**2. Polyphenols**

Polyphenols include various different compounds divided into several classes: phenolic acids, flavonoids, stilbenoids, tannins, coumarins, and polymeric lignans (Figure 1) [18]. Their chemical structure may vary from simple to complex. The largest group of polyphenols is that of flavonoids, divided into several subgroups (flavonols, flavones, flavanols, flavanones, anthocyanidins, isoflavonoids, and chalcones) [19]. Their structure consists of two phenyl groups linked with a three-carbon bridge. According to the degree of oxidation and unsaturation of the three-carbon segment, they differ from each other. Different sugar molecules can be attached to the hydroxyl groups of flavonoids. They are usually in a glycosidic form which improves their solubility in water. Acylation of the glycosides where sugar hydroxyls are derivatized with acid (such as ferulic and acetic acids) is also common. The interconnection between several basic units of polyphenols makes larger and more complex structures, such as hydrolysable tannins and condensed tannins [20]. Phenolic acids represent a large group of hydrophilic polyphenols and they are constituted of a single phenyl ring [14]. Due to the diversity in the structures of polyphenols, they possess different properties (such as solubility and polarity) [20].

Polyphenols exist ubiquitously in vegetables and fruits and their consummation is very desirable [14]. Various positive bioactivities of polyphenols toward pathologic conditions are known, through their antioxidant properties. These molecules are capable of donating hydrogen atoms and electrons [14]. One such activity is the anticancer property of polyphenols [21]. Hollman et al. [22] reviewed the antioxidant activities of polyphenols within the organism and their positive impact on cardiovascular health. In addition, polyphenols contribute to the sensory quality of the products (wine, jellies, juices, chocolate, etc.). They affect color, bitterness, turbidity, etc. [6,23]. Due to their many functional properties, polyphenols are of great interest to the food, chemical and pharmaceutical industries [2].

**Figure 1.** Classification of polyphenols and the most important representatives of individual groups (adapted from Dobson et al. [18]).

### **3. Encapsulation**

Encapsulation is the process of entrapment of active compounds into particles to enable isolation or controlled release of given compounds. The main role of encapsulation is the protection of sensitive active compounds from degradation [17]. Various techniques can be used for encapsulation depending on core material, size of required particles, physical state, or sensitivity to high temperature [24]. These techniques include freeze-drying, spraydraying, extrusion, fluidized bed coating, spray-cooling/chilling, coacervation, liposome entrapment, cocrystallization, vacuum-drying, centrifugal suspension separation, nanoencapsulation, molecular inclusion and emulsification [13]. Encapsulation with coacervation is based on phase separation of a hydrocolloid from an initial solution and subsequent deposition of the formed coacervate phase around an active ingredient suspended in media. It is considered an expensive technique but very beneficial for high-value compounds [25]. Extrusion is a process based on passing a polymer solution with active ingredients through a nozzle (or syringes) into a gelling solution. Usually the used wall material is sodium alginate while calcium chloride solution serves for capsule forming. It is easy to execute on a laboratory scale with long shelf-life capsules, while scale-up of this technique is expensive and demanding with a limited choice of wall materials [26]. Emulsification includes the dispersion of one liquid into the other (two immiscible liquids) in the form of droplets. An emulsifier is required for stabilization and by application of drying, a powder form of encapsulates can be achieved [26]. The molecular inclusion method is also known as host-guest complexation. Apolar guest molecules are trapped inside the apolar cavity of host molecules (such as cyclodextrins) through non-covalent bonds [26]. Cocrystallization techniques include modification of the crystalline structure of sucrose to an irregular agglomerated crystal with porous structure in which active ingredient can be incorporated [25]. Fluidized bed coating is also referred to as fluidized bed processing, air suspension coating, or spray coating. The principle is that coating is applied to the particles

suspended in the air. For this technique a wide range of coating materials such as aqueous solutions of cellulose, starch derivates, gums, or proteins is suitable [27]. Spray chilling is also known as spray cooling, prilling, or spray congealing. The basic principle is similar to spray drying with the key difference of using a cooling chamber instead of a drying chamber. Regarding coating materials, only lipid-based materials (fats, waxes, fatty acids, fatty alcohols and polyethylene glycols) are used [27]. Nanoencapsulation is an innovative trend in the field of food technologies. The final results are particles of diameters ranging from 1 to 1000 nm. The term nanoparticles includes nanospheres and nanocapsules. The first one has a matrix-type structure where the active ingredients can be adsorbed at the sphere surface or encapsulated in particles. In nanocapsules, the active ingredient is limited to a cavity with an inner liquid core surrounded by a polymeric membrane. Nanoparticles have a larger surface area, increased solubility, enhanced bioavailability and improved controlled release [25]. Freeze-drying and spray-drying are the most frequently employed methods for removing water from foods with encapsulation effects [28]. A schematic view of these methods is presented in Figure 2.

**Figure 2.** Schematic view of freeze-drying and spray-drying process, characteristics of particles and advantages and disadvantages of methods (adapted from Fang and Bhandari, [25]; Grgi´c et al., [26]).

The wide range of polyphenol biological activities can be restricted due to their low stability, low bioavailability, and unpleasant flavor [14]. Encapsulation of polyphenols improves their stability during storage and can also be used to achieve the masking of unpleasant flavors in foods (such as bitter taste and astringency) [2]. Food with high intensity of bitterness and astringency elicit negative consumer reactions. Compounds responsible for that are flavanols and flavonols. Flavan-3-ol monomers such as catechin, epicatechin, epigallocatechin, epicatechin gallate and epigallocatechin gallate as well as their oligomers proanthocyanidins (condensed tannins) are abundant in wine and tea. It can be said that flavanols are the main compounds that cause bitterness and astringency in tea and red wine (with the exception of caffeine in tea) [29]. The method selected for encapsulation must be based on the polyphenol's characteristics such as chemical structure, thermophysical stability, solubility, affinity to coating material, target properties such as particle size and morphology, among others [14].

#### *3.1. Freeze-Drying*

Freeze-drying, also known as lyophilization, is a drying technique based on the phenomenon of sublimation. This technique allows the long-term preservation of heatsensitive and oxidation-prone compounds, as well as foods and other biological materials, since it is conducted at low temperatures and under vacuum [30]. This method does have disadvantages such as higher unit cost and processing time [17]. Despite those issues, freeze-drying is widely used to obtain high-value food products and is considered a standard method for encapsulation in most research studies [30].

The freeze-drying process includes the complete freezing of samples, ice sublimation (primary drying) and desorption of remaining unfrozen/bond water (secondary drying). In the first step, the freezing rate determines the formation and size of ice crystals. Large ice crystals formed by the slow rate of freezing can sublimate easily and increase the primary drying rate. In primary drying, the shelf temperature will increase using a vacuum to start the sublimation. It is necessary that product temperature is 2–3 ◦C below the temperature level of collapse, at which the product can lose its macroscopic structure. The endpoint of the primary drying phase is a key parameter to determine because the increased temperature (in the secondary drying) before the sublimation of all ice could collapse the final product quality [30].
