Three further triterpenoid saponins from Gleditsia caspica fruits and protective effect of the total saponin fraction on cyclophosphamide-induced genotoxicity in mice
-
Farouk R. Melek
,
Mona A.M. Abo-Zeid
Abstract
Three triterpenoidal saponins were isolated from the saponin fraction derived from a Gleditsia caspica Desf. methanolic fruit extract. The isolated saponins were identified as gleditsiosides B, C, and Q based on spectral data. The saponin-containing fraction was evaluated in vivo for genotoxic and antigenotoxic activities. The fraction caused no DNA damage in Swiss albino male mice treated with a dose of 45 mg/kg body weight for 24 h, although it significantly inhibited the number of chromosomal aberrations induced by cyclophosphamide (CP) in bone marrow and germ cells when applied before or after CP administration. The inhibitory indices in chromosomal aberrations were 59% and 41% for bone marrow and 48% and 43% for germ cells, respectively. In addition, the saponin fraction was found to reduce the viability of the human tumor cell line MCF-7 in a dose-dependent manner with an extrapolated IC50 value in the range of 220 μg/mL.
1 Introduction
The genus Gleditsia (family Fabaceae) comprises 14 species of deciduous trees [1]. Gleditsiacaspica (Caspian locust), a tree that grows up to 12 m, is cultivated in public gardens in Egypt mainly for ornamental purposes due to its graceful habit, elegant form, and delicate fern-like foliage. Gleditsia species have been widely used in folk medicine. The thorns of G. sinensis have been used for the treatment of carbuncles, scabies, and suppurate skin diseases, whereas the mature pods and anomalous fruits are mainly used for treating apoplexy, headache, productive cough, and asthma. The dried fruits of G. japonica Miq. have long been known in oriental medicine as diuretic and expectorant [2]. Saponins, the main constituents of Gleditsia fruits, were previously reported from fruits of different Gleditsia species [3–10]. From G. caspica, we recently reported the isolation and characterisation of 11 new triterpenoidal and acylated triterpenoidal saponins named caspicaosides A–K [11, 12], as well as the known Gleditsia saponins C′ and E′ and gleditsioside I [13].
As a part of our continuous interest in bioactive saponins from plants cultivated in Egypt [12, 14–19], we report here the isolation and identification of three further saponins from the saponin fraction of G. caspica fruits (SFGC). Also, the genotoxic and antigenotoxic activities of SFGC are presented.
2 Materials and methods
2.1 General
1H (400 MHz) and 13C (100 MHz) NMR spectra were recorded on an Jeol α-400 FT-NMR spectrometer (Tokyo, Japan), and chemical shifts are given as δ values with tetramethylsilane (TMS) as internal standard at 35 °C in pyridine-d5. High-performance liquid chromatography (HPLC) was performed on a Jasco system 800 instrument (Tokyo, Japan).
2.2 Chemicals
Cyclophosphamide (CP) and all other material used in cell culture were purchased from Sigma-Aldrich (St. Louis, MO, USA). All other chemicals used in extraction and isolation of saponins were purchased from ADWIC (Cairo, Egypt).
2.3 Plant material
Fruits of G. caspica were collected from El-Orman public garden, Giza, Egypt, in November 2012. A voucher specimen was deposited in the herbarium of the National Research Centre (CAIRC), Giza, Egypt.
2.4 Preparation of the saponin fraction of G. caspica (SFGC) and isolation of saponins
The air-dried fruits of G. caspica (2.0 kg) were defatted with n-hexane, then extracted twice with CHCl3, followed by MeOH until exhaustion. The combined MeOH extract was evaporated under reduced pressure to dryness. The residue (66 g) was dissolved in the least possible amount of MeOH, and the solution was diluted with the 10-fold amount of acetone to precipitate 18 g of a crude saponin mixture. The mixture was dissolved in H2O (0.2%), and the aqueous solution passed through a chromatographic column packed with 500 g Diaion HP-20 polymer gel (Mitsubishi, Tokyo, Japan). After washing the column with distilled water for several times, elution was carried out with 25%, 50%, 60%, 75% aqueous MeOH and finally with 100% MeOH. The collected fractions were examined by silica gel thin layer chromatography (TLC) (Merck, Darmstadt, Germany) using the solvent systems CHCl3/MeOH/H2O (60:30:5, v/v/v) and n-BuOH/EtOH/NH4OH (7:2:5) and visualized by spraying with 20% sulfuric acid in MeOH followed by heating at 110 °C. Based on TLC analysis, similar fractions were combined. Fractions eluted with 75% and 100% MeOH were found similar and contained saponin constituents. The two fractions were combined, and part of the combined fraction (SFGC) (3 g) was kept in a refrigerator until used for the biological study. The remainder (2.5 g) was applied onto a chromatographic column packed with 120 g PSQ 100B silica gel (Fuji Silysia, Nagoya, Japan) and eluted with CHCl3/MeOH/H2O with increasing polarity (70:27:3–58:35:7). A total of 50 fractions, 50 mL each, were collected. Similar fractions were combined after TLC analysis to yield 20 subfractions, A–T. Subfractions A (605 mg) and B (410 mg) were subjected to repeated HPLC (column, TSK gel ODS-80TS, 5 mm×60 cm; solvent, 30%–45% CH3CN in H2O, linear gradient; flow rate, 45 mL/min; detection, UV at 205 nm). Part of fraction A (150 mg) yielded gleditsioside Q (1; 22 mg). Part of fraction B (180 mg) afforded gleditsioside B (2; 20 mg) and gleditsioside C (3; 10 mg).
2.5 Gleditsioside Q (1)
Amorphous powder (22 mg). – 1H NMR (C5H5N, 400 MHz): aglycone: δ=0.92, 0.95, 1.00, 1.09, 1.10, 1.32, 1.85 (each 3H, s), 5.46 (1H, br t, J=3.0 Hz, H-12), 5.21 (1H, br s, H-16); sugar units: δ=1.71 (3H, d, J=5.8 Hz, Rha Me-6), 4.88 (1H, d, J=7.5 Hz, Glc H-1), 4.98 (1H, d, J=6.8 Hz, Xyl H-1), 5.13 (1H, d, J=7.5 Hz, Xyl′ H-1), 5.13 (1H, d, J=5.0 Hz, Ara H-1), 5.19 (1H, d, J=7.9 Hz, Xyl″ H-1), 6.11 (1H, d, J=7.9 Hz, Glc′ H-1), 6.34 (1H, d, J=1.5 Hz, Rha H-1); monoterpene unit (MT): δ=1.44 (3H, s, Me-10), 5.15 (1H, dd, J=10.5, 2.0 Hz, H-8a), 5.53 (1H, dd, J=17.0, 2.0 Hz, H-8b), 6.11 (1H, dd, J=17.0, 10.5 Hz, H-7), 7.20 (1H, t, J=8.0 Hz, H-3). – 13C NMR (C5H5N, 100 MHz): [6].
2.6 Gleditsioside B (2)
Amorphous powder (20 mg). – 1H NMR (C5H5N, 400 MHz): aglycone: δ 0.87, 0.91, 0.97, 0.99, 1.08, 1.33, 1.36 (each 3H, s), 5.47 (1H, br t, J=3.0 Hz, H-12); sugar units: δ 1.76 (3H, d, J=6.1 Hz, Rha Me-6), 4.97 (1H, d, J=7.0 Hz, Xyl H-1), 5.06 (1H, d, J=7.0 Hz, Xyl′ H-1), 5.14 (1H, d, J=5.2 Hz, Ara H-1), 5.17 (1H, d, J=7.8, Xyl″ H-1), 6.13 (1H, d, J=9.0 Hz, Glc′ H-1), 6.35 (1H, d, J=1.3 Hz, Rha H-1); monoterpene unit (MT): δ 1.44 (3H, s, Me-10), 5.14 (1H, dd, J=10.8, 1.8 Hz, H-8a), 5.53 (1H, dd, J=17.3, 1.8 Hz, H-8b), 6.10 (1H, dd, J=17.3, 10.8 Hz, H-7), 7.23 (1H, t, J=7.9 Hz, H-3). – 13C NMR (C5H5N, 100 MHz): [5].
2.7 Gleditsioside C (3)
Amorphous powder (10 mg). – 1H NMR (C5H5N, 400 MHz): aglycone δ 0.92, 0.94, 0.99, 1.08, 1.11, 1.31, 1.84 (each 3H, s), 5.19 (1H, br s, H-16), 5.64 (1H, br t, J=3.0 Hz, H-12); sugar units: δ 1.62 (3H, d, J=6.5 Hz, Rha Me-6), 4.92 (1H, d, J=7.5 Hz, Glc H-1), 4.99 (1H, d, J=6.9 Hz, Xyl H-1), 5.10 (1H, d, J=7.5 Hz, Xyl′ H-1), 5.11 (1H, d, J=7.0 Hz, Xyl″ H-1), 5.12 (1H, d, J=5.0 Hz, Ara H-1), 5.15 (1H, d, J=10.5 Hz, Gal H-1), 5.93 (1H, d, J=9.0 Hz, Glc′ H-1), 6.43 (1H, d, J=1.3 Hz, Rha H-1); monoterpene unit (MT): δ 1.45 (3H, s, Me-10), 5.15 (1H, dd, J=10.8, 1.8 Hz, H-8a), 5.53 (1H, dd, J=17.3, 1.8 Hz, H-8b), 6.10 (1H, dd, J=17.3, 10.8 Hz, H-7), 7.22 (1H, t, J=7.8 Hz, H-3). – 13C NMR (C5H5N, 100 MHz): [5].
2.8 Animals
Laboratory-bred strain Swiss albino male mice, 10–12 weeks old with an average weight of 25±2.5 g were obtained from the National Research Centre, Giza, Egypt. Animals were housed in groups (five animals/group) and maintained under standard conditions of temperature, humidity, and light. The animals were given standard food and water ad libitum.
2.9 In vitro study
2.9.1 Cell culture:
The breast cancer cell line (MCF-7) (ATCC, Rockville, MD, USA) was routinely cultured in RPMI-1640 media supplemented with 10% fetal bovine serum (FBS), 2 mM glutamine, containing 100 U/mL penicillin G sodium, 100 U/mL streptomycin sulfate, and 250 ng/mL amphotericin B. Cells were maintained in humidified air containing 5% CO2 at 37 °C. SFGC was dissolved in deionized distilled water before being added to cultured cells.
2.9.2 MTT Assay of cell viability:
The proliferation of the MCF-7 cells was estimated by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay [20]. Cells were cultured in 6-wells plates for 24 h in medium (5·105 cells/well), then SFGC was added at concentrations from 12.5 to 100 μg/mL in triplicate. The plates were incubated for 24 h at 37 °C in humidified air containing 5% CO2, before being submitted to the MTT assay. The absorbance was measured with an ELISA reader (BioRad, Munich, Germany) at 570 nm. The relative cell viability was determined by the amount of MTT converted to the insoluble formazan precipitate. The data were expressed as the mean percentage of viable cells as compared to the respective control untreated cultures.
2.10 In vivo study
2.10.1 Animals and treatment:
Animals were divided into nine groups of five animals each. Group I received distilled water and was used as negative control. Group II, the positive control, was administered CP [20 mg/kg body weight (b.w.)] intraperitonealy (i.p.) in two instalments within a 24-h interval. Group III was treated orally with 45 mg/kg b.w. SFGC, for 24 h. Groups IV through VI were treated orally with 4.5, 9.0, and 45 mg/kg b.w. SFGC 24 h before CP administration, and groups VII through IX were treated orally with 4.5, 9.0, and 45 mg/kg b.w. SFGC 24 h after CP administration. Animals were sacrificed 24 h after the last treatment. For preparation of somatic and germ cells, animals from the different groups were i.p. injected with colchicine (10 mg/kg b.w.) 2–3 h before sacrifice.
2.10.2 Chromosomal aberrations in somatic cells
Chromosome preparations from bone marrow (somatic cells) were carried out according to the method of Yosida and Amano [21]. One hundred well-spread metaphases were analysed per mouse. Metaphases with gaps, chromosome or chromatid breakage, fragmentation, deletions, Robertsonian translocation, as well as numerical aberrations (polyploidy) under 100-fold magnification with a light microscope (Olympus, Saitama, Japan), were recorded.
2.10.3 Chromosomal abnormalities in germ cells
Chromosome preparations from spermatocytes (germ cells) were made according to the technique of Evans et al. [22]. One hundred well-spread diakinase-metaphase I cells were analysed per animal for chromosomal aberrations. Metaphases with univalents, fragments, and chromosome translocations were recorded.
