الفهرس يوجد فقط 14 صفحة متاحة للعرض العام
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المستخلص
General summary br Citrus fruits are one of the most popular foods in the world, with worldwide agricultural production over 100 million metric tons per year. br They have been shown to possess many constituents which have important effects on the human health: vitamin C, carotenoids (β–carotene and lycopene), limonoids, flavonoids, coumarins, acridone alkaloids, high quality soluble fiber, vitamin–B complex and related nutrients (thiamine, riboflavin, nicotinic acid/ niacin, pantothenic acid, pyridoxine, folic acid, biotin, choline, and inositol). br Citrus limonoids are biologically active, displaying anti-carcinogenic activity in laboratory animals, cultured human cancer cell lines. In addition, antitumor activity against benzo[α] pyrene induced neoplasia in the fore – stomach of mice and against azoxymethane induced colon carcinogenesis in rats. Moreover they have insecticidal activity against mosquito Culex quinquejasciatus larvae and possessed anti-inflammatory activity against bradykinin – induced paw edema and arachidonic acid induced ear swelling and antifungal activity against Puccinia arachidis. Also, it was reported that limonoids possessed hypoglycemic activity br Flavonoids show a great variety of functions, ranging -#102;-#114;-#111;-#109; their ecological and biological functions in the accumulating organisms to their exogenous biological activity. Among these, antioxidant, antiviral, antiallergic, cardioprotective, and anticarcinogenic effects have been investigated. br Coumarins isolated -#102;-#114;-#111;-#109; Citrus plants were found to possess antibacterial, antifