Research background. Outer scales of common onion (Allium cepa L.) are often discarded as biological waste but can be an interesting source of polyphenols with antioxidant and antidiabetic properties that can be used in the food and pharmaceutical industries. Experimental approach. The study goal is to use dried chamomile flowers (Matricaria camomilla L.) containing 6 % flavonoid apigenin in the form of water extract to improve the polyphenol profile, antioxidant and antiglycation activity of onion outer scales. “Red Carmen” onion bulbs with roots were exposed to deionized water (control) and chamomile water extract (test treatment) in a phytotron (23 °C/16 h day/8 h dark). From dried and pulverized outer scales and chamomile, Fourier transform infrared (FTIR) analysis was performed. Also, pure standards of polyphenols (apigenin, quercetin and caffeic acid) was used for FTIR spectra comparison. Results and conclusions. In the initial phase, the salivary, gastric, and intestinal phase; THA (total hydroxycinnamic acids), TFL (total flavonols), antioxidant (ABTS; (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonicacid and FRAP; ferric reducing antioxidant power) activity, and inhibition of BSA (bovine serum albumin) glycation activity of on-ion scales extract was measured. Chamomile treatment improved THA, TFL, antioxidant (FRAP) and antiglycation activity (BSA method) in almost all in vitro digestion phases. Treatment of onions with chamomile water extract is good method of enrichment of biological waste. This is confirmed by FTIR analysis. Enricht onion scales represent good sources of bioaccessible polyphenols with high antioxidant (ABTS>85 % and FRAP>95 %) and moderate antiglycation activity (43–62 %). Novelty and scientific contribution. The enrichment of the phytochemical composition and biological potential of onions had potential applications of this technique for health and nutritional purposes. Since the method does not require genetic modification but relies on natural absorption mechanisms and stress induction, it could become a valuable tool for enriching the nutritional and therapeutic composition of herbs. These results represent a foundation for future research, which should further elucidate the mechanisms of transmission, long-term effects, and potential industrial applications of this phenomenon.

Since honey has a therapeutic role in the treatment of many diseases, we investigated the content of phenolic compounds and the antioxidant activity in acacia (Robinia pseudoacacia L.), chestnut (Castanea sativa Mill.) and lime-tree (Tilia spp.) honey originating from Croatia and Germany. Total phenols, flavonols, and flavanols contents were observed at higher levels in Croatian Castanea honey compared to German Castanea honey. Significant higher values of total flavanols and hydroxycinnamic acids were measured in Croatian Tilia honey compared to German Tilia honey. For Robinia honey, significantly higher values of total phenols and flavonols were observed in almost all Croatian honey samples compared to German honey. Croatian honey samples had higher antioxidant activity compared to German honey samples with most tested methods. The highest total phenols, total flavanols, ABTS, DPPH, and FRAP values were measured in Castanea honey, then in Robinia honey, and the lowest values in Tilia honey samples. With new developed HPLC method, pinobanksin, pinocembrin, and chrysin were identified in the majority of honey samples. Our results imply that both botanical and geographical origin influence the final quality of phenolic compounds and antioxidant activity in honey. A high positive correlation between the results of antioxidant activity and polyphenols was detected.

The goal of this research is improved polyphenol profile, antioxidant and antiglycation activity of onion bulbs using Matricariae flos (Matricaria chamomilla L.) water extract (12 g L-1). The incubation of 24 h of onion roots was in glass tubes filled with deionized water (control) and in aqueous extract of Matricariae flos (test treatment), in a climate chamber. Total phenols, total hydroxycinnamic acids, total flavonols, antioxidant activity and antiglycation activity were measured in ethanolic (7.5 g L-1) extract before (initial phase) and after in vitro digestion (salivary, gastric and intestinal phase). In almost all phases of in vitro digestion Matricariae flos treatment statistically improved total phenols, total hydroxycinnamic acids, total flavonols, antioxidant activity (ABTS and FRAP) and antiglycation activity (BSA method). The results presented positive influence of Matricariae flos treatment of onion bulbs. Aqueous chamomile extract increases the proportion of polyphenols and antioxidant and antiglycation activity before and after in vitro digestion.

