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Currently, there is a growing market demand for products derived from industrial hemp for food, recreational and medical use. This plant is widely known to be the major source of cannabinoids, including cannabidiol (CBD), cannabidiolic acid (CBDA), tetrahydrocannabinol (THC), cannabichromene (CBC), cannabigerol (CBG), and cannabinol (CBN). Cannabinoids are very significant group with strong remedial potential against inflammation, depression, nausea and epilepsy. On the other hand, THC is mostly responsible for psychotropic activities, so it is a controlled substance. Given that, cannabinoids are carriers of biological activity of industrial hemp, which is increasingly used in various forms (food, cosmetics or tinctures), it is important not just to develop a fast and reliable method of quantification of cannabinoids in hemp samples but to find the most suitable solvent and extraction conditions. The aim of this paper was to select the most suitable solvent and conditions among conventional (aqueous ethanol solutions (30-70% (v/v)), ethanol, methanol, acetonitrile) and hydrophobic deep eutectic solvents (hDESs), as well as the development of HPLC method for the detection of cannabinoids in prepared extracts. Depending on the conditions used and the solvent, different cannabinoids were extracted. The solvents show different ability to extract cannabinoids, so the conclusion about a universal solvent for all cannabinoids cannot be made. However, in terms of CBD, as the best known cannabinoid and its precursor, CBDA, the most suitable solvent is ethanol when extracted at 70 °C for 30 min. On the other hand, the most suitable hDES is a menthol:acetic acid (1: 1) for 60 min at 70 °C. Using ethanol as solvent 7.57 ug/mg of plant CBD and 100.25 ug/mg of plant CBDA was extracted, while by using menthol: acetic acid(1:1) 10.0 ug/mg of plant CBD and 128.67 ug/mg of plant CBDA was extracted proving that hDES can be a good substitute in the extraction of bioactive components from industrial hemp.

Due to the growing use of bioactive components in the food and in the pharmaceutical industry, rapid development of various methods for their extraction and separation is required. In the last few years, deep eutectic solvents (DESs), composed of a hydrogen bond acceptor (HBA) and hydrogen bond donor (HBD) components, stand out as an appropriate substitution to conventional solvents due to their properties, with emphasis on non- toxicity, thermal stability and biodegradability, with easy preparation and low price of components. Alchemilla vulgaris L. is a plant that is often used in folk medicine as it possesses numerous health-promoting benefits due to its specific chemical composition with emphasis on phenolic acids and elagitannins. The most important components with antioxidant activity in sage are phenolic acids, such as gallic and ellagic acid as well as elagitannins which can be extracted by various extraction techniques, most commonly solid-liquid extraction. The aim of this paper was to select the most suitable choline chloride based deep eutectic solvent for the extraction of gallic and ellagic acid as well as elagitannins from lady's mantle (Alchemilla vulgaris L.) and then optimize extraction conditions for extraction with stirring and heating, ultrasound-assisted extraction and mechanoextraction. Seventeen different choline chloride-based DESs with different volumes of water (10%, 30%, and 50% (v/v)) were used for the extraction at different temperatures (30, 50, and 70 °C) to investigate the influence on this extraction parameters on yield of desired components. The results obtained were compared with the results obtained by extraction with conventional solvents. Gallic and ellagic acid content was obtained by high- performance liquid chromatography with diode-array detector (HPLC- DAD) and total ellagitannins were determined spectrophotometrically in the obtained extracts. Screening revealed that among 17 different DESs, the most suitable solvent is choline chloride: urea (1:2) and the most suitable extraction technique is extraction with stirring and heating. According to the results, the prepared solvents prove to be a good substitute for organic solvents in the extraction of bioactive components from lady's mantle plant.

Cinnamon, a commonly used spice as well as a part of many commodities, such as breakfast cereals, teas and bakery products, represents an important part of the diet. Cinnamon, among many bioactive components, contains an important active compound coumarin. Numerous studies have shown that coumarin has a beneficial health effect on the body in optimal consumption, while increased long-term consumption can lead to adverse health effects. In this paper, the amount of coumarin in ten different products found on the Croatian market, including ground spices and teas, was analyzed. In addition, different extraction parameters were analyzed in order to find the best method of extraction of coumarin from cinnamon and products containing cinnamon.

Marine algae are known for their numerous benefits and pharmacological effect due to the presence of various bioactive compounds such as terpenoids, meroterpenoids, lipids, phlorotannins and phenolic compounds. Phlorotannins usually serve as a plant defense system against different adverse conditions and are often associated with different biological activities of macroalgae extracts, such as antifungal against Candida sp., antibacterial, anti-inflammatory, antioxidant, antitumor and neuroprotective. Phlorotannins possess numerous - OH groups in their structure, making them a highly hydrophilic molecules which can be extracted using polar deep eutectic solvents (DES).Fifteen different choline chloride based DESs were used for extraction of phlorotannins from Padina pavonica and Cytoseira compressa. Phlorotanin content and antioxidant activity (DPPH method) were determined spectrophotometrically in obtained extracts. Phlorotannines content is much higher in most of the C. compressa extracts compared to P. pavonica. For C. compressa, choline chloride:lactic acid (1:2) showed the best results, considering both phlorotannin yield and DPPH scavenging activity. In the case of P. pavonica extracts, choline chloride:urea (1:2) was the most efficient for the extraction of phlorotannins and the best DPPH scavenging activity was obtained with choline chloride:thiourea (1:2).

Extraction of rutin from Ruta graveolens L. leaves was performed in choline chloride based deep eutectic solvents (DESs). Firstly, rutin extraction was performed in five different DESs ; pure DESs and with different amounts of water added (25% and 50% v/v), for different times (1h, 3h and 6h) in order to find the best DES for further optimization. Choline chloride plus citric acid (1:1) was shown to be best DES and optimization of rutin extraction was performed using response surface methodology (RSM) based on the selected solvent and applying different extraction parameters: extraction time, temperature and water content. The rutin content was determined by HPLC and varied from 0.55 g/100 g of plant to 1.81 g/100 g of plant depending on the applied conditions. The best conditions for rutin extraction were found to be: a temperature of 70°C, an extraction time of 52 min and a water content of 27% for the extraction of 1.88g /100 g plant.

The genus Helichrysum consists of 600 species, among which the most significant are dwarf everlast (Helichrysum arenarium) and curry plant (Helichrysum italicum). The H. arenarium spreads from the Mediterranean to Northern Europe, while H. italicum grows exclusively in the Mediterranean countries. Only H. italicum is cultivated for the production of essential oil. As a perennial plant, H. italicum is cultivated on the same location for 5-8 years. Therefore, a lot of care should be taken when choosing the plot because this plant thrives on sandy and rocky soil with a lot of sunshine. Alkaline soil (with a lot of calcium) that retains water well is the best type of soil for this plant. H. italicum can reproduce generative, by seed through seedlings, and vegetative, by cuttings.

Cultivation of medicinal and aromatic plants has increased in recent years and more and more arable land in Croatia is designed specifically for this type of farming. Medicinal and aromatic plants are grown primarily because they contain biologically active components that make up the basic raw material for the production of drugs, cosmetics and flavors for foodstuff. Chamomile (Matricaria chamomilla) has been known as a medicinal plant for a long time, mainly due to its anti-inflammatory, antiseptic and antispasmodic activity. For this reason, chamomile is used as the starting material for consideration in this paper. Among many biologically active compounds found in chamomile, a coumarin derivative, umbelliferone, was also found. The aim of this study was to compare chamomile extracts, produced with various extraction techniques (soxhlet, hydrodestillation, macaretion, supercritical CO2 extraction, high pressure maceration system) based on their antioxidant activity and umbelliferone content determined by high performance liquid chromatography (HPLC). The following samples of chamomile were analyzed: unprocessed chamomile flowers first class, processed chamomile flowers first class, pulvis and processing waste obtained from the company Matricia Ltd. The greatest extraction yield (22.3%) was obtained with 50% ethanol in high pressure maceration system and the highest yield of umbelliferone (41.03 mg/100g) was obtained using high pressure maceration system. Extracts have also proven to possess antioxidant activity (45.4 - 73.7% DPPH scavenging activity). The significance of this study offers a possibility of potential utilization of waste from chamomile processing, applying different extraction techniques and attaining the final natural product by using supercritical CO2 extraction without any residues of organic solvents and with potential use in the cosmetic industry. Furthermore, a new developed system of maceration under pressure significantly reduces the time of extraction providing extracts with higher antioxidant activity.