Evaluation of the activity of SFGC to reduce chromosomal aberrations induced by CP was carried out according to the formula of Madrigal-Bujaidar et al. [23] as follows:
2.11 Statistical analysis
The significance of the results from the data of the negative control and between SFGC with CP compared to CP alone was calculated using the t-test for chromosomal aberrations. Results in case of SFGC 24 h before CP were compared with CP after 24 h (first instalment), while results in case of SFGC 24 h after CP was compared with CP after 48 h (second instalment).
3 Results
3.1 Phytochemistry
The NMR data of the isolated compounds are recorded in the materials and methods section.
3.2 In vitro study
3.2.1 Cell viability
Cells of the breast cancer cell line (MCF-7) were incubated with increasing concentrations of SFGC for 24 h. The MTT assay revealed that cell viability decreased in a dose-dependent manner but statistically significant only with doses of 50 and 100 μg/mL. By linear extrapolation, the IC50 value would be expected in the range of 220 μg/mL.
3.3 In vivo study
3.3.1 Chromosomal aberrations in somatic cells
Table 1 shows the number and percentage of the chromosomal aberrations in control animals and animals treated with SFGC. The percentage of aberrant cells in animals treated with SFGC alone was not significantly different from that in the control group. But SFGC was found to reduce the number of chromosomal aberrations when administered 24 h either before or after administration of CP. Upon treatment with a dose of 45 mg/kg b.w. of SFGC, the reduction of chromosomal abnormalities, excluding gaps, was 59% when added before and 41% when added after CP administration, respectively. The reduction was highly significant (p<0.01) in comparison with CP alone.
The effect of SFGC on cyclophosphamide-induced chromosomal aberrations in mouse bone marrow cells in vivo.
| Treatments, mg/kg b.w. | Total abnormal metaphases | Number of different types of metaphases | Inhibitory index (excluding gaps) | |||||||
|---|---|---|---|---|---|---|---|---|---|---|
| Number | Mean±SE, % | G. | Frag. and/or Br. | Del. | C.F. | M.A. | Polyp. | |||
| Including gaps | Excluding gaps | |||||||||
| I. Control | 24 | 4.80±0.38 | 2.80±0.40 | 10 | 8 | 6 | 0 | 0 | 0 | – |
| II. CP (20) | 117a | 23.40±0.42c | 18.80±0.60c | 23 | 55 | 8 | 5 | 20 | 6 | – |
| 104b | 20.80±0.54c | 16.60±0.50c | 21 | 55 | 6 | 4 | 15 | 3 | – | |
| III. SFGC (45) | 23 | 4.60±0.50 | 2.40±0.62 | 11 | 8 | 4 | 0 | 0 | 0 | – |
| IV. SFGC (4.5) | ||||||||||
| Before CP | 82 | 16.40±0.30c,d | 13.00±0.62c,d | 17 | 51 | 5 | 2 | 7 | 0 | 30 |
| V. SFGC (9.0) | ||||||||||
| Before CP | 68 | 13.60±0.50c,d | 11.40±0.50c,d | 11 | 52 | 3 | 0 | 2 | 0 | 46 |
| VI. SFGC (45) | ||||||||||
| Before CP | 57 | 11.40±0.58c,d | 9.40±0.70c,d | 10 | 45 | 2 | 0 | 0 | 0 | 59 |
| VII. SFGC (4.5) | ||||||||||
| After CP | 92 | 18.40±0.42c | 14.60±0.44c | 19 | 56 | 5 | 1 | 10 | 1 | 14 |
| VIII. SFGC (9.0) | ||||||||||
| After CP | 80 | 16.00±0.56c,d | 12.00±0.68c,d | 20 | 51 | 4 | 0 | 5 | 0 | 33 |
| IX. SFGC (45) | ||||||||||
| After CP | 63 | 12.60±0.40c,d | 11.00±0.42c,d | 8 | 47 | 6 | 0 | 1 | 1 | 41 |
Total number of examined metaphases, 500 (five animals/group). G., gap; Frag., fragments; Br., breaks; Del., deletions; C.F., centric fusions; M.A., multiple aberrations; Polyp., polyploidy. aSamples taken after 24 h. bSamples taken after 48 h. cSignificant compared to vehicle control (p<0.01); dSignificant compared to CP treatment (p<0.01) (t-test).
