**2.1 Botanical description**

*Cissus sicyoides* L. or *Cissus verticillata* L., which belongs to the Vitaceae family, is also known as vegetal insulin, cipó-pucá, or puci. It is considered a plant of the Neotropical region and is usually found in the Amazon region. It is a climbing plant, which can reach up to 6 m in length, and presents fleshy articulated branch, alternating leaf of ovate format, pale or yellowish-green flowers, and round fruit, with variations of color from violet to black (**Figure 1**) [19, 20].

#### **2.2 Chemical composition**

The bioactive compounds present in the leaf and stem are represented by carotenoids (α-carotene and β-carotene) [39] and phenolic compounds such as

**Figure 1.** *Description of C. sicyoides parts and the main chemical structure of the antioxidant compounds.*

flavonoids (quercetin 3-O-rhamnoside and kaempferol 3-O-rhamnoside) [21]. But also, three new flavonoid glycosides were found, denominate cissosides I, II, and III (kaempferol 3-O-α-L-(5"-O-acetyl)-arabinofuranosyl-7-O-α-L-rhamnopyranoside, quercetin 3-O-α-L-arabinofuranosyl-7-O-α-L-rhamnopyranoside, and quercetin 3-O-α-L-(5"-O-acetyl)-arabinofuranosyl-7-O-α-L-rhamnopyranoside) [22].

Recently, three different flavonoids were identified (quercetin-3-O-hexoside, quercetin-3-O-deoxyhexoside, and kaempferol-3-O-deoxyhexoside) [23]. In addition, resveratrol (3,5,4′-trihydroxystilbene) [40] and a new benzofuran-type stilbene (cissusin) [22], tannins, coumarins (glycoside 5,6,7,8-tetrahydroxycoumarin-5β-xylopyranoside and sabandin), and steroids (β-sitosterol and 3β-O-β-Dglucopyranosyl sitosterol) were found [21]. The presence of essential oils was also detected [41]. In the supercritical extracts of *C. sicyoides* phytochemical screening obtained from leaves and stems by high-performance thin-layer chromatography (HPTLC), the presence of terpenes, phenolic compounds, and flavonoids was evidenced [24]. In the fruit composition analysis, three anthocyanins were found (delphinidin-3-rutinoside, cyanidin-3-rhamnosyl-arabinoside, and delphinidin-3-rhamnoside) [42]. Therefore, the fruit of this plant may have potential use as a food coloring. **Figure 1** shows the chemical structures of the main antioxidant compounds found in *C. sicyoides*.

#### **2.3 Antioxidant and biological activity**

In *C. sicyoides* polyphenolic profile, we can find flavonoids: the quercetin and kaempferol as the majority and its various isomers. The mechanism of quercetin antioxidant action has been associated mainly with the reduction of ROS/RNS, which is a compound that prevents or retards oxidative stress, which enables the prevention of various chronic diseases [6, 43]. In the study conducted by Crespo et al., treatment with quercetin and kaempferol prevented the production of ROS such as peroxides, superoxide anion, and nitric oxide. These results confirmed the differential protective effect of these flavonoids in the diet against oxidative stress induced by proinflammatory stimuli in parenchymal liver cells [44]. Resveratrol plays an important antioxidant role in reducing hydroxyl radicals, superoxide, and metal-induced radicals, as well as showing antioxidant abilities in ROS-producing cells. Also, it has a protective effect against lipid peroxidation in cell membranes and DNA damage caused by ROS/RNS [45]. Benzofuran is a potent radical scavenger capable of inhibiting lipid peroxidation, its FRS capacity being greater than α-tocopherol [46].

**Figure 2** shows the results of the qualitative analysis of antioxidant activity by HPTLC of C. sicyoides extracts obtained by supercritical extraction (essays 1–15), hexane extract (HE), and ethanolic extract (EE) obtained with conventional extraction by Soxhlet. The plaque was derivatized with DPPH (2,2-diphenyl-1-picrylhydrazyl), and it was possible to detect the presence of yellow spots on the plaque purple bottom resulting from the reduction of the DPPH•; in the presence of antioxidant substances, 2,2-diphenyl-picryl-hydrazine is reduced, losing its purple coloration. The study results confirmed the presence of chemical constituent characteristic of this plant, with antioxidant activity. In the same study, a quantitative determination of the antioxidant activity by the DPPH method was carried out. It was demonstrated that with the extractive methodologies (SFE and Soxhlet) used it was possible to extract with low EC50 values, related to a high antioxidant activity; for EE, the value of EC50 (325.67 g of extract/g of DPPH) is similar to the value of EC50 (404.81 g of extract/g of DPPH) obtained with SFE [24].

In the in vitro antioxidant activity determination by the ABTS method of the *C. sicyoides* aqueous extract obtained by decoction, it was evidenced that the extract has an antioxidant activity of IC50 = 13.0 ± 0.2 μg/ml. These results indicate that

**319**

*Antioxidant and Biological Activity of* Cissus sicyoides and Rosmarinus officinalis *Extracts*

the extract is a potential source of natural antioxidant and may be useful in the prevention of diseases associated with oxidative stress [47]. Thus, the antioxidant activity results of *C. sicyoides* extracts are related to the extraction methods and to

*Qualitative analysis of antioxidant activity by HPTLC of C. sicyoides extracts obtained by supercritical* 

Due to antioxidant properties, *C. sicyoides* has been used by folk medicine to treat rheumatism, epilepsy, stroke, abscesses, arthritis, and diabetes; it has also been used to treat respiratory diseases. Some biological activities are attributed to the plant as anti-inflammatory, antirheumatic, antiepileptic, antihypertensive, antimicrobial, antipyretic, antioxidant, antiallergic, anticancer, and antidiabetic

Several studies point to the application of *C. sicyoides* in the treatment of various diseases, in order to demonstrate the cytotoxic activity of the *C. sicyoides* aqueous extract obtained by decoction against cells human epidermoid carcinoma no. 2 (HEp-2 cells), showing complete inhibition of cell division after 24 h of treatment [51]. Also, the antitumor activity of *C. sicyoides* hydroalcoholic extract obtained by maceration, in animals at doses of 300 and 600 mg/kg in weight, was investigated, being demonstrated that the extract showed an inhibition of tumor activity in sarcoma-180 of 49 and 62% and Ehrlich carcinoma of 69 and 84% [52].

Regarding the plant anti-inflammatory activity, it has been demonstrated that the oral administration of 300 and 500 mg/kg of the *C. sicyoides* stem's aqueous extract obtained by decoction in mice with induced edema has a potent anti-inflammatory activity, and administration of the extract produced an approximately 50% reduction of the induced edema [53]. Resveratrol was indicated as one of the constituents responsible for the anti-inflammatory and antiallergic properties presented by *C. sicyoides* alcoholic extract obtained by maceration [40]*.* However, modern ethnopharmacological use reports that the *C. sicyoides* hydroalcoholic extract by percolation has anti-inflammatory and antidiarrheal actions, due to the abundant presence of flavonol-O-glycoside derivatives of quercetin and kaempferol, which are mainly

In addition, pharmacological effects were detected in the treatment and/or prevention of dysfunctions such as hypertension and vasoconstriction of arteries, veins, and capillaries with aqueous extract of *C. sicyoides*. These compounds act at the membrane level, increasing the calcium entry through the membrane as well as

In the evaluation of the antidiabetic potential of *C. sicyoides*, the effects of leaf tea from the plant were studied; the in vivo experimental model chosen proved to be an appropriate treatment, reducing blood and urine glucose levels [48]. Later, it

responsible for the plant pharmacological effects [23].

acting on the internal calcium deposits [54].

*DOI: http://dx.doi.org/10.5772/intechopen.83733*

*extraction and conventional extraction (Soxhlet) [29].*

the solvent used.

**Figure 2.**

activities [23, 40, 48–50].

*Antioxidant and Biological Activity of* Cissus sicyoides and Rosmarinus officinalis *Extracts DOI: http://dx.doi.org/10.5772/intechopen.83733*

**Figure 2.**

*Antioxidants*

compounds found in *C. sicyoides*.

