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Oleanolic acid from black raisins, Vitis vinifera with antioxidant and antiproliferative potentials on HCT 116 colon cancer cell line

Abstract

Vitis vinifera (black raisin) is commonly used in traditional medicine for the treatment of various ailments. In the present study, anti-oxidative and anti-cancer efficacy of oleanolic acid from ethyl acetate fraction of black raisins was evaluated and oleanolic acid was isolated without using of any chromatographic techniques and subjected to spectral assessment using UV-Vis spectrophotometer, 1H NMR, 13C NMR, MS and FT-IR for structural confirmation. Antiproliferative efficacy of oleanolic acid against human colon adenocarcinoma HCT-116 cells was assessed using cell viability assay. The minimum inhibitory concentration (IC50) was determined and found to be 40 µg/mL at 48h incubation. Furthermore, antioxidant property of oleanolic acid was analyzed using DPPH method (IC50 is 61.5µg/mL) by compared to standard antioxidants ascorbic acid, gallic acid, pyrogallol and butylated hydroxytoluene. Hence, the present study aims to establish the use of oleanolic acid as a potential therapeutic agent against human colon cancer.

Keywords:
Vitis vinifera; Oleanolic acid; Anti-oxidant; Anti-proliferative


INTRODUCTION

Black raisin (Vitis vinifera) belongs to the Vitaceae family. Many berries make up a cluster or bunch of grapes. Polyphenol is an important phytochemical in the grape which promotes many biological activities and has immense health-promoting benefits (Xia et al., 2010Xia EQ, Deng GF, Guo YI, Li HB. Biological activities of polyphenols from grapes. Int J. Mol Sci. 2010;11(2):622-646.). Grapes are available in the form of fresh fruit, raisins, juice and wine. In India, V. vinifera has been used as a traditional medicine for human health over 2000 years ago in ‘Darakchasava,’ a well-known Indian herbal preparation. This ‘Ayurvedic’ medicine is prescribed as a cardiotonic (Prasad, Tyagi, 2015Prasad S, Tyagi AK. Traditional medicine: The Goldmine for modern drugs. Adv Tech Biol Med. 2015;3:1-2.) and is also used in the prescriptions for cough, problems in the respiratory tract, catarrh, sub-acute cases of enlarged liver and spleen, as well as being used in alcohol-based tonics (Aasavs) (Wang, 2014Wang L, Waltenberger B, Wenzig EMP, Blunder M, Liu X, Malainer C, et al. Natural product agonists of peroxisome proliferators activated receptor gamma (PPARg): a review. Biochem Pharmacol. 2014;92(1):73-89.).

The triterpenoids are natural compounds that are widely distributed in the skin and seeds of different edible fruits, such as olives and grapes (Allouche et al., 2009Allouche Y, Jimenez A, Uceda M, Aguilera MP, Gaforio JJ, Beltran G. Triterpenic content and chemometric analysis of virgin olive oils from forty olive cultivars. J AgricFoodChem.2009;57(9):3604-3610.).Oleanolic acid (OA) is a major triterpenoid acid found in the fruits and is also present in both virgin olive oils and red wine (Pensec et al., 2016Pensec F, Paczkowski C, Grabarczyk M, Wozniak A, Gellon MB, Bertsch C, et al. Changes in the triterpenoid content of cuticular waxes during fruit ripening of eight grape (Vitis vinifera) cultivars grown in the upper Rhine valley. J Agric Food Chem. 2014;62(32):7998-8007.). The role of triterpenoids in the diet helps in the prevention of certain cancers, including breast cancer (Patlolla, Rao, 2012Patlolla JM, Rao CV. Triterpenoids for cancer prevention and treatment: Current status and future prospects. Curr Pharm Biotechnol. 2012;13(1):147-155.). OA also has the potential to be used as an antifungal (Tang et al., 2000Tang HQ, Hu J, Yang L, Tan RX. Terpenoids and flavonoids from Artemisia species. Planta Med. 2000;66(4):391-393.), anti-HIV (Khasiwada et al., 2000Kashiwada Y, Nagao T, Hashimoto A, Ikeshiro Y, Okabe H, Cosentin LM, et al. Anti-HIV activity of 3-O-acyl ursolic acid derivatives. J Nat Prod. 2000;63(12):1619-1622.), diuretic (Alvarez, Maria, Saad, 2002Alvarez ME, Maria AO, Saad JR. Diuretic activity of Fabiana patagonica in rats. Phytother Res. 2002;16(1):71-73.), and anti-inflammatory effects (Singh et al., 1992Singh GB, Singh S, Bani S, Gupta BD, Banerjee SK. Anti-inflammatory activity of oleanolic acid in rats and mice. J Pharm Pharmacol. 1992;44(5):456-8.). However there is little literature currently available with respect to their effects on colon carcinoma cells.

