A Comparative Pharmacognostical Study of Certain Jasminum Species

Home , Jasminum azoricum, Jasminum humile, Jasminum mesnyi, Jasminum officinale

English Summary A comparative Pharmacognostical Study of certain Jasminum Species

A thesis Presented by

The master Fatma Alzahraa Mokhtar Ali

For the degree of Ph.D in Pharmaceutical Sciences

(Pharmacognosy)

Under the supervision of

Prof. Dr. Seham Salah El-din El-Hawary Prof. of Pharmacognosy, Pharmacognosy Department, Faculty of Pharmacy, Cairo University

Prof. Dr. Hala M. El-Hefnawy Prof. of Pharmacognosy, Head of Pharmacognosy Department, Faculty of Pharmacy, Cairo University

Dr. Samir M. Osman Dr. Mohamed A. ELRaey

Assoc. Prof. of Pharmacognosy, Assoc. Prof. of Pharmacognosy, pharmacognosy Department, Phytochemistry and Plant Faculty of Pharmacy, October 6 Systematic Department, National University Research Centre

Pharmacognosy Department Faculty of Pharmacy 1

English Summary Cairo University 2019

English Summary

Abstract

Jasminum is a genus of flowering climbing shrubs and vines (family Oleaceae). Genus Jasminum composed of over 2000 plants distributed all over the world. They are native to Eurasia, India, and the Mediterranian region. Jasminum. azoricum L., Jasminum humile L., Jasminum multiflorum Burm.f.., Jasminum officinale L., Jasminum sambac L. (Arabian nights) and Jasminum sambac L. (Grand Duke of Tuscany) were collected from AL-Keram farms, Al-Beheira Government. The plants; were subjected to DNA fingerprinting using SCOT and ISSR primers for genetic characterization. The study was performed on biologically guided bases, so four in-vitro biological studies were performed; antioxidant, antimicrobial, cytotoxic effect on three cell lines and antihepatic C virus. The comparative biological studies showed that J. multiflorum leaves have higher antioxidant effects by DPPH assay, while J. officinale and J. sambac (G) flower extracts showed more antimicrobial effects than other species and cultivars. The cytotoxicity study showed that all the species possessed cytotoxic effects in different degrees, the higher cytotoxic effects observed in J. humile and J. sambac. In-vitro HCV activity was performed in addition to cell genes analysis by western blot technique, proved that J. multiflorum leaves and flowers can decrease the viral load and inhibit its replication by 85-88%. They inhibit viral replication by affecting NS5A protein and inhibit viral penetration and expansion of infection by affecting the EPAH2 gene. Comparative phytochemical investigations were performed on the six Jasminum plants, starting with phytochemical screening for the flowers and leaves. The HPLC-PDA-ESI-MS/MS technique was used to explore the chemical composition of the leaves, a total of 115 compounds were determined tentatively with interspecies variations, different classes of compounds were identified; 5 phenylethanoids, 19 simple phenols, 18 flavonoids, 7 lignans, and 66 secoiridoids. Estimation of flavonoids and phenolic component in each plant showed the highest phenolic contents in J. multiflorum, while similarity in flavonoid contents was observed in the six plants. Volatile components of the six plants were investigated using solid phase microextraction technique, that showed the interspecies differences of volatile oil composition among selected species and cultivars, the major components were; E,E-α-Farnesene, benzoyl acetate and cis-3- hexenyl benzoate. The quantitative determination of some phenolic acids and flavonoids of the methanolic extracts of leaves were performed using HPLC-DAD method.

