الفهرس 
INTRODUCTIONOnly 14 pages are availabe for public view
 |  | from | 231 |  |  |
| | | from | 231 | | |
Abstract
Fabaceae (Leguminosae) is one of the largest and most economically important families of flowering plants. It is the third-largest plant family in a number of species. Genus Erythrina belongs to family Fabaceae and is widely distributed in tropical and subtropical regions of the world. Erythrina comprises over 130 species of orange and/or red flowering trees, shrubs, and herbaceous plants. The plants of this genus exhibit significant role in traditional medicine targeting different diseases. Alkaloids and flavonoids were reported as the chief bioactive constituents of this genus with a wide range of biological activities. Anti-inflammatory, antimicrobial, anticonvulsant, anxiolytic, curare-like activity, insecticidal, and cytotoxic activities were reported for Erythrina sp. extracts, and compounds.
E. speciosa var. Rosea and E. crista-galli cultivated in Egypt were choosen for preliminary phytochemical studies. The results of the preliminary studies revealed that E. speciosa var. Rosea contain more promising compounds. Additionally, numerous phytochemical and biological studies were reported for E. crista-galli and only few reports were traced on E. speciosa var. Rosea. We, therefore, conducted the present work to evaluate the potential antiulcer and gastroprotective activity of E. speciosa var. Rosea as well as to reveal the underlying mechanism of action. Additionally, we were interested in finding new antibacterial and antiviral secondary metabolites from Erythrina sp. Although many phytochemical and biological studies were conducted on different plants belonging to this genus, very few reports were found on E. speciosa var. Rosea. This encouraged us to carry out in-depth biological and phytochemical studies on this selected plant.
The present study comprised two parts:
1. Phytochemical investigation on selected Erythrina species.
2. Biological investigation of E. speciosa var. Rosea leaves and flowers extracts
PART I
Phytochemical investigation on selected Erythrina species
1- Preliminary phytochemical studies on selected Erythrina species
1.1. Phytochemical screening of E. speciosa var. Rosea var. Rosea and E. crista-galli leaves
The phytochemical screening indicated the presence of alkaloids, flavonoids, saponins, sterols and/or triterpenes and carbohydrates and/or glycosides. The results revealed that these species most probably does not contain tannins or anthraquinone.
1.2. Solvent fractionation of E. speciosa var. Rosea var. Rosea and E. crista-galli leaves extract and preliminary chromatographic studies
The methanol extract of E. speciosa var. Rosea and E. crista-galli leaves were fractionated using n-hexane, followed by CH2CL2, EtOAc, and finally n-butanol. Preliminary 2D PC and TLC profiling of E. speciosa var. Rosea and E. crista-galli fractions demonstrated that E. speciosa var. Rosea comprised a variety of compounds in quite high concentrations.
2. Phytochemical studies on E. speciosa var. Rosea leaves and flowers
2.1. Phytochemical screening of E. speciosa var. Rosea leaves and flowers
The phytochemical screening indicated the presence of alkaloids, flavonoids, saponins, sterols and/or triterpenes and carbohydrates and/or glycosides.
2.2. Solvent fractionation of E. speciosa var. Rosea leaves and flowers extracts and preliminary chromatographic studies.
The methanol extract of E. speciosa var. Rosea leaves and flowers were fractionated using n-hexane, followed by CH2CL2, EtOAc, and finally n-butanol. Preliminary 2D PC and TLC profiling of E. speciosa var. Rosea leaves and flowers fractions demonstrated that E. speciosa var. Rosea comprised a variety of compounds in quite high concentrations, with the alkaloids concentrated in the dichloromethane fraction of both the leaves and flowers
2.3. Chemical investigation of the dichloromethane soluble fraction of E. speciosa var. Rosea leaves methanol extract (ESLDF)
In order to detect the phytochemical constituent responsible for ESLDF antibacterial activity. The chemical investigation was performed on ESLDF and resulted in the isolation of eleven compounds (1-11) including one novel alkaloid (Figure 84):
Compound 1: Genistein.
Compound 2: Lupiwighteone.
Compound 3: chryseriol.
Compound 4: Daidzein.
Compound 5: Protocatechuic acid methyl ester.
Compound 6 and 7 (Mixture): β-Sitosterol β-D-glucoside and β-stigmasterol β-D-glucoside.
Compound 8: Erysotrine.
Compound 9: 11α-Hydroxyerysotrine-8β-methyl acetate (New Natural Product).
Compound 10: 11-α-Hydroxyerysotrine.
Compound 11: Erybidine.
2.4. Chemical investigation of the ethyl acetate soluble fraction of E. speciosa var. Rosea leaves methanol extract (ESLEF)
Different chromatographic techniques were applied to detect compounds responsible for the antiulcer and antiviral activity of the ethyl acetate fraction of E. speciosa var. Rosea (Figure 84).