The present study was designed to isolate and purify OA in a limited resource laboratory and investigate its inhibitory effects on human colon carcinoma cell line HCT116 and also as an antioxidant potential.

MATERIAL AND METHODS

General experiments

UV-Vis Spectrophotometric analysis was run on a Shimadzu UV-2401 PC Spectrometer. FT-IR spectrum analysis was performed in KBr pellets on a Perkin Elmer FT-IR spectrum BX Spectrophotometer. A 1H and 13C NMR spectra were recorded on BrunkerAvance FT-NMR spectrometer operating at 300 MHz and100 MHz respectively.

Plant materials

V. vinifera was sourced locally from the Vellore market. It was deseeded manually using a knife and the peel was dried in an oven at 50 ºC for around 48 h and stored at 4 ºC.

Extraction and isolation of oleanolic acid

20 g of black raisin (V. vinifera) was taken and dried at 50 ºC for 48 h, pulverized and extracted with MeOH under reflux (6 h). After removal of the solvent using a rotary evaporator, the thick paste of MeOH extract was obtained. The oleanolic acid was isolated from methanolic extract using modified solvent-solvent fractionation without using any sophisticated chromatographic techniques (Sasikumar, Ghosh, 2017Sasikumar K, Ghosh AR. Isolation of myristyl alcohol from Hybanthus enneaspermus. Bangladesh J Pharmacol. 2017;12(2):113-14.; Shoba, Sasikumar, Sathiavelu, 2018Shoba S, Sasikumar K, Sathiavelu M. Isolation of isosativenetriol from endophytic fungus Cochliobolus spp. of Aerva lanata. Bangladesh J Pharmacol. 2018;13(1):57-58.). The methanol extract was then resuspended with Mili Q water and the aqueous insoluble extract was further washed with EtOAc. The washed soluble fraction of EtOAc was collected, dried and stored for further isolation process. The diethyl ether soluble fraction was separated from the EtOAc dried extract. The dried diethyl ether fraction was washed with hexane and the hexane insoluble extract was dissolved with the acetone followed by chloroform. The insoluble content of both the solvents were removed and the soluble fraction of chloroform was completely dried and the same was dissolved in dichloromethane. The solvent was removed under reduced pressure which afforded compound 1 (175mg).

DPPH Radical Scavenging Assay

The radical scavenging activity of OA was measured using the DPPH method (Ghafoor, 2014Ghafoor K. Antioxidant properties of oleanolic acid from grape peel. Agro Food Ind Hi-Tech. 2014;25(2):54-57.). In brief, 1 mL solution of antioxidants (0.5 mg/mL DMSO) was mixed with 2 mL of 10 mg/L methanolic solution of DPPH. This solution was then mixed thoroughly and kept at an ambient temperature for 5 min followed by absorbance measurement at 517 nm. The control solution for this test was made by mixing the reagent solution with methanol which did not contain sample or standard. The radical scavenging activity (RSA) was obtained using the following formula:

RSA % = Control OD Sample OD Control OD × 100

Cell proliferation assay

HCT-116 cells were procured from the National Centre for Cell Science, Pune, India. The cells were cultured in Dulbecco’s modified eagle’s medium (DMEM) supplemented with 10% heat-inactivated Foetal bovine serum (FBS), penicillin (10 U/mL), streptomycin (10 µg/mL) and 0.2 mM sodium pyruvate. The cultures were incubated in the presence of 5% CO2 at 37 ºC in humidified atmosphere. HCT-116 cells were seeded in 96-well micro plates at a density of 1×103 cells/well and incubated overnight. The medium was replaced with serum free DMEM supplemented with oleanolic acid at different concentrations (5-1000 µg/mL). After 48 h of incubation, the cell viability was determined using the colorimetric MTT assay as described earlier (Encalada et al., 2011Encalada MA, Hoyos KM, Rehecho S, Berasategi I, Ciriano MGID, Ansorena D, et al. Antiproliferative effect of Melissa officinalis on human colon cancer cell line. Plant Foods Hum Nutr.2011;66(4):328-334.). 15 µL of MTT solution (stock 5mg/mL) was added to each well and subsequently incubated for 4 h. The formazan crystals were dissolved using DMSO and absorbance was recorded at 570nm. The untreated cells (vehicle alone) were chosen as the negative control. Three independent assays were performed and the % cell viability was calculated.

All experiments were carried out in triplicates and expressed as Mean ± SD. The statistical analysis for each experiment was done by using the one-way ANOVA at significance level <0.05.

RESULTS AND DISCUSSION

The methanolic extract of the black raisins was used for the isolation of OA. The methanolic extract was partitioned with water and ethyl acetate to obtain two distinct fractions. The ethyl acetate fraction was subjected to various solvent-solvent fractionation (Figure 1). Without using any chromatographic technique, compound 1 was isolated and the yield (8.7 mg/g) was obtained. The yield was significantly high when compared to other optimized methods (Ghafoor, 2014Ghafoor K. Antioxidant properties of oleanolic acid from grape peel. Agro Food Ind Hi-Tech. 2014;25(2):54-57.).

FIGURE 1
Flow chart of isolation of oleanolic acid from the black raisins.

  • Compound (1): White powder with slight green pigmentation; mp 302-305 ºC; UV (MeOH) λmax (log ϵ) 235, 282 nm; IR (KBr) νmax 3446 (OH), 2926, 2852, 1687, 1641, 1452, 1388, 1271, 1178, 1083,1035, 997 cm1; HREIMS m/z 456.7032 (calculated for C30H49O3, m/z 342.348); 1H NMR(CDCl3) δ 5,27 (1H, t, H-12), δ 3.18 (1H, dd, J = 11.0, 5.0 Hz, H-3), δ 2.85 (1H, dd, J =4.0, 14.0 Hz,H-18), δ 0.93 (3H, s), δ 0.839 (3H, s), δ 0.963 (3H, s), δ 0.80 (3H, s), δ 0.996 (3H, s), δ 1.08 (3H, s), δ 0.964 (3H, s); 13C NMR δ 39.16 (C-1), δ 27.44 (C-2), δ 78.31 (C-3), δ38.43 (C-4), δ 55.36 (C-5), δ 18.1 (C-6), δ 33.5 (C-7), δ 46.23 (C-8), δ 48.11 (C-9), δ 36.77 (C-10), δ 23.12 (C-11), δ 122.25 (C-12), δ 143.79 (C-13), δ 41.49 (C-14), δ 26.47 (C-15), δ 22.66 (C-16), δ 45.85 (C-17), δ 32.16 (C-18), δ 41.33 (C-19), δ 30.20 (C-20), δ 32.41 (C-21), δ 22.57 (C-22), δ 28.79 (C-23), δ 14.9 (C-24), δ 14.48 (C-25), δ 16.32 (C-26), δ 25.0 (C-27), δ 180.4 (C-28), δ 32.62 (C-29) and δ 27.34 (C-30). The FT-IR data (Onoja, Ndukwe, 2013Onoja E, Ndukwe IG. Isolation of oleanolic acid from chloroform extract of Borreri astachydea (DC) Hutch. J Nat Prod Plant Resour. 2013;3(2):57-60.) and NMR data of 1 were identical to the published data of 3-β-Hydroxyolean-12-en-18α-28-oic acid (18α- Oleanolic acid) (Seebachar et al., 2003Seebacher W, Simic N, Weis R, Saf R, Kunert O. Complete assignments of 1H and 13C NMR resonance of oleanolic acid, 18α-oleanolic acid, ursolic acid and 11-oxo derivatives. Magn Reson Chem. 2003;41(8):636-638.; Zhang et al., 2004Zhang Y, Jayaprakasam B, Seeram NP, Olson LK, Witt DD, Nair MG. Insulin secretion and cyclooxygenase enzyme inhibition by cabernet sauvignon grape skin compounds. J Agric Food Chem. 2004;52(2):228-233.; Gohari et al., 2009Gohari AR, Saeidnia S, Hadjiakhoondi A, Abdoullahi M, Nezafati M.Isolation and quantificative analysis of oleanolic acid from Satureja mutica Fisch.& C.A. Mey. J Med Plants. 2009;8(5):65-69.; Maillard, Adewunmi, Hostettmann, 1992Maillard M, Adewunmi CO, Hostettmann KA. Triterpene glycoside from the fruits of Tetrapleura tetraptera. Phytochem,1992;31(4):1321-1323.).

