Coccoloba uvifera Leaves: Polyphenolic Profile, Cytotoxicity, and Antioxidant Evaluation

Abstract

This study aimed to investigate the chemical composition of Coccoloba uvifera leaves and evaluate the antioxidant and antitumor effects of the total extract and its major metabolites. Four assays were used to determine the antioxidant activity, including radical scavenging abilities of 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS), radical cation, and ferric-reducing power. Additionally, vincristine was used as a reference medication to examine the anticancer activity on Ehrlich aesthete carcinoma cells (EACC). Nine compounds were isolated from C. uvifera leaves aqueous methanol extract. Their structures were identified as gallic acid (1), methyl gallate (2), protocatechuic acid methyl ester (3), protocatechuic acid (4), quercetin 3-O-β-d-glucopyranoside (isoquercitrin, 5), kaempferol 3-O-β-D-neohespridoside (6), myricitrin 4″-O-gallate (7), myricetin 3-O-β-d-glucopyranoside (8), and myricetin 3-O-arabinopyranoside (9). The majority possess noticeable antioxidant and antitumor properties. However, compounds 1, 5, 4, 2, and 7 displayed a strong antioxidant potential in terms of DPPH radical scavenging activity, with values of 85.72 ± 0.30, 82.16 ± 0.20, 81.34 ± 0.20, 79.62 ± 0.29, and 79.34 ± 0.20%, respectively. Compounds 4, 1, 5, 7, and 2 revealed high reducing power activity, with respective values of 1.348 ± 0.043, 1.303 ± 0.011, 1.154 ± 0.020, 1.058 ± 0.032, and 1.056 ± 0.019. Compounds 4 and 1 showed the highest ABTS radical scavenging capabilities (91.90 ± 0.24 and 91.83 ± 0.74%) and ferric-reducing power ability (1979 ± 14.53 and 1965 ± 26.86 μmol Trolox/100 g, respectively). Compound 4 has the highest level of cytotoxicity, resulting in 78.710.21% dead cells.

1. Introduction

Polygonaceae, among the most numerous families of medicinal plants, has about 1,200 species in 48 genera that are widely spread worldwide. Meanwhile, some species stretch from the tropics to the arctic; most species are found in the northern temperate area.¹ Regarding biological processes and secondary metabolites, one of the most intriguing genera in Polygonaceae is Coccoloba. Coccoloba is derived from the Spanish term “coccolobis,” which refers to a particular variety of grape, or the Greek words “kokkos” and “lobos,” which allude to the grape-like fruits’ pods or lobes.²Coccoloba plants are commonly identified by their simple, alternate leaves and characteristic ochrea.³ This genus of plants is used all around the world in conventional folk medicine.

Numerous different kinds of secondary metabolites, such as volatile oils, anthraquinones, phenolic acids, flavonoids, triterpenes, diterpenes, and anthraquinones, were identified from this genus. Different biological effects have been reported from Coccoloba spp., such as antioxidant, antimicrobial, cytotoxic, genotoxic, mutagenic, anti-inflammatory, hypoglycemic, and photoprotective properties. In Brazil, certain Coccoloba species are employed as an astringent for treating gonorrhea, hemorrhoids, uterine hemorrhages, diarrhea, and menstrual irregularities.⁴ In folk medicine, it is reported that C. mollis is helpful for various conditions, including sexual impotence, anemia, insomnia, memory loss, diminishing eyesight, stress, and tension.⁵ Native Americans prepared medicinal teas from the leaves, bark, and roots of C. uvifera. The roots, bark, and wood of the plant were all used to make astringent decoctions and juices to cure diarrhea, hemorrhages, dysentery, and venereal infections. They are used externally to treat skin disorders like rashes. Asthma and hoarseness were treated with leaves, and they were also used to cleanse wounds. The bark resinous gum was utilized to treat throat conditions.⁴ Furthermore, the antibacterial, antifungal, poisonous, and phototoxic properties of the methanol extract of C. uvifera seeds were evaluated.⁶ The interest in C. uvifera has substantially expanded recently as a result of various research studies demonstrating its medicinal benefits and the fact that it contains a variety of active compounds. Ascorbic acid, anthocyanins, phenolic compounds, and flavonoids have all been found to be present in the fruit of C. uvifera and to function as antioxidants by scavenging free radicals,⁷ while the plant’s seeds contain gallic acid, an organic acid called hexenedioic acid, and a benzopyran with both antifungal and antibacterial properties.⁸ From the leaves of C. uvifera, physcion, rhein, royleanone, α-amyrin, and β -sitosterol have been isolated,⁹ along with myricetin 3-O-rhamnoside, quercetin 3-O-rhamnoside, and quercetin 3-O-arabinoside as flavonoids.¹⁰