Importance of pharmacologically active natural compounds from plant sources has been re- evaluated in recent years and it has become one of the most active research fields. Despite great advances in medicine, plants still play important roles in healthcare due to their useful natural components such as antioxidants. The main purpose of this review was to present updated information about chemical composition of Vitex agnus-castus (VAC) and its role in healthcare. According to several studies, chemical composition of VAC is characterized by presence of different secondary metabolites, among which iridoids, flavonoids, diterpenoids, essential oils (EO) and ketosteroids are dominant. Differences in chemical composition are observed as a result of different geographic origin and applied extraction method. In this review, the chemical composition of EO and VAC extracts obtained by different extraction methods, including conventional and non-conventional methods, are given and discussed.

Importance of biologically active natural compounds from plant sources has become one of the most active research in the recent years. Biologically active compounds are often present in the plants in low concentrations, they are chemically sensitive and make up the basic raw material for the production of drugs, cosmetics and flavors for foodstuff. Seven medicinal plants, namely Helichrysum italicum (Roth) G. Don, Angelica archangelica L., Matricaria chamomilla L., Lavandula officinalis L., Salvia officinalis L., Melilotus officinalis L. and Ruta graveolens L. were used in this study. The aim of this study was to compare obtained extracts of medicinal plants, produced with various extraction techniques (soxhlet, macaretion, supercritical CO2 extraction) based on their antioxidant activity and coumarin content determined by high performance liquid chromatography (HPLC). Melilotus officinalis ethanol extract was found to contain the highest yield of coumarin, while Salvia officinalis and Helichrysum italicum ethanol extracts showed the highest antioxidant activity. Generally, ethanol extracts showed the best antioxidant activity and in the most cases the highest yield of extracted coumarin

Cilj ovog rada bilo je odrediti koja metoda i uvjeti ekstrakcije su najbolji za ekstrakciju bioaktivnih komponenti prisutnih u vrkuti ili gospinom plaštu (Alchemilla vulgaris) pomoću niskotemperaturnih eutektičkih otapala. Vrkuta je višegodišnja zeljasta biljka koja se koristi za liječenje kožnih bolesti, probavnih problema i slično. Bogata je bioaktivnim komponentama zbog čega ima antivirusno, antibakterijsko, antioksidativno i antiproliferativno djelovanje. Niskotemperaturna eutektička otapala se koriste kao nova održiva, sigurnija i zelenija alternativa komercijalnim otapalima, te pokazuju različitu funkcionalnost i topljivost za komponente koje se nalaze u biljci. Ekstrakcija niskotemperaturnim eutektičkim otapalima je provedena miješanjem i na ultrazvučnoj (UZV) kupelji. Bioaktivne komponente s naglaskom na hidrolizirane tanine, flavonoide i fenolne kiseline, identificirane su spektrofotometrijski odnosno tekućinskom kromatografijom visoke djelotvornosti (HPLC). Nakon identifikacije i kvantifikacije može se zaključiti da je ekstrakcija miješanjem na magnetskoj miješalici učinkovitija u odnosu na ekstrakciju potpomognutu ultrazvukom, te da prilikom ekstrakcije na visokoj temperaturi i kroz duže vrijeme dolazi do degradacije komponenti i smanjenog prinosa. The aim of this work was to determine which extraction method and conditions are best for the extraction of bioactive components present in lady's mantle (Alchemilla vulgarisI) using deep eutectic solvents. Lady’s mantle is a perennial herb that is used to treat skin diseases, digestive problems and similar conditions. It is rich in bioactive components, possessing antiviral, antibacterial, antioxidant and antiproliferative effects. Deep eutectic solvents are used as a new, sustainable, safer and greener alternative to commercial solvents and they show different functionality and solubility for components found in the plant. Extraction with deep eutectic solvents was performed using by stirring on magnetic stirrer and in ultrasonic (UZV) bath. Bioactive components, with an emphasis on hydrolyzed tannins, flavonoids and phenolic acids, were identified spectrophotometricaly or by high performance liquid chromatography (HPLC). After identification and quantification, it can be concluded that the extraction using a magnetic stirrer is more efficient compared to the ultrasound-assisted extraction and that high temperature and long extraction time cause degradation of components and a yield decrease.

Uzgoj ljekovitog i aromatičnog bilja posljednjih godina poprima sve veće razmjere i sve je više obradivih površina u Hrvatskoj namijenjenih upravo za ovu vrstu uzgoja. Kamilica (Matricaria chamomilla) je od davnina u narodu poznata kao ljekovita biljka, uglavnom zbog svog protuupalnog, antiseptičkog te spazmolitičkog djelovanja, te sadrži mnoštvo različitih biološki aktivnih tvari. Cilj rada bio je usporediti ekstrakte kamilice dobivene različitim tehnikama ekstrakcije na osnovu njihove antioksidacijske aktivnosti te sadržaja umbeliferona određenih primjenom tekućinske kromatografije visoke djelotvornosti (HPLC). U tu svrhu analizirani su sljedeći uzorci kamilice dobiveni tijekom različitih faza prerade: nedorađeni cvijet kamilice I. klase, dorađeni cvijet kamilice I. klase, pulvis i otpad od dorade. Uz tradicionalne tehnike ekstrakcije (hidrodestilacija, soxhlet ekstrakcija, maceracija) u radu je korištena i ekstrakcija pomoću superkritičnog CO2 (SC-CO2), te je razvijen i novi sustav maceracije pod visokim tlakom. Najveći prinos ekstrakta (26,52 g/100 g) i najveći udio umbeliferona (44,82 g/100 g) dobiven je maceracijom pod visokim tlakom. Antioksidacijska aktivnost u dobivenim uzorcima kretala se između 45,4 - 73,7% DPPH. Značaj istraživanja očituje se u pružanju mogućnosti korištenja otpada iz procesa prerade kamilice, primjene različitih tehnika ekstrakcije te u mogućnosti dobivanja krajnjeg proizvoda pomoću ekološki prihvatljive SC-CO2 ekstrakcije. Nadalje, novo razvijeni sustav maceracije pod tlakom značajno smanjuje vrijeme ekstrakcije uz dobivanje ekstrakata s većim udjelom komponenata koje doprinose antioksidacijskoj aktivnosti. Cultivation of medicinal and aromatic plants is on the increase in recent years and more and more arable land in Croatia is designed specifically for this type of farming. Chamomile (Matricaria chamomilla) has been known as a medicinal plant for a long time, mainly due to its anti-inflammatory, antiseptic and antispasmodic action. The aim of this study was to compare chamomile extracts produced with various techniques of extraction based on their antioxidant activity and umbelliferone content applying high performance liquid chromatography (HPLC). The following samples of chamomile were analyzed: unprocessed chamomile flowers first class, processed chamomile flowers first class, pulvis and processing waste. In addition to traditional extraction techniques (hydrodistillation, Soxhlet extraction, maceration), supercritical CO2 extraction (SC-CO2) was performed and a new system of maceration under high pressure has been developed. The greatest extraction yield (26.52 g/100 g) and the highest yield of umbelliferone (44.82 mg/100 g) were obtained using high pressure maceration system. Extracts have also proven to possess antioxidant activity (45.4 – 73.7% DPPH scavenging activity). The significance of this study is reflected in enabling the use of waste from processing chamomile, applying different extraction techniques and attaining products by using environmentally friendly SC-CO2 extraction. Furthermore, a newly developed system of maceration under pressure significantly reduces the time of extraction thereby obtaining extracts that are high in components that contribute to the antioxidant activity.

Svaka industrija pa tako i prehrambena ima cilj proizvesti što kvalitetniji proizvod uz što bolje iskorištenje sirovine tj. uz minimalnu količinu otpada, gdje otpad ne mora nužno biti ''otpad'' već može predstavljati nusproizvod ili sirovinu u nekom budućem procesu. Danas postoji velik interes u iskorištenju nusproizvoda prehrambene industrije u različite svrhe jer sadrže mnoge potencijalno korisne tvari i zbog toga bi mogli predstavljati značajne sirovine u proizvodnji/razvoju novih proizvoda. U ovom radu proizvedeni su ekstrakti različitih nusproizvoda prehrambene industrije biljnoga podrijetla: unutarnje pregrade oraha, kakao ljuska, pljevica kave, sjemenke grožđa Teran i Merlot, kožica grožđa, kora nara, koštice lubenice, koštice i peteljke trešnje, te sjemenke marelice, te je određeno njihovo antioksidacijsko i antibakterijsko djelovanje. Antioksidacijska aktivnost ekstrakata određena je DPPH metodom. Antibakterijska aktivnost provedena je na četiri soja test bakterija: Bacillus subtilis, Staphylococcus aureus, Pseudomonas aeruginosa i Escherichia coli, te je određena s obzirom na minimalne koncentracije inhibitora modificiranom metodom mikrodilucije supstrata. Najveću antioksidacijsku aktivnost pokazao je ekstrakt unutarnjih pregrada oraha (18,80 mg/mL), a najveću antibakterijsku aktivnost ekstrakti unutarnjih pregrada oraha i pljevica kave (0,0156 mg/mL - E. coli; 0,0156 mg/L - P. aeruginosa; 0,0078 mg/mL - B. subtilis; 0,0625 mg/mL - S. aureus). Nadalje, u radu je ispitan i utjecaj metode ekstrakcije (hladno prešanje i superkritična \(CO_{2}\) ekstrakcija) na izolaciju ulja iz sjemenki marelice i sjemenki grožđa. U sjemenkama marelice naglasak je dan na udio amigdalina, a u sjemenkama grožđa na udjele polifenola koji su određeni primjenom tekućinske kromatografije visoke djelotvornosti (HPLC). Sastav masnih kiselina u dobivenim uljima je određen metodom plinske kromatografije s plamenim detektorom GC/FID. Ovaj rad doprinosi rješenju stvaranja velike količine organskog otpada koji danas predstavlja ogromni ekološki i financijski teret u gotovo svim granama prehrambene industrije. Every industry, including food industry has the goal of obtaining the most quality product with the best possible use of raw materials, with the minimum amount of waste, where waste does not necessarily have to be "waste" but may represent a by-product or raw material in some future process. Nowadays, a tendency for reducing food industry waste is becoming more prominent due to a growing interest for utilization of food industry by-products for various purposes. These by-products contain many potentially useful substances and they could become significant raw materials in the production/development of new products. In this research, extracts of various food industry by-products of the plant origin have been produced: walnut partitions, cocoa bean shells, coffee parchment, Teran and Merlot grape seeds, grape skin, cherry seeds, watermelon seeds, apricot kernels, and their antioxidative and antibacterial activity is determined. Antioxidant activity of extracts was determined by DPPH method. Antibacterial activity was conducted against four test bacteria strains: Bacillus subtilis, Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli in terms of minimum inhibitory concentrations by a modified broth microdilution method. The highest antioxidant activity was determined in the extract of the walnut partitions (18.80 mg/mL), and the extracts of walnut partitions and coffee parchment (0.0156 mg/mL - E. coli, 0.0156 mg/mL - P. aeruginosa, 0.0078 mg/mL - B. subtilis, 0.0625 mg/mL - S. aureus) showed the highest antibacterial activity. Furthermore, the effect of the extraction method (cold pressing and supercritical \(CO_{2}\) extraction) on the isolation of apricot and grape seed oil was tested. In apricot seeds, emphasis is given to the content of amygdalin, and in grape seeds on polyphenols that are determined by the high performance liquid chromatography (HPLC). The fatty acid composition in the obtained oils was determined by the gas chromatographic method with the flame detector GC/FID. This paper offers a potential solution against generation of large amounts of organic waste, which represents a huge ecological and financial burden in almost all branches of food industry.