Structural elucidation of compound (1)

The Infrared (IR) analysis of compound 1 showed a peak at 3446cm-1, which suggests the presence of a hydroxyl group, while the peak at 1687cm-1indicates the presence of a carbonyl group. These two peaks may be due to the presence of carboxylic acid functional group (COOH). The peak at 1388cm-1 may be due to the presence of tri-substituted olefinic group, while the peak at 1462cm-1 may represent the presence of -CH3 and -CH2- signals. The peak at 2926 cm-1 showed the presence of C-H stretch for alkanes. This signal suggests that the molecule may be highly saturated. The molecular ion peak was established by HRMS spectrum which showed a molecular ion peak at m/z 456.7032, corresponding to the molecular formula C30H48O3 (calcd. 456.3603). In the HRMS, the pronounced peak at m/z 418.9207 represents the loss of 38 m. u. corresponding to the COOH group. The prominent baseline peak at m/z 248.6462 represents a typical RDA fragmentation characteristic of ∆12-oleanane type triterpenes with COOH group, at C-17 (Ayatollahi et al., 2011Ayatollahi AM, Ghanadian M, Afsharypour S, Abdella OM, Mirzai M, Askari G. Pentacyclic triterpenes in Euphorbia microsciadia with their T-cell proliferation activity.Iran J Pharm Res. 2011;10(2):287-294.). Another baseline peak at m/z 203.6032 can be attributes to the loss of COOH from the fragment at m/z 248. The1H-NMR spectrum of compound 1 shows seven singlets methyl groups at δH1.08, 0.83, 0.99, 0.80, 0.96, 0.96 and 0.93 (H-23, 24, 25, 26, 27, 29 and 30). The carbonyl proton at C-3 position resonated at δ 3.18 (J = 11.0, 5.0 Hz,) suggesting its α- and axial configuration and the olefinic proton at C-12 appeared at δ 5.27 as a triplet (J=3.5Hz). A one proton double doublet at δ 2.85 was assigned to H-18 on the basis of its chemical shift as well as the multiplicity pattern reported for H-18 with β-stereochemistry. The 13C-NMR spectrum of compound 1 exhibited thirty carbon peaks. The peak at δ 180.4 may be due to the presence of the carbonyl group assigned to C-28. The two peaks at δ 122.25 and δ-143.79 may be due to the presence of a pair of sp2 hybridized carbon- carbon atoms assigned to C-12 and C-13. While the seven peaks at δ 28.79, δ 14.9, δ 14.48, δ 16.32, δ 25.0, δ 180.4, δ 32.62 and δ 27.34 can be attributed to the seven methyl groups which are assigned to C-23, C-24, C-25, C-26, C-27, C-29 and C-30 respectively. Therefore compound 1 could be assigned as 3-β-Hydroxyolean-12-en-18α-28-oic acid (18α - Oleanolic acid) (Figure 2).

FIGURE 2
Structure of compound 1 is oleanolic acid (OA).