**2.3 Antioxidant and biological activity**

flavonoids (quercetin 3-O-rhamnoside and kaempferol 3-O-rhamnoside) [21]. But also, three new flavonoid glycosides were found, denominate cissosides I, II, and III (kaempferol 3-O-α-L-(5"-O-acetyl)-arabinofuranosyl-7-O-α-L-rhamnopyranoside, quercetin 3-O-α-L-arabinofuranosyl-7-O-α-L-rhamnopyranoside, and quercetin 3-O-α-L-(5"-O-acetyl)-arabinofuranosyl-7-O-α-L-rhamnopyranoside) [22]. Recently, three different flavonoids were identified (quercetin-3-O-hexoside, quercetin-3-O-deoxyhexoside, and kaempferol-3-O-deoxyhexoside) [23]. In addition, resveratrol (3,5,4′-trihydroxystilbene) [40] and a new benzofuran-type stilbene

(cissusin) [22], tannins, coumarins (glycoside 5,6,7,8-tetrahydroxycoumarin-5β-xylopyranoside and sabandin), and steroids (β-sitosterol and 3β-O-β-Dglucopyranosyl sitosterol) were found [21]. The presence of essential oils was also detected [41]. In the supercritical extracts of *C. sicyoides* phytochemical screening obtained from leaves and stems by high-performance thin-layer chromatography (HPTLC), the presence of terpenes, phenolic compounds, and flavonoids was evidenced [24]. In the fruit composition analysis, three anthocyanins were found (delphinidin-3-rutinoside, cyanidin-3-rhamnosyl-arabinoside, and delphinidin-3-rhamnoside) [42]. Therefore, the fruit of this plant may have potential use as a food coloring. **Figure 1** shows the chemical structures of the main antioxidant

In *C. sicyoides* polyphenolic profile, we can find flavonoids: the quercetin and kaempferol as the majority and its various isomers. The mechanism of quercetin antioxidant action has been associated mainly with the reduction of ROS/RNS, which is a compound that prevents or retards oxidative stress, which enables the prevention of various chronic diseases [6, 43]. In the study conducted by Crespo et al., treatment with quercetin and kaempferol prevented the production of ROS such as peroxides, superoxide anion, and nitric oxide. These results confirmed the differential protective effect of these flavonoids in the diet against oxidative stress induced by proinflammatory stimuli in parenchymal liver cells [44]. Resveratrol plays an important antioxidant role in reducing hydroxyl radicals, superoxide, and metal-induced radicals, as well as showing antioxidant abilities in ROS-producing cells. Also, it has a protective effect against lipid peroxidation in cell membranes and DNA damage caused by ROS/RNS [45]. Benzofuran is a potent radical scavenger capable of inhibit-

ing lipid peroxidation, its FRS capacity being greater than α-tocopherol [46].

EC50 (404.81 g of extract/g of DPPH) obtained with SFE [24].

**Figure 2** shows the results of the qualitative analysis of antioxidant activity by HPTLC of C. sicyoides extracts obtained by supercritical extraction (essays 1–15), hexane extract (HE), and ethanolic extract (EE) obtained with conventional extraction by Soxhlet. The plaque was derivatized with DPPH (2,2-diphenyl-1-picrylhydrazyl), and it was possible to detect the presence of yellow spots on the plaque purple bottom resulting from the reduction of the DPPH•; in the presence of antioxidant substances, 2,2-diphenyl-picryl-hydrazine is reduced, losing its purple coloration. The study results confirmed the presence of chemical constituent characteristic of this plant, with antioxidant activity. In the same study, a quantitative determination of the antioxidant activity by the DPPH method was carried out. It was demonstrated that with the extractive methodologies (SFE and Soxhlet) used it was possible to extract with low EC50 values, related to a high antioxidant activity; for EE, the value of EC50 (325.67 g of extract/g of DPPH) is similar to the value of

In the in vitro antioxidant activity determination by the ABTS method of the *C. sicyoides* aqueous extract obtained by decoction, it was evidenced that the extract has an antioxidant activity of IC50 = 13.0 ± 0.2 μg/ml. These results indicate that

**318**

*Qualitative analysis of antioxidant activity by HPTLC of C. sicyoides extracts obtained by supercritical extraction and conventional extraction (Soxhlet) [29].*

the extract is a potential source of natural antioxidant and may be useful in the prevention of diseases associated with oxidative stress [47]. Thus, the antioxidant activity results of *C. sicyoides* extracts are related to the extraction methods and to the solvent used.

