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Lovor (Laurus nobilis L.) je zimzeleni mediteranski grm čije se lišće stoljećima koristilo u narodnoj medicini zbog svojih blagotvornih učinaka na zdravlje koji se uvelike mogu pripisati sadržaju polifenola. Budući da su polifenoli nestabilni i skloni gubljenju svojih aktivnih svojstava tijekom skladištenja, od velike je važnosti poboljšati njihovu stabilnost kako bi bili primjenjivi u industriji. To se može postići različitim metodama inkapsulacije među kojima je najčešće korištena metoda sušenja raspršivanjem (SR) u kojoj se tekući ekstrakt s otopljenim nosačem propušta kroz struju vrućeg zraka prilikom čega dolazi do isparavanja otapala te nastaje prah s bioaktivnim molekulama inkapsuliranim u zaštitni omotač nosača. Elektrostatska ekstruzija (EE) je metoda u kojoj se pomoću elektrostatske sile ekstrakt s otopljenim nosačem propušta kroz mlaznicu te u kontaktu sa želirajućom otopinom nastaju uniformne kuglice gela s inkapsuliranim bioaktivnim spojevima. U ovom istraživanju, 50% etanolni ekstrakt lovorovog lista inkapsuliran je primjenom SR pri prethodno definiranim optimalnim uvjetima (120°C ; mješavina β- ciklodekstrin:maltodekstrin (1:1) ; omjer uzorak: nosač 1:2), a vodeni ekstrakt je inkapsuliran primjenom EE također pri prethodno definiranim optimalnim uvjetima (1, 0% otopina alginata u ekstraktu ; 1, 5% CaCl2 uz 0, 5% kitozana kao želirajuća otopina). Dobiveni prah je otopljen u 80% metanolu, a gel-kuglice u 5% otopini natrijeva citrata te su zajedno s inicijalnim ekstraktima analizirani primjenom tekućinske kromatografije ultra-visoke djelotvornosti s masenom spektrometrijom (UPLC/MS-MS). Učinkovitost inkapsulacije ukupnih polifenola kod SR iznosila je 88, 32%, a kod EE 68, 46%. Kod SR najuspješnije inkapsulirana skupina bili su flavan-3-oli (95, 67%), a kod EE flavoni (90, 82%). Najslabije inkapsulirana skupina kod SR bile su fenolne kiseline (76, 10%), a kod EE proantocijanidini (6, 38%). Rezultati su pokazali prednost primjene SR nad EE u cilju postizanja veće učinkovitosti inkapsulacije, a još jedna prednost se nalazi u mogućnosti primjene vodenih otopina etanola za ekstrakciju polifenola čime se kombiniraju maksimalna učinkovitost ekstrakcije i inkapsulacije, dok se EE primjenjuje samo na vodenim ekstraktima. Unatoč tome, obje metode pokazale su relativno visoke postotke inkapsulacije te će budući odabir metode ovisiti prije svega o konačnoj namjeni kapsuliranih sustava.

Medicinal plant extracts can serve as the functional constituents for the dairy products enrichment due to the presence of bioactive compounds. Except beneficial effect on human health, plant extracts also have antimicrobial activity which can extend shelf life of foods. Plant extracts can be dried and added to milk raw materials in the powder form and the choice of extraction technique and conditions significantly affect the yield of target groups of bioactive molecules in powder production. In this study the yields of total phenolics (TP), flavonoids (TF), hydroxycinnamate acids (THCA), flavonols (TFL) and pigments (TPIG) of carob pod (Ceratonia siliqua L.), bay (Laurus nobilis L.) and sage (Salvia officinalis L.) leaves were evaluated using different extraction techniques and solvents, respectively. The selected plants were successive extracted with hexane, 80%-acetone and 96%-ethanol in shaking water bath (SWB) and under ultrasound-assisted (UAE) and accelerated solvent extraction (ASE). Mass fraction of TP, TF, THCA, TFL, TPIG were determined by spectrophotometric methods. The highest TP, TF and THCA content was determined in sage extracts obtained by UAE and the highest TFL content by SWB technique. Laurel extracts had the highest TPIG content using a SWB and ASE technique. In general, sage was richer source of phenolics and laurel of pigments while UAE technique was better method for phenolic and ASE for pigments extraction.

Multifunctional packaging systems have a high potential for commercial packaging applications. By applying the layer of the biopolymer coating on the polymer surface, it is possible to change its barrier properties and can serve as a transporter to deliver active substances having a special function to packed food. Life style and market demands pushes food industry towards the development of new materials that consequently leads to the increased product shelf life while minimising the impact on the nature and climate. The main aim of this study was to investigate the possibility of applying naturally friendly chitosan coating on the surface of commercially available oriented poly(ethylene- terephthalate)/polypropylene (OPET/PP). This bi- layer material is used as the cover film for PP buckets ; in which bakery fruit fillings are stored. Moreover, orange peel essential oil was used as the active compound that was nanoencapsulated in the coating. The essential oil was prepared by microwave assisted extraction coupled with ETHOS. This method is environmentally friendly providing green production concept by minimising the use of naturally unfriendly solvents. Also, by reusing the orange peel as a food waste, this study contributes to a sustainable material production. Produced materials were tested for their gas and water vapour barrier performance, physico-chemical properties (thickness, UV transmission and colour), and antimicrobial properties. The results show the possibility of improving the commercial material properties. It is possible to couple the traditional with green extraction technology to reuse natural product considered as a food waste for food packaging and thus minimise the negative impact on the nature. In order to better check the functional character, further investigation is needed on real food products during their storage and impact on their shelf-life.