Free radical scavenging assay

The free radical scavenging activity of oleanolic acid was investigated by DPPH assays and the result exhibited a good free radical inhibited activity when compared to standard antioxidants ascorbic acid (AA), gallic acid (GA), pyrogallol (PG) and butylated hydroxytoluene (BHT). DPPH involves hydrogen atoms transfer and electrons transfer (Kaviarasan et al., 2007Kaviarasan S, Naik GH, Gangabhagirathi R, Anuradha CV, Priyadarsini KI.In vitro studies on anti-radical and antioxidant activities of fenugreek (Trigonella foenumgraecum) seeds. Food Chem. 2007;103(1):31-37.). The result of the present investigation illustrated that oleanolic acid may reduce/inhibit the production of free radicals by donating hydrogen molecules and result in the decolourization in the DPPH assays. Figure 3 demonstrates the abilities of the different antioxidants to scavenge free DPPH radicals. It is evident that oleanolic acid has 88.30±1.84% DPPH-scavenging activity, which is comparable with that of AA (93.11±2.14%), GA (90.85±1.32%), PG (93.03±1.12%) and BHT (93.20±2.33%). The minimum inhibitory concentration (IC50) of oleanolic acid was found to be 61.5 µg/mL.

FIGURE 3
Antioxidant activity (DPPH method) of oleanolic acid.

Anti-proliferative assay

The anti-cancer efficacy of OA was determined using cell-based in vitro techniques. The proliferation of HCT-116 was found to be inhibited which was evident from reduced cell viability of OA treated HCT-116 cells when compared with the untreated control cells. This inhibitory effect was concentration dependent and was found to lower the cell viability at the concentration 50 µg/mL of OA. The required concentration for the 50% inhibition (IC50) of cells was 40 µg/mL (Figure 4).The similar class of pentacyclic triterpenoids like Asiatic acid (AA), betulinic acid (Bet A), boswellic acid (BA), glycyrrhizin, lupeol, ursolic acid (UA) and their derivatives were also reported previously to have a potent anticancer effect (Paduch, Kandefer-Szerszen, 2014Paduch R, Kandefer-Szerszeń M. Antitumor and antiviral activity of pentacyclic triterpenes. Mini-Rev Org Chem.2014;11(3):262-268.). Asiatic acid (AA) caused about a 50% reduction in the viability of ovarian cancer cells SKOV3 and OVCAR-3 at the concentration of 40 µg/mL (Ren et al., 2016Ren L, Cao QX, Zhai FR, Yang SQ, Zhang HX. Asiatic acid exerts anticancer potential in human ovarian cancer cells via suppression of PI3K/Akt/mTOR signaling. Pharm Biol. 2016;54(11):2377-2382.). Betulinic acid was active against breast cancer cell lines with an IC50 value on MDA-MB-231 (21.9 µM) and MDA-MB-468 (46.0 µM) cell lines (Weber et al., 2014Weber D, Zhang M, Zhuang P, Zhang Y, Wheat J, Currie G, Al-Eisawi Z. The efficacy of betulinic acid in triple-negative breast cancer. SAGE Open Med. 2014;2:2050312114551974.). It also showed concentration-dependent anti-proliferative activity on paclitaxel-resistant lung H460 cells and IC50 was found to be 50 µM (Zhan et al., 2018Zhan XK, Li JL, Zhang S, Xing PY, Xia MF. Betulinic acid exerts potent antitumor effects on paclitaxel-resistant human lung carcinoma cells (H460) via G2/M phase cell cycle arrest and induction of mitochondrial apoptosis. Oncol Lett. 2018;16(3):3628-3634.).Glycyrrhizin exhibited a significant inhibition at 100 µM on the human lung carcinoma