Abstract
A large common species, Syzygium aqueum, belonging to the genus Syzygium possesses numerous bioactive phytochemical constituents. Moreover, the different parts of this species have been used as folk medicine since centuries ago. In this study, a phytochemical exploration was carried out on the plant’s stem bark. Isolation of the compounds was carried out through the extraction step with some organic solvents, followed by separation and purification using chromatography techniques until the two triterpenoids were isolated from nonpolar and semipolar extracts. Structure elucidation was done using spectroscopic methods. These compounds were identified as alphitolic acid and arjunolic acid. Subsequently, these two compounds were used in anticancer tests against human cancer cells HeLa, T47D, and A549 using colorimetric assay. The result showed that both compounds showed more inhibition of the growth of HeLa and T47D than A549 cancer cells, with the highest activity shown by arjunolic acid against HeLa cell lines.
1 Introduction
One of the probable medicament manufacturing countries, Indonesia, between the Pacific and Indian Oceans, is commonly known as a country with the second richest biodiversity of medicinal plants on the earth. Medicinal plants have therapeutic properties or provide beneficial pharmacological effects for the human body. Some examples are given here. The chemical composition of the essential oils extracted from Elaeagnus umbellata Thunb fruit demonstrated antioxidant, anticholinesterase, and antidiabetic activities that could be used as an alternative drug to treat oxidative stress-related diseases [1]. The essential oil of Teucrium stocksianum possesses a strong antinociceptive potential, which is further used as a topical analgesic [2]. The study of Woodwardia unigemmata (Makino) Nakai plant extracts, which were rich in polyphenolic compounds (total phenolic compounds and total flavonoid compounds) with efficient antioxidant activity, also showed remarkable antibacterial activity against plant and animal pathogenic bacteria [3]. The diverse Indonesian medicinal plants have been used as the sources of herbal medicines for the inhibition and treatment of human diseases for several thousand years [4].
The sixteenth largest genus of flowering plants in the Myrtaceae family, Syzygium, comprises about 1,200–1,800 species. Moreover, Syzygium species of tree and shrub types are distributed in tropical areas worldwide, especially Indonesia, Malaysia, Philippines, and Thailand. Some species of Syzygium are widely cultivated for economic importance, edible fruits, spices, flavoring agents, and pharmacological proprieties. Syzygium contains abundant diversity of bioactive phytochemical constituents such as terpenoids, steroids, chalcones, flavonoids, lignans, alkyl phloroglucinols, hydrolysable tannins, and chromone derivatives [5].
A medicinal species of Syzygium found in Indonesia, Syzygium aqueum, is a large common species in this genus found in Malaysia [6]. Fruits, leaves, and roots of this species have been used as folk medicine, especially for antibiotic properties, diabetic treatment, the treatment for a cracked tongue, relieving itches, stomach aches, dysentery, and reducing of swelling, respectively [7,8,9,10,11].
Some researchers were focused on bioactive phytochemical constituents based on the above medicinal values of this plant. Therefore, bioactive phytochemical constituents such as tannins, flavonoids, steroids, triterpenoids, and phenols from leaves and stem bark of this plant have already been reported in some research articles [12,13,14,15,16,17]. These articles also showed that some flavonoids from S. aqueum (leaves) strongly showed antidiabetic properties using α-glucosidase and α-amylase inhibition assays and showed their cytotoxicity against MCF-cell line using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Moreover, some triterpenoids and steroids from S. aqueum (stem bark) were active with high cytotoxicity on HeLa, T47D, and A549 cancer cell lines using MTT and 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) assays.
Based on the literature study, the stem bark of S. aqueum was designated for this research. Primarily, some extracts of stem bark of S. aqueum were separated to collect bioactive constituents with vacuum liquid chromatography and column chromatography. The structure of isolated compounds was identified with spectroscopic methods, including UV, Fourier transform infrared (FTIR), nuclear magnetic resonance (NMR [1D and 2D]), and HR-DART-MS. After this, their anticancer activity was determined with colorimetric assay (MTT and XTT assays). The isolated bioactive constituents were triterpenoid compounds, first isolated from S. aqueum.