Impatiens glandulifera Royle (I. glandulifera) in the EU presents a threat to the ecosystem, and is thus on the list of invasive alien species. The aim of this study was to clarify if an allelopathic effect of I. glandulifera involves the activation of oxidative stress in target plants. First, I. glandulifera leaf extract was prepared and levels of 2-hydroxy-1,4-naphtoquinone (2-HNQ) and 2-methoxy-1,4-naphthoquinone (2-MNQ), as main allelochemicals of I. glandulifera, determined by HPLC. Next, seeds of white mustard (Sinapis alba) or radish (Raphanus sativus) were exposed to the extract or to 2-MNQ (in the concentration range of 1–30 µg/mL) for 3 days and parameters of germination and oxidative stress were assessed. Both the leaf extract and 2-MNQ inhibited germination of white mustard and radish; however, the effect of 2-MNQ was more pronounced. Furthermore, the extract and 2-MNQ induced activation of antioxidative defense and caused oxidative damage to lipids and proteins in white mustard and radish seedlings. It was observed that radish seedlings were less susceptible to negative effect of I. glandulifera extract and 2-MNQ. This study’s obtained results demonstrated that 2-MNQ is the main allelochemical of I. glandulifera and that the mechanism by which I. glandulifera and 2-MNQ induce negative effects to target plants involves oxidative stress. In addition, species-dependent susceptibility to I. glandulifera and 2-MNQ was shown.

Extracts obtained from common butterbur (Petasites hybridus), standardized to petasins, are existing pharmaceutical options for the treatment and/or prevention of allergic rhinitis (leaves) and migraine (rhizomes). In this study, the total phenolic content, flavonoid content, and antioxidant potential of ten samples of Croatian Petasites species (four P. hybridus, four P. albus, one P. kabli kianus, and one P. paradoxus) obtained by ultrasound-assisted extraction of leaves were compared. The total phenolic content (Folin-Ciocalteu assay) of methanolic leaf extracts ranged from 4.43 ± 0.09 to 10.76 ± 0.60 mg gallic acid equivalent g-1 dry mass (mg GAE g-1 DM) for P. hybridus and from 6.66 ± 0.43 to 19.92 ± 2.90 mg GAE g-1 DM for P. albus samples, while those of P. kablikianus and P. paradoxus were equal to 7.56 ± 0.17 mg GAE g-1 DM and 10.22 ± 0.46 mg GAE g-1 DM, respectively. Flavonoid content (AlCl3 assay) varied between 2.51 ± 0.10 and 4.03 ± 0.08 mg quercetin equivalent g-1 dry mass (mg QE g-1 DM) for P. hybridus and between 2.21 ± 0.09 and 5.22 ± 0.02 mg QE g-1 DM for P. albus samples, while those of P. kablikianus and P. paradoxus were equal to 5.59 ± 0.05 mg QE g-1 DM and 5.50 ± 0.09 mg QE g-1 DM, respectively. Antioxidant potential was in high correlation with total phenolic content (r = 0.93, p < 0.001). Due to the expected contribution of plant polyphenols and flavonoids to the activity of butterbur extracts and their observed great variabilities, determining the content of these compounds may be of interest to the pharmaceutical industry.

This study evaluates the antihyperlipidemic (pancreatic lipase inhibition assay), antiglycation (inhibition of bovine serum albumin glycation, BSA glycation), and antioxidant activity (ABTS, DPPH and FRAP assays) of ethanolic extracts from flowering parts of five widely distributed plant species in Croatia - Crocus heuffelianus Herb. (tepals), Nicotiana tabacum L. (petals), Malva sylvestris L. (petals), Calendula officinalis L. and Helianthus annuus L. (both sterile ligulate flowers). An in vitro-simulated system of human digestion was employed to assess the bioaccessibility of the selected phenolics and the stability of the extracts' antioxidant, hypolipi demic, and antiglycation potential following each digestion phase. The concentrations of l-ascorbic acid, individual flavonoids, and phenolic acids were determined using RP-HPLC analysis. Principal component analysis revealed significant differences in the content of bioactive compounds and their biological activity among the investigated plant species. All original extracts exhibited high antioxidant capacity (> 70 %) in at least one assay, with N. tabacum and H. annuus demonstrating the strongest anti-oxidant capacity throughout digestion. H. annuus contained the highest levels of total identified phenolic acids, total identified phenols, and total identified compounds, while N. tabacum and C. heuffelianus exhibited the highest total flavonoid content. Among individual compounds, protocatechuic acid, quercetin, and ferulic acid significantly contributed to antioxidant activity. N. tabacum had the strongest antihyperlipidemic potential in the original extracts, as well as in the most digestion phases. Strong BSA glycation inhibition (70-100 %) was observed in all plant extracts across various digestion phases, with the exception of C. heuffelianus, which exhibited mode rate inhibitory effects. These findings suggest that the analyzed flower-derived plant materials, some of which are often considered agricultural waste, can serve as sustainable and valuable resources of bioactive compounds for functional food, dietary supplements, and pharmaceutical applications.

The present work evaluates the effect of casein, glucose, and olive oil on phytochemical bioaccessibility, antioxidant potential (DPPH and FRAP), antidiabetic potential (inhibition of amylase, α-glucosidase, and BSA glycation), and antihyperlipidemic potential (inhibition of lipase) of gingko standardized leaf extract in the form of tablets after in vitro digestion. Gingko extract formulations with protein, carbohydrates, and oil had high (>70%) in vitro bioaccessibility of quercetin, kaempferol, and isorhamnetin after each phases of digestion in comparison to moderate (35–70%) in vitro bioaccessibility from gingko water extract. Formulation with the highest in vitro bioaccessibility of the majority of the tested polyphenolic groups and terpene lactones after oral and intestinal phases was ginkgo with olive oil. High (>70%) antioxidant (DPPH and FRAP), antidiabetic (α-glucosidase and BSA glycation), and antihyperlipidemic potential were detected in almost all ginkgo formulations. Based on the results, we conclude that the in vitro bioaccessibility of individual compounds or groups of compounds depends on whether the tablets are taken with water or with foods (protein, carbohydrates, and oil).

Iris taxa are sources of valuable essential oils obtained from aged rhizomes used by various industries, including pharmacy, cosmetic, perfume, and food industry, in which irones are the most important aroma components. In this study, volatile organic compounds (VOCs) obtained from dried rhizomes of three endemics from Croatia, Iris pseudopallida, I. illyrica, and I. adriatica, were studied. The VOCs were isolated by three different methods: headspace solid-phase microextraction (HS–SPME) using divinylbenzene/carboxene/polydimethylsiloxane (DVB/CAR/PDMS) fiber or polydimethylsiloxane/divinylbenzene (PDMS/DVB) fiber, and hydrodistillation (HD). The samples were analyzed by gas chromatography–mass spectrometry (GC–MS). In five out of six samples, the main compounds detected by HS–SPME were perilla aldehyde, butan-2,3-diol, acetic acid, 2-phenylethanol, benzyl alcohol, hexanal, and nonanal, while 6-methylhept-5-en-2-one, trans-caryophyllene, and ethanol were common for all studied samples. The former VOCs were absent from the oldest, irone-rich I. pseudopallida sample, mainly characterized by cis-α-irone (43.74–45.76%). When using HD, its content was reduced (24.70%), while docosane prevailed (45.79%). HD yielded predominantly fatty acids, including myristic, common for all studied taxa (4.20–97.01%), and linoleic (40.69%) and palmitic (35.48%) as the major VOCs of I. adriatica EO. The performed GC–MS analyses of EOs, in combination with HS–SPME/GC–MS, proved to be useful for gaining a better insight into Iris VOCs.

The study aimed to develop an HPLC method for a simultaneous determination of 2-hydroxy-1,4-naphthoquinone (2-HNQ) and 2methoxy-1,4-naphthoquinone (2-MNQ) in plant material.The method was implemented to determine naphthoquinones (NQs) in leaves and flowers of the invasive alien species Impatiens glandulifera and I. balfourii collected in Croatia.Two NQs extraction methods from plant material were compared: decoction and ultrasonic extraction.Validation parameters indicate that the developed method is reliable for the simultaneous determination of 2-HNQ and 2-MNQ in plant material.The decoction extraction obtained a higher NQs yield than ultrasonic extraction.In the leaves and flowers of I. glandulifera and I. balfourii only 2-MNQ was detected.Both, I. glandulifera and I. balfourii, had higher 2-MNQ concentrations in flowers than in leaves.A significantly higher 2-MNQ concentration was found in I. glandulifera compared to I. balfourii.Therefore, I. glandulifera can be considered a rich source of 2-MNQ whose antitumor potential is established.

