Phytochemical and cytotoxicity studies on Arbutus pavarii, Asphodelus aestivus, Juniperus phoenicea and Ruta chalepensis growing in Libya

The work incorporates systematic bioassay-guided phytochemical and cytotoxicity/anticancer studies on four selected medicinal plants from the Libyan flora. Based on information on their traditional medicinal uses and the literature survey, Juniperus phoenicea L. (Fam: Cupressaceae), Asphodelus aestivus Brot. (Fam: Asphodelaceae), Ruta chalepensis. L (Fam: Rutaceae) and Arbutus pavarii Pampan. (Fam: Ericaceae) have been selected for investigation in the current endeavour. The four plants are well-known Libyan medicinal plants, which have been used in Libyan traditional medicine for the treatment of various human ailments, including both tumours and cancers. The cytotoxic activity of the n-hexane, dichloromethane (DCM) and methanol (MeOH) extracts of these plants were assessed against five human tumour cell lines: urinary bladder cancer [EJ-138], liver hepatocellular carcinoma [HEPG2], lung cancer [A549], breast cancer [MCF7] and prostate cancer [PC3] cell lines. The cytotoxicity at different concentrations of these extracts (0, 0.8, 4, 20, 100 and 500 µg/mL) was evaluated by the MTT assay. The four plants showed notable cytotoxicity against the five aforementioned human tumour cell lines with different selectivity indexes on prostate cancer cells. Accordingly, the cytotoxic effect of various chromatographic fractions from the different extracts of these plants at different concentrations (0, 0.4, 2, 10, 50 and 250 µg/mL) revealed different cytotoxic properties. Twenty-nine compounds were isolated from different fractions of these plants: three bioflavonoids, amentoflavone (25), cupressoflavone (24) and sumaflavone (76); four diterpenes. 13-epi-cupressic acid (42), imbricatholic acid (41), 3-hydroxy sandaracopimaric acid (44) and dehydroabietic acid (46), one alkanol (heptacosanol) and two lignans, deoxypodophyllotoxin (29) and β-peltatin methyl ether (28)] from J. phoenicea leaves; one flavonoid, luteolin (22), four anthraquinones [aleo-emodin (13), chrysphanol anthrone (79), 10, 10`chrysphanol bianthrone (80) and C-α-rhamnopyranosyl bianthracene-9, 9’, 10 (10'H)-trione glycoside (81) and p-hydroxy-phenethyl trans-ferulate (82) from A. aestivus leaves and tubers; three alkaloids, kokusaginine (61), graveoline (60), and 4-hydroxy-2-nonyl-quinoline (85), three coumarins, bergapten (63), chalepin (64) and chalepensin (65), one alkane, tetradecane, two flavonoid glycosides, rutin (52) and methoxy rutin (83) and 3``, 6`-disinapoylsucrose (84) from R. chalepensis aerial parts; one hydroquinone--D-glucopyranoside, arbutin (53) and two pentacyclic triterpenes, methyl betulinate (89) and ursolic acid (88) from A. pavarii leaves. Twenty-three isolated compounds were tested for their cytotoxicity against the most sensitive cancer cell lines. Eight compounds revealed good cytotoxic activity: cupressoflavone (24), sumaflavone (76), epicupressic acid (42), luteolin (22), chalepin (64) and 4-hydroxy-2-nonyl-quinoline (85) were cytotoxic against the A549 with IC50 values of 65, 77, 159, 76.9, 92 and 97.6 µM, respectively, whilst, compound 64 showed toxicity also against EJ138 with an IC50 value of 117 µM. C-α-rhamnopyranosyl bianthracene-9, 9’, 10 (10'H)-trione (81) and ursolic acid (88) were toxic against the prostate cancer cell line with IC50 values of 62 µM and 8.22 µM, respectively. The study findings also indicated that compounds 24, 64 and 88-induced cell death might involve the plasma membrane damage resulting in the release of LDH enzyme from the necrotic cells

Arbutin: Occurrence in Plants, and Its Potential as an Anticancer Agent

Arbutin, a hydroquinone glucoside, has been detected in ca. 50 plant families, especially in the plants of the Asteraceae, Ericaceae, Proteaceae and Rosaceae families. It is one of the most widely used natural skin-whitening agents. In addition to its skin whitening property, arbutin possesses other therapeutically relevant biological properties, e.g., antioxidant, antimicrobial and anti-inflammatory, as well as anticancer potential. This review presents, for the first time, a comprehensive overview of the distribution of arbutin in the plant kingdom and critically appraises its therapeutic potential as an anticancer agent based on the literature published until the end of August 2022, accessed via several databases, e.g., Web of Science, Science Direct, Dictionary of Natural Products, PubMed and Google Scholar. The keywords used in the search were arbutin, cancer, anticancer, distribution and hydroquinone. Published outputs suggest that arbutin has potential anticancer properties against bladder, bone, brain, breast, cervix, colon, liver, prostate and skin cancers and a low level of acute or chronic toxicity

Chalepin and Chalepensin: Occurrence, Biosynthesis and Therapeutic Potential

Dihydrofuranocoumarin, chalepin (1) and furanocoumarin, chalepensin (2) are 3-prenylated bioactive coumarins, first isolated from the well-known medicinal plant Ruta chalepensis L. (Fam: Rutaceae) but also distributed in various species of the genera Boenminghausenia, Clausena and Ruta. The distribution of these compounds appears to be restricted to the plants of the family Rutaceae. To date, there have been a considerable number of bioactivity studies performed on coumarins 1 and 2, which include their anticancer, antidiabetic, antifertility, antimicrobial, antiplatelet aggregation, antiprotozoal, antiviral and calcium antagonistic properties. This review article presents a critical appraisal of publications on bioactivity of these 3-prenylated coumarins in the light of their feasibility as novel therapeutic agents and investigate their natural distribution in the plant kingdom, as well as a plausible biosynthetic route

Growth inhibitory activity of biflavonoids and diterpenoids from the leaves of the Libyan Juniperus phoenicea against human cancer cells

Three biflavonoids [cupressuflavone (1), amentoflavone (2)and sumaflavone (3)], four diterpenoids [13-epi-cupressic acid (4), imbricatholic acid (5), 3-hydroxy-sandaracopimaric acid (6)and dehydroabietic acid (7)]and onelignan [β-peltatin methyl ether (8)],were isolated from the cytotoxicfractions of the extracts of the leaves of the Libyan Juniperus phoeniceaL. The structures of these compounds were elucidated by spectroscopic means.Cytotoxicity of the compounds 1-6were assessedagainst the human lung cancer cell lineA549 using the MTT assay. Compounds 1and 3showed cytotoxicityagainst the A549cells(IC50= 65 µMand 77 µM, respectively), whereas, compound 2did not show any activity. Diterpenes4-6exhibited weak cytotoxicity against the A549 cells with the IC50values of 159 µM, 263 µMand 223 µM, respectively. The cytotoxicity of each compound was compared with the anticancer drug,etoposide (IC50=61 µM).Cupressuflavone (1)wasevaluatedalso for cytotoxicity against both the human PC3 cancer cell lineand the normal prostatecell line (PNT2), and this compoundrevealed a high degreeof cytotoxic selectivity towards the prostate cancer cells (PC3), with IC50value of 19.9 µM, without any evidence of cytotoxicity towards the normal prostatecell line(PNT2)

Cytotoxic properties of the stem bark of Citrus reticulata Blanco (Rutaceae)

The bioassay-guided fractionation of the n-hexane extract of Citrus reticulata Blanco (Rutaceae) stem bark yielded scoparone (1), xanthyletin (2), lupeol (3), β-amyrin (4), stigmasterol (5), β-sitosterol (6) and palmitic acid. The structures of these compounds were determined by comprehensive spectroscopic analyses, i.e., 1D and 2D NMR and EI-MS, and by comparison with the reported data. Extracts, fractions and isolated compounds 1-6 were assessed for cytotoxicity by the MTT assay against three human cancer cell lines, i.e., human lung adenocarcinoma cell line A549, human breast adenocarcinoma cell line MCF7 and human Caucasian prostate adenocarcinoma cell line PC3. Significant activity of the n-hexane and the dichloromethane extracts was observed against the breast cancer cell line MCF7 with IC50s of 45.6 and 54.7 μg/mL, respectively. Moreover, the 70% ethyl acetate in n-hexane chromatographic fraction showed significant activity displaying IC50 values of 53.0, 52.4 and 49.1 μg/mL against the cancer cell lines A549, MCF7 and PC3, respectively. Encouragingly, an IC50 of 510.0 µg/mL against the human normal prostate cell line PNT2 indicated very low toxicity, and hence favourable selectivity indices for the 70% ethyl acetate in n-hexane fraction in the range of 9.6-10.4 towards cell lines A549, MCF7 and PC3. Since compounds isolated from the above fraction only delivered IC50 values in the range of 18.2-96.3, 9.2-34.1 and 7.5-97.2 μg/mL against A549, MCF7 and PC3 cell lines, respectively, synergistic action between compounds is suggested. Bioassay results valorize the anticancer effectivity of the stem bark of this plant in Cameroonian pharmacopeia

Bioassay-guided Isolation of Ursolic Acid as the Major Cytotoxic Compound Present in the Methanolic Extract of the Leaves of Arbutus pavarii Pamp

Arbutus pavarii Pamp. (Fam. Ericaceae), an endemic Libyan medicinal plant, is commonly known as “Shmeri”, “Shmar” or “Libyan Strawberry”, and almost exclusively found in the Al-Jabel Al-Akhdar mountainous region in Libya. This plant is a forage species for honeybees and thus, is important for honey production. A. pavarii has long been used in Libyan traditional medicine to treat both gastritis and kidney diseases. Previous limited phytochemical studies on this plant furnished the presence of simple phenolics like arbutin and gallic acid, and polyphenolics including flavonoids and tannins, for example, apigenin, epicatechin, hesperidin, kaempferol, naringin, quercetin and rutin, as well as some triterpenes and sterols. This paper describes, for the very first time, a cytotoxicity assay-guided isolation of the major active compound from the methanol extract of the leaves of A. pavarii, and its identification as ursolic acid (1) by comprehensive spectroscopic analyses

Dichloromethane Extract of the Leaves of Arbutus pavarii Pamp. Exhibits Cytotoxicity Against the Prostate Cancer Cell Line PC3: A Bioassay-guided Isolation and Identification of Arbutin and Betulinic Acid Methyl Ester

Objectives: To assess the cytotoxicity of Arbutus pavarii Pamp. (fam. Ericaceae), a Libyan medicinal plant, against human cancer cell lines and to carry out bioassay-guided isolation and identification of compounds. Materials and Methods: Shed-dried and ground leaves of A. pavarii were Soxhlet-extracted, successively, with n-hexane, dichloromethane (DCM) and methanol (MeOH), and assessed for cytotoxicity against several human cancer cell lines using the MTT assay. The cytotoxicity of the DCM extract against the normal human prostate cell line PNT2 was also assessed to determine the selectivity index (SI). The DCM extract was subjected to vacuum liquid chromatography (VLC) to produce eight fractions, which were then tested against the prostate cancer cell line PC3, as the DCM extract was considerably cytotoxic to this cell line. A reversed-phase preparative HPLC analysis of the active VLC fraction was carried out to purify the major compounds present in the active fraction, and the structures of the compounds were elucidated by spectroscopic means. Results: The DCM extract displayed most prominent cytotoxicity against the PC3 cell line with an IC50 value of 26 g/mL. However, this extract was much less cytotoxic to the normal human prostate cell line PNT2 (IC50 = 90 g/mL) with a selectivity index of 3.5. VLC analysis produced eight fractions, and among them, fraction VLC-5 was most active against the PC3 cell line. Prep-HPLC-based purification of VLC-5 afforded the isolation of arbutin (1) and betulinic acid methyl ester (2), the structures of which were elucidated by spectroscopic means. Conclusion: The DCM extract of the leaves of A. pavarii exhibited significant cytotoxicity to PC3 cells, but much less cytotoxicity against normal human prostate cell line. The isolated compounds from the active fraction, arbutin (1) and betulinic acid methyl ester (2), which were previously shown to possess cytotoxic properties, could be responsible for the cytotoxicity of the DCM extract

