**4.1 Food**

274 Biochemistry

Flavonoid acylation with aromatic acids was reported to improve physiological activities, such as UV-absorbing capacity, radical scavenging ability (Delazar et al., 2005; Ishihara & Nakajima, 2003; Harborne & Williams, 2000; Alluis & Dangles 1999; Jungblut et al., 1995) pigment stabilization (especially anthocyanins) (Ishihara & Nakajima 2003), and interaction

Flavonoid esters acylated with *p*-coumaric acid were found to increase antioxidant (Pajero et al., 2005) and anti-inflammatory activities (Harborne & Williams, 2000), as well as antiproliferative and cytotoxic effects on various cancer cell lines (Mitrokotsa et al., 1993). Moreover, *p*-coumaroyl esters of quercetin and kaempferol were reported to have positive effects on cerebrovascular disorders (Calis et al., 1995). Similarly, flavonoid esters esterified with cinnamic acid were shown to exhibit antiproliferative activity against several human cancer cell lines (Duarte-Almeida et al., 2007). Flavonoid acylation with caffeic acid contributes to the enhancement of antioxidant properties (Pajero et al., 2005). Flavonolignans acylated with truxinic acid were shown to possess hepatoprotective as well

Biological activities of aliphatic acids are not of a big importance in comparison with aromatic acids. These compounds are mainly accepted as energy storage and components of several compartments of cells, such as membranes, enzymes, surfactants, etc. In the literature, more studies can be found describing changes in biological activities of flavonoids

The aliphatic acylation of anthocyanins with malonic acid is important for enhancing the pigment solubility in water, protecting glycosides from enzymatic degradation and stabilizing anthocyanin structures (Nakayama et al., 2003). Several *in vitro* observations suggest that acylation with malonic acid or sinapic acid is crucial for efficient flavonoid

Fatty acid esters of catechins were reported to display antitumor, antibacterial and 5-α reductase inhibiting activity (Fukami et al., 2007) as well as antioxidant properties (Sakai et al., 1994). Lee et al. (2003) reported anti-atherogenic activity of two naringenin derivatives,

Acylation of the flavonoid molecule with polyunsaturated fatty acids introduces potential antitumor and antiangiogenic properties (Mellou et al., 2006). Anticarcinogenic effects were observed also in silybin esters acylated with butyric and lauric acid (Xanthakis et al., 2010). Recently, we found that acylation of rutin with unsaturated fatty acids, such as oleic, αlinoleic and linolenic, increased the antioxidant potential of the initial compound (Viskupicova et al., 2010). This observation is in accordance with the results of Mellou et al.

In the field of fatty acid ester synthesis, information on the photoprotective effectiveness of new quercetin derivatives acylated with acetic, propionic and palmitic acids, has been reported. The authors found that esterification with a short side-chain (such as acetate or propionate) may improve migration through the aqueous environment and interaction with

or penetration into phospholipid membranes (Saija et al., 2003).

with cellular targets (Ferrer et al., 2008).

as anticancer activity (Sharma et al., 2003).

after their acylation with aliphatic acids.

**3.2 Esters with aliphatic acids** 

accumulation in plants.

7-*O*-oleic ester and 7-*O*-cetyl ether.

(2006) and Katsoura et al. (2006).

The major contribution of acylated flavonoids in the food industry lies in the improvement of stability and solubility of initial molecules, e.g. by reducing lipid oxidation in oil/fat based food systems, desirable modification of unwanted sensory properties of certain flavonoids, taking advantage of pigment stabilization by the means of flavonoid acylation, or other food characteristics. Furthermore, selectively acylated flavonoids may cause significant changes in their bioavailability and bioactivity, and when consumed, may thus play a role in preventing diseases.

Flavonoid acylation is a useful tool for modification of sensory properties of food. While flavonoids provide a variety of health benefits, flavonoid-containing food often suffers from bitter and astringent taste. Degenhardt et al. (2007) found that certain glycosylation and acylation patterns can effectively modulate these negative taste factors in edible preparations, pharmaceutical preparations and cosmetics with mouth contact (i.e. tooth paste, mouth wash). Both the taste intensity and the taste profile perception are improved by the novel compounds. Ghoul et al. (2006) introduced a process for the selective preparation of acylated flavonoid glycosides with improved stability and solubility in various preparations with their antioxidant effect remaining intact or being improved.

Another particular advantage obtained by these modified flavonoids is the bifunctional character of their molecule with higher biological activity. Free unsaturated fatty acids represent a potential risk because they are highly reactive and by creating free radicals they cause undesirable damage in food. Enzymatic synthesis of flavonoids with unsaturated fatty acids was found to be a useful solution for the stabilization of these highly oxidizable acids (Viskupicova et al., 2010; Mellou et al., 2006).