2 Materials and methods
2.1 Plant materials
The stem bark of S. aqueum was collected in December 2018 from Wage, Taman, Sidoarjo, East Java, Indonesia. The plant material was identified at the Department of Biology, Faculty of Science and Technology, Universitas Airlangga, and the voucher specimen (UA-MSa050918) was deposited at the Herbarium of Universitas Airlangga, Laboratory of Biosystematic, Department of Biology, Faculty of Science and Technology, Universitas Airlangga. The collected stem bark was cleaned, chopped, and crushed into small pieces of coarse powder.
2.2 Instrumentations
A polarimeter with φ 3.5 mm diameter (cell size) × 100 mm (cell length) was used to measure the specific rotation. For structure elucidation, some of the instruments used are a UV-vis 1800 spectrometer (Shimadzu), FTIR spectra on a Tracer-100 spectrophotometer (Shimadzu), NMR spectra on Bruker Avance III HD 600, and HR-DART-MS on an Exactive Plus Orbitrap DART Mass Spectrometer. Enzyme-linked immunosorbent assay (ELISA) reader was used for the anticancer test.
2.3 General procedure
For extraction, fractionation, isolation, and purification of a sample, organic solvents such as n-hexane (n-Hex), ethyl acetate (EtOAc), dichloromethane (DCM), and methanol (MeOH) were used. In analytical thin-layer chromatography (TLC), a precoated silica gel 60 F254 (Merck) was used and an anisaldehyde-sulfuric acid reagent was sprayed on a TLC plate for visualization. Silica gel 60 (700–200 mesh ASTM) was applied for column chromatography. Then, vacuum liquid chromatography (VLC) was performed using Kieselgel 60 (F254, Merck). The melting point of pure compounds was measured on a Fisher-Johns melting point apparatus (Stuart SMP30). Spectroscopic methods, including UV-vis, IR, NMR, and MS, were used to determine the structure of isolated compounds. An anticancer test was carried out using MTT and XTT reagents (colorimetric assay), and the absorbance for an anticancer test was recorded on an ELISA reader.
2.4 Extraction and isolation
At room temperature, 2 kg of coarse powder of stem bark was extracted with a polar solvent (MetOH, 40 L) for 3 × 24 h. This crude extract (450 g) was then mentioned to partition with a nonpolar solvent (n-hexane). It was separated into a nonpolar fraction and a methanol fraction. Then, 400 mL of aqueous was added to methanol fractions before partitioning with a semipolar solvent, EtOAc. After this, this mixture was separated into a semipolar fraction and an aqueous methanol residue. First, the nonpolar fraction was chromatographed with gradient polarity solvents, n-Hex:EtOAc. Separation procedure was carried out for three times using column chromatography to obtain a pure compound. Compound 1 (30 mg) was acquired from a fraction eluted with n-Hex:EtOAc = 7:3. The semipolar fraction was separated by VLC using gradient solvent mixtures of n-Hex:DCM and DCM:EtOAc. Separation procedure using column chromatography was carried out twice to obtain a pure compound, Compound 2 (50 mg), that was eluted with DCM:EtOAc = 6:4.
2.5 Anticancer activity
2.5.1 Cell culture
Three cancer cell lines, HeLa (cervical cancer), T47D (breast cancer), and A459 (lung cancer), were obtained from American Type Culture Collection. Cells were cultured at 37°C in a CO2 incubator for 24 h and 100% humidity in medium supplemented with 10% FBS, 1% l-glutamine, and 1% penicillin/streptomycin. The cell culture process was carried out in the Cancer Chemoprevention Research Center, Faculty of Pharmacy at Universitas Gadjah Mada, Indonesia and in Physic Laboratory, Faculty of Science of University Putra Malaysia (UPM), Malaysia.
2.5.2 MTT assay
An anticancer test of pure compounds was performed using MTT assay. Cells were cultured in 96 well plates at 213 × 104 cell/well density and were incubated at 37°C in a CO2 incubator for 24 h. The cells were then treated with 100 µL of the prepared sample with different concentration series (1.5625–100 µg/mL) at 37°C in a CO2 incubator for 24 h. As much as 100 µL of the MTT reagent (50 mg in 10 mL of PBS) was filled in each well after incubation, and it was incubated again at 37°C in a CO2 incubator for 2–4 h until purple formazan crystals are formed. After the crystal formation, cells were observed with a microscope, and 100 µL of 10% SDS stopper in 0.1 N HCl was added, and it was kept in a dark place overnight (room temperature). Next, the plate’s absorbance value was read at a wavelength of 560 nm by using an ELISA reader [18,19].