In recent times, increasing evidence has demonstrated that polyphenols, secondary plant metabolites, can be utilized by the gut microbiota which results in promising effects in the prevention and treatment of various diseases such as diabetes, obesity, autoimmune diseases, inflammatory bowel disease, colorectal cancer and other. Laurus nobilis L. is an evergreen Mediterranean shrub whose leaves have traditionally been used in folk medicine due to their beneficial health effects that can largely be attributed to the presence of polyphenols which include flavonoids, phenolic acids, tannins and lignans. Since polyphenols are unstable which limits their biological activities and aggravates the process of studying their association with gut microbiota, encapsulation technologies focused on protection against unfavorable conditions in the stomach are emerging. Microencapsulation by spray- drying is the most commonly used method for encapsulating bioactive molecules in which the liquid extract with the dissolved carrier is passed through a stream of hot air during which the solvent evaporates and a powder with bioactive molecules encapsulated in the protective coating of the carrier is formed. In the present study, hydroalcoholic (70% ethanol) extract of L. nobilis leaf polyphenols was encapsulated by spray-drying on Büchi Mini Spray-Dryer B-290 at 180°C using three different carriers (β-cyclodextrin ; β- cyclodextrin:maltodextrin (50:50) ; β- cyclodextrin:gum arabic (50:50)) at a ratio 1:2 (extract dry matter:carrier). The obtained powders were subjected to in-vitro simulated digestion and three phases (gastric, absorbed and intestinal) were analyzed by Folin-Ciocalteu spectrophotometric method for total phenolic content (TPC). The TPC in the intestinal phase, which is considered as bioavailable for the gut microbiota, ranged from 46.9–54.5% compared to the TPC in initial powders prior to digestion. β- cyclodextrin:maltodextrin (50:50) resulted in highest, and β-cyclodextrin:gum arabic in lowest bioavailability. The integrity of individual polyphenols present in the intestinal phase was confirmed by ultra-high performace liquid chromatography tandem mass spectrometry (UPLC/MS- MS).

Medicinal and aromatic plants are a rich source of structurally diverse bioactive molecules (BAM) (polyphenols, dietary fiber, sterols, pigments, polysaccharides, vitamins, minerals, etc.) which possess various biological activities, and have a great potential for the application in food, pharmaceutical and cosmetic industry. Extraction is one of the most important steps in BAM isolation and can be affected by the applied extraction techniques, conventional or advanced with lower organic solvent consumption (microwave assisted, ultrasound assisted, pressurized liquid extraction, etc.) as well as various extraction conditions (pH, time, temperature, pressure, particle size, solvent, sample/solvent ratio, microwave power, etc.). Furthermore, for the dairy industry and production of enriched functional dairy products, liquid plant extracts are often processed into a more stable powder form by using appropriate encapsulation techniques. Therefore, this paper presents a systematic and comprehensive review of techniques used in the production of liquid plant extracts and powders, which have a great perspective for the application in the production of functional and/or enriched dairy products. Besides selection of the efficient extraction techniques and optimal extraction conditions, it is also important to conduct the economic feasibility study to determine which technique is the best choice for sustainable plant extracts and powder production from technical and economic aspects.

Both nettle and fennel are valuable plants with high content of bioactive molecules. Beneficial effects of nettle are mainly attributed to polyphenols, while fennel is recognized for its essential oil (EO) content. Although chemically different, both nettle extract and fennel EO are characterized with loss of stability and bioactivity during prolonged storage and processing. Spray drying, as a method for encapsulation of both hydrophilic and lipophilic compounds, can provide stabilization, preservation as well as controlled release and delivery of both nettle bioactives and fennel EO. Therefore, the aim of this study was to evaluate and optimize the spray drying encapsulation of nettle leaves extract and fennel EO using different carrier materials and drying temperatures. Nettle leaves extract was obtained by microwave-assisted extraction and encapsulated using maltodextrin (MD) and β-cyclodextrin (CD) as single carriers or in mixtures with gum arabic (GA) in proportion 1:1 and 3:1, respectively. Carriers were used in sample:carrier ratios 1:1, 1:2 and 1:3, while spray drying was performed at 120, 160 and 200 °C. Fennel EO was spray dried at same temperatures and emulsified with following carrier mixtures, MD:CD, MD:GA and CD:GA in proportions 1:1, 1:3 and 3:1 at fixed EO:carrier ratio of 1:3. The highest yield and encapsulation capacity for nettle leaves extract was achieved with addition of MD as a carrier agent in ratio 1:3 at 120 °C, while optimal encapsulation conditions for fennel EO were mixture of CD:GA in proportion 3:1 at 200 °C which resulted in the highest EO retention in microcapsules and the highest encapsulation efficiency. The differences in obtained encapsulation conditions arise from the differences in chemical properties of encapsulated materials, showing that carrier agents such as CD and GA are suitable for non-polar constituents of EO, while MD is more efficient in encapsulation of polar compounds such as polyphenols. The encapsulation of nettle extract and fennel EO provided preservation and stabilization of respective bioactive compounds and increased their bioavailability in encapsulated form, indicating the great potential of such microcapsules for further application in food industry.

Thyme (Thymus vulgaris L.) is a Mediterranean herb belonging to the Lamiaceae family well known in the folk medicine for its various health beneficial properties. These properties can be attributed to the high content of biologically active molecules, such as polyphenols. In recent years, the growing demand on functional products enriched with naturally derived polyphenols has brought the need for more sustainable isolation technologies that would allow for their fast and effective isolation. Microwave-assisted extraction has emerged as a green extraction technology due to its main advantage of reducing the extraction time and solvent consumption while achieving higher extraction efficiency and lower degradation of targeted compounds compared to conventional extraction techniques. In the present study, the influence of different microwave-assisted extraction parameters on the isolation efficiency of Thymus vulgaris L. leaf polyphenols was examined in two phases. In the first phase, different percentages of aqueous ethanol (0, 30, 50, 70 and 96%) and sample:solvent ratios (1:10, 1:20 and 1:30) were varied while other extraction parameters (temperature 60°C ; microwave power 800W ; irradiation time 10 min) were kept constant. The total phenolic content (TPC) of the extracts was determined by Folin-Ciocalteu spectrophotometric method. Both ethanol percentage and sample:solvent ratio have shown significant influence on the TPC of the Thymus vulgaris L. leaf extracts, and the highest TPC (10.59 mg GAE/g leaf) was achieved when 30% ethanol and a ratio of 1:20 were applied. In the second research phase, the ethanol percentage and sample:solvent ratio were kept constant (30% ethanol and 1:20), while the temperature (40, 60 and 80°C) and irradiation time (5, 10 and 15 min) were varied. The results of Folin-Ciocalteu analysis showed that temperature had a significant influence on the TPC of the Thymus vulgaris L. leaf extracts, while the influence of irradiation time was not significant. The highest TPC (11.99 mg GAE/g leaf) was achieved at 60°C after 5 min of irradiation. The results showed how different parameters of microwave-assisted extraction influence the TPC of Thymus vulgaris L. extracts, proving the importance of establishing optimal isolation methodologies that would result in the highest recovery effectiveness and utilization of the plants’ beneficial properties.

Komorač (Foeniculum vulgare Mill.) je višegodišnja, ljekovita i aromatična biljka koja pripada obitelji štitarki (lat. Apiaceae). Od davnina se koristi u narodnoj medicini i kulinarstvu, a danas se sve više koristi u farmaceutskoj, kozmetičkoj i prehrambenoj industriji. Sjemenke komorača obiluju eteričnim uljem (5-6%) koje karakterizira bogata aroma te je značajno zbog svojih antimikrobnih i antioksidativnih svojstava [1]. Međutim, visoka hlapljivost eteričnog ulja i podložnost oksidaciji u prisutnosti svjetlosti, zraka i vlage te nestabilnost tijekom prerade ograničava njegovu upotrebu u industriji. Slijedom navedenog, inkapsulacija sušenjem raspršivanjem predstavlja pogodnu metodu za produljenje stabilnosti eteričnih ulja i zadržavanje njihovih funkcionalnih svojstava tijekom manipulacije, obrade i skladištenja. Stoga je cilj ovog rada bio ispitati utjecaj omjera dvokomponentne kombinacije nosača maltodekstrina i arapske gume (1:1, 1:3 i 3:1) te temperature sušenja (120, 160 i 200 °C) na iskorištenje procesa, zadržavanje ulja i učinkovitost inkapsulacije. Porastom udjela arapske gume dolazi do povećanja vrijednosti sva tri promatrana parametra, dok porast temperature do 160 °C dovodi do povećanja zadržavanja i učinkovitosti. Daljnjim porastom temperature dolazi do povećanja iskorištenja. Optimalni uvjeti za učinkovitu inkapsulaciju eteričnog ulja iz sjemenki komorača su omjer maltodekstrina i arapske gume 1:3 te temperatura sušenja 160 °C.

Due to the rich source of biologically active molecules (BAM), extracts from algae and plants are an excellent potential for the production of functional foods with added value. Despite the fact that today there are many researches on different plant species, their application in the production of functional products is not sufficient. Studies have shown that the brown alga Halopteris scoparia is a source of various BAM's such as amino acids, polysaccharides, vitamins, sterols, minerals, dietary fibers, fatty acids and pigments (El-Din & Alagawany, 2019., Mišurcová et al., 2014), which have enormous functional and nutraceutical potential, while on the other hand, the analysis of mastic tree (Pistacia lentiscus L.) leaves, stems, fruits and roots revealed significantly higher concentration of the main groups of secondary metabolites (flavonoids, phenolic acids and tannins), indicating that the plant has great biological potential (Dragović et al., 2020). Due to the different composition of BAM, these two remarkable natural sources show a greater synergistic effect in the production of functional products by mixing the extracts. Given the particular sensitivity to degradation of BAM by temperature, pH, enzymes, and oxygen, encapsulation has proven to be an effective way to protect these compounds, especially in the production of functional foods. Therefore, the aim of this work was to optimize the spray-drying conditions of mixed extracts from the algae H. scoparia and P. lentiscus using two different carriers (maltodextrin DE 13-17 and gum arabica) on the phenolic and pigment content and physical properties of the obtained powder at spray-drying temperature of 120, 150 and 180 °C and wall-to- core ratio of 1:20. The powders prepared with MD had lower product yield but better other physical properties and retention of BAC. The obtained results showed a strong synergistic effect of phenols from Mediterranean plants and algal pigments and their encapsulation by spray drying, which can be an excellent basis for further research and production of new value-added functional foods.

The growing demand on naturally derived ingredients has brought the need for finding new sources of biologically active molecules which can be isolated and used as functional ingredients in food. Bay leaf is a Mediterranean shrub well known in the folk medicine for its many health beneficial effects that can be largely attributed to the content of polyphenols. In order to enable efficient industrial utilization of these valuable compounds, it is necessary to isolate them from plant material. Various green extraction technologies, such as pressurized liquid extraction (PLE), have emerged in recent years due to many advantages over conventional techniques including the shorter extraction time, lower energy and solvent consumption as well as higher extraction efficiency and lower degradation of targeted compounds. Combined chromatographic and spectral techniques, such as ultra-performance liquid chromatography tandem mass spectrometry (UPLC/MS-MS) allow effective polyphenolic characterization of the obtained extracts, enabling the optimal choice of the extraction methodology as well as plant material whose chemical composition may vary significantly depending on the environmental factors, harvesting season and storage conditions. The aim of this research was to compare the polyphenolic content of bay leaves collected in November 2021 in the region of Rijeka, Croatia and Dubrovnik, Croatia and stored in the same conditions. The extracts were obtained from finely grinded dry leaf samples by PLE at previously established optimal conditions (50% ethanol, 150°C, 1 extraction cycle and 5 min static time) and analyzed by UPLC/MS-MS. The results showed that the sample collected in the region of Dubrovnik, Croatia had a higher content of total polyphenols (45, 09 202 ± 0, 21 mg g-1) than the sample collected in the region of Rijeka, Croatia (36, 12 ± 0, 17 mg g1). Flavonols were the most abundant group of compounds in both samples, followed by flavan-3-ols, phenolic acids, proanthocyanidins and flavones. Significant differences were observed in the content of individual polyphenols, showing that environmental growth factors have an important effect on the polyphenolic profile of bay leaf and should be further researched in order to produce extracts of maximum quality, while using less plant material, resulting in a more sustainable process.