Due to antioxidant properties, *C. sicyoides* has been used by folk medicine to treat rheumatism, epilepsy, stroke, abscesses, arthritis, and diabetes; it has also been used to treat respiratory diseases. Some biological activities are attributed to the plant as anti-inflammatory, antirheumatic, antiepileptic, antihypertensive, antimicrobial, antipyretic, antioxidant, antiallergic, anticancer, and antidiabetic activities [23, 40, 48–50].

Several studies point to the application of *C. sicyoides* in the treatment of various diseases, in order to demonstrate the cytotoxic activity of the *C. sicyoides* aqueous extract obtained by decoction against cells human epidermoid carcinoma no. 2 (HEp-2 cells), showing complete inhibition of cell division after 24 h of treatment [51]. Also, the antitumor activity of *C. sicyoides* hydroalcoholic extract obtained by maceration, in animals at doses of 300 and 600 mg/kg in weight, was investigated, being demonstrated that the extract showed an inhibition of tumor activity in sarcoma-180 of 49 and 62% and Ehrlich carcinoma of 69 and 84% [52].

Regarding the plant anti-inflammatory activity, it has been demonstrated that the oral administration of 300 and 500 mg/kg of the *C. sicyoides* stem's aqueous extract obtained by decoction in mice with induced edema has a potent anti-inflammatory activity, and administration of the extract produced an approximately 50% reduction of the induced edema [53]. Resveratrol was indicated as one of the constituents responsible for the anti-inflammatory and antiallergic properties presented by *C. sicyoides* alcoholic extract obtained by maceration [40]*.* However, modern ethnopharmacological use reports that the *C. sicyoides* hydroalcoholic extract by percolation has anti-inflammatory and antidiarrheal actions, due to the abundant presence of flavonol-O-glycoside derivatives of quercetin and kaempferol, which are mainly responsible for the plant pharmacological effects [23].

In addition, pharmacological effects were detected in the treatment and/or prevention of dysfunctions such as hypertension and vasoconstriction of arteries, veins, and capillaries with aqueous extract of *C. sicyoides*. These compounds act at the membrane level, increasing the calcium entry through the membrane as well as acting on the internal calcium deposits [54].

In the evaluation of the antidiabetic potential of *C. sicyoides*, the effects of leaf tea from the plant were studied; the in vivo experimental model chosen proved to be an appropriate treatment, reducing blood and urine glucose levels [48]. Later, it was demonstrated that treatment of diabetic rats with *C. sicyoides* aqueous extract obtained by decoction, for 7 days (100 and 200 mg/kg), reduced blood glucose levels by 22 and 25%, respectively [49]. However, *C. sicyoides* leaf tea was used to investigate the plant therapeutic efficacy in volunteers who are diabetic and intolerant to glucose. A single dose of tea (1 g of dried leaf powder in 150 ml of water) was used for a period of 7 days. It was observed in people intolerant to glucose that the tea had antidiabetic activity [55].

*C. sicyoides* has antibacterial activity, showing inhibitory capacity against bacteria that cause food poisoning [56], which causes acute effects in the gastrointestinal tract and, in some cases, a high severity that patients come to death (*Bacillus cereus*, *Bacillus subtilis*, *Bacillus megaterium*, *Staphylococcus aureus*, and *Escherichia coli*). In addition, the antifungal activity of plant leaf and stem alcoholic extracts was demonstrated, inhibiting the growth of fungi *Cladosporium sphaerospermum* and *Cladosporium cladosporioides* [57].

Recently, Salazar et al. carried out the biological activity determination of *C. sicyoides* supercritical extract; an in vivo test using a focal cerebral ischemia model was performed, and the extract had shown to have a neuroprotective and anti-inflammatory effect, justifying the use in traditional folk medicine for central nervous system diseases. These effects were associated to the presence of phenolic compounds in the extract [24]. Therefore, the results of these studies justify the traditional use of *C. sicyoides*, pointing to the plant extract potential benefit as a possible alternative medicine in disease treatment.