2.5.3 XTT assay
The anticancer test was carried out using XTT assay. Cells were seeded (104–105 cells/well) into 96 well plates and were incubated at 37°C in a CO2 incubator for 24 h. As much as 100 µL of the prepared sample with various concentrations (1.5625–100 µg/mL) was inserted to treat cancer cells. The plate was incubated at 37°C in a CO2 incubator for 24 h. And then, PMS solution (10 mM PMS solution and PBS (3 mg PMS + 1 mL PBS)) was prepared and 10 µL of this solution was added in 4 mL of XTT (4 mg of XTT dissolved in 4 mL of 37°C cells culture medium). After incubation, 25 µL of the XTT reagent (XTT/PMS solution) was filled in each well. After this, it was incubated for 2–4 h in a CO2 incubator at 37°C until the changing of the orange formazan. The next step was to read the absorbance value at a wavelength of 450 nm using an ELISA reader [19,20]. Each experiment was carried out in triplicate, and the number of viable cells was calculated using the following formula:
3 Results and discussion
3.1 Isolation of compounds and structure elucidation
Nonpolar and semipolar fractions of S. aqueum (stem bark) gave one bioactive compound each, Compound 1 and Compound 2. Structures of these compounds were identified and elucidated using spectroscopic methods.
Compound 1 was acquired as white powder,
DQF-COSY, NOESY, and HMBC correlations of alphitolic acid from S. aqueum.
NMR spectra of alphitolic acid by comparing to literature
| Position | DEPT | Experiment | Literature [21] | ||||
|---|---|---|---|---|---|---|---|
| δ C (ppm) | δ H (ppm) {mult, J (Hz)} | δ C (ppm) | δ H (ppm) {mult, J (Hz)} | ||||
| 1 | CH2 | 46.9 | 0.82 | (1H, m) | 46.5 | — | — |
| 2.00 | (1H, m) | ||||||
| 2 | CH | 68.4 | 3.60 | (1H, m) | 68.6 | 3.46 | (1H, dd, J = 4.0, 9.5 Hz) |
| 3 | CH | 83.0 | 2.89 | (1H, d, J = 9.6 Hz) | 83.2 | 2.74 | (1H, d, J = 9.5 Hz) |
| 4 | C | 39.1 | — | 40.6 | — | ||
| 5 | CH | 55.2 | 0.82 | (1H, m) | 55.2 | — | |
| 6 | CH2 | 18.1 | 1.54 | (1H, m) | 18.2 | — | |
| 1.43 | (1H, m) | ||||||
| 7 | CH2 | 34.1 | 1.43 | (2H, m) | 34.1 | — | |
| 8 | C | 40.6 | — | 39.2 | — | ||
| 9 | CH | 50.6 | 1.39 | (1H, m) | 50.3 | — | |
| 10 | C | 38.1 | — | 38.2 | — | ||
| 11 | CH2 | 20.8 | 1.43 | (2H, m) | 20.9 | — | |
| 12 | CH2 | 25.4 | 1.73 | (1H, m) | 25.3 | 1.52 | (1H, m) |
| 1.08 | (1H, m) | 1.64 | (1H, m) | ||||
| 13 | CH | 38.2 | 2.32 | (1H, m) | 38.1 | — | |
| 14 | C | 42.3 | — | 40.6 | — | ||
| 15 | CH2 | 29.4 | 1.30 | (1H, m) | 30.4 | — | |
| 1.15 | (1H, m) | ||||||
| 16 | CH2 | 32.0 | 0.90 | (2H, m) | 32.1 | — | |
| 17 | C | 56.1 | — | 56.1 | — | ||
| 18 | CH | 49.1 | 1.61 | (1H, m) | 48.1 | — | |
| 19 | CH | 47.1 | 3.03 | (1H, m) | 48.9 | 2.86 | (1H, m) |
| 20 | C | 150.6 | — | 150.7 | — | ||
| 21 | CH2 | 30.3 | 1.39 | (2H, m) | 29.5 | — | |
| 22 | CH2 | 36.7 | 1.91 | (1H, m) | 37.0 | — | |
| 1.43 | (1H, m) | ||||||
| 23 | CH3 | 27.7 | 0.99 | (3H, s) | 28.2 | 0.81 | (3H, s) |
| 24 | CH3 | 15.8 | 0.78 | (3H, s) | 16.4 | 0.60 | (3H, s) |
| 25 | CH3 | 16.5 | 0.92 | (3H, s) | 17.1 | 0.71 | (3H, s) |
| 26 | CH3 | 15.3 | 0.97 | (3H, s) | 15.7 | 0.80 | (3H, s) |
| 27 | CH3 | 13.7 | 1.01 | (3H, s) | 14.5 | 0.75 | (3H, s) |