Reactive oxygen species (ROS) accumulation in the intracellular environment causes oxidative stress, which damages biomolecules and is a defining feature of serious diseases, such as cancer, aging, diabetes, heart disease, and neurological disorders. Because of the mitochondria’s electron transport chain, ROS are continuously produced.¹¹ Oxidative stress has been implicated in physiological aging,¹² diabetes,¹³ the development of neurodegenerative conditions like Parkinson’s and Alzheimer’s,¹⁴ cardiovascular diseases,¹⁵ and cancer.¹⁶ Antioxidants are crucial in ensuring the survival of aerobic species by combating oxidative stress. When antioxidants are present, they serve as electron donors, converting highly reactive species into their stable reduced forms.¹⁷ The three synthetic antioxidants butylated hydroxyanisole (BHA), propyl gallate (PG), and butylated hydroxytoluene (BHT) have only been used rarely owing to their insufficient solubility, adverse health effects, and moderate antioxidant capacity.¹⁸ Accordingly, there is a growing market for natural antioxidants because of their probable biosafety and positive benefits for human health.^19,20 An efficient alternate supply of antioxidant and anticancer minerals may be a herbal medicine. Many people prefer herbal treatments to conventional artificial treatments because of their low cost, effectiveness, and safety. Different chemical components found in the genus Coccoloba were discovered by a literature review on that genus. However, since C. uvifera is a rich source of bioactive constituents that contribute to a wide range of medicinal activities and there is a shortage of scientific data regarding the antioxidant and cytotoxic activities of C. uvifera leaves, more extensive phytochemical and biological investigation is required. Therefore, the primary goal of this study is to assess the antioxidant and anticancer effects of the significant polyphenolic metabolites and total leaf extract of C. uvifera.

2. Results and Discussion

2.1. Chemistry

2.1.1. Identification of Isolated Compounds

Using consecutive column chromatography, nine compounds were isolated from C. uvifera leaves and expected to be four derivatives of phenolic acid (1–4) besides five flavonol 3-O-glycosides (5–9) consistent with their chromatographic characteristics (fluorescence in short/long UV light, R_f values and their changes with NH₃ vapors, AlCl₃, FeCl₃, and Naturstoff spray reagents).²¹⁻²⁴ The first group was expected to be of gallic and protocatechuic acid-like structures, appearing only in short UV as blue-shining violet spots that enhanced in NH₃ vapors, AlCl₃, and Naturstoff spray reagents and transformed into blue color with FeCl₃ reagent. Unambiguously, the separated compounds were established as gallic acid (1), methyl gallate (2), protocatechuic acid methyl ester (3), and protocatechuic acid (4) based on the splitting patterns and their values in ¹H and APT ¹³C NMR that sorted quaternary (Q)/CH₂ carbons upward and CH/CH₃ carbons downward (Figures 1 and S1–S7; Table 1).

Figure 1.

Structural formulas of isolated metabolites (1–9).

Table 1. ¹H and APT ¹³C NMR Spectral Data of Compounds 1–4 (400/100 MHz, DMSO-d₆).