3.3.2 Chromosomal abnormalities in germ cells
There were no significant differences between the animals treated with SFGC alone and the control animals (Table 2). The mean percentage of diakinesis-metaphase I cells were (21.60±0.70) % and (18.20±0.44) % (p<0.01) with 20 mg/kg b.w. of CP administered after 24 h and 48 h, respectively. This percentage was decreased after treatment with SFGC. Upon treatment with a dose of 45 mg/kg b.w. of SFGC, the maximum reductions were 48% and 43% when added before and after CP administration, respectively. Table 2 illustrates the protective effect of SFGC on the different types of aberrations, such as XY-univalents and/or autosomal univalents, fragments, and chain (IV).
The effect of SFGC on cyclophosphamide-induced chromosomal abnormalities in mouse spermatocyte cells in vivo.
| Treatment, mg/kg b.w. | Total abnormal metaphases | Number of different types of metaphases | Inhibitory index | |||||
|---|---|---|---|---|---|---|---|---|
| Number | Mean±SE, % | XY-uni. | Auto. uni. | XY-uni.+Auto. uni. | Frag. | Chain (IV) | ||
| I. Control | 15 | 3.00±0.48 | 9 | 6 | 0 | 0 | 0 | – |
| II. CP (20) | 108a | 21.60±0.70c | 73 | 21 | 4 | 2 | 8 | – |
| 91b | 18.20±0.44c | 58 | 24 | 3 | 1 | 5 | – | |
| III. SFGC (45) | 22 | 4.40±0.40 | 13 | 9 | 0 | 0 | 0 | – |
| IV. SFGC (4.5) | ||||||||
| Before CP | 75 | 15.00±0.40c,d | 54 | 21 | 0 | 0 | 0 | 35 |
| V. SFGC (9.0) | ||||||||
| Before CP | 67 | 13.40±0.48c,d | 55 | 10 | 1 | 1 | 0 | 44 |
| VI. SFGC (45) | ||||||||
| Before CP | 54 | 10.80±0.64c,d | 45 | 9 | 0 | 0 | 0 | 48 |
| VII. SFGC (4.5) | ||||||||
| After CP | 83 | 16. 60±0.45c | 60 | 18 | 1 | 0 | 4 | 11 |
| VIII. SFGC (9.0) | ||||||||
| After CP | 74 | 14.80±0.57c,d | 52 | 18 | 1 | 1 | 2 | 22 |
| IX. SFGC (45) | ||||||||
| After CP | 58 | 11.60±0.40c,d | 48 | 10 | 0 | 0 | 0 | 43 |
Total number of examined metaphases, 500 (five animals/group). XY-uni., XY- univalent; Auto. uni., autosomal univalent; XY-uni.+Auto. uni., XY-univalent+autosomal univalent; Frag., fragment. aSamples taken after 24 h. bSamples taken after 48 h. cSignificant compared to vehicle control (p<0.01). dsignificant compared to CP treatment (p<0.01) (t-test).
4 Discussion
In the present work, the crude saponin fraction from the Gleditsia caspica fruit extract was subjected to Diaion HP-20 polymer and silica gel column chromatography, respectively, followed by repeated HPLC to afford three triterpenoidal saponins (Fig.1). The isolated saponins were characterised based on their spectral data (materials and methods section) as gleditsiosides B, C [5] and Q [6]. Antigenotoxicity of SFGC was established in bone marrow and spermatocyte cells in CP treated mice.
Chemical structures of the isolated gleditsiosides B,C, and Q.
Glc, β-D-glucopyranosyl; Ara, α-L-arabinopyranosyl; Xyl, β-D-xylopyranosyl; Rha, α-L-rhamnopyranosyl; Gal, β-D- galactopyranosyl; MT, monoterpene unit.
SFGC at a dose of 45 mg/kg b.w. had no apparent genotoxic effect, as the proportion of aberrant cells was not significantly different from that of the negative control in both somatic and germ cells, whereas conversely, it displayed significant antigenotoxic activity against CP-induced mutagenesis in bone marrow cells.
Cyclophosphamide is a bi-functional alkylating agent. It is used to treat a wide range of cancers, in addition to its use as an immunosuppressive agent. However, it is a known carcinogen in humans and transforms into secondary metabolites. Its cytotoxic effects result from these reactive metabolites that alkylate DNA, RNA, and proteins [24]. Cyclophosphamide can cause DNA single strand breaks, as well as DNA-DNA and DNA-protein cross-links in post-implantation rat embryos and in testicular cells. CP treatment can induce structural chromosomal aberrations and sister-chromatid exchanges in embryos, Chinese hamster cells, human chorionic villi, and germ cells at various stages of spermatogenesis [25, 26]. DNA breaks induced by CP are important markers of genotoxicity [27].