English Summary As J. multiflorum showed a very high activity towards Virus C infection, J. multiflorum was the selected plant for the isolation of biologically active compounds. A total of 14 compounds were isolated from the leaves and flowers methanolic extracts include; secoiridoids, flavonoids, phenylethanoids and phenolic acid. Virtual docking screening was performed as antiviral and cytotoxic activities, that showed compound sambacoside and molihausides are cytotoxic with no carcinogenicity, while jasfloroside A and Jasfloroside B have the antiviral activity theoretically. Green synthesis of silver nanoparticles using leaves methanolic extracts of the selected Jasminum species and cultivars was performed and study their cytotoxic activities towards three cell lines; breast cancer (MCF-7), bladder (5637), and hepatocellular (Huh-7) cell lines. Keywords: Anti HCV; Docking; Jasminum; Oleaceae; Silver nanoparticles; Secoiridoids

English Summary

1. Introduction Jasminum is a genus of flowering climbing shrubs and vines (family Oleaceae). Genus Jasminum composed of over 2000 pants distributed all over the world, the flowers are mostly white in color except few are yellow. They are native to Eurasia especially Italy, Iran, India and Mediterranian region (Joshi, 2000). Jasminum flowers are characterized by a very pleasant aroma, used for thousands of years in perfume industry (Yang et al, 2008). Among Jasminum species; Jasminum azoricum L., Jasminum humile L, Jasminum multiflorum (Burm.f.), Jasminum officinale L, Jasminum sambac L. Ait. (Arabian nights) and Jasminum sambac L. Ait. (Grand Duke of Tuscany) are cultivated in Egypt for commercial purposes. Jasmine flowers are sensitive flowers of great importance in industrial, food, and medicinal applications. Small differences in morphological characteristics of the flowers are present especially number and arrangement of petals. Industrially, J. officinale and J. sambac are the major sources of raw material for the perfume industry as Jasmine oil (Groom, 1997). In fact, Egypt is considered the top producer of J. officinale concrete worldwide. The concrete is exported to France, England, and USA. (Wickens et al., 1989) for the production of high grade perfumes. The Egyptian concrete is synthesized mainly from the flowers of Jasminum officinale L by solvent extraction technique (Braun et al., 2009) and have a valuable contribution in the national income. Jasminum officinale and Jasminum sambac flowers are important edible flowers used worldwide as in infusion water, Jasmine tea in USA, Europe and Asia. Jasminum officinale leaves are used as a spice in Asian foods and the flowers are used as flavoring agent in high fancy desserts (Wickens et al., 1989). Phytochemical studies of different Jasminum species attracted the attention of many authors for their wide variety of constituents. These include mainly volatile constituents (Kaviani et al., 2014; Peyrot et al., 1995); iridoids (Hao et al., 2013); secoiridoids (Takenaka et al., 1998a; Tanahashi et al., 1999b), lignans (Ali, 2018) and flavonoids (Wang et al., 2012; Zhao et al., 2012). Extracts of the plant were reported to possess many biological activities as; anticancer (Akter et al., 2014; Kalaiselvi et al., 2011), antimicrobial activity (Nagarajappa et al., 2015), anthelmintic activity (Dullu, 2014), antioxidant (Alali et al., 2007), effects on CNS (Pal et al., 2007). Reviewing the current literature, nothing was reported concerning the different Jasminum species cultivated in Egypt.

English Summary The aim of work: Differentiate between the different Jasminum species cultivated in Egypt A- Genetically B- Chemically C- Biologically The present study includes: 1. DNA profiling of the selected Jasminum species and cultivarsto establish genetic criteria for their characterization 2. Biological study:  Evaluation of the antioxidant activity of the leaves extracts of the selected Jasminum species and cultivars  Evaluation of the antimicrobial activity of the flower extracts of the selected Jasminum species and cultivars.  Cytotoxic activity of the selected Jasminum species and cultivars towards breast cancer (MCF-7), hepatic cancer (Huh-7) and bladder cancer (5637) cell lines.  Antiviral activity of the selected Jasminum species and cultivars towards hepatic viral C infection. 3. Phytochemical study  Preliminary phytochemical screening of methanolic extracts of leaves and flowers of the selected Jasminum species and cultivars  HPLC-PDA-ESI-MS/MS analysis of chemical composition of the leaves of the selected Jasminum species and cultivars  Estimation of volatile oil composition flowers of the selected Jasminum species and cultivars by solid phase microextraction (SPME) technique and chemometric analysis  Qualitative and quantitative detrermination of phenolics and flavonoids in the selected Jasminum species and cultivars  Isolation of chemical constituents from the most bioactive fractions.  Virtual screening of some identified and isolated compounds using molecular docking. 4. Application Green synthesis of silver nanoparticles using leaves methanolic extracts of the selected Jasminum species and cultivars.