	1		2	3		4
no.	δH	δC	δH	δH	δC	δH	δC
1		121.2(Q)			122.3(Q)		122.1(Q)
2	6.91 s	109.4c	6.92 s	7.34 br s	116.6(CH)	7.33 d (2.0)	116.9(CH)
3		145.9(Q)			145.6(Q)		145.9(Q)
4		138.5(Q)			150.9(Q)		150.9(Q)
5		145.9(Q)		6.80 d (8.5)	116.8(CH)	6.80 d (8.5)	116.7(CH)
6	6.91 s	109.4(CH)	6.92 s	7.42 br d (8.5)	121.1(CH)	7.29 dd (8.5, 2.0)	122.3(CH)
7		168.2(Q)			166.7(Q)		167.5(Q)
-CH3			3.70	3.72 s	52.1(CH3)		-

However, the second group was detected on the PC chromatogram as deep purple spots that changed into yellow fluorescence with NH₃ vapors and AlCl₃, while all were transformed into green color with FeCl₃ to expect flavonol 3-O-glycoside structures. In addition, their spots changed into orange (5), greenish-yellow (6), or red (7–9) fluorescence with the Naturstoff spray reagent to confirm the aglycones as quercetin, kaempferol, or myricetin, respectively. Like in the case of the first group, their structures were confirmed as quercetin 3-O-β-d-glucopyranoside (isoquercitrin, 5), kaempferol 3-O-β-D-neohespridoside (6), myricitrin 4″-O-gallate (7), myricetin 3-O-β-d-glucopyranoside (8), and myricetin 3-O-arabinopyranoside (9), by complete interpretation of the corresponding spectral information (Figures 1 and S8–S17, Table 2) (using Bruker spectrometer, 500, 400 MHz for ¹H and 125, 100 MHz for ¹³C NMR) together with the literature.²⁵⁻²⁷

Table 2. ¹H and APT ¹³C NMR Spectral Data of Compounds 5–9 (400/100 MHz, DMSO-d₆).

	5		6		7		8		9
no.	δH	δC	δH	δC	δH	δC	δH	δC	δH	δC
2		156.9 (Q)		156.4 (Q)		157.9 (Q)		156.72 (Q)		156.6 (Q)
3		133.7 (Q)		133.2 (Q)		134.7 (Q)		133.9 (Q)		133.0 (Q)
4		177.8 (Q)		177.4 (Q)		178.2 (Q)		177.9 (Q)		177.8 (Q)
5		161.5 (Q)		161.2 (Q)		161.7 (Q)		161.7 (Q)		161.7 (Q)
6	6.20 d (2.0)	99.2 (CH)	6.22 d (1.8)	98.8 (CH)	6.20 d (1.5)	99.1 (CH)	6.18 br s	99.1 (CH)	6.19 d (2.0)	99.0 (CH)
7		164.4 (Q)		164.6 (Q)		164.6 (Q)		164.5 (Q)		164.3 (Q)
8	6.41 d (2.0)	94.0 (CH)	6.44 d (1.8)	93.7 (CH)	6.38 d (1.5)	94.0 (CH)	6.36 br s	93.8 (CH)	6.38 d (2.0)	93.5 (CH)
9		156.8 (Q)		156.2 (Q)		156.9 (Q)		156.67 (Q)		156.5 (Q)
10		104.4 (Q)		103.8 (Q)		104.5 (Q)		104.4 (Q)		104.5 (Q)
1′		123.1 (Q)		121.1 (Q)		120.1 (Q)		120.5 (Q)		120.0 (Q)
2′	7.58 m	115.7 (CH)	8.02 d (9.0)	130.8 (CH)	6.88 s	108.4 (CH)	7.19 s	109.0 (CH)	7.19 s	108.8 (CH)
3′		145.1 (Q)	6.86 d (9.0)	115.1 (CH)		145.8 (Q)		145.8 (Q)		145.9 (Q)
4′		148.8 (Q)		159.9 (Q)		136.9 (Q)		137.1 (Q)		136.7 (Q)
5′	6.85 d (9.0)	116.6 (CH)	6.86 d (9.0)	115.1 (CH)		145.8 (Q)		145.8 (Q)		145.9 (Q)
6′	7.58 m	122.1 (CH)	8.02 d (9.0)	130.8 (CH)	6.88 s	108.4 (CH)	7.19 s	109.0 (CH)	7.19 s	108.8 (CH)
1″	5.48 d (7.0)	101.4 (CH)	5.48 d (7.5)	101.1 (CH)	5.19 br s	102.4 (CH)	5.47 d (7.7)	101.2 (CH)	5.34 d (6.9)	101.2 (CH)
2″	3.30–3.80 m	74.3 (CH)	3.30–3.80 m	76.9 (CH)	3.30–3.80 m	70.8 (CH)	3.30–3.80 m	74.4 (CH)	3.30–3.80 m	74.2 (CH)
3″		76.7 (CH)		76.2 (CH)		71.0 (CH)		77.0 (CH)		76.0 (CH)
4″		70.1 (CH)		70.4 (CH)		74.7 (CH)		70.4 (CH)		69.0 (CH)
5″		77.5 (CH)		77.1 (CH)		70.4 (CH)		78.1 (CH)		65.8 (CH)
6″		61.2 (CH2)		60.7 (CH)	0.84 d (6.0)	17.9 (CH3)		61.5 (CH2)
1‴			5.26 br s	103.9 (CH)		120.9 (Q)
2‴			3.30–3.80 m	71.1 (CH)	6.91 s	109.2 (CH)
3‴				71.3 (CH)		146.2 (Q)
4‴				72. Nine (CH)		138.4 (Q)
5‴				68.8 (CH)		146.2 (Q)
6‴			1.00 d (6.0)	16.9 (CH3)	6.91 s	109.2 (CH)
7‴						167.9 (Q)