Essential oils are volatile mixtures of compounds with characteristic flavor obtained from plant material. One of the plants rich in essential oil is fennel (Foeniculum vulgare Mill.). It is a perennial plant with a pleasant and aromatic odor, whose seeds are rich in essential oil (5-6%). Since the essential oil of fennel seeds has antimicrobial and antioxidant properties, nowadays there is a growing interest in its use in the food and pharmaceutical industries. However, its high volatility and susceptibility to oxidation in the presence of moisture, light and oxygen limit its use. Thus, encapsulation represents a suitable method to prolong the stability of essential oils and preserve their functional properties. One of the most commonly used encapsulation methods is spray drying, in which the suspension is dried in a stream of hot air with the addition of a suitable carrier to form stable powder in which flavor and aroma are preserved. Therefore, the aim of this research was to investigate the impact of temperature (120, 160 and 200 °C) and ratio of maltodextrin and β-cyclodextrin carriers (1:1, 3:1 and 1:3) on process yield, oil retention and encapsulation efficiency, and to determine the optimal conditions for producing powders with the best properties. The spray drying yield of fennel essential oil powder ranged from 60.89- 74.51%, oil retention ranged from 27.40-57.66%, and encapsulation efficiency ranged from 18.99- 42.96%. An increase in temperature increased the values of all three observed parameters, and an increase in maltodextrin content led to an increase in oil retention and encapsulation efficiency. However, the yield was highest at a higher β-cyclodextrin content. The optimal conditions for the production of powders with the best properties were a drying temperature of 200°C and a ratio of maltodextrin and β-cyclodextrin of 3:1. The application of the spray drying process for fennel essential oil encapsulation proved to be effective in producing powders with desirable properties.

Sea buckthorn (Eleagnus Rhamnoides (L.) A. Nelson) berry oil (SBO) as a rich source of bioactive compounds (carotenoids, tocopherols, omega-3, 6, 7, and 9 fatty acids) is susceptible to oxidative deterioration and unstable under processing and storage conditions, limiting its use in the food industry. Encapsulation by spray drying (SD) is a reliable technique widely used to overcome the above-mentioned problems. However, to achieve high encapsulation efficiency (EE) and the desired powder quality, the SD parameters such as drying temperatures, carriers and carrier-oil ratios should be optimized. Successful encapsulation of oils should result in an encapsulated powder with minimum surface oil content and maximum retention of oil in the powder particles (Jafari et al., 2008). Gum arabic is the most commonly used carrier for the encapsulation of oils. Therefore, the objective of this study was to determine the influence of SD conditions on EE and product yield (PY) using gum arabic as a carrier at different drying temperatures (120 to 180 °C) and carrier-oil ratios (2 to 4). The EE of all SBO powders ranged from 79.33 to 93.18%, and the highest EE was obtained at a drying air temperature of 180 ˚C and a carrier-oil ratio of 4. Increasing the drying temperature and adding more carrier to the initial emulsion resulted in a higher EE of SD, probably due to less diffusion of the oil to the surface, as the time to form a crust around the droplets is reduced and the amount of unencapsulated surface oil decreases (Di Giorgio, Salgado & Mauri, 2019 ; Tonon & Hubinger, 2014). The PY of all SBO powders ranged from 35.79 to 52.28 % and the highest PY was obtained at a drying air temperature of 120 ˚C and a carrier-oil ratio of 3. Higher drying temperatures resulted in lower PY which could be due to extreme vaporization at high inlet air temperatures and degradation or loss of encapsulated cores. Drying temperature, carrier oil ratio and their combined effect significantly influenced EE and PY. The SD is an effective method for producing SBO powders with great potential as functional food ingredients.

In recent years, the demand for novel antioxidant agents derived from natural products is rising in the food industry. Medicinal plants such as bay laurel (Laurus nobilis L), sage (Salvia Officinalis L.) and thyme (Thymus vulgaris L.) contain a variety of bioactive molecules that contribute to the plants' antioxidant potential. While the antioxidant potential of plant extracts and essential oils has been well researched, the potential of hydrosols secondary products of hydrodistillation process, has recently raised research interest. Hydrosols consist of distillation water with small amounts of dispersed essential oils which provide them antioxidant and antimicrobial properties potentially applicable in the food industry. The aim of this study was to evaluate the antioxidant potential of hydrosols of bay laurel, sage and thyme, and their mixture (1:1:1) obtained during hydrodistillation on Clevenger apparatus, and to monitor their antioxidant stability through 28 days of storage at room temperature by DPPH Radical Scavenging and Ferric Reducing Antioxidant Power (FRAP) assays. Bay laurel hydrosol showed the highest antioxidant activity on the first day with a value of 2000 µM of Trolox equivalents (TE) as determined by both DPPH and FRAP followed by the plant mixture (DPPH 1000 µM TE, FRAP 621 UM TE), thyme (DPPH and FRAP 370 µM TE) and sage whose antioxidant activity was at the bottom of detection level. Both FRAP and DPPH showed that the antioxidant activity of all hydrosols has gradually decreased down to approximately 50% after 28 days. Nevertheless, bay laurel hydrosol has shown the highest potential for application in the food industry, for example in minimal processing of fruits and vegetables with the goal of prolonged shelf life, since the DPPH and FRAP values were still relatively high (1000 µM TE) after 28 days.