Ekstrakcija pomoću superkritičnog CO2 kao nova zelena tehnologija razmatra se intenzivno posljednjih godina jer se CO2 pokazao kao vrlo poželjno otapalo u separacijskim procesima budući da je neotrovan, nezapaljiv, bez okusa i mirisa, jeftin i lako dostupan u većim količinama, kao i zbog toga što je ekološki prihvatljivo otapalo. Stoga su ciljevi ovoga rada sljedeći: (1) proizvodnja ekstrakata iz ploda konopljike (Vitex agnus castus L.) pomoću superkritičnog CO2 pri različitim procesnim parametrima (tlak, temperatura, protok otapala, veličina čestica), (2) dobivanje eteričnog ulja konopljike postupkom vodene destilacije, (3) ispitivanje kemijskog sastava dobivenih ekstrakata primjenom plinske kromatografije, (4) proizvodnja suhih ekstrakata postupkom liofilizacije i formuliranje tableta. Promjena uvjeta ekstrakcije značajno utječe na prinos ekstrakta. Najveći prinos ekstrakta 12,27% ostvaren je pri tlaku od 225 bar, temperaturi od 60°C, protoku CO2 od 1,95 kg/h i veličini čestica 0,393 mm. Utvrđeno je da ekstrakt ploda konopljike ima veoma kompleksan kemijski sastav. Sastav eteričnog ulja se bitno razlikuje od sastava ekstrakta dobivenog superkritičnim CO2, posebice u količini pojedinih spojeva poput viridiflorola, koji je glavni sastojak eteričnog ulja i α-humulena, koji također dominira u eteričnom ulju, dok je u superkritičnom ekstraktu zastupljen u znatno manjoj količini. Pronađena je i potencijalna primjena dobivenog ekstrakta u farmaceutske svrhe budući da sadrži brojne bioaktivne komponente. Supercritical CO2 extraction as a new green technology is discussed extensively in recent years and it has proven that CO2 is highly desirable solvent in the separation process since it is non-toxic, non-flammable, odorless, tasteless, it is cheap and readily available in large quantities, thus being an environmentally friendly solvent. The aims of this work were as follows: (1) production of extracts from Vitex agnus-castus fruit by supercritical CO2 extraction, under different process parameters (pressure, temperature, solvent flow rate, particle size), (2) production of essential oil of Vitex agnus-castus fruit by hydrodistillation, (3) investigation of chemical composition of obtained extracts applying gas-chromatography, (4) production of dry extracts using freeze drying and formulation of tablets. Extraction conditions change significantly affects the extraction yield. The highest extraction yield of 12.27% was achieved at pressure of 225 bar, temperature of 60 °C, CO2 flow rate of 1.95 kg /h and particle size of 0.393 mm. It was found that an extract of the fruit of Vitex agnus castus L. has a very complex chemical composition. Obvious differences were found between the composition of essential oil and extracts obtained by supercritical CO2 in the amount of certain compounds such as viridiflorol, which is the main component of essential oil and α-humulene, which also dominates in essential oil, while in supercritical extract it is present in much lower quantities. Resulting extracts have found potential application as a pharmaceuticals since they contain a number of bioactive components.