| 28 | COOH | 178.7 | — | 179.3 | — | ||
| 29 | CH2 | 108.7 | 4.60 | (1H, s) | 109.4 | 4.42 | (1H, s) |
| 4.71 | (1H, s) | 4.54 | (1H, s) | ||||
| 30 | CH3 | 18.2 | 1.70 | (3H, s) | 19.1 | 1.51 | (3H, s) |
Compound 2 was acquired as white powder, m.p. 338–340°C. The UV spectrum in MeOH appeared at λ max 220 nm. In FTIR spectra (KBr), hydroxyl group (3,371 cm−1), asymmetric and symmetric sp3 C–H stretching vibration (2,941 cm−1), carbonyl (COOH) group (1,693 cm−1), CH2 bending (1,462 cm−1) and CH3 bending (1,388–1,365 cm−1), and C–O stretching (1,049 cm−1) were observed. In the 1H NMR spectrum, four carbinol protons were shown at δ H: 3.68 ppm (H-2β), δ H: 3.35 ppm (H-3α), δ H: 3.27 ppm (H-23), and δ H: 3.50 ppm (H-23). The H-3α and H-2β were coupled through the DQF-COSY spectrum. The trans configuration was confirmed due to the 10.2 Hz coupling constant of H-3α and NOESY correlation of H-2β and H-24β. Thus, H-23 was correlated to C-3 (δ C: 76.8 ppm) according to the HMBC spectrum. The olefinic protons were shown at δ H: 5.25 ppm (H-12) and coupled to H-11 (δ H: 1.95 ppm) in the DQF-COSY spectrum. Moreover, six methyl protons were appeared at δ H: 0.70 ppm (H-24), 1.03 ppm (H-25), 0.82 ppm (H-26), 1.18 ppm (H-27), 0.91 ppm (H-29), and 0.94 ppm (H-30). These methyl protons have correlations observed on the HMBC spectrum, especially between H-24 and C-23 (δ C: 65.0 ppm), H-26 and C-8 (δ C: 39.2 ppm) and C-14 (δ C: 41.6 ppm), H-27 and C-8 (δ C: 39.2 ppm), C-14 (δ C: 41.6 ppm) and C-15 (δ C: 27.4 ppm), and H-29 and H-30 and C-20 (δ C: 30.2 ppm). On the 13C NMR spectrum, the three carbinol carbons were observed at δ C: 68.8 ppm (C-2), 76.8 ppm (C-3), and 65.0 ppm (C-23). The carbons of C-2 and C-3 were identified as methine alcohol (R2-CH-OH), and C-23 was suggested as methylene alcohol (R-CH2-OH) according to DEPT (DEPT 135 and DEPT 90) experiments. Chemical shifts at δ C: 180.7 ppm in the 13C NMR spectrum indicated the presence of the carboxylic acid functional group (C-28), which was correlated with H-18 (δ H: 2.86 ppm) on the HMBC spectrum. The double bond carbons at positions C-12 and C-13 were appeared at δ C: 121.9 ppm (methine carbon) and δ C: 144.1 ppm (quaternary carbon). The remaining quaternary carbon signals were shown at δ C: 42.7 ppm (C-4), 39.2 ppm (C-8), 41.6 ppm (C-14), 46.3 ppm (C-17), and 30.2 ppm (C-20). Carbon signals at δ C: 12.4 ppm (C-24), 16.2 ppm (C-25), 16.4 ppm (C-26), 25.1 ppm (C-27), 32.1 ppm (C-29), and 22.6 ppm (C-30) were suggested as primary carbons. The presence of eight quaternary carbons, six methine carbons, ten methylene carbons, and six methyl carbons was also confirmed by the DEPT experiment. According to the above data, the structure of Compound 2 was verified as arjunolic acid, and its molecular formula was C30H48O5. DQF-COSY (1H–1H), NOESY (1H–1H), and HMBC (1H–13C) correlations of the structure are demonstrated in Figure 2. And then, the spectral data of both 1H NMR (600 MHz, methanol-d 4) with their coupling constants and 13C NMR (150 MHz, methanol-d 4) are shown in Table 2, completed with the reported literature data.
DQF-COSY, NOESY, and HMBC correlations of arjunolic acid from S. aqueum.
NMR spectra of arjunolic acid by comparing to literature
| Position | DEPT | Experiment | Literature [22] | ||||
|---|---|---|---|---|---|---|---|
| δ C (ppm) | δ H (ppm) (mult, J in Hz) | δ C (ppm) | δ H (ppm) (mult, J in Hz) | ||||
| 1 | CH2 | 46.5 | 0.89 | (1H, m) | 47.8 | — | |
| 1.95 | (1H, m) | ||||||
| 2 | CH | 68.8 | 3.68 | (1H, m) | 69.6 | 3.66 | (1H, m) |
| 3 | CH | 76.8 | 3.35 | (1H, d, J = 10.2 Hz) | 78.1 | 3.36 | (1H, m) |
| 4 | C | 42.7 | — | 43.0 | — | ||
| 5 | CH | 46.8 | 1.28 | (1H, m) | 48.3 | — | |
| 6 | CH2 | 17.7 | 1.39 | (1H, m) | 19.1 | — | |
| 1.48 | (1H, m) | ||||||
| 7 | CH2 | 32.0 | 1.28 | (1H, m) | 33.8 | — | |
| 1.64 | (1H, m) | ||||||
| 8 | C | 39.2 | — | 40.7 | — | ||
| 9 | CH | 47.6 | 1.69 | (1H, m) | 48.1 | — | |
| 10 | C | 37.7 | — | 38.5 | — | ||
| 11 | CH2 | 23.2 | 1.95 | (1H, m) | 24.0 | — | |
| 1.24 | (1H, m) | ||||||
| 12 | CH | 121.9 | 5.25 | (1H, m) | 123.4 | 5.23 | (1H, m) |
| 13 | C | 144.1 | — | 144.3 | — | ||
| 14 | C | 41.6 | — | 42.7 | — | ||
| 15 | CH2 | 27.4 | 1.07 | (1H, m) | 29.1 | — | |
| 1.78 | (1H, m) | ||||||
| 16 | CH2 | 22.6 | 1.59 | (1H, m) | 24.4 | — | |
| 2.24 | (1H, m) | ||||||
| 17 | C | 46.3 | — | 46.6 | — | ||
| 18 | CH | 41.4 | 2.86 | (1H, dd, J = 3.9, 13.7 Hz) | 43.3 | — | |
| 19 | CH2 | 45.9 | 1.13 | 2H (m) | 44.1 | — | |
| 20 | C | 30.2 | — | 31.6 | — | ||
| 21 | CH2 | 33.5 | 1.39 | (1H, m) | 34.8 | — | |
| 1.21 | (1H, m) | ||||||
| 22 | CH2 | 32.5 | 1.75 | (1H, m) | 33.5 | — | |
| 1.54 | (1H, m) | ||||||
| 23 | CH2 | 65.0 | 3.50 | (1H, d, J = 11.0 Hz) | 66.2 | 3.50 | (1H, s) |
| 3.27 | (1H, d, J = 11.0 Hz) | 3.31 | (1H, s) | ||||
| 24 | CH3 | 12.4 | 0.70 | (3H, s) | 13.9 | 0.69 | (3H, s) |
| 25 | CH3 | 16.2 | 1.03 | (3H, s) | 17.7 | 0.81 | (3H, s) |
| 26 | CH3 | 16.4 | 0.82 | (3H, s) | 17.5 | 0.91 | (3H, s) |
| 27 | CH3 | 25.1 | 1.18 | (3H, s) | 26.8 | 1.17 | (3H, s) |
| 28 | COOH | 180.7 | — | 181.8 | — | ||
| 29 | CH3 | 32.1 | 0.91 | (3H, s) | 25.3 | 1.03 | (3H, s) |
| 30 | CH3 | 22.6 | 0.94 | (3H, s) | 24.6 | 0.94 | (3H, s) |
3.2 Anticancer activity
Isolated compounds in this study were evaluated their anticancer activity against the human cancer cells as HeLa (cervical cancer), T47D (breast cancer), and A549 (lung cancer) cell lines using MTT and XTT assays, which are the colorimetric measurement of cell viability. In the present anticancer activity test, doxorubicin was used as the standard. The pure compound is deemed highly toxic if the half-maximal inhibitory concentration (IC50) is ≤4 µg/mL and toxic if the IC50 is ≤20 µg/mL. The compound is moderately toxic if the IC50 is 20–100 g/mL and if IC50 above 100 g/mL is recommended as nontoxic [23,24].