Salvia brachyodon, the short-tooth sage, is one of the rarest plant species and endemic in the Adriatic area of the Balkan Peninsula. As aside from its essential oil, only limited information on its phytochemical composition is known, a more detailed study of the leaves was undertaken. From its leaves two diterpenes, agastanol (2), and a new natural compound 1, i.e., 3-methyl-4-methylen-11,12,14-trihydroxy-8,11,13-abietatrien-7-one, were isolated and identified by NMR spectroscopy and mass spectrometry. In addition, caffeic acid, isoquercitrin, luteolin 7-O-glucoside and rosmarinic acid were identfied by comparison with reference compounds. The fraction containg the diterpenes as well as the isolated compound 1 showed significant antimycobacterial activity against Mycobacterium smegmatis. The diterpenes of S. brachyodon represent promising antimycobacterial substances for further evaluation. Due to the endangered nature of the plant, the wide use of S. brachyodon and its bioactive compounds could be achieved by growing the plants in culture.

The objective of this study was to spectrophotometric determinate the total phenolic, flavonoid, hydroxycinnamic acid, and flavonol content of orange, pineapple, and apple juices fortified with wild thyme (Thymus serpyllum L.), Dalmatian sage (Salvia officinalis L.), and wild thyme-Dalmatian sage (3 : 1, v / v) extracts, and to evaluate their DPPH radical scavenging activity as a contribution to the development of a new functional beverage.The plant extracts addition increased the amount of phenolic compounds in fruit juices and improved their antioxidant properties.The highest concentrations of bioactive compounds and the greatest DPPH radical activity were obtained by adding Dalmatian sage extract to orange juice.Our study provides the novelty of fortifying fruit juices with wild thyme and Dalmatian sage extracts and offers significant potential for the creation of functional beverages.

The aim of this study was to compare the effects of dexmedetomidine and dexamethasone as adjuvants to preoperative epidural administration of local anesthetic (ropivacaine) in thoracic surgery on the postoperative level of pain, use of analgesics, inflammation, and oxidative stress. The study enrolled 42 patients who underwent elective thoracic surgery in a one-year period at the University Hospital Dubrava (Zagreb, Croatia). Based on a computer-generated randomization list the patients were assigned to the dexmedetomidine (n = 18) or dexamethasone (n = 24) group. Postoperatively, patients of dexmedetomidine group reported lower pain (VAS value 1 h post surgery, 3.4 ± 2.7 vs. 5.4 ± 1.8, dexmedetomidine vs. dexamethasone, p < 0.01) and had lower anal-gesic requirements in comparison with dexamethasone group. Thus, dexmedetomidine in comparison with dexamethasone was more efficient in lowering pain and analgesia requirements 24 h after the surgery. On the contrary, dexamethasone had better anti-inflammatory properties (CRP level 24 h post surgery, 131.9 ± 90.7 vs. 26.0 ± 55.2 mg L−1, dexmedetomidine vs. dexamethasone, p < 0.01). Both dexmedetomidine and dexamethasone exhibited antioxidant effects, however, their antioxidant properties should be further explored. The results of this study improve current knowledge of pain control in thoracic surgery.

Species from the genus Globularia L. have been used as healing agents for various ailments, with utilization of Globularia alypum L. being most frequently reported. The aim of this study was to evaluate the antidiabetic, antioxidant, anti-inflammatory, antibacterial and anticancer potential of G. alypum and three related species, G. punctata Lapeyr., G. cordifolia L. and G. meridionalis (Podp.) O.Schwarz, in relation to their phytochemical compositions. Globularin and verbascoside were identified using LC-PDA-ESI-MSn as the major metabolites of G. alypum with known biological activities. G. alypum demonstrated the greatest α-glucosidase inhibitory activity and DPPH radical scavenging activity (IC50 = 17.25 μg/mL), while its anti-inflammatory activity was not significantly different from those of related species. All investigated species showed considerable antibacterial activity against methicillin-resistant Staphylococcus aureus in the broth microdilution method (MIC = 1.42–3.79 mg/mL). G. punctata also showed antibacterial activities against Escherichia coli (MIC = 1.42 mg/mL), Bacillus subtilis (MIC = 1.89 mg/mL), B. cereus (MIC = 2.84 mg/mL) and Enterococcus faecalis (MBC = 5.68 mg/mL). G. punctata, G. cordifolia and G. meridionalis showed greater anticancer potential than G. alypum. Obtained results indicate investigated Globularia species could serve as sources of diverse bioactive molecules, with G. punctata having the greatest antibacterial potential.

Acacetin, apigenin, chrysin, and pinocembrin are flavonoid aglycones found in foods such as parsley, honey, celery, and chamomile tea. Flavonoids can act as substrates and inhibitors of the CYP3A4 enzyme, a heme containing enzyme responsible for the metabolism of one third of drugs on the market. The aim of this study was to investigate the inhibitory effect of selected flavonoids on the CYP3A4 enzyme, the kinetics of inhibition, the possible covalent binding of the inhibitor to the enzyme, and whether flavonoids can act as pseudo-irreversible inhibitors. For the determination of inhibition kinetics, nifedipine oxidation was used as a marker reaction. A hemochromopyridine test was used to assess the possible covalent binding to the heme, and incubation with dialysis was used in order to assess the reversibility of the inhibition. All the tested flavonoids inhibited the CYP3A4 enzyme activity. Chrysin was the most potent inhibitor: IC50 = 2.5 ± 0.6 µM, Ki = 2.4 ± 1.0 µM, kinact = 0.07 ± 0.01 min−1, kinact/Ki = 0.03 min−1 µM−1. Chrysin caused the highest reduction of heme (94.5 ± 0.5% residual concentration). None of the tested flavonoids showed pseudo-irreversible inhibition. Although the inactivation of the CYP3A4 enzyme is caused by interaction with heme, inhibitor-heme adducts could not be trapped. These results indicate that flavonoids have the potential to inhibit the CYP3A4 enzyme and interact with other drugs and medications. However, possible food–drug interactions have to be assessed clinically.

Propolis is flavonoid rich source that has shown numerous beneficial biological effects such as antibacterial, antiviral, antitumoral, antiaggregatory, and hepatoprotective. Ginkgo has been used as a dietary supplement for enhancement of cognitive functions. The aim of this study was assessment of the effect of propolis and ginkgo extracts on primary hemostasis and verify whether the use of a combination of propolis and ginkgo extracts in the prevention of cardiovascular diseases is justified. Commercially available extracts were assessed by impedance aggregometry on the whole blood samples of healthy subjects collected at the Croatian Institute of Transfusion Medicine. Two aggregation inducers were used: ADP (adenosine diphosphate, weak agonist of platelet aggregation) and TRAP (hexapeptide surrogate of thrombin, strong inducer of platelet aggregation). In an ADP induced aggregation assay, propolis showed an antiaggregatory effect at a concentration of 10 μM. In the same test, the antiplatelet effect of ginkgo extract was absent. In the hexapeptide-induced thrombin mimetic (TRAP) aggregation test, propolis showed an antiplatelet effect at a concentration of 20 μM as well as ginkgo extract. Combinations of propolis and ginkgo in a ratio of 1:25 and 1:50 showed an antiplatelet effect at 20 μM concentrations of extracts. Combination of propolis and ginkgo is justified from a pharmacodynamic aspect, as propolis and ginkgo have different mechanisms of action because they act on different signaling pathways of aggregation.

The present paper gives basic data on vaccination and vaccine production using recombinant DNA technology with emphasis put on the possibilities, advantages and expected challenges of recombinant vaccine production in higher plants. The history and present situation considering utilization of plants in recombinant protein production, as well as its future perspectives, including the potential of plants in the development of vaccines against SARS-CoV-2, are also considered.