Another important benefit of acylated anthocyanins lies in the use as food colorants which can serve as a useful alternative to synthetic additives (Giusti & Wrolstad, 2003; Fox, 2000; Asen et al., 1979). The discovery of acylated anthocyanins with increased stability has shown that these pigments may provide food products with the desirable color and stability at a wide pH range. Examples of suitable acylated anthocyanin sources may be radishes, red potatoes, red cabbage, black carrots, and purple sweet potatoes (reviewed in Giusti & Wrolstad, 2003). The invention of Asen et al. (1979) refers to a stable food colorant from a natural source. It relates to an anthocyanin isolated from the Heavenly Blue Morning Glory (*Ipomoea tricolor* Cav cv), peonidin 3-(dicaffeylsophoroside)-5-glucoside, which is characterized by the stability of colors ranging from purplish-red to blue produced in food and beverage products at pH values from about 2.0 to about 8.0. Fox (2000) reported the invention referring to a stable, ruby red natural colorant (anthocyanins acylated with chlorogenic acid) derived from purple sunflower hulls, useful as a coloring agent in food products, cosmetics, pharmaceuticals and other materials.

#### **4.2 Pharmaceuticals**

In recent years, coronary artery diseases, such as atherosclerosis and hypercholesterolemia, represent a major cause of death, exceeding even oncological causes or infectious diseases. Novel acylated flavanone derivatives are effective in the treatment or prevention of elevated blood lipid level-related diseases, e.g. hyperlipidemia, arteriosclerosis, angina pectoris, stroke and hepatic diseases since they exert inhibitory effects on acylcolicholesterol acyl transferase activity and HMG-CoA reductase activity. In spite of their potent efficacies, the flavanone derivatives exhibited no toxicity or mitogenicity in tests using mice (Bok et al., 2001).

Mellou et al. (2005) carried out enzymatic acylation on Greek endemic plants and reported that this modification increased both their antioxidant activity towards isolated low-density lipoproteins (LDL) and serum model and antimicrobial activity against two Gram-positive bacteria, *Staphylococcus aureus* and *Bacillus cereus*. Katsoura et al. (2006) also found that biocatalytic acylation of rutin with various acyl donors affected its antioxidant potential towards both isolated LDL and total serum model *in vitro*. A significant increase in antioxidant activity was observed for rutin-4'''-oleate.

The 6''-*O*-esterification of kaempferol-3-*O*-glucoside (astragalin) with *p*-coumaric acid was found to increase its anti-inflammatory activity eight times compared to the initial flavonoid, while addition of another *p*-coumaroyl group at 2'' position gave an activity 30 times greater than that of astragalin (Harborne & Williams, 2000). Another kaempferol derivative, kaempferol 3-(2'',3''-di-E-*p*-coumaroylrhamnoside), was found to possess a cytotoxic effect. It significantly modulated the proliferation of promyelocytic cell line HL60 and MOLT3 (a T-ALL with phenotypic characteristics of cortical thymocytes) (Mitrokotsa et al., 1993). Also Demetzos et al. (1997) synthesized novel flavonoid esters with cytotoxic activity. These acetylated esters of tiliroside exhibited a strong cytotoxic effect against four leukemic cell lines (HL60, DAUDI, HUT78 and MOLT3), whilst the maternal compound had no effect (Demetzos et al., 1997). Tricin-7-*O*-β-(6''-methoxycinnamic) glucoside, a flavone from sugarcane, was found to exhibit antiproliferative activity against several human cancer cell lines, with higher selectivity toward cells of the breast resistant NIC/ADR line (Duarte-Almeida et al., 2007). Mellou et al. (2006) provided evidence that flavonoid derivatives esterified with polyunsaturated fatty acids were able to decrease the production of vascular endothelial growth factor by K562 human leukemia cells unlike the initial flavonoids, indicating that these novel compounds might possess improved anti-angiogenic and anti-tumor properties. Anticancer acitivity was established also in two *O*-acylated flavonoids, daglesiosides I and II, which were isolated from the leaves of *Pseudotsuga menziesii* (Sharma et al., 2003).

Parejo et al. (2005) examined quercetagetin glycosides acylated with caffeic and *p*coumaric acid for antioxidant activity. They found that these compounds exhibited a high radical scavenging activity in comparison with reference compounds. Fatty acid derivatives of catechins are described as having antitumorigenesis promoting activity or 5-α reductase inhibiting activity, as well as antibacterial activity (Fukami et al., 2007). Since these acylated catechin compounds have a greatly superior solubility in fats and oils than any catechins previously known, they may be used as a highly effective antioxidative agents (Sakai et al., 1994).

A different catechin derivative, 3-*O*-octanoyl-(+)-catechin, was synthesized by Aoshima et al. (2005) by incorporation of an octanoyl chain into (+)-catechin. This ester was found to be more efficient than catechin in inhibiting the response of ionotropic gamma-aminobutyric acid receptors and Na+/glucose cotransporters expressed in *Xenopus oocytes* in a noncompetitive manner. Moreover, it induced a nonspecific membrane current and decreased the membrane potential of the oocyte. This newly synthesized catechin derivative possibly binds to the lipid membrane more strongly than do catechin, (-)-epicatechin gallate, or (-)-epigallocatechin-3-gallate, and as a result it perturbs the membrane structure (Aoshima et al., 2005).