English Summary Review of literature Several reports focused on the chemical constituents of different Jasminum species and isolation of chemical components viz; iridoids, secoiridoids, phenolic acids, flavonoids and volatile oils, which have been reported as major components of these spp. 2.1. Chemical constituents of genus Jasminum 2.1.1. Iridoids and secoiridoids: Analysis performed on Jasminum tortuosum Willd, led to the isolation and the identification of four compounds, namely ginkgool, olivil-4′-O-β- glucopyranoside. oleoside dimethyl ester, and oleoside 11-methyl ester. The presence of these compounds is significant from a chemotaxonomic point of view (Tomassini et al., 2018). Eight compounds including four caffeoyl phenylpropanoid glycosides, jasnervosides A–D, one monoterpenoid glycoside, jasnervoside E, and three secoiridoid glycosides, jasnervosides F–H, were isolated from the stems of Jasminum nervosum Lour. (Oleaceae), along with four known compounds, poliumoside, verbascoside, α-l-rhamnopyranosyl-(1→3)-O-(α-l-rhamnopyranosyl (1→6)-1-O-E-caffeoyl-β-d-glucopyranoside, and jaspolyanthoside (Guo et al., 2014). Twelve compounds were isolated from Jasminum lanceolarium Roxb. two lignanoids and ten iridoids were identified. The existence of iridoids and lignanoids might be regarded as valuable chemotaxonomic markers for further classification and subdivision of the genus of Jasminum (Ning et al., 2013). Establishing the quality standards for total iridoid glycosides from Jasminum officinale L.var.grandiflorum. Color reaction and TLC for qualitative identification of total iridoid glycosides from Jasminum officinale L. var. grandiflorum. The effective part of total iridoid glycosides content was determined by UV and oleuropein content was estimated using HPLC (Hao et al., 2013). 2.1.2. Phenolic compounds Reviewing the literature, several classes of phenolic compounds such as phenylpropanoids, phenylethanoids, phenolic acids and flavonoids especially kaempferol and apigenin derivatives were reported in different Jasminum spp. Four phenylpropanoid glycosides, 9-O-(E-cinnamoyl)-coniferin, 6′-O-(E- cinnamoyl)-coniferin, 6′-O-(E-cinnamoyl)-syringin and 2′-O-(E-cinnamoyl)- syringin, together with two known phenylpropanoid glucosides, coniferin and ethylsyringin were obtained from the roots of Jasminum girialdii (Yue et al., 2017). Ten compounds were isolated from the barks of Jasminum giraldii; (+)- medioresinol, (+) -syringaresinol, syringaresinol-4'-O-beta-D-glucopyranoside, oleanic acid, 3-methoxy-4-hydroxy-trans-cinnamaldehyde, trans-sinapaldehyde, 7

English Summary syringaldehyde, 1-(4-methoxy -phenyl) -ethanol, trans-cinnamic acid, and 4-(1- methoxyethyl) -phenol (Zhang et al., 2014). 2.1.3. Volatile oils Jasminum multiflorum concrete extraction was carried out by solvent extraction with hexane. The chemical composition of the concrete was analysed by gas chromatography- mass spectrometry (GC-MS). The major chemical components detected were Triacontane; Tetratriacontane; 1-Pentanol, 4- methyl-2-propyl and Nonacosane (Ranchana et al., 2017a). The chemical composition of the Jasminum nitidum concrete was analysed by (GC-MS). The major chemical components detected are 1- Pentanol, 4-methyl- 2-propyl; Pentane, 3-ethyl-2,2-dimethyl-; Pentane, 2,2,3,4- tetramethyl (Ranchana et al., 2017b). 2.1.4. Terpenoids Six triterpenoid saponins were identified as 3-O-α-L-rhamnopyranosyl (1→2)-β-D-xylopyranosyl-hederagenin 28-O-β- D-galactopyranosyl (1→6)-β-D- galactopyranosyl ester, hederagenin -3-O-β- D-glucopyranosyl (1→3)α-L- arabinopyranoside, 2α,3β,23-trihydroxyolean-12-en-28-oic-O-β-D-glucopyranosyl ester, hederagenin-3-O-β-D-xylopyranosyl (1→3)-α-Lrhamnopyranosyl (1→2)-α- L-arabinopyranoside, 2α,3β,23-trihydroxyolean-12-en-28-oic-O-α-L-rhamnopyranosyl (1→4) -β-D-glucopyranosyl (1→6)-β-D-glucopyranosyl ester, hederagenin-3- O-α-L-rhamnopyranosyl (1→2)-α-L-arabinopyranoside (Zhao and Dong, 2008). Leaves of J. fexile, J.humie, J. multiflorum and J. sambac contained friedelin, lupeol, botulin, betulinic acid, α-amyrin, ursolic acid, oleanolic acid and β-sitosterol (Dan and Dan, 1985). 2.2. Biological studies of genus Jasminum