Liquid chromatography mass spectrometry analysis and cytotoxicity of Asparagus adscendens roots against human cancer cell lines

Background: Asparagus adscendens Roxb. (Asparagaceae), is native to the Himalayas. This plant has been used in the prevention and effective treatment of various forms of cancers. Objective: This paper reports, for the first time, on the cytotoxicity of the methanol (MeOH) extract of the roots of A. adscendens and its solid‑phase extraction (SPE) fractions against four human carcinoma cell lines and LC‑ESI‑QTOF‑MS analysis of the SPE fractions. Materials and Methods: Finely powdered roots of A. adscendens were macerated in methanol and extracted through SPE using gradient solvent system (water: methanol) proceeded for analysis on LC‑ESI‑QTOF‑MS and cytotoxicity against four human carcinoma cell lines: breast (MCF7), liver (HEPG2), lung (A549), and urinary bladder (EJ138), using the 3‑(4,5‑dimethylthiazol‑2‑yl)‑2,5‑diphenyltetrazoliumbromide assay. Results: The MeOH extract and four SPE fractions exhibited cytotoxicity against all cell lines with the IC50 values ranging from 6 to 79 μg/mL. As observedin other Asparagus species, the presence of saponins and sapogenins in the SPE fractions was evident in the liquid chromatography‑mass spectrometry data. Conclusion: It is reasonable to assume that the cytotoxicity of the MeOH extract of the roots of A. adscendens and its SPE fractions, at least partly, due to the presence of saponins and their aglycones. This suggests that A. adscendens could be exploited as a potential source of cytotoxic compounds with putative anticancer potential

Cytotoxicity of the Roots of Trillium govanianum Against Breast (MCF7), Liver (HepG2), Lung (A549) and Urinary Bladder (EJ138) Carcinoma Cells.

Trillium govanianum Wall. (Melanthiaceae alt. Trilliaceae), commonly known as 'nag chhatri' or 'teen patra', is a native species of the Himalayas. It is used in various traditional medicines containing both steroids and sex hormones. In folk medicine, the rhizomes of T. govanianum are used to treat boils, dysentery, inflammation, menstrual and sexual disorders, as an antiseptic and in wound healing. With the only exception of the recent report on the isolation of a new steroidal saponin, govanoside A, together with three known steroidal compounds with antifungal property from this plant, there has been no systematic pharmacological and phytochemical work performed on T. govanianum. This paper reports, for the first time, on the cytotoxicity of the methanol extract of the roots of T. govanianum and its solid-phase extraction (SPE) fractions against four human carcinoma cell lines: breast (MCF7), liver (HEPG2), lung (A549) and urinary bladder (EJ138), using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide cytotoxicity assay and liquid chromatography and electrospray ionization quadrupole time-of-flight mass spectrometry analysis of the SPE fractions. The methanol extract and all SPE fractions exhibited considerable levels of cytotoxicity against all cell lines, with the IC50 values ranging between 5 and 16 µg/mL. Like other Trillium species, presence of saponins and sapogenins in the SPE fractions was evident in the liquid chromatography mass spectrometry data. Copyright © 2016 John Wiley & Sons, Ltd