Production of natural compounds from medicinal plants with numerous biological effects has become very popular due to increasing concerns about the safety of using synthetic substances and the effects of COVID -19 on health. Sea buckthorn (Elaeagnus rhamnoides (L.) A. Nelson), (SB) is known as an important source of natural antioxidant and antimicrobial bioactive molecules (BAM), but also, when added to foods, improves sensory properties, microbiological stability, and shelf life of products. The best-known sea buckthorn product is berry oil (SBO) (Yu et al., 2017), which contains large amounts of lipophilic BAM such as unsaturated fatty acids, tocopherols, carotenoids, and sterols (Koskovac e al., 2017), but differences in composition have been found depending on the cultivar, growing conditions and extraction methods used (Christe et al., 2020 ; Cenkowski, 2006). Supercritical CO2 extraction of oil as an advanced method is gaining popularity due to its better extraction efficiency compared to conventional extraction methods. Moreover, CO2 as an environmentally friendly and safe solvent (GRAS) can be easily removed from oil. Therefore, the aim of this study was to determine the content of fatty acids, sterols, and α-tocopherols, as well as the antioxidant capacity (AC) of SBO obtained by supercritical CO2 extraction from SB berries from Croatia. The tocopherol content of the SBO extracts was determined by HPLC/FLD, the content of fatty acids and sterols by GC-MS and AC by the lipophilic ORAC method. A total of 16 sterols were determined, with sitosterol (589.28 mg/100 g oil) being the most abundant, followed by campesterol (31.97 mg/100 g oil), uvaol (21.79 mg/100 g oil), and obtusifoliol (12.99 mg/100 g oil). High content of unsaturated fatty acids such as omega-7 palmitoleic acid (34.54%) and omega-6- γ-linolenic acid (10.78%) was also found. SBO was characterized by high content of α-tocopherol (275.63 mg/100 g oil) and high antioxidant ORAC capacity (1676.90 μmol TE /100 g oil). The results suggest that the SBO obtained by supercritical CO2 extraction is a rich source of various lipophilic BAM with high antioxidant capacity, which could be used for the production of natural products, functional foods and as food additives.

Fennel (Foeniculum vulgare Mill.) is a perennial aromatic and medicinal plant belonging to the Apiaceae family. It originates from the Mediterranean area, but it has spread worldwide through naturalization and cultivation. The use of fennel in culinary purposes and traditional medicine is known from ancient times, but nowadays its application has been extended to the pharmaceutical, cosmetic and food industries. Although different parts of fennel (leaves, young shoots, roots and seeds) can be utilized, fennel seeds are mostly exploited due to their abundance of essential oil (up to 5-6%) which is characterized by a strong and pleasant aroma. Fennel essential oil has been recognized to possess antimicrobial, antifungal and antioxidant properties, therefore it is widely used in various areas. The chemical composition of fennel essential oil showed up to 80 volatile compounds, among which trans-anethole, fenchone and estragole are major compounds, being the key compounds of the characteristic fennel flavor. Besides these compounds, the presence of other compounds such as α- and β-pinene, myrcene, α-phellandrene, limonene and p-anisaldehyde have also been reported. The isolation of essential oil can be accomplished by various extraction techniques, such as hydrodistillation (HD) and steam distillation (SD) which are known as conventional ones and still are the most common in use. They are effective for the extraction of essential oils from spices and herbs from which such oils are difficult to isolate and they do not include the use of chemical solvents. On the other hand, their main drawbacks are long duration, difficult regulation of heat and possible loss of thermolabile and light volatiles. Therefore, in order to overcome these deficiencies, advanced extraction techniques have been developed, including microwave-assisted extraction (MAE), enzyme-assisted extraction (EAE), ultrasound-assisted extraction (UAE), subcritical water extraction (SWE) and supercritical fluids extraction (SFE), namely extraction with supercritical carbon dioxide (SC-CO2) which is considered “green” showing the most promising beneficial effects due to being sustainable, environmentally friendly and cost-effective. However, regardless of the selected extraction technique, it is necessary to define optimal process conditions taking into account all process parameters in accordance with the raw material used, and optimize the extraction process with the intention to achieve the maximum yield of essential oil of the best quality and unaltered composition.

Recently, bioactive molecules (BAM) from plant- based sources have gained interest of both consumers and scientific community due to the spreading knowledge of their health beneficial properties. Fruits, vegetables, herbs and spices have shown to be a source of various bioactives. Furthermore, food industry produces a significant amount of waste rich in BAM which can be revalorized for application in food and nutraceutical industry. Generally, extraction method is the first and the most crucial step in the research and utilization of bioactives. Recent studies emphasize the advantages of “green” extraction techniques for BAM isolation versus conventional ones. New technologies include microwave-assisted, ultrasound-assisted, enzyme- assisted, pressurized liquid extraction, pulsed electric field extraction, supercritical-fluid extraction, etc. Although based on different mechanisms, all these techniques have similar advantages, mainly enhancement of the extraction yield, shortening of the extraction time, reduced solvent consumption and minimizing the degradation of thermosensitive compounds. Another important segment in bioactives’ utilization is their stability. Most of the plant BAM are prone to degradation due to the exposure to light, oxygen and elevated temperatures, which has negative impact on their applicability in food industry as can significantly decrease their bioactivity. One of the most reliable methods for protecting the bioactives is encapsulation. Additionally, encapsulation provides controlled delivery to the target place in organism and protection in the GI tract and has been reported to increase the bioavailability of plant BAM. Furthermore, encapsulated BAM are easier to incorporate into food matrix which is especially important for lipophilic compounds. Various encapsulation techniques have been developed, such as spray drying, freeze drying, extrusion, ionic gelation, coacervation, liposomes, molecular inclusion, etc. The selection of the proper procedure depends upon the properties of the BAM to be encapsulated and the final purpose and designation place of the product.

In recent years, a growing demand on functional products enriched with naturally derived ingredients has raised interest in finding new sources of bioactive molecules as well as establishing optimal isolation technologies that would allow their fast and efficient isolation. Various green extraction technologies, such as ultrasound-assisted extraction (UAE), microwave-assisted extraction (MAE) and pressurized liquid extraction (PLE) have emerged due to their main advantages of reducing the extraction time, energy and solvent consumption while achieving higher extraction efficiency and lower degradation of targeted compounds compared to conventional extraction techniques. Carob tree (Ceratonia siliqua L.) is a Mediterranean plant belonging to the Fabiaceae family which has recently drawn attention due to the nutritional value of its pods, while other plant parts, such as leaves, have been researched to a lesser extent. Since the health beneficial properties of polyphenols are well known, the aim of this research was to evaluate carob leaves grown in south Croatia as a source of polyphenols by analyzing the phenolic content and antioxidant activity of carob leaf extracts obtained by three different green extraction techniques (UAE, MAE and PLE). The extractions were carried out on a sample of grinded dry carob leaves for 10 min at a temperature of 80°C, sample: solvent ratio 1:20 and 50% aqueous ethanol as a solvent. The frequency in UAE was 37 kHz, microwave power in MAE 500 W and PLE was set to 1 static cycle. The total phenolic content (TPC), hydroxycinnamic acids content (HAC) and flavonol content (FC) were determined by spectrophotometric methods, while the antioxidant activity was determined by DPPH Radical Scavenging and Ferric Reducing Antioxidant Power (FRAP) assays. It was shown that carob leaves can be considered a source of phenolic compounds since the TPC was in the range of 51.86–64.81 mg gallic acid equivalents (GAE) g-1 . The extracts have also shown high antioxidant activity with values of 25, 10–28, 90 mM Trolox equivalent (TE) g-1 and 22, 92– 51, 23 mM TE g-1 as determined by DPPH and FRAP, respectively. The extraction techniques have significantly influenced all of the examined parameters and MAE was shown to be the most suitable for achieving maximum TPC, HAC, FC and antioxidant activity as determined by DPPH. The results highlight the importance of research of different extraction techniques that would allow the highest recovery effectiveness and utilization of the plants’ beneficial properties.

Sea buckthorn (Hippophae rhamnoides L.) is a valuable and unique plant, rich in various bioactive compounds contained in the different parts of the plant, especially in the fruits and leaves. Due to its great potential, the scientific community is showing a great interest in this plant, and numerous studies have reported beneficial health effects, such as antioxidant, anticancer, antiinflammatory, etc. Compounds of interest can be isolated by various extraction methods, either conventional or advanced, however it is of great importance to optimize the process conditions to obtain maximum yield of target compounds. Therefore, this study examined the influence of temperature (80, 100, and 120 °C), static extraction time (5, 10, and 15 min) and number of extraction cycles (1, 2, and 3) during the pressurized liquid extraction (PLE) of phenols from freeze-dried sea buckthorn leaves using an aqueous ethanol solution (70%) as extraction solvent. The extracts obtained were analyzed spectrophotometrically for the content of total phenols (TPC) using the Folin-Ciacolteau method, while the FRAP method was used to determine antioxidant capacity (AC). All results were statistically analyzed by multifactorial analysis of variance and Tukey's HSD test. The TPC was determined in a range of 69.05-112.96 mg GAE g-1 of dry matter (DM) and the values of AC ranged from 766.72-1182.02 µmol TE g-1 DM. Statistical analysis showed a significant influence of all tested PLE parameters on the TPC as well as on the AC of the extracts. The highest TPC were measured in extracts obtained at the highest temperature and cycle number, while there was no significant difference in the TPC extracted during a static extraction time of 10 or 15 min. On the other hand, the AC values of the extracts were highest at the highest level of all three PLE parameters tested. In conclusion, PLE using 70% ethanol showed to be an effective method for the successful extraction of phenols from sea buckthorn leaves, where PLE conditions of 120 °C/10 min/3 cycles provided the highest TPC, while the highest value of AC was obtained at 120 °C/15 min/3 cycles.

Dictyota dichotoma is a species of brown algae found in rocky littoral and sublittoral zones in a wide range of habitats, from the Atlantic Ocean and the Mediterranean Sea, to the western Indian Ocean. Research has shown that Dictyota dichotoma contains a variety of chemical compounds, including pigments, phenols, flavonoids, alkaloids, steroids, tannins, coumarins, quinones, and glycosides. The presence of these biologically active molecules, especially polyphenols, is often associated with numerous health-promoting properties, which is why they are frequently used to fortify functional foods. Microwave-assisted extraction (MAE) is one of the more sustainable isolation techniques that enable rapid and more effective isolation of these substances. In this study, MAE was applied to isolate Dictyota dichotoma polyphenols in two research phases. In the first phase, the influence of different percentages of aqueous ethanol (0, 30, 50, 70, and 96%) and sample-to- solvent ratio (1:10, 1:20, and 1:30) were investigated, while temperature (60°C), irradiation time (10 min), and microwave power (800W) were kept constant. The total phenolic content (TPC) of the extracts was determined by the Folin-Ciocalteu spectrophotometric method. The choice of solvent and sample-to-solvent ratio showed significant influence on Dictyota dichotoma TPC, and the highest TPC (6.42 mg GAE g-1) was achieved when 30% ethanol and a ratio of 1:60 were applied. In the second phase of the research, temperature (40, 60, and 80°C) and irradiation time (5, 10, and 15 min) were varied, while the ethanol content (30%) and sample-to-solvent ratio (1:60) were kept constant. The results of the statistical analysis showed that temperature had a significant influence (p£0.05) on Dictyota dichotoma TPC, while the influence of irradiation time was not significant (p³0.05). The highest TPC (3.33 mg GAE g-1) was achieved at 40°C after 5 min of irradiation. The results of this study showed the influence of various MAE parameters on Dictyota dichotoma TPC and confirmed the importance of developing optimal isolation method that can lead to the most effective polyphenol recovery.