2.4.1. HPLC-ESI/MS/MS and HPLC-PDA profiling of the ethyl acetate soluble fraction of E. speciosa var. Rosea leaves methanol extract (ESLEF)
Chemical investigation of ESLE fraction resulted in the tentative identification of fourteen compounds. Most of the identified compounds were alkaloids, flavonoids, and saponins:
Hypaphorine
Abrine
11-Hydroxyerysodine hexouronic acid or it’s derivative
Apigenin-di-C-hexoside (vicenin-2)
Apigenin-C-hexoside-C-pentoside (schaftoside/isoschaftoside)
Luteolin-C-hexoside (orientin/isoorientin)
Apigenin-C-hexoside (vitexin/isovitexin)
Genistein hexoside
Genistein-deoxyhexoside-hexoside
Luteolin
Deoxyhexoside-hexoside-hexouronicA-soyasapogenol B
Deoxyhexoside-hexoside- hexuronicA-soyasapogenol E
Deoxyhexoside-hexoside-hexuronicA-soyasapogenol E
2.4.2. Isolation and structural elucidation of the major compounds of the ethyl acetate soluble fraction of E. speciosa var. Rosea leaves methanol extract (ESLEF)
The biologically active fraction obtained from ESLEF was subjected to further phytochemical investigation to isolate a pure single compound:
Compound 12: Vitexin
2.5. Chemical investigation of the dichloromethane soluble fraction of E. speciosa var. Rosea flowers methanol extract (ESFDF)
The dichloromethane soluble fraction of the flowers extract was subjected to further fractionation using different chromatographic techniques to isolate two pure compounds (Figure 85).
Compound 13: Erysotrine
Compound 14: Erysotrine N-oxide
As a result of this intensive study, fourteen compounds were isolated and identified from E. speciosa var. Rosea leaves and flowers extracts, one compound was reported for the first time from nature, two compounds were reported for the first time from the genus Erythrina, and seven compounds were reported for the first time from E. speciosa var. Rosea. Additionally, fourteen other compounds were tentatively identified from the leaves.
PART II
Biological investigation of E. speciosa var. Rosea leaves and flowers extracts
The total extract and fractions of E. speciosa var. Rosea leaves and flowers were subjected in the present study to in vitro and in vivo biological screening. In-depth biological investigation was subsequently carried out on the active fractions and pure isolated compounds.
1. In vitro biological activity
1.1. Cytotoxic activity
The total extract and the fractions of both the leaves and the flowers showed no cytotoxic activity at 20 μg/ml against Molt 4, K 562, LNcap, PC 3, and MCF-7 cell lines.
1.2. Antibacterial activity
The results revealed that the dichloromethane fraction of the leaves (ESLDF) was active against Gram-positive bacteria particularly Staphylococcus aureus ATCC9027 (inhibition zone diameter = 14.2 ± 0.217 mm). Fractions ESLD-VIII and ESLD-IX showed moderate antibacterial activity, while compounds 10 and 11 isolated from these fractions exhibited the most potent activity against the tested bacteria.
1.3. Antiviral activity
The results revealed that the hexane fraction from the flowers (ESFHF) and the ethyl acetate fraction from the leaves (ESLEF) exhibited the most potent antiviral inhibitory percentage against HSV-1 with inhibitory percentage 77% ± 2.7 and 65% ± 1.9, respectively. Bioactivity-guided fractionation resulted in the isolation of a bioactive antiviral drug compound 12 (EC50 = 35 ± 2.7 µg/ml against HAV-H10 and 18 ± 3.3 µg/ml against HSV-1 virus).
2. In vivo gastroprotective activity
2.1. In vivo gastroprotective, pilot study of E. speciosa var. Rosea leaves fractions
Preliminary in vivo gastroprotective pilot study was performed on E. speciosa var. Rosea leaves fractions in order to detect the most active fraction for further in-depth in vivo biological and phytochemical studies. A significant reduction in the percentage of the ulcerated areas was observed in the ESLEF treated group when compared with the ulcer control group.
2.2. Acute toxicity study on ESLEF
The median lethal dose (LD50) of ESLEF was higher than 1000 mg/kg body weight. These findings indicated that ESLEF has a low toxicity profile.
2.3. In vivo gastroprotective activity of the ethyl acetate soluble fraction of E. speciosa var. Rosea leaves methanol extract (ESLEF)
The gastroprotective activity of ESLEF was studied against ethanol-induced gastric ulceration in rats. The rats were pretreated with ESLEF (25, 50, and 100 mg/kg) 1 h before the administration of absolute ethanol to induce acute gastric ulceration. Pre-treatment with ESLEF at different doses resulted in a significant reduction in gastric lesions and significant elevation in the mucin production. These effects could be partially mediated by the potent anti-inflammatory activity of ESLEF as evidenced by the significant reduction in the immunoexpression of NF-κB, COX-2, iNOS and the reduction in the pro-inflammatory marker, TNF-α. ESLEF counteracted the ethanol-induced oxidative stress by increasing the levels of the depleted GSH, catalase and significantly attenuating the ethanol-induced lipid peroxidation tissue levels.