Extracts of the plants of genus were reported to possess cytotoxic, anti- inflammatory, anthelmintic, antiviral, hepatoprotective and CNS effects in addition to wound healing activity.

2.2.1. Cytotoxic activity:

Chemical investigation of Jasminum pentaneurum Hand.-Mazz led to the isolation and identification of 12 compounds. All the isolates were assayed for their inhibitory activity on three human cancer cells; lung cancer SK-MES-1, liver cancer SMMC-7721 and gastric cancer SGC-7901 with IC50 values ranging from 83.0 to 172.0 μM. (Wei et al., 2019). Ethanol extracts of Jasminum grandiflorum L. and Ficus religiosa L. were screened for cytotoxicity and antibacterial activity. Cytotoxic potential of the extracts were measured by using in vivo brine shrimp lethality assay. (Rahman et al., 2014).

English Summary 2.2.2. Anti-inflammatory activity Four new sesquiterpenoids，including three nor-cinalbicane type sesquiterpenoids, named Jasminol A, G, H and one eremophilene-type sesquiterpenoid, named Jasminol B with nine known compounds were isolated from the stems of Jasminum officinale L. The anti-inflammatory activity of isolated compounds was evaluated using lipopolysaccharide (LPS)-induced murine macrophage RAW264.7, and the new compounds exhibited a moderate inhibition of LPS-induced nitric oxide (NO) production in RAW264.7 cells with IC50 values of 20.56 ± 1.31, 30.12 ± 0.89, 30.35 ± 2.72 and 31.60 ± 1.69 μM, respectively, and CC50 values >200 uM (Lu et al., 2019). 2.2.3. Anthelmintic activity Anthelmintic activity of ethanolic extract of leaves of Jasminum mesnyi was carried out on adult Indian earthworm Eisenia fetida. In this study, 20 mg/mL and 40 mg/mL concentrations of plant extract were tested which involved the time of paralysis and time of death of worm. The extract exhibited significant anthelmintic acitivity which was more in higher concentration (Dullu, 2014). 2.2.4. Effect on GIT The investigation and evaluation of the gastroprotective effects of ethanolic extracts of Jasminum sambac leaves against acidified ethanol-induced gastric ulcers in rats. Immunohistochemistry showed overexpression of Hsp70 protein and down expression of Bax protein in rats pretreated with extract. Significant increase in the pH, mucus of gastric content and high levels of PGE2, SOD and reduced amount of MDA were observed (AlRashdi et al., 2012). 2.2.5. Antiviral activity Evaluation of the effect of 8-epi-kingiside (8-Epik) derived from the buds of Jasminum officinale var. grandiflorum (JOG) on hepatitis B virus (HBV) replication in Hep-G2 cell line in vitro and duck hepatitis B virus (DHBV) replication in ducklings in vivo. 8-Epik effectively blocked HBsAg secretion in HepG2 2.2.15 cells in a dose-dependent manner [IC50 = (19.4 ± 1.04) μg/mL]. 8-Epik (40 or 80 mg/kg, ip, twice daily) also reduced viremia in DHBV-infected ducks (Zhao et al., 2013). 2.2.6. Activities on CNS The Jasminum multiflorum Andr. extract showed significant analgesic properties as evidenced by the significant reduction in the number of writhes and stretches induced in mice by 1.2% acetic acid solution. Pretreatment with the extract caused significant protection against pentylene tetrazole induced convulsion. The behavioural studies on mice indicate CNS depressant activity of the ethanol extract of J. multiflorum (Pal et al., 2007)