cell line, HCC827 cells after 48 h (Ma et al., 2016Ma YF, Guo NN, Chu J, Jin S, Yang B, Li J, Zhang T, Guo JT, Chen L, Liang CY, Wang LH, Liu Y. Glycyrrhizin treatment inhibits proliferation and invasive potential of lung cancer cells. Int J Clin Exp Med. 2016;9(6):10592-10596.) but the cell viability was not affected on the primary mouse mammary epithelial cells at 50, 100 and 200 µg/mL (Fu et al., 2014Fu Y, Zhou E, Wei Z, Liang D, Wang W, Wang T, Guo M, Zhang N, Yang Z. Glycyrrhizin inhibits the inflammatory response in mouse mammary epithelial cells and a mouse mastitis model. FEBS J. 2014;281(11):2543-57.).Lupeol also inhibited the growth of the MCF-7 breast cancer cells with a dose-dependent increase providing an IC50 value of the compound as 80 µM (Pitchai, Roy, Ignatius, 2014Pitchai D, Roy A, Ignatius C. In vitro evaluation of anticancer potentials of lupeol isolated from Elephantopus scaber L. on MCF-7 cell line. J Adv Pharm Technol Res. 2014;5(4):179-84.). Oleanolic acid from Coleus tuberosus, has an antiproliferative activity through induced apoptosis in the MCF-7 cells and was in a dose-dependent manner. IC50 of OA was found at 48.61µg/mL after 48h (Nugraheni et al., 2011Nugraheni M, Santoso U, Suparmo H, Wuryastuti H. In vitro antioxidant, antiproliferative and apoptosis effect of Coleus tuberosus L. Afr J Food Sci. 2011;5(4):232-241.). Colon carcinoma HCT15 cells were treated with various concentrations of ursolic acid (UA) and OA for 48 h. The cell viability was significantly decreased in a dose-dependent manner and IC50 of UA and OA was found to be 30 µM/L and 60 µM/L respectively (Li, Gai, Yang, 2002Li J, Gai WJ, Yang QY. Effects of urosolic acid and oleanolic acid on human colon carcinoma cell lines HCT15. World J Gasteroenterol. 2002;8(3):493-495.). OA administered in a dose and time-dependent manner exhibits a significant decrease in the cell viability of GBC-SD and NOZ (human gallbladder cancer) cell lines. The IC50 of the GBC-SD and NOZ cells was found to be 50µM/L at 48 hr (Li et al., 2015Li HF, Wang XA,Xiang SS,Hu YP, Jiang L,Shu YJ,Li ML, Wu XS, Zhang F,Ye YY, Weng H,Bao RF,Cao Y,Lu W,Dong Q, Liu YB. Oleanolic acid induces mitochondrial-dependent apoptosis and G0/G1 phase arrest in gallbladder cancer cells. Drug Des Devel Ther. 2015;9:3017-3030.). The previous report found that OA has similar IC50 of 47µg/mL against the same cell line HCT 116 (Abdelwahab, Hussein, Kadry, 2015Abdelwahab MF, Hussein MH, Kadry HH. Cytotoxicity and antioxidant activity of new biologically active constituents from Micromeria nervosa grown in Egypt. Bull Fac Pharm Cairo Univ. 2015;53(2):185-194.). According to morphological changes in HCT116 cells, OA has a direct cytotoxic effect which is concurrent with a previous study on it and it has been reported that OA has a cytotoxic activity in HCT15 (Li, Gai, Yang, 2002Li J, Gai WJ, Yang QY. Effects of urosolic acid and oleanolic acid on human colon carcinoma cell lines HCT15. World J Gasteroenterol. 2002;8(3):493-495.). Some studies have also supported that oleanane-type triterpenoids had inhibitory effects on DNA polymerase beta and DNA topoisomerases (Wada, Iida, Tanaka, 2001Wada S, Iida A, Tanaka R. Screening of triterpenoids isolated from Phyllanthus flexuosus for DNA topoisomerase inhibitory activity. J Nat Prod.2001;64(12):1545-1547.).