Compounds 1 and 8 were previously isolated from the plant, whereas compound 2 was isolated from Coccoloba parimensis. Compounds 3, 4, 5, 6, 7, and 9 have been isolated for the first time from C. uvifera leaves.²⁸

2.2. Biological Evaluation

2.2.1. Antioxidant Activity of Total Extract and Isolated Compounds

The antioxidant effect of the C. uvifera leaf total extract and its isolated pure compounds was determined using different assays (Figure 2A–D). All investigated samples were found to have good antioxidant activity. The IC₅₀ values of DPPH radical scavenging activity (Figure 2A, Table 3) showed that compounds 1, 5, 4, 2, and 7 possess a high antioxidant potential with 14.58, 15.21, 15.36, 15.70, and 17.15 μg/mL at a concentration of 25 μg/mL. The mean percentage of DPPH radical scavenging activity of the standard synthetic BHA was 91.44 ± 0.29 with IC₅₀ (13.67 μg/mL). On the total extracts, 6 and 3 recorded moderate DPPH radical scavenging activity (44.36 ± 0.19, 51.31 ± 0.36, and 69.34 ± 0.22%).

Figure 2.

Antioxidant activity of the total extract and compounds 1–9 of C. uvifera using different antioxidant assays: (A) Scavenging ability on DPPH radical, (B) reducing power, (C) scavenging ability on ABTS radicals, and (D) antioxidant capacity FRAP assay. Data are the mean ± standard deviation of triplicate experiments.

Table 3. IC₅₀ Values of DPPH and ABTS Radical Scavenging Activity of the Total Extract and Compounds 1–9 of C. uvifera.

	IC50 (μg/mL)
samples	DPPH	ABTS
1	14.58	13.61
2	15.70	17.80
3	18.03
4	15.36	13.60
5	15.21	16.49
6	48.72	40.64
7	17.15	21.02
8	20.34
9
total extract
BHA	13.67	14.13

The current paper describes an in vitro antioxidant investigation that was conducted utilizing four different assays; DPPH, ABTS, ferric-reducing power, and radical cation. All data are the mean ± standard deviation of triplicate experiments. IC₅₀ of DPPH and ABTS radical scavenging activity were calculated and are illustrated in Table 3.