English Summary The English summary Jasmine is a genus of flowering climbing shrubs and vines (Oleaceae family). Genus Jasminum composed of over 2000 pants distributed all over the world, the flowers mostly white in color except few are yellow, they are native to Eurasia especially Italy, Iran, India and mediterranian region. Jasminum flowers are characterized by very pleasant aroma, used for thousands of years in perfume industry. Among Jasminum species; Jasminum azoricum L., Jasminum humile L, Jasminum multiflorum (Burm.f.), Jasminum officinale L, Jasminum sambac Ait. (Arabian nights) and Jasminum sambac Ait. (Grand Duke of Tuscany) are cultivated in Egypt. The present stud includes: 1. Genetic Identification of the selected Jasminum species and cultivars based on SCoT and ISSR. In the present study, five SCoT primers were successful out of ten SCoT primers to determine SCoT polymorphism of the six the selected Jasminum species and cultivars. The resulted amplified bands and their densitometric analysis are illustrated. In the present study, five ISSR primers were successful out of fifteen ISSR primers to determine ISSR polymorphism of the selected Jasminum species and cultivar sunder study Results from this analysis showed two main groups: the first main group was included two species J. sambac (A) and J. sambac (G) the second main group was divided into two sub main groups: the first sub main group included specie J. azoricum alone and the second sub main group was divided into two sub sub group: the first sub sub group was included specie J. humile alone and the second sub sub group was included each of J. multiflorum and J. officinale species Part 1 Biological study In-vitro studies . Chapter 1: Estimation of the antioxidant activity of the leaves extracts of the selected Jasminum species and cultivars . Chapter 2: Estimation of the antimicrobial activity of the flower extracts of the selected Jasminum species and cultivars . . Chapter 3: Cytotoxic activity of the selected Jasminum species and cultivars towards breast cancer (MCF-7), hepatic cancer (Huh-7) and bladder cancer (5637) cell lines. . Chapter 4: Antiviral activity of the selected Jasminum species and cultivars towards hepatic viral C infection

English Summary Chapter 1: Estimation of the antioxidant activity of the leaves extracts of the selected Jasminum species and cultivars Studying the scavenging activities of J. azoricum L., Jasminum humile L., Jasminum multiflorum Burm.f.., Jasminum officinale L., Jasminum sambac L. (Arabian nights) and Jasminum sambac L. (Grand Duke of Tuscany) is a basic study to determine the antioxidant characters of these species. The antioxidant activity of each extract was determined by the DPPH free radical scavenging assay J.azoricum extract show no scavenging acticities with IC50 of 199.21 µg/ml, while other Jasminum species show moderate –high activities with IC50 values of 94.6, 34.8, 76.6, 130.7 and 155.5 for J. humile, J. multiflorum, J. officinale, J. sambac (A) and J. sambac (G) respectively, that indicate high antioxidant characters of J. multiflorum total methanolic extract compared to other Jasminum species and ascorbic acid reference standard Chapter 2: Estimation of the antimicrobial activity of the flower extracts of the selected Jasminum species and cultivars. The potential antimicrobial activities of the hexane extracts of J. azoricum, J. humile, J. multiflorum, J. officinale, J. sambac (A) and J. sambac (G) flowers were studied by determination of the zones of inhibition induced by agar well diffusion techniques as mentioned under. MICs were also determined by using serial microplate dilution technique using clinical isolates including three-gram negative bacteria; Enterobacter cloacae, Escherichia coli, Proteus vulgaris, three gram– positive bacteria; Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus mutant and filamintus fungi; Aspergillus fumigatus, Aspergillus flavus, Aspergillus niger and one yeast Candida albicans using Ketoconazole and Gentamycin Positive control for fungi and bacteria respectively. Proteus vulgaris is inhibited by J. sambac (A) with MIC of 7.5 µg/ml Chapter 3: Cytotoxic activity of the selected Jasminum species and cultivars towards breast cancer (MCF-7), hepatic cancer (Huh-7) and bladder cancer (5637) cell lines. The Cytotoxicity of the total methanolic extracts of of leaves of J. azoricum, J. humile, J. multiflorum, J. officinale, J. sambac (A) and J. sambac (G) were investigated using the neutral red uptake (NRU) assay. The IC50 values were defined from the obtained dose-response curves and expressed in mean ± SD. All compounds were tested in duplicate. Etoposide was used as positive control for MCF-7 breast cancer and Huh-7 hepatocellular cancer cell lines, and use Vincristine as positive