FIGURE 4
Anti-proliferative efficacy of oleanolic acid against HCT116 colon cancer cells.

CONCLUSION

OA obtained from V. vinifera can be established as a suitable drug candidate for anti-colon cancer from based on the inferences drawn from the antioxidant and cytotoxic assays. These findings of the present study show black raisin as a hopeful source to inhibit colorectal cancer. Though OA metabolites are present in low quantity in raisins they possess good anticancer activity due to the synergetic effect of the various metabolites that are present in the black raisins. Further refinements may attribute to elucidate the molecular mechanism of interaction for anti-colon cancer activity of OA. Considering the clinical efficacy and toxicity of numerous anticancer agents which are unknown and uncertain, understanding the fundamental role of herbal extracts has been found to play an essential role in the development of herbal drugs and its use in the treatment of cancer.

Supplementary data: UV- Vis spectra, FT-IR spectra, NMR spectra (1H and 13C NMR) and HR-ESI-MS spectra for the compound 1 (oleanolic acid).

ACKNOWLEDGEMENT

The authors are grateful to the management of VIT University for their support in pursuing research and to the Sophisticated Analytical Instrument Facility, Indian Institute of Madras for MS and NMR spectral analysis.

REFERENCES

  • Abdelwahab MF, Hussein MH, Kadry HH. Cytotoxicity and antioxidant activity of new biologically active constituents from Micromeria nervosa grown in Egypt. Bull Fac Pharm Cairo Univ. 2015;53(2):185-194.
  • Allouche Y, Jimenez A, Uceda M, Aguilera MP, Gaforio JJ, Beltran G. Triterpenic content and chemometric analysis of virgin olive oils from forty olive cultivars. J AgricFoodChem.2009;57(9):3604-3610.
  • Alvarez ME, Maria AO, Saad JR. Diuretic activity of Fabiana patagonica in rats. Phytother Res. 2002;16(1):71-73.
  • Ayatollahi AM, Ghanadian M, Afsharypour S, Abdella OM, Mirzai M, Askari G. Pentacyclic triterpenes in Euphorbia microsciadia with their T-cell proliferation activity.Iran J Pharm Res. 2011;10(2):287-294.
  • Encalada MA, Hoyos KM, Rehecho S, Berasategi I, Ciriano MGID, Ansorena D, et al. Antiproliferative effect of Melissa officinalis on human colon cancer cell line. Plant Foods Hum Nutr.2011;66(4):328-334.
  • Fu Y, Zhou E, Wei Z, Liang D, Wang W, Wang T, Guo M, Zhang N, Yang Z. Glycyrrhizin inhibits the inflammatory response in mouse mammary epithelial cells and a mouse mastitis model. FEBS J. 2014;281(11):2543-57.
  • Ghafoor K. Antioxidant properties of oleanolic acid from grape peel. Agro Food Ind Hi-Tech. 2014;25(2):54-57.
  • Gohari AR, Saeidnia S, Hadjiakhoondi A, Abdoullahi M, Nezafati M.Isolation and quantificative analysis of oleanolic acid from Satureja mutica Fisch.& C.A. Mey. J Med Plants. 2009;8(5):65-69.
  • Kashiwada Y, Nagao T, Hashimoto A, Ikeshiro Y, Okabe H, Cosentin LM, et al. Anti-HIV activity of 3-O-acyl ursolic acid derivatives. J Nat Prod. 2000;63(12):1619-1622.
  • Kaviarasan S, Naik GH, Gangabhagirathi R, Anuradha CV, Priyadarsini KI.In vitro studies on anti-radical and antioxidant activities of fenugreek (Trigonella foenumgraecum) seeds. Food Chem. 2007;103(1):31-37.
  • Li J, Gai WJ, Yang QY. Effects of urosolic acid and oleanolic acid on human colon carcinoma cell lines HCT15. World J Gasteroenterol. 2002;8(3):493-495.
  • Li HF, Wang XA,Xiang SS,Hu YP, Jiang L,Shu YJ,Li ML, Wu XS, Zhang F,Ye YY, Weng H,Bao RF,Cao Y,Lu W,Dong Q, Liu YB. Oleanolic acid induces mitochondrial-dependent apoptosis and G0/G1 phase arrest in gallbladder cancer cells. Drug Des Devel Ther. 2015;9:3017-3030.
  • Ma YF, Guo NN, Chu J, Jin S, Yang B, Li J, Zhang T, Guo JT, Chen L, Liang CY, Wang LH, Liu Y. Glycyrrhizin treatment inhibits proliferation and invasive potential of lung cancer cells. Int J Clin Exp Med. 2016;9(6):10592-10596.