The DPPH method is based on the spectrophotometric measurement of its concentration change resulting from the reaction with an antioxidant species. This test is easy, quick, affordable, and sensitive. It mostly serves the purpose of measuring antioxidants in complex systems or with low activity. However, the fundamental drawback of the test is that DPPH radicals are absent in human cells, which causes an unphysiological likeness to radicals.²⁹

Concerning the reducing power assay, increasing the absorbance at 700 nm reflects the increase in the reducing power capacity (Figure 2B). Compounds 4, 1, 5, 7, and 2 exhibited high reducing power activity 1.348 ± 0.043, 1.303 ± 0.011, 1.154 ± 0.020, 1.058 ± 0.032, and 1.056 ± 0.019, respectively, that was higher than that recorded by the equivalent concentration (25 μg/mL) of the reference BHA (0.975 ± 0.013). The effects of the ABTS process were expressed as a percentage of ABTS radical scavenging activity.

With a relatively minimal concentration (25 μg/mL), compounds 4 and 1 showed very high ABTS radical scavenging capacities of 91.90 ± 0.24 and 91.83 ± 0.74% with IC₅₀ 13.61 and 13.60 μg/mL, respectively (Figure 2C, Table 3), which were found to be higher than that recorded by the standard BHA (88.42 ± 0.24%, 14.13 μg/mL). On the other hand, good ABTS radical scavenging ability was noticed in compounds 5, 2, and 7; they recorded 75.79 ± 0.21, 70.22 ± 0.24, and 59.44 ± 0.21% radical scavenging capacities at the concentration of 25 μg/mL, whereas compound 6 gave 61.51 ± 0.46% at 50 μg/mL. Concerning ferric-reducing power capacity, the results were at the same trend as DPPH radical scavenging activity (Figure 2A), where compounds 4 and 1 exhibited the highest ferric-reducing power ability (1979 ± 14.53 and 1965 ± 26.86 μmol Trolox/100 g). Compounds 5, 2, and 6 also possess a relatively high ferric-reducing power activity (1567 ± 19.04, 1131 ± 21.73, and 1113 ± 17.95 μmol Trolox/100 g, respectively). The lowest ferric-reducing power activity was recorded by compound 9 (643 ± 19.08) at a concentration of 100 μg/mL. It can be concluded that most of the isolated compounds of C. uvifera possess high antioxidant activity and could be considered to be promising antioxidant agents. A FRAP test is employed to assess the metabolites reducing power. Unlike the DPPH assay, this assay is based on the idea of raising the reaction mixture’s absorbance. Greater absorbency translates into a greater extract redox potential.³⁰ It has been found that phenolics are potent antioxidants that can neutralize free radicals by offering an electron or an atom of hydrogen. By preventing or inactivating the creation of active species precursors, they can reduce the oxidation rate and suppress the production of free radicals. Phenolics are known as metal chelators, in addition to their ability to neutralize free radicals. Chelation of transition metals, such as Fe²⁺, can stop the Fenton reaction from occurring as quickly, preventing the oxidation brought on by highly reactive hydroxyl radicals.^31,32

Additionally, flavonoids have powerful chelating and antioxidant effects. Flavonoids’ ability to deliver electrons to free radicals, bind metal catalysts, initiate antioxidant enzymes, and suppress oxidases is thought to be the cause of their protective actions in biological systems.³³ Notably, glycosylation reduces the antioxidant action of flavonoids, such as quercetin. Although the amount of antioxidant activity is not dependent on the number of galloyl groups, the presence of a galloyl group in the molecule significantly influences this activity.³⁴