English Summary control (reference) for 5637 bladder cancer cell line with comparison to normal keratinocyte cell line (Ha Cat). We observed effects of J. azoricum towards (MCF-7, Huh-7 and 5637) cell lines but with no selectivity towords tumors indicated by low therapeutic index to all tested cell lines which indicate a toxic effect of J.azoricum to both normal and cancer cell lines so obviously we can neglect its results. All Jasminum species show moderate to high cytotoxicity toward MCF-7 breast cancer cell line except J. sambac (G) While all species show high cytotoxicity towards Huh-7 hepatic cancer cells, the efficiency are arranged in the descending order J.officinale > J. sambac (A) > J. humile > J. multiflorum > J. sambac (G). Although 5637 bladder cell line is considered a resistance cell line to medication treatments, Jasminum species show moderate to high cytotoxic effects especially J. sambac (A) Chapter 4: Antiviral activity of the selected Jasminum species and cultivars towards hepatic viral C infection. Total viral clearance in addition to effect on either positive stand (HCV genome) or negative (infective strand) in both Huh-7 infected cells with HCV was evaluated. The qualitative PCR for both total viral or differential strands was performed and the results showed reduction in viral load especially J. multiflorum leaves and flowers Western blot analysis of cell cycle genes proved that Jasminum multiflorum Inhibition of viral entry process was achieved through inhibition of receptor protein kinase EPHA2 (SAM domain) which is attractive target for antiviral therapy through inhibition of initiation and expansion the viral infection and control of hepatocellular carcinoma (HCC) related HCV infection oncogenesis. Part 2 Phytochemical study . Chapter 1: Preliminary Phytochemical Screening of methanolic extracts of leaves and flowers of the selected Jasminum species and cultivars . Chapter 2: HPLC-PDA-ESI-MS/MS analysis of chemical composition of the leaves of the selected Jasminum species and cultivars . Chapter 3: Estimation of volatile oil composition of flowers of the selected Jasminum species and cultivarsby solid phase microextraction (SPME) technique and chemometric analysis 12

English Summary . Chapter 4: Qualitative and Quantitative Estimation of Phenolics and Flavonoids in the selected Jasminum species and cultivars . Chapter 5: Isolation of chemical constituents of Jasminum multiflorum extracts as the most bioactive species against HCV. . Chapter 6:V irtual screening of some compounds using molecular docking. Chapter 1: Preliminary Phytochemical Screening of methanolic extracts of leaves and flowers of the selected Jasminum species and cultivars The air dried leaves and flowers of J. azoricum, J. humile, J. multiflorum, J. officinale, J. sambac (A) and J. sambac (G) were, separately, screened for: volatile substances, carbohydrates and/or glycosides, tannins, flavonoids, saponins, unsaturated sterols and/or triterpenes, alkaloids and/or nitrogenous bases, anthraquinones and cardiac glycosides to confirm the presence or absence of plant constituents in the leaves and flowers. Chapter 2

