, can easily be transferred to malting and brewing by-products. Besides fungal metabolites, other toxins originating from plants can be harmful to animal health. Precise and accurate analytical techniques broadened the spectrum of known toxins originating from microorganisms and plants that can pose a threat to animal health. Multi-(myco)toxin analyses are advanced and useful tools for the assessment of product safety, and legislation should follow up and make some important changes to regulate yet unregulated, but highly occurring, microbial and plant toxins in malting and brewing by-products used for animal feed.

Aspergillus flavus is the most important mycotoxin-producing fungus involved in the global episodes of aflatoxin B1 contamination of crops at both the pre-harvest and post-harvest stages. However, in order to effectively control aflatoxin contamination in crops using antiaflatoxigenic and/or antifungal compounds, some of which are photosensitive, a proper understanding of the photo-sensitive physiology of potential experimental strains need to be documented. The purpose of the study is therefore to evaluate the effect of visible (VIS) light illumination on growth and conidiation, aflatoxin production ability and modulation of A. flavus oxidative status during in vitro experiment. Aflatoxigenic A. flavus strain was inoculated in aflatoxin-inducing YES media and incubated under three different VIS illumination regimes during a 168 h growth period at 29 °C. VIS illumination reduced A. flavus mycelia biomass yield, both during growth on plates and in liquid media, promoted conidiation and increased the aflatoxin production. Furthermore, aflatoxin production increased with increased reactive oxidative species (ROS) levels at 96 h of growth, confirming illumination-driven oxidative stress modulation activity on A. flavus cells.

A series of Schiff’s bases (E)-N-2-aryliden-2-(4-methyl-2-oxo-2H-chromen-7-yloxy)acetohydrazides 2a-l and N-(2-(substituted phenyl)-4-oxo-thiazolidin-3-yl)-2-(4-methyl-2-oxo-2H-chromen-7-yloxy)acetamides 3a-l were synthesized and evaluated for their antioxidant activity by the phosphomolybdenum method. Most of the Schiff’s bases and thiazolidine-4-ones bearing two hydroxyl groups on the phenyl ring showed excellent antioxidant activity in comparison with ascorbic acid. Preliminary investigation on cytotoxic and antifungal activity was done on some representative samples.

Versatile microbiota are inevitably naturally present on cereals. Fungi, yeasts and bacteria and their metabolites all contribute to the quality and safety of the final products derived from most common beer cereals—barley and wheat. The microorganisms that are most often associated with the safety and quality of cereals for beer production belong to the Fusarium spp. They greatly influence yields from the field, and can modify and diminish economic success for farmers. However, the real problem is their harmful metabolites—mycotoxins—that affect the health of humans and animals. In the era of emerging analytical methodologies, the spectrum of known toxins originating from microorganisms that can pose a threat to humans has grown tremendously. Therefore, it is necessary to monitor microflora throughout the productive “barley to beer” chain and to act suppressive on the proliferation of unwanted microorganisms, before and during malting, preventing the occurrence of mycotoxins in final products and by-products. Multi-mycotoxin analyses are very advanced and useful tools for the assessment of product safety, and legislation should follow up and make some important changes to regulate as yet unregulated, but highly occurring, microbial toxins in malt and beer.