Posljednjih godina, uslijed povećanja svjesnosti potrošača o brizi za zdravlje, potražnja na tržištu funkcionalnih proizvoda prirodnog podrijetla koji nose ekološke i bio oznake sve je veća. Ljekovito i aromatično bilje predstavlja važan izvor bioaktivnih molekula, koje, osim povoljnih učinaka na ljudski organizam, također mogu djelovati kao prirodni antioksidansi, konzervansi i mikrobiocidi te omogućiti formulaciju proizvoda koji će udovoljavati zahtjevima potrošača. Među mnogobrojnim biljnim vrstama ističe se lovor (L. nobilis L.), mediteranska biljka tradicionalno upotrebljavana u narodnoj medicini čiji se brojni biološki učinci u značajnoj mjeri pripisuju različitim grupama fenolnih spojeva. Ubrzana ekstrakcija pri povišenom tlaku (ASE) pokazala se vrlo učinkovitom u ekstrakciji fenolnih spojeva iz različitih biljnih vrsta te je stoga cilj ovog istraživanja bio optimirati ASE ekstrakciju fenolnih spojeva iz lista lovora s ciljem postizanja većih prinosa. Monitoring fenolnih spojeva u ekstraktima lista lovora proveden je spektrometrijskom Folin-Ciocoltaeu metodom. Varirani parametri bili su koncentracija etanola koji je korišten kao otapalo (50-70% w/v), temperatura ekstrakcije (90-150°C), broj ciklusa (1-3) i statičko vrijeme (5 i 10 minuta). Definirani su optimalni uvjeti ekstrakcije (50% etanol, 150°C, 3 ciklusa, 10 minuta) pri kojima je postignuta najveća koncentracija fenolnih spojeva koja je iznosila 49, 30 mg GAE g-1.

Ljekovito i aromatično bilje te napredne tehnike ekstrakcije predstavljaju važan segment u pronalasku alternativnih i održivih rješenja za proizvodnju sirovina prehrambene, farmaceutske i kozmetičke industrije. Različite porodice ljekovitog i aromatičnog bilja bogat su izvor bioaktivnih molekula, sekundarnih metabolita koji se potencijalno mogu upotrijebiti u različite svrhe kao konzervansi, mikrobiocidi i antioksidansi, što može biti održiva alternativa postojećim rješenjima. Među mnogobrojnim biljnim vrstama ističe se lovor (L. nobilis L.), mediteranska biljka čiji su listovi tradicionalno korišteni u narodnoj medicini zbog brojnih bioloških učinaka koji se u značajnoj mjeri pripisuju različitim grupama fenolnih spojeva. U fokusu brojnih istraživanja su i napredne tehnike ekstrakcije koje za cilj imaju postizanje što većih prinosa uz smanjenje upotrebe organskih otapala, skraćivanje vremena ekstrakcije, provođenje ekstrakcije pri nižim temperaturama te posljedično smanjenje negativnih utjecaja na okoliš. Stoga je fokus istraživanja stavljen na optimiranje postupaka ekstrakcije fenolnih spojeva iz lista lovora primjenom naprednih tehnika ekstrakcije kao što su ekstrakcija potpomognuta ultrazvukom (UAE), mikrovalovima (MAE) te ubrzana ekstrakcija pri povišenom tlaku (ASE) s ciljem postizanja većih prinosa. U skladu s načelima „zelene kemije“ kao otapala za ekstrakciju koriste se voda te 30% do 70% vodene otopine etanola. Monitoring fenolnih spojeva u ekstraktima lista lovora provodi se tekućinskom kromatografijom ultra visoke djelotvornosti sa spektrometrijom masa (UPLC/MS-MS). Za svaku od primijenjenih tehnika definirani su optimalni uvjeti, a 70% etanol pokazao se najučinkovitijim otapalom u svim tehnikama ekstrakcije. Primjenom UAE najveća koncentracija fenolnih spojeva postignuta je nakon 15 minuta pri amplitudi od 100% i iznosila je 277, 44 mg GAE/g, dok je primjenom MAE pri temperaturi od 80°C, snazi mikrovalova 800 W nakon 15 minuta ekstrakcije najveća postignuta koncentracija iznosila 262, 11 mg GAE/g. Ubrzana ekstrakcija pri povišenom tlaku pokazala se najučinkovitijom metodom s najvećom koncentracijom fenolnih spojeva od 355, 97 mg GAE/ g, postignutom pri temperaturi 150°C, statičkom vremenu 5 minuta i dva ciklusa ekstrakcije. Sve napredne tehnike pokazale su se učinkovitijima od konvencionalne refluks ekstrakcije gdje je koncentracija fenolnih spojeva u 70% etanolu iznosila 168, 89 mg GAE/g.