  • Maillard M, Adewunmi CO, Hostettmann KA. Triterpene glycoside from the fruits of Tetrapleura tetraptera Phytochem,1992;31(4):1321-1323.
  • Nugraheni M, Santoso U, Suparmo H, Wuryastuti H. In vitro antioxidant, antiproliferative and apoptosis effect of Coleus tuberosus L. Afr J Food Sci. 2011;5(4):232-241.
  • Onoja E, Ndukwe IG. Isolation of oleanolic acid from chloroform extract of Borreri astachydea (DC) Hutch. J Nat Prod Plant Resour. 2013;3(2):57-60.
  • Paduch R, Kandefer-Szerszeń M. Antitumor and antiviral activity of pentacyclic triterpenes. Mini-Rev Org Chem.2014;11(3):262-268.
  • Patlolla JM, Rao CV. Triterpenoids for cancer prevention and treatment: Current status and future prospects. Curr Pharm Biotechnol. 2012;13(1):147-155.
  • Pensec F, Paczkowski C, Grabarczyk M, Wozniak A, Gellon MB, Bertsch C, et al. Changes in the triterpenoid content of cuticular waxes during fruit ripening of eight grape (Vitis vinifera) cultivars grown in the upper Rhine valley. J Agric Food Chem. 2014;62(32):7998-8007.
  • Pitchai D, Roy A, Ignatius C. In vitro evaluation of anticancer potentials of lupeol isolated from Elephantopus scaber L. on MCF-7 cell line. J Adv Pharm Technol Res. 2014;5(4):179-84.
  • Prasad S, Tyagi AK. Traditional medicine: The Goldmine for modern drugs. Adv Tech Biol Med. 2015;3:1-2.
  • Sasikumar K, Ghosh AR. Isolation of myristyl alcohol from Hybanthus enneaspermus Bangladesh J Pharmacol. 2017;12(2):113-14.
  • Seebacher W, Simic N, Weis R, Saf R, Kunert O. Complete assignments of 1H and 13C NMR resonance of oleanolic acid, 18α-oleanolic acid, ursolic acid and 11-oxo derivatives. Magn Reson Chem. 2003;41(8):636-638.
  • Shoba S, Sasikumar K, Sathiavelu M. Isolation of isosativenetriol from endophytic fungus Cochliobolus spp. of Aerva lanata Bangladesh J Pharmacol. 2018;13(1):57-58.
  • Singh GB, Singh S, Bani S, Gupta BD, Banerjee SK. Anti-inflammatory activity of oleanolic acid in rats and mice. J Pharm Pharmacol. 1992;44(5):456-8.
  • Ren L, Cao QX, Zhai FR, Yang SQ, Zhang HX. Asiatic acid exerts anticancer potential in human ovarian cancer cells via suppression of PI3K/Akt/mTOR signaling. Pharm Biol. 2016;54(11):2377-2382.
  • Tang HQ, Hu J, Yang L, Tan RX. Terpenoids and flavonoids from Artemisia species. Planta Med. 2000;66(4):391-393.
  • Wada S, Iida A, Tanaka R. Screening of triterpenoids isolated from Phyllanthus flexuosus for DNA topoisomerase inhibitory activity. J Nat Prod.2001;64(12):1545-1547.
  • Wang L, Waltenberger B, Wenzig EMP, Blunder M, Liu X, Malainer C, et al. Natural product agonists of peroxisome proliferators activated receptor gamma (PPARg): a review. Biochem Pharmacol. 2014;92(1):73-89.
  • Weber D, Zhang M, Zhuang P, Zhang Y, Wheat J, Currie G, Al-Eisawi Z. The efficacy of betulinic acid in triple-negative breast cancer. SAGE Open Med. 2014;2:2050312114551974.
  • Xia EQ, Deng GF, Guo YI, Li HB. Biological activities of polyphenols from grapes. Int J. Mol Sci. 2010;11(2):622-646.
  • Zhan XK, Li JL, Zhang S, Xing PY, Xia MF. Betulinic acid exerts potent antitumor effects on paclitaxel-resistant human lung carcinoma cells (H460) via G2/M phase cell cycle arrest and induction of mitochondrial apoptosis. Oncol Lett. 2018;16(3):3628-3634.
  • Zhang Y, Jayaprakasam B, Seeram NP, Olson LK, Witt DD, Nair MG. Insulin secretion and cyclooxygenase enzyme inhibition by cabernet sauvignon grape skin compounds. J Agric Food Chem. 2004;52(2):228-233.

Publication Dates

  • Publication in this collection
    16 Mar 2020
  • Date of issue
    2020

History

  • Received
    07 Apr 2017
  • Accepted
    17 Dec 2018
Universidade de São Paulo, Faculdade de Ciências Farmacêuticas Av. Prof. Lineu Prestes, n. 580, 05508-000 S. Paulo/SP Brasil, Tel.: (55 11) 3091-3824 - São Paulo - SP - Brazil
E-mail: bjps@usp.br