HPLC-PDA-ESI-MS/MS analysis of chemical composition of the leaves of the selected Jasminum species and cultivars The HPLC-PDA-ESI-MS/MS technique was used to explore the chemical composition of the leaves extracts of J. azoricum, J. humile, J. multiflorum, J. officinale, J. sambac (A) and J. sambac (G) and the compounds were tentatively identified on the bases of values mass peaks degradation and UV absorbance values in comparison to the previously published data, a total of 115 compounds were determined tentatively with interspecies variations, different classes of compounds were identified; 5 phenylethanoids, 19 simple phenols, 18 flavonoids, 7 lignans, and 66 secoiridoids. Chapter 3: Estimation of volatile oil composition of flowers of the selected Jasminum species and cultivarsby solid phase microextraction (SPME) technique and chemometric analySolid phase micro-extraction coupled to gas chromatography- mass spectroscopy (SPME-GC-MS) led to identification of 71 volatile compounds distributed among Jasminum species with the following findings:  The major volatile constituents in J. azoricum are Benzyl acetate  The identification of 29 volatile compound from J. humile, The major volatile constituents are E,E-α-Farnesene  The Identification of 30 volatile compounds from J. multiflorum, the major compounds are E,E-α-Farnesene

English Summary  The identification of 42 volatile compounds from J.officinale, the major compounds are Benzyl acetate  The identification of 45 volatile compounds from J. sambac (M), the major compounds are Benzyl acetate  The identification of 26 volatile compounds from J. sambac (G), the major compounds are E,E-α-Farnesene  All species showed the presence of cis-3-Hexenyl benzoate, E,E-α- Farnesene, trans-α-Bergamotene, β-(Z)-Farnesene in different concentrations. Chapter 4:

Qualitative and Quantitative Estimation of Phenolics and Flavonoids in the selected Jasminum species and cultivars A. Spectroscopic method Total phenolics were expressed as gallic acid equivalent using Folin- Ciocalteu method and total flavonoids were expressed as Quercetin equivalent using aluminum chloride colorimetric method. J. multiflorum have the highest phenolics value, extremely different comparable to other planwhile small differences in total flavonoids were observed among the selected Jasminum species and cultivars. B- Quantitative determination of phenolic compounds by HPLC-DAD The quantitative determination of phenolic acids and flavonoids of the methanolic extracts of leaves of J. azoricum, J. humile, J. multiflorum, J. officinale, J. sambac (A) and J. sambac (G) was performed using high performance liquid chromatography coupled to diode-array detector using, the samples were injected against 20 reference standards for the quantitative determination of detected compounds the analysis revealed the following results: Hydroxytyrosol is a phenylethanoid compound present in high concentration in J. multiflorum Protocatechuic acid is present with high concentration in J. multiflorum followed by J. officinale P-hydroxybenzoic acid is detected in a high concentrations in J. multiflorum higher than other species, followed by J. officinale Chlorogenic acid is detected in high concentration in J. officinale Syringic acid was not detected in J. multiflorum but detected in other species in low concentrations. Rutin flavonoid was determined in the six species; it was detected in high concentration in J. azoricum followed by J. sambac (G) P-coumaric acid was determined in the selecyed species; it was detected in high concentration in J. azoricum followed by J. humile 14

English Summary Chapter 5: Isolation of chemical constituents of Jasminum multiflorum extracts as the most bioactive species against HCV. A- Isolation of the constituents of the ethyl acetate Fraction of J. multiflorum leaves The ethyl acetate fraction 22.5 g was loaded with silica gel for VLC on VLC column led to fifteen fractions, fractions four, seven, eight and ten were subjected to column chromatography and paper chromatography, led to the isolation of eight compounds: Fraction 4 led to isolation of three compounds  Compound J1: hydroxyoleuropein  Compound J2: Jasmultiside  Compound J3: Multifloroside Fraction 7 led to the isolation of one compound  Compound J4: Kempferol 3-O-neohesperidoside Fraction 8 led to the isolation of three compounds  Compound J5: Astragalin (Kaempferol-3-O-glucoside)  Compound J6: Hydroxytyrosol  Compound J7: tyosol-6-O-glucoside Fraction 10 led to isolation of one compound  Compound J8: Protocatechuic acid B- Isolation of the constituents of the butanol fraction of J. multiflorum leaves Extraction and isolation The butanol fraction 15 g was loaded on the HP20 column, led to 10 fractions. Fraction 3 was subjected to column chromatography and paper chromatography to isolate three compounds Compound J9: Jasfloroside A Compound J10: Jasfloroside B Compound J11: Kaempferol 3,7,4`triglucoside C- Isolation of the constituents of the ethyl acetate fraction of the J. multiflorum flowers The ethyl acetate fraction 20.0g was loaded with silica gel for VLC on VLC column, led to 13 fractions. Fraction 6 was subjected to paper chromatography and column chromatography, led to isolation of eight compounds