Sea buckthorn or sea berry (Hippophae rhamnoides L. ; Elaeagnus rhamnoides L.) (SB) berries contain a wide spectrum of lipophilic and hydrophilic bioactive molecules which possess nutritional and health-promoting properties. Therefore, the aim of this study was to evaluate the content of carotenoids, fatty acids, phenolics, α-tocopherol and L-ascorbic acid in SB berries grown in Croatia. Lipophilic and hydrophilic extracts were obtained by ultrasound-assisted extraction with n-hexane (lipophilic) or 70% ethanol (v/v) (hydrophilic), and analyzed using liquid and gas chromatography. Results showed that analyzed SB extracts were characterized with high content of ascorbic acid (123.89 mg/100 g dw), carotenoids (53.93 mg/100 g dw), flavonoids (50.87 mg/100 g dw) and α-tocopherol (29.85 mg/100 g dw). Moreover, high amounts of unsaturated fatty acids, such as omega-7 palmitoleic fatty acid (34.54%) and omega-6 γ-linolenic acid (10.78%) were also determined. Carotenoids were mainly represented by lutein and zeaxanthin fatty acid esters (35.02 and 15.13 mg/100 g dw), respectively. Among the flavonoids, the most abundant were flavanol glycosides (25.06 mg/100 g dw) such as isorhamnetin-3-hexoside, rutin and isorhamnetin-3-rutinoside. SB berries certainly showed to be a rich source of various antioxidants and they could be used in the production of functional and value added products.

Sea buckthorn (Hippophaë rhamnoides L.) oil (SBO) as the most valuable part of berries receives increasing attention worldwide due to health effects. It is rich source of bioactive compounds like tocopherols, carotenoids, flavonoids, essential and polyunsaturated fatty acids and essential amino acids. These compounds are relatively unstable and susceptible to degradation and it is important to improve their stability during handling, processing and storage. Spray drying method is the most commonly used microencapsulation method in the food industry for oil protection. Therefore, the objective of this study was to evaluate the influence of the spray drying conditions at different temperatures (120 to 180 °C) and wall to core ratio (2 to 4) on the total carotenoid (TC) and tocopherol (TT) content and antioxidant capacity (AC) of sea buckthorn oil microencapsulated using gum Arabic as wall material. The TC and TT was determined using HPLC methods and AC by the ORAC method, respectively. The carotenoid retention in powders compared to SBO was in range from 8.19 to 47.13 % and tocopherol retention from 34.75 to 45.33 %, respectively. The results showed that increase in a wall to core ratio favors carotenoid and tocopherol retention, while carotenoids were more susceptible on increase of drying temperature than tocopherols. Temperature and wall to core ratio significantly affected the AC. The highest retention of carotenoids (88.36 mg/100 g dm) and tocopherols (50.79 mg/100 g dm) showed sea buckthorn powder obtained at 120 °C and wall to core ratio of 2 while the highest ORAC value (305.8 μmol TE/L) was determined in powder obtained at same drying temperature but wall to core ratio of 4. The sea buckthorn oil powders obtained by spray drying showed carotenoid and tocopherol stability with antioxidant properties and potential to be used for the development of functional products.

Mastic tree (Pistacia lentiscus L.) is an evergreen shrub with a strong characteristic aroma and green leaves, widely spread along Mediterranean countries. Different parts of P. lentiscus (aerial part, leaves, fruit and resin) have been traditionally used in the treatment of hypertension, gastrointestinal diseases and throat infections due to their antioxidant, anti-inflammatory, hepatoprotective and antimicrobial effects. These health promoting properties of mastic tree have been attributed to the presence of various bioactive compounds (phenolics, triterpenoids, pigments). Chlorophylls are pigments responsible for the green colour of plants and are largely determining photosynthetic capacity and plant growth. Apart from their functional role in photosynthesis, chlorophylls act as protective factors against excessive UV radiation and oxidative stress. Their content varies upon genetic factors, season, phenological stage, harvest time, geographical origin and environment in which the leaves have developed. Therefore, the aim of this study was to evaluate the content of chlorophylls present in P. lentiscus leaves grown in four locations in Croatia (Barbariga, Lun, Hvar and Korčula) and harvested during three phenological stages (flowering stage in May, early fruiting in August and late fruiting stage in October). The extraction and analysis of chlorophyll content was performed conventionally with 80% aqueous solution of methanol and by UV-VIS spectrophotometry, respectively. Results of statistical analysis showed that influence of growing location and phenological stage as well as their interaction had a significant impact on chlorophylls content in mastic tree leaves samples (p

Sea buckthorn (Hippophae rhamnoides L.) is rich source of bioactives such as phenolics and lipophilic compounds and used as food and medicinal plant. Higher phenolic contents (PC) and antioxidant capacities (AC) were determined in sea buckthorn leaves (SBL) than in berries. Phenolic extraction from plants is the first step in utilization of these compounds and more efficient extraction processes such as microwave assisted extraction (MAE) have been developed in recent years due to reduction of extraction time and solvent. Therefore, the aim of this study was to evaluate the PC and AC in SBL extracts obtained under different MAE conditions of microwave power (300, 500 and 700 W) and extraction time (5, 10 and 15 minutes) at 60 ºC using 70% ethanol (v/v) as the extraction solvent. Phenolic profile of SBL extracts was determined using UPLC-MS2 and AC by ORAC method, respectively. A total of 30 phenolic compounds were determined comprising flavonols (FOL), flavones (FON), flavan-3-ols (F-3-OL) and phenolic acids (PA). The FOL (372.35 to 538.69 mg 10-2g-1 dm) showed to be the predominant phenolic group in SBL represented by rutin and kaempferol as the most abundant. Considerably lower content of total PA (71.29-132.29 mg 10-2g-1 dm), FON (7.99-22.49 mg 10-2g-1 dm) and F-3-ON (2.70-3.72 mg 10-2g-1 dm) were determined. The all SBL extracts had high AC (8779.17-9186.42 μmol TE 10- 2g-1 dm). Applied MAE conditions significantly affected the concentration of FOL, FON, PA and AC. Generally, the highest content of FOL, FON and AC was observed in extracts obtained under microwave power of 500 W and extraction time of 15 minutes, while the highest content of PA was observed under microwave power of 700 W and shorter extraction time of 5 minutes. Therefore, MAE is effective method for the production of valuable extracts from SBL with potential beneficial applications.