Based on anticancer activity data obtained, alphitolic acid was suggested as a toxic compound against HeLa and T47D cell lines, with IC50 being 16.12 µg/mL and 7.37 µg/mL, respectively. However, its activity against the A549 cell line was presented as moderate inhibition (IC50 = 84.41 µg/mL). Moreover, arjunolic acid with an IC50 of 6.74 µg/mL was considered a toxic compound against HeLa cell lines. Arjunolic acid was a moderate compound against the T47D cancer cell line with an IC50 of 27.15 µg/mL, although it was a nontoxic compound against the A549 cancer cell line. According to the data mentioned earlier, the two types of triterpenoids inhibited higher toxicity against HeLa and T47D cell lines than the A549 cell line, and their activities may improve to obtain the natural origin drug from this species. The IC50 values of the two compounds are listed in Table 3.
Anticancer activity against HeLa, T47D, and A549 cell lines
| Compound name | MTT assay (IC50 ± SD µg/mL) | XTT assay (IC50 ± SD µg/mL) | |
|---|---|---|---|
| HeLa | T47D | A549 | |
| Alphitolic acid | 16.12 ± 0.681 | 7.37 ± 0.388 | 84.41 ± 0.048 |
| Arjunolic acid | 6.74 ± 0.015 | 27.51 ± 0.036 | 168.22 ± 0.090 |
| Doxorubicin | 2.67 ± 0.25 | 0.035 ± 0.012 | NT |
The meaning of NT is not test, SD = standard deviation.
Many terpene compounds have interesting bioactivities, so it is necessary to carry out other activity tests such as agar-well diffusion methods to evaluate the minimum inhibitory concentrations, minimum bactericidal concentration, IC50, and zone of inhibitions that could determine the potential antimicrobial and antifungal efficacy. The significant antioxidant potential could be evaluated through 1,1-diphenyl-2-picryl-hydrazyl and 2,2'-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) assays. The inhibiting of the α-glucosidase represents an antidiabetic activity that also could be visualized through molecular docking simulations. The effective anti-inflammatory agent could be determined via the carrageenan-induced assay, while an appreciable analgesic activity could be observed through the acetic acid-induced writhing bioassay [25].
4 Conclusion
This study reports the success of isolating two bioactive triterpenoid compounds from stem bark of S. aqueum, an Indonesian medicinal plant, namely alphitolic acid and arjunolic acid. These two isolated triterpenoids were significantly toxic against cervical cancer and breast cancer, shown by small IC50 values. Based on this result, we recommended continuing exploring the potential of this plant so that it can be used as a source of secondary metabolite compounds with interesting bioactivity.
Acknowledgments
All authors express big thanks to the Indonesian Ministry of Education and Culture for the Research Grant (Penelitian Dasar, 2021), to the Faculty of Pharmacy, Meijo University, Tempaku, Nagoya, Japan, for NMR and MS data, to the Faculty of Pharmacy, Gajah Mada University, Indonesia, and the Faculty of Science, UPM, Malaysia for an anticancer test.
-
Funding information: This work was supported by the Indonesian Ministry of Education and Culture for the Research Grant (Penelitian Dasar, 2021).
-
Author contributions: Alfinda Novi Kristanti: conceptualization, formal analysis, funding acquisition, investigation, resources, supervision, and writing – review and editing; Ei Ei Aung: data curation, formal analysis, investigation, project administration, validation, visualization, and writing – original draft; Nanik Siti Aminah: conceptualization, formal analysis, funding acquisition, methodology, resources, and supervision; Yoshiaki Takaya: data curation, resources, software, validation, and visualization; Hnin Thanda Aung: data curation, formal analysis, methodology, and supervision; and Rico Ramadhan: data curation, formal analysis, software, and validation.
-
Conflict of interest: Authors state no conflict of interest.
