**2. Phytochemical content of areca nut**

Determination of areca plant as follows:



*Antioxidant Activity of Areca Nut to Human Health: Effect on Oral Cancer Cell Lines… DOI: http://dx.doi.org/10.5772/intechopen.96036*


Areca nut contains several alkaloids that belong to the pyridine group, especially arecoline. Arecoline affects the oculomotor nerve which can cause mydriasis, mild

**Figure 1.** *Areca plants [8].*

**Figure 2.** *The ripe areca nuts*.

paralysis, and pupillary dilatation [15]. Several studies showed that arecoline can induce neurotoxicity through its action in oxidative stress and generating Reactive Oxygen Species (ROS), hepatotoxicity, testicular toxicity, oropharyngeal cancers, and oral submucous fibrosis [16, 17]. Besides alkaloids, ent-catechin, and jacareubin are the major compound in areca nut [7]. Catechin and its analogs are antioxidant, anti-allergic, anticancer, anti mutation, and anti-inflammatory. It can improve liver function and scavenge free radicals [18]. The beneficial effect of catechins is reported in the treatment of cancer, cardiovascular diseases, diabetes, neurodegenerative diseases, and liver diseases [19]. Apparently, catechins have function not only as a powerful antioxidant [20], preventing oxidative damage in healthy cells [21], but also as an antiangiogenic, antitumor agent [22], and a modulator of tumor cell response to chemotherapy [23]. It can induce apoptosis by increasing caspases [24] and promotes cell growth arrest by altering the expression of cell cycle regulatory proteins [25]. Jacareubin is reported to have an anti-inflammatory property and increases the activity of H<sup>+</sup> , K<sup>+</sup> -ATPase which is needed to gain the energy of the cells [26, 27].

The majority of phytochemical ingredients in areca nut extract are phenolic compounds such as flavonoids, tannins, and alkaloids [28]. It was established that a great quantity of tannins has been found in dark dry fruits such as tea. The areca nut also has a reddish-brown color. The levels of tannin found in areca nut have different contents in different areas or regions [29]. It could be caused by climate conditions and environmental factors where it grows. Climatic factors such as temperature, weather, and rainfall. The environmental factors such as soil type and fertility, the height of growing, and plant maintenance.

The presence of catechin and quercetin mostly were identified through High-Performance Liquid Chromatography (HPLC) analysis. These two compounds are well-known antioxidants and could have contributed to the observed antioxidant activity [30]. Previous studies have identified several phenolic compounds in the areca nuts including trimer procyanidin, dimer procyanidin (B1 dan B2), catechin, and isorhamnetin 3-O-rutinoside [31]. Many of these compounds have antioxidants activities [20, 32]. Catechin was proved to have strong antioxidant activity which could contribute towards the anti-cancer effects [33–36]. It has also been shown that polymerized catechin suppresses the activity of *Staphylococcus aureus* α-toxin and as an effective urease inhibitor in *Staphylococcus saprophyticus* strains. Although the level of quercetin is not much in areca nut extract, it also has been proved as an antioxidant. It possesses an anti-inflammatory potential that can be expressed in different cell types, both in animal and human models [37]. Quercetin is also able to inhibit the growth of cancer cells through induction of apoptosis and inhibitory proliferation in gastrointestinal, breast, esophagus, and ovary cancer [38].

### **3. Quantitative data of polyphenolic content in areca nut**

Various studies have been conducted to calculate the content of polyphenolic areca nut from various regions around the world. The results also showed various results. This difference shows that the variation in polyphenolic content depends on the geographical conditions in which the plant grows so that it affects the quality and quantity of the phytochemical composition in areca nut. This variation is also influenced by the species and type of the areca nuts used such as freshness, maturity, and form of the nut drying process. One of the polyphenolic content derived from areca nut on the island of Sumatra in Indonesia is shown in **Table 1**. The total phenolic and flavonoid contents of areca nut are expressed as Tannin Acid Equivalents (TAE) and Catechin Equivalents (CE).


*Antioxidant Activity of Areca Nut to Human Health: Effect on Oral Cancer Cell Lines… DOI: http://dx.doi.org/10.5772/intechopen.96036*

#### **Table 1.**

*Quantitative data of polyphenolic content in areca nut from Aceh, Indonesia.*

#### **4. Molecular structure of bioactive compounds in areca nut**

#### **4.1 Phenolic compounds**

Phenolic compounds are the largest part of the phytochemicals in plants, especially fruit, seeds, and leaves. This collection of compounds provides many health benefits because they contain many antioxidants that can scavenge oxidative stress due to reactive oxygen species. Intake of fruit, vegetables, and whole grains that are rich in phenolics can lower the risk of cardiovascular disease, chronic inflammation, cancer, and neurodegenerative diseases. Polyphenolics contain an aromatic hydroxyl ring derived from L-phenylalanine. Phenolic compounds comprise one (phenolic acids) or more (polyphenols) aromatic rings with attached hydroxyl groups in their structures. Several types of polyphenols that are classified as phenolic acids are hydrolyzed tannins, lignans, stilbene, and flavonoids. The intact polyphenolic form is usually absorbed directly from the digestive tract.