Our findings are in agreement with previous reports demonstrating the antimutagenic activity of saponins. Amara-Mokrane et al. [28] suggested a desmutagenic activity of the triterpenoid saponin α-hederin isolated from Hedera helix against doxorubicin at all doses tested. This was later confirmed by Villani et al. [29] who discussed their mechanism of action. Furthermore, 13 saponins from Hedera helix, Calendula officinalis, and Calendula arvensis were assayed for their mutagenic and antimutagenic activity using the known promutagen benzo-[a]-pyrene (BaP) and a mutagenic urine concentrate from a smoker (SU). All the saponins were found to be nontoxic and nonmutagenic up to doses of 400 μg. Four saponins from Calendula arvensis and three saponins from Hedera helix counteracted the mutagenic activity of BaP (1 μg) and SU (5 μL) as a function of their concentration. A modified liquid incubation technique of the Salmonella/microsomal assay (Ames test) was used in this study [30]. The triterpenoid saponin ginsenoside Rb1 from Panax ginseng also provided significant protection against DNA damage and apoptosis induced by CP [31]. Soy saponins were proven to be antimutagens in Salmonella typhimurium TA98 against mutagenic heterocyclic amines and arylamines [32]. The maesasaponin mixture B, consisting of six homologous oleanane-type triterpenoid saponins isolated from Maesa lanceolata, was reported to exhibit moderate antimutagenic activity [33]. The antimutagenic mechanism of saponins may be related to the promotion of DNA repair [34].
The antigenotoxic property of SFGC provides additional health supplemental value to the other claimed therapeutic properties of Gleditsia plants. Saponins from G.sinensis fruits have been suggested for the therapy of rheumatoid arthritis [35]. The cytotoxic activity of G. sinensis fruit extract (GSE) against breast cancer [36] and nasopharyngeal carcinoma [37] has also been reported. Moreover, GSE was suggested as a potential chemotherapeutic drug to treat patients with acute and chronic myelogenous leukemia [38], and it was also proposed to be of use as an angiogenic inhibitor in both solid tumor and leukaemia therapy [39], as well as a novel anticancer agent for esophageal squamous cell carcinoma. Gleditsia saponin C, from G. japonica fruits, was reported to have significant anti-HIV activity [40]. The saponins from G. triacanthos exerted moderate oncostatic activity against sarcoma 180 and Ehrlich carcinoma [41].
In conclusion, SFGC proved to be nontoxic and to exhibit antigenotoxic potential.
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©2015 by De Gruyter
Artikel in diesem Heft
- Frontmatter
- Larvicidal activity of the essential oil of Youngia japonica aerial parts and its constituents against Aedes albopictus
- Antiviral and antitumor activities of the lectin extracted from Aspidistra elatior
- Susceptibility of Microsporum canis arthrospores to a mixture of chemically defined essential oils: a perspective for environmental decontamination
- Central analgesic activity of the aqueous and ethanolic extracts of the leaves of Albizia lebbeck: role of the GABAergic and serotonergic pathways
- Three further triterpenoid saponins from Gleditsia caspica fruits and protective effect of the total saponin fraction on cyclophosphamide-induced genotoxicity in mice
- Acylated flavonol diglucosides from Ammania auriculata
- Synthesis and antimicrobial properties of N-substituted derivatives of (E)-2′,3″-thiazachalcones
Artikel in diesem Heft
- Frontmatter
- Larvicidal activity of the essential oil of Youngia japonica aerial parts and its constituents against Aedes albopictus
- Antiviral and antitumor activities of the lectin extracted from Aspidistra elatior
- Susceptibility of Microsporum canis arthrospores to a mixture of chemically defined essential oils: a perspective for environmental decontamination
- Central analgesic activity of the aqueous and ethanolic extracts of the leaves of Albizia lebbeck: role of the GABAergic and serotonergic pathways
- Three further triterpenoid saponins from Gleditsia caspica fruits and protective effect of the total saponin fraction on cyclophosphamide-induced genotoxicity in mice
- Acylated flavonol diglucosides from Ammania auriculata
- Synthesis and antimicrobial properties of N-substituted derivatives of (E)-2′,3″-thiazachalcones