-
Ethical approval: The conducted research is not related to either human or animal use.
-
Data availability statement: All data generated or analyzed during this study are included in this published article [and its supplementary information files].
References
[1] Nazir N, Zahoor M, Uddin F, Nisar M. Chemical composition, in vitro antioxidant, anticholinesterase, and antidiabetic potential of essential oil of Elaeagnus umbellata Thunb. BMC Complement Med Ther. 2021;21:73. 10.1186/s12906-021-03228-y.Search in Google Scholar PubMed PubMed Central
[2] Shah SMM, Ullah F, Shah SMH, Zahoor M, Sadiq A. Analysis of chemical constituents and antinociceptive potential of essential oil of Teucrium stocksianum bioss collected from the North West of Pakistan. BMC Compl Altern Med. 2012;12:244. 10.1186/1472-6882-12-244.Search in Google Scholar PubMed PubMed Central
[3] Takuli P, Khulbe K, Kumar P, Parki A, Syed A, Elgorban AM. Phytochemical profiling, antioxidant and antibacterial efficacy of a native Himalayan Fern: Woodwardia unigemmata (Makino) Nakai. Saudi J Biol Sci. 2020;27:1961–7. 10.1016/j.sjbs.2020.06.006.Search in Google Scholar PubMed PubMed Central
[4] Elfahmi Y, Woerdenbag HJ, Kayser O. Jamu: Indonesia traditional herbal medicine towards radional phytopharmacological use. J Herb Med. 2016;4(2):51–73. 10.1016/j.hermed.2014.01.002.Search in Google Scholar
[5] Memon AH, Ismail Z, Al-Suede FSR, Aisha AFA, Hamil MSR, Saeed MAA. Isolation, characterization, crystal structure elucidation of two favanones and simultaneous RP-HPLC determination of five major compounds from Syzygium campanulatum Korth. Molecules. 2015;20:14212–33. 10.3390/molecules200814212.Search in Google Scholar PubMed PubMed Central
[6] Abera B, Adane L, Mamo F. Phytochemical investigation the root extract of Syzygium guineense and isolation of 2,3,23-trihydroxy methyl oleanate. J Pharmacogn Phytochem. 2018;7(2):3104–11.Search in Google Scholar
[7] Sobeh M, Mahmoud MF, Petruk G, Rezq S, Ashour ML, Youssef FS. Syzygium aqueum: a polyphenol-rich leaf extract exhibits antioxidant, hepatoprotective, pain-killing and anti-inflammatory activities in animal models. Front Pharmacol. 2018;9:1–14. 10.3389/fphar.2018.00566.Search in Google Scholar PubMed PubMed Central
[8] Osman H, Rahim AA, Isa NM, Nornaemah M, Bakhir NM. Antioxidant activity and phenolic content of Paederia foetida and Syzygium aqueum. Molecules. 2009;14:970–8. 3390/molecules14030970.Search in Google Scholar
[9] Sujanapal P, Sankaran KV. Common plants of maldives. Bangkok: FAO; 2016.Search in Google Scholar
[10] Costa JS, Cruz ENS, Setzer WN, Silva JKRS, Maia JGS, Figueiredo PLB. Essentials oils from brazilian Eugenia and Syzygium species and their biological activities. Biomolecules. 2020;10(1155):1–36. 10.3390/biom10081155.Search in Google Scholar PubMed PubMed Central
[11] Palanisamy UD, Ling LT, Manaharan T, Sivapalan V, Subramaniam T, Helme MH. Standard extract of Syzygium aqueum: a safe cosmetic ingredient. Int J Cosmet Sci. 2011;33:269–75. 10.1111/j.1468-2494.2010.00637.x.Search in Google Scholar PubMed
[12] Nonaka G, Aiko Y, Aritake K, Nishioka I, Samarangenins AB. novel proanthocyanidins with doubly bonded structures, from Syzygium samarangense and Syzygium aqueum. Chem Pharm Bull. 1992;40(10):2671–3.10.1248/cpb.40.2671Search in Google Scholar