Microorganisms can cause the degradation of polyphenolics into aglycones and aromatic acids. Polyphenolics are detected in all tissues, especially the digestive tract and oral mucosa. All types of polyphenolics are excreted in the urine and bile ducts. The bioavailability of polyphenolics is low, and the remaining at urinary excretion is 0.3% for anthocyanins and 43% for isoflavones. The absorption of phenolic compounds in food is determined by the chemical structure which influences the degree of glycosylation and acylation, basic structure, conjugation with other phenolic compounds, molecular size, polymerization, and solubility. The maximum concentration in plasma rarely exceeds 1 mM after ingestion of 10–100 mg of a single phenolic compound. Polyphenolics that are mostly absorbed by the body are isoflavones, gallic acid, flavanones, catechins, and quercetin glycosides, while proanthocyanidins, anthocyanins, and gallolylated catechins are less widely absorbed.

#### **4.2 Flavonoid**

The total flavonoids in ripe areca nuts were found to be the highest in levels. Flavonoids belong to a group of natural substances with variable phenolic structures and are found in vegetables, tea, flowers, fruit, grains, vegetables, grains, and wine. This natural ingredient is known to have many health benefits. Flavonoids are a group of more than 4000 polyphenolic compounds that occur naturally in foods of plant origin. These compounds possess a common phenylbenzopyrone structure (C6-C3-C6), and they are categorized according to the saturation level and opening of the central pyran ring, mainly into flavones, flavanols, isoflavones, flavonols, flavanones, and flavanonols. Flavonoids are polyphenolic compounds that consist of several types based on their structure, namely flavonols (quercetin and kaempferol), flavones (luteolin and wogonin), flavanols (catechin, gallocatechin), isoflavones (genistein), flavanones, and flavanonols.

Flavonoids have an antibacterial, anti-inflammatory, antioxidant, allergy, antimutagenic, antiviral, antineoplastic, anti-thrombotic, vasodilatory, and antihepatotoxic activities. Flavonoids can cause cell cycle arrest, thereby inhibiting the activity of Cyclin-Dependent Kinases (Cdks), Phosphorylation Kinases (PKs), and signal transduction of cell proliferation. Flavonoids are also able to modulate Mitogen-Activated PKs (MAPKs). The Cdk enzyme is an enzyme that functions to control the activity of the phosphorylase enzyme which regulates every phase of the cell cycle, especially during DNA replication and chromosome formation. The anti-tumor properties of flavonoids can inhibit the release of prooxidant enzymes, modulate carcinogen metabolism, suppress protein tyrosine kinase activity, antiproliferation, anti-metastasis, inhibit some drug resistance, are antioxidant and anti-angiogenesis, induce apoptosis and cell cycle retention. The chemical structures of flavonoids in areca nut can be seen in **Figure 3**.

## **4.3 Catechin**

Catechins are flavonoids (flavanols) that are included in the polyphenol group, which have high concentrations in vegetables, fruits, and beverages. Catechins contain two aromatic rings and several hydroxyl groups. Catechins are divided into two groups, namely free catechins and esterified catechins. Catechin, gallocatechin, epicatechin, epigallocatechin are non-esterified catechins, whereas epigallocatechin gallate, epicatechin gallate, gallocatechin gallate are classified as esterified. The chemical structure of catechins can be seen in **Figure 4**.

Several *in vitro* studies have proven the role of catechins as protection for several diseases such as degenerative, heart disease, and bacterial infection. The catechins in green tea are able to inhibit carcinogenesis of the skin, lungs, esophagus, stomach, liver, small intestine, colon, and mammary glands in animal experiments.

### **5. The side effect of crude extract of areca nut**

Among the benefits of consuming areca nut, other studies have shown that these nuts are carcinogens so that they can cause oral malignancy lesions. Arecoline is the most common alkaloid found in areca nut which is known to cause cytotoxicity in mammalian cells *in vivo* and cause carcinogenicity. Some of the oral lesions that can be caused by crude areca nuts include:

#### **5.1 Oral submucous fibrosis**

Oral submucous fibrosis (OSF) is a chronic disease that produces hyperkeratosis, epithelial atrophy, tissue fibrosis, and precancerous lesions. Pathological characteristics include chronic inflammation, excessive collagen deposition in the connective tissues below the oral mucosal epithelium, local inflammation in the lamina propria *Antioxidant Activity of Areca Nut to Human Health: Effect on Oral Cancer Cell Lines… DOI: http://dx.doi.org/10.5772/intechopen.96036*

*Chemical structures of the flavonoid family. The areca nut contains flavonol (quercetin) and flavanol (catechin).*

or deep connective tissues, and degenerative changes in the muscles. OSF patients experience a severe burning sensation in the mouth after ingesting spicy foods. Other symptoms of OSF include dry mouth, pain, taste disorders, restricted tongue mobility, trismus, dysphagia, and altered tone. This disease contributes significantly to mortality because of its high malignant transformation rate (1.5–15%). Previous research has shown that the arecoline in areca nuts can induce COX-2 expression in humans. It also suggests contributing to pathogenesis of OSF in betel chewers. Consistently, the elevated expression level of COX-2 has also observed in arecolinetreated HGF-1 cells and primarily cultured HGF cells in the study, suggesting these cells of different origins derived from oral cavity might have similar inflammatory responses upon exposure to arecoline, which in turn promote oral lesions and tumorigenesis.

**Figure 4.** *Chemical structures of catechin. The areca nut contains (+)-Catechin.*