A study on the headspace volatile organic compounds (VOCs) profile of native populations of Sideritis romana L. and Sidertis montana L., Lamiaceae, from Croatia is reported herein, to elucidate the phytochemical composition of taxa from this plant genus, well-known for traditional use in countries of the Mediterranean and the Balkan region. Headspace solid-phase microextraction (HS-SPME), using divinylbenzene/carboxene/polydimethylsiloxane (DVB/CAR/PDMS) or polydimethylsiloxane/divinylbenzene (PDMS/DVB) fiber, coupled with gas chromatography-mass spectrometry (GC-MS) was applied to analyze the dried aerial parts of six native populations in total. Furthermore, principal component analysis (PCA) was conducted on the volatile constituents with an average relative percentage ≥1.0% in at least one of the samples. Clear separation between the two species was obtained using both fiber types. The VOCs profile for all investigated populations was characterized by sesquiterpene hydrocarbons, followed by monoterpene hydrocarbons, except for one population of S. romana, in which monoterpene hydrocarbons predominated. To our knowledge, this is the first report on the VOCs composition of natural populations of S. romana and S. montana from Croatia as well as the first reported HS-SPME/GC-MS analysis of S. romana and S. montana worldwide.

Himalayan balsam, Impatiens glandulifera Royle, and Balfour’s touch-me-not, Impatiens balfourii Hooker f., are invasive weeds that are threatening habitat diversity in many European countries, including Croatia. Various bioactive secondary metabolites from species of genus Impatiens L. have been reported and some species are traditionally used as herbal medicines. Th e aim of this research was to determine total polyphenol (TP) and total fl avonoid (TF) content, as well as antioxidant activity of leaves and fl owers of species I. glandulifera and I. balfourii. Spectrophotometric methods were used to obtain TP, TF content and inhibition of 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals to measure antioxidant activity. Total polyphenol quantity ranged from 7.9 to 18.6 mg equivalent of gallic acid per g of dried plant material, while total fl avonoid content ranged from 4.6 to 20.5 mg equivalent of quercetin per g of dried plant material. Leaves of I. glandulifera contain more polyphenols than fl owers of the species (18.5 vs. 13.1 mg EGA/g DPM) but less fl avonoids comparing to fl owers (5.7 vs. 8.3 mg EQ/g D PM). However, in the case of I. balfourii both polyphenols and fl avonoids are more abundant in fl owers and have the highest quantity of polyphenols and fl avonoids overall (18.6 mg EGA/g DPM and 20.5 mg EQ/g DPM). Both TP and TF content showed no signifi cant correlation with antioxidant activity, except in the case of TP and DPPH test where correlation was found (r=0.67). Despite diff erences in phenolics, fl owers of I. glandulifera exhibit the highest antioxidant activity in both tests, 3.8 and 3.8 mg EGA/g DPM, for DPPH and ABTS test respectively. In sum, fl owers seem to possess higher quantity of antioxidant compounds than do leaves in case of both species.

An adequate level of low molecular weight thiols (LMW-SH, especially glutathione (GSH)) protects cellular macromolecules against toxic agents, and is used as a sensitive biomarker of exposure to toxic compounds. During sample collection, storage and preparation, non-enzymatic and enzymatic oxidation of LMW-SH can occur leading to analytical inaccuracy. The aim of this study was to optimize a fast and reliable screening method for the determination of LMW-SH, mainly GSH, in blood and plasma samples as well as to investigate the impact of storage conditions on the LMW-SH stability. Based on our results, the described spectrophotometric method allows fast and reliable determination of LMW-SH in blood and plasma samples. Results on incubation of samples at 37 degrees C imply that synthesis of LMW-SH (probably GSH) as well as dynamic interexchange among various thiols forms can be induced in blood cells in in vitro conditions. Importantly, the level of LMW-SH in blood and plasma stored at -20 degrees C was constant, indicating that they can be stored at -20 degrees C for at least 30 days. Therefore, the method is suitable for assessment of LMW-SH in long-term human biomonitoring as well as environmental field studies, especially those involving a large number of samples such as epidemiological studies.

Chemistry is the art of creating and knowing molecules, what they do. Each day a lot of chemists around the world produce and invent new molecules. Fungi, plants, insects and bacteria produce thousands of chemical compounds and many of these compounds are bio-active, it means that they interact with living matter to cause disease or, even cure disease.