2.2.2. In Vitro Antitumor Activity

The antitumor activity of the investigated pure compounds (100 μg/mL) was determined on Ehrlich ascites carcinoma cells (EACC). Total extract and most of the tested compounds exhibited antitumor activity and affected the ability of Ehrlich ascites carcinoma cells to survive (Figure 3). Compound 4 possesses the greatest impact on EACC viability in terms of cytotoxicity, with 78.71 ± 0.21% dead cells, followed by compounds 1, 5, and total extract; they recorded 75.36 ± 0.25, 73.26 ± 0.36, and 66.47 ± 0.31%, respectively, in comparison to the standard drug vincristine (Figure 3), which recorded 90.64 ± 0.39% dead cells at the same concentration (100 μg/mL). Compound 7 displayed a medium-level tumor-fighting capacity with a 61.45 ± 0.27% reduction in the viability of EACC, while the lowest cytotoxicity effect was found with compounds 9 and 6 (8.32 ± 0.20 and 12.26 ± 0.12%). Flavonoids’ double bond (C2=C3) significantly participates in ring C and A/B conjugation and the molecular planarity necessary for effective tumor suppression. Thus, flavonol compounds such as kaempferol have been shown to involve different molecular mechanisms to contribute to anticancer activity.³⁵ But by introducing a hydroxyl group at C3, the cytotoxicity was reduced when comparing the flavonol’s activity to other flavonoids. The causes of the antiproliferative effect’s reverse and null results are still unknown.³⁶

Figure 3.

Effect of the total extract and compounds 1–9 of C. uvifera on the viability of EACC compared with vincristine. Values are the averages and standard deviations for three independent experiments.

Although it has been discovered that phenolic substances have antioxidant and antitumor activities, their effectiveness varies due to differences in their structural nature. For instance, investigations on the link between structure and activity have shown the importance of aromatic rings and hydroxyl groups in demonstrating phenolic compounds’ powerful antioxidant and anticancer properties. Better anticancer activity was seen in compounds with more hydroxyl groups than those with –OCH₃ moieties or no hydroxyl groups. For instance, it has been claimed that gallic acid, which has three hydroxylic groups, is more efficient than monohydroxy benzoic acid.^37,38 Moreover, it was shown that methyl gallate, which has (3OH) groups besides a group of carboxylic acids ester, is less active than gallic acid, which has a carboxyl group plus (3OH) groups.

Additionally, methyl gallate’s methoxy group may prevent the hydroxyl groups from scavenging by forming intra- or intermolecular hydrogen bonds.³⁹ Protocatechuic acid (3,4-dihydroxy benzoic acid) reportedly has antioxidant, anti-inflammatory, and antibacterial properties and protects the liver.⁴⁰ However, it was shown that protocatechuic acid methyl ester is less active than protocatechuic acid.

3. Experimental Section

3.1. Chemistry

3.1.1. Plant Material

Coccoloba uvifera fresh leaves were collected in March 2019 from the Orman Botanical Garden, Giza, Egypt. The plant was authenticated by Dr. Therisa Labib, doctor of plant taxonomy, Orman Botanical Garden, Giza, Egypt. A voucher specimen (Reg. no. Cu-3-2019) was kept at the herbarium of the Pharmacognosy Department, Faculty of Pharmacy, AL-Azhar University (Girls), Cairo, Egypt.

3.1.2. Extraction and Isolation

Briefly, 1.3 kg of powdered, air-dried C. uvifera leaves was thoroughly extracted with 70% methanol (4 × 3 L) under reflux. A sticky crude extract (700 g) (53.8% of the dried powder) was produced by vacuum-evaporating (Buchi G., Switzerland) the combined methanol extract at 50 °C. It was defatted by CH₂Cl₂ to produce an amount of 98 g (7.5% of the dried powder) dry-soluble portion and 590 g (45.4% of the dried powder) residue that is chromatographed on a polyamide column using water first and then a gradient of water and methanol (MeOH) up to 100%. Eight collective fractions were created via TLC inspection of the individual fractions, detection with VIS/UV light, and spray reagents. These were successively fractionated and purified using Sephadex LH-20, silica gel, or cellulose columns with several suitable solvent systems, yielding nine pure metabolites (1–9) as mentioned in the isolation scheme (Supporting Information).

3.2. Biological Evaluation

3.2.1. Materials for Biology

The following chemicals were obtained from Sigma Chemical Company: DPPH, BHA, 2,4,6-tripyridyl-s-triazine (TPTZ), potassium ferricyanide (C₆N₆FeK₃), trolox, ferric chloride (FeCl₃), and 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonate), while ABT was obtained from St. Louis, MO. All other materials employed in the current in vitro tests were of analytical grade and obtained from Sigma, Merck, and Aldrich.

3.2.2. Antioxidant Activity Investigation Assays

The antioxidant activity (DPPH, reducing power capability, FRAP, and ABTS radical scavenging) assays of the investigated compounds (1–9) and the total extract of C. uvifera were determined according to techniques outlined by Rice-Evans et al. and Re et al.⁴¹⁻⁴⁵

3.2.3. In Vitro Antitumor Activity/Cytotoxicity Assay Using EACC

Microscopically counting viable cells, cytotoxicity, and cell viability tests were performed. Cytotoxicity was measured in tumor cells suspended from the peritoneal cavities of mice with tumors. The optimal concentration of the plant extract was assessed. Once the tumor cells were incubated with the isolated pure compounds, the viability percentages of the cells were calculated. Vincristine was used as a standard drug. The antitumor effect of examined compounds and total extract (100 μg) was determined according to Ahmed et al. and Haroun and Taie (2014).^45,46

4. Conclusions

Nine polyphenols were identified in the C. uvifera leaves, four of which are phenolic acid derivatives, including gallic and protocatechuic acids and their methyl esters. In addition, five flavonol 3-O-glycosides were isolated based on quercetin, kaempferol, and myricetin aglycones. The biological findings proved that after applying safety studies, most of the isolated compounds and total extract of C. uvifera leaves possess noticeable antioxidant and antitumor activities, and compounds 1, 5, 4, 2, and 7 displayed a strong antioxidant potential in terms of DPPH radical scavenging activity, with values of 85.72 ± 0.30, 82.16 ± 0.20, 81.34 ± 0.20, 79.62 ± 0.29, and 79.34 ± 0.20%, respectively. Compounds 4, 1, 5, 7, and 2 revealed high reducing power activity, with respective values of 1.348 ± 0.043, 1.303 ± 0.011, 1.154 ± 0.020, 1.058 ± 0.032, and 1.056 ± 0.019. Compounds 4 and 1 showed the highest ABTS radical scavenging capabilities (91.90 ± 0.24 and 91.83 ± 0.74%) and ferric-reducing power ability (1979 ± 14.53 and 1965 ± 26.86 μmol Trolox/100 g, respectively). Compound 4 has the highest level of cytotoxicity, resulting in 78.710.21% dead cells. This study’s findings expand the pharmaceutical industry’s usage of C. uvifera as a therapeutic agent.

Glossary

Abbreviation

ABTS

2,2′-azinobis-3-ethylbenzothiazoline-6-sulfonic acid

APT

attached proton test

BHA

butylated hydroxyanisole

DPPH

2,2-diphenyl-1-picrylhydrazyl

EACC

Ehrlich ascites carcinoma cells

FRAP

ferric-reducing antioxidant power

PC

paper chromatogram

TLC

thin-layer chromatography

Data Availability Statement

The manuscript includes all of the information needed to support the study’s conclusions.

Supporting Information Available

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acsomega.3c04025.

• ¹H and APT ¹³C NMR spectra for all compounds (Figures S1–S17) and additional isolation scheme (PDF)

Author Contributions

S.A.E., N.A.S., H.A.T., and M.M. conceived and constructed the study proposal and wrote the original draft; H.T.A. performed the antioxidant and antitumor investigation; S.A.E. and N.A.S. conducted the practical separation section and helped with data gathering; S.A.E., N.A.S., H.A.T., and M.M. analyzed the data during the investigation and interpreted the results. All authors have read and permitted the final submitted manuscript.

This research did not receive any external funding.

The authors declare no competing financial interest.

Notes

Totally techniques were accompanied in obedience to the animal care and use committee’s ethical standards and guidelines at the Faculty of Pharmacy, Al-Azhar University, Egypt (340).