English Summary  Compound J12: Jaspolyanthoside B  Compound J13: kaempferide-3-O-glucoside  Compound J14: oleuropein aglycone In addition to compounds previously isolated from the flowers: J2, J3, J4, J5, J6 Chapter 6: Virtual screening of some compounds using molecular docking. The virtual screening was performed for the most common compounds identified in Jasminum extracts. By using CDOCKER protocol to predict the proposed binding mode, affinity, preferred orientation of each docking pose and binding free energy (∆G) of the tested compounds with MST3 protein, NS5A protein and EphA2 All tested compounds isolated from J. multiflorum leaves and flowers were subjected to molecular docking for the detection of the possible potent compounds toward HCV-NS5A and SAM domain in EPHA2, Jasfloroside A followed by multifloroside and jasfloroside B showed a high theoretical affinity against HCV- NS5A suggesting they are possible potent molecules for drug development against HCV infection.

Part 3

Application Green synthesis of silver nanoparticles using leaves methanolic extracts of the selected Jasminum species and cultivars. The formed AgNPs synthezised by J. azoricum, J. humile, J. multiflorum, J. officinale, J. sambac (A) and J. sambac (G). were examined for their cytotoxic activities towards three cell lines; breast cancer (MCF-7), bladder (5637) and hepatocellular (Huh-7)S cell lines.

English Summary

Conclusion and Recommendations

Conclusion:

The present study had led to the following results:

By combining SCoT and ISSR, the highest similarity was between each of J. sambac (A) and J. sambac (G) at 1.00 and the lowest similarity was between species Jasminum azoricum and J. humile species The Jasminum extracts showed antioxidant, antimicrobial, cytotoxicity and antiviral activities in different degrees J. multiflorum leaves possess a higher antioxidant activity followed by J. officinale Jasminum flowers have antimicrobial activities toward some gram –ve and gram +ve bacteria in different degrees J. officinale have the higher cytotoxic effects on MCF-7 and Huh-7 cell lines. J. sambac (A) have the higher cytotoxic effect on 5637 bladder cancer cell line. J. multiflorum leaves have the higher cytotoxicity toward Hep-G cell line Extracts of the genus Jasminum after defatting could be a rich source of bioactive compounds especially, secoiridoids, phenylethanoids, phenolic acids and flavonoids Species of Jasminum under investigations are rich sources for phenolic compounds and secoiriodoids especially J. multiflorum as evidenced by HPLC-PDA-MS/MS. The chemical composition of the volatile aroma from the J. azoricum flowers is completely not correlated to other species. J. multiflorum leaves and flowers possess a remarked anti HCV activity in cancer infected cells; Jasfloroside A, multifloroside and jasfloroside B showed high theoretical affinity against HCV-NS5A and EPHA2 suggesting they are possible potent molecules for drug development against HCV infection (initiation, penetration and replication of virus) J. sambac (A) AgNPs and J. humile AgNPs are considered powerful cytotoxic agents against breast (MCF-7), bladder (5637) and hepatic (Huh-7) cancer cell lines

English Summary

Recommendations:

 Individual isolated compounds from J. multiflorum (especially jasfloroside A, jasfloroside B, and multifloroside) should be subjected to in-vitro and in-vivo studies on virus C to discover the actual mode of action experimentally.  Isolation of active compounds of J. officinale and J. sambac (A) responsible for the cytotoxic effects.  Further experimental in vivo studies to are needed to explore the cytotoxicity mechanism of the formed nanoparticles