**3.2. The structure of cyclodextrins**

**3. Chemistry and structure and cyclodextrins**

• Culture of the producing microorganism CGTase enzyme

The CDs are produced by degradation of the prehydrolyzed starch and their subsequent cyclization-mediated cyclodextrin glucosyltransferase enzyme (CGTase, EC 2.4.1.19) produced by bacteria that belong to the genus *Bacillus.* Due to the helical structure of the starch molecules, the primary cleavage product undergoes an intermolecular reaction forming cyclic products joined by α-1,4 linkages, generally designated by cyclodextrins. To distinguish them, Greek letters are used to specify the number of D-glucose units (in brackets): α (6) β (7) γ (8) δ (9) ε (10) ξ (12) η (13). The shapes α, β, and γ are the natural cyclodextrins and most commonly used (**Figure 2 (c)**). Higher numbers of counterparts of glucose units also exist but are difficult to purify, with weaker inclusion properties. Cyclodextrins with a number of glucose units less than 6 do not

The preparation of cyclodextrins can be subdivided into the following main stages:

• Separation of the enzyme from the medium, their concentration, and purification

• Enzymatic conversion of prehydrolyzed starch a mixture of cyclic and noncyclic dextrins

In industrial production of cyclodextrins, the most frequently used source of enzyme is *Bacillus macerans*, renamed as *Paenobacillus macerans*. Other enzymatic sources used are *Klebsiella pneumonia* and *Alkalophilic bacterium* 38–2. The forms α, β, and γ are dependent from the source of CTGase enzyme. The *Bacillus macerans* and *Klebsiella pneumonia* CTGase mainly produce the α form. *Alkalophilic bacterium* 38-2 mainly produces β-cyclodextrin. However, the relationship between the CD formed also depends on the incubation time of the enzyme in starch medium culture because most CTGases initially produce the α form, while the synthesis of other forms is slower [3].

**Figure 2.** Structure of a cyclic oligosaccharide, cyclodextrin, CD (a); The "donut" Molecular (b); Paragraph equal to six, seven or eight rings of D-glucopyranose, joined by glycosidic linkages of the type α-1.4, representing α-CD, β-CD and

• Separation of the CDs are from the conversion mixture, purification and crystallization

**3.1. Biological synthesis of cyclodextrins**

122 Cyclodextrin - A Versatile Ingredient

exist, probably due to steric hindrance.

γ-CD, respectively (c); white crystalline powder β-CD (d).

The native cyclodextrin molecules (α-CD, β-CD, and γ-CD) have the shape of a short truncated cone with a cavity inside, i.e., a toroidal shape. The length is determined by the height of the glucose unit (7.9 Å = 0.79 nm), and the diameter of the cavity is determined by the number of glucose units (**Figure 2 (a)** and (**c)**).

The glucose rings linked together by α-1.4 linkages as in amylose. They are oriented in the same direction, and thus, the narrow end of the torus is formed by the primary hydroxyl groups (O (6) H), while the wider edge of the truncated cone is occupied by the secondary hydroxyl (O (2) H, O (3) H) groups. These peripheral hydroxyl groups confer hydrophilic properties to the CD surface. Moreover, the internal cavity has mainly hydrophobic characteristics due to the methine group (CH) and the oxygen atoms of the ether type (O (4) and (5)).

The CDs may crystallize in the form of hydrate or inclusion compound, and the crystal structure was mainly determined by the following factors:


The interstices between the CD units are occupied by water molecules incorporated in the overall structure (see **Figure 3**) [3, 4].

The CDs cavity in the center, with predominantly hydrophobic character, is large enough to hold, accommodate, or include other molecules. When this occurs, there is the formation of an inclusion compound. These compounds, or complexes, may be described as a molecular-level nanoencapsulation. Food ingredients formulated with cyclodextrins become stable to heat and oxidation processes and are not affected by dispersion forces and are readily dispersed for use in liquid products [5].

**Figure 3.** The crystalline hydrate of β-CD. The blue are represented statistical locations of the interstices water molecules; red are represented statistical locations of cavity water molecules [3, 4].

The food industry uses the native cyclodextrins in different ways owing to the above-described properties, being used in various applications due to their ability to form inclusion compounds. The α-cyclodextrin acts even as prebiotic. Thus, formulations with CDs are used in food and also in the designated functional food markets in order to circumvent the problems of stability, taste, and flavors of special ingredients. In this context, natural functional foods are food systems enriched, e.g., with bacterial cultures, omega 3 fatty acids, anthocyanins, dietary fibers, etc., which can contribute to the maintenance of health and reduction of disease risk. [6]