[13] Aung EE, Kristanti AN, Amina NS, Takaya Y, Ramadhan R. Isolation of 2,4-di-tert-butylphenol and butyrospermol 3-β-O-palmitate from Syzygium aqueum stem bark. MOJ Eco Environ Sci. 2020;5(4):193–7. 10.15406/mojes.2020.05.00193.Search in Google Scholar
[14] Manaharan T, Appleton D, Cheng HM, Palanisamy UD. Flavonoids isolated from Syzygium aqueum leaf extract as potential antihyperglycaemic agents. Food Chem. 2012;132:1802–7. 10.1016/j.foodchem.2011.11.147.Search in Google Scholar
[15] Subarnas A, Diantini A, Abdulah R, Zuhrotun A, Hadisaputri YE, Puspitasari IM. Apoptosis induced in MCF-7 human breast cancer cells by 2’,4’-gihydroxy-6-methoxy-3,5-dimethylchalcone isolated from Eugenia aquea burm f. leaves. Oncol Lett. 2015;9:2303–6. 10.3892/ol.2015.2981.Search in Google Scholar PubMed PubMed Central
[16] Aung EE, Kristanti AN, Amina NS, Takaya Y, Ramadhan R. Cytotoxicity of butyrospermol and sitosterone from the stem bark of Syzygium aqueum. Trop. J Nat Prod Res. 2020;4(11):899–904. 10.26538/tjnpr/v4i11.10.Search in Google Scholar
[17] Aung EE, Kristanti AN, Amina NS, Takaya Y, Ramadhan R, Aung HT. Anticancer activity of isolated compounds from Syzygium aqueum stem bark. Rasayan J Chem. 2021;14(1):312–8. 10.31788/RJC.2021.1416106.Search in Google Scholar
[18] Cancer Chemoprevention Research Centre (CCRC); 2014. Prosedur tetap uji sitotoksik metode MTT, Faculty of Pharmacy Gadjah Mada University (UGM), Yogyakarta, Indonesia (in Indonesian). http://ccrc.farmasi.ugm.ac.id/wp-content/uploads/10_sop-uji-sitotoksik-metode-mtt.pdf.Search in Google Scholar
[19] Mutiah R, Indradmojo C, Dwi HH, Griana TP, Listyana A, Ramdhani R. Induction of apoptosis and phase-cell cycle inhibition of G0-G1, S, G2-M of T47D breast cancer cells on treatment with ethyl acetate fraction of jackfruit parasite leaves (Macrosolen cochinensis). J Appl Pharm Sci. 2017;7(10):138–43. 10.7324/JAPS.2017.71020.Search in Google Scholar
[20] American Type Culture Collection (ATCC). XTT cell proliferation assay Kit Instruction Manual. https://www.atcc.org/products/30-1011k.Search in Google Scholar
[21] Marzouk AM. Hapatoprotective triterpenes from hairy root cultures of Ocimum basilicum L. Z Naturforsch. 2009;64c(3–4):201–9. 10.1515/znc-2009-3-409.Search in Google Scholar PubMed
[22] Nono RN, Barboni L, Teponno RB, Quassinti L, Bramucci M, Vitali LA. Antimicrobial, antioxidant, anti-inflammatory activities and phytoconstituents of extracts from the roots of Dissotis thollonii Cogn. (Melastomataceae). S Afr J Bot. 2014;93:19–26.10.1016/j.sajb.2014.03.009Search in Google Scholar
[23] Sriwiriyajan S, Ninpesh T, Sukpondma Y, Nasomyon T, Graidist P. Cytotoxicity screening of plants of genus piper in breast cancer cell lines. Trop J Pharm Res. 2014;13(6):921–8. 10.4314/tjpr.v13i6.14.Search in Google Scholar
[24] Fikroh RA, Matsjeh S, Anwar C. Synthesis and anticancer activity of (E)-2’-hydroxy-2-bromo-4,5-dimethoxychalcone against breast cancer (MCF-7) cell line. Molekul. 2020;15(1):34–9. 10.20884/1.jm.2020.15.1.558.Search in Google Scholar
[25] Shah M, Murad W, Rehman NU, Mubin S, Al-Sabahi JN, Ahmad M, et al. GC-MS analysis and biomedical therapy of oil from n-hexane fraction of Scutellaria edelbergii Rech. f.: in vitro, in vivo, and in silico approach. Molecules. 2021;26:7676. 10.3390/molecules26247676.Search in Google Scholar PubMed PubMed Central
© 2022 Alfinda Novi Kristanti et al., published by De Gruyter
This work is licensed under the Creative Commons Attribution 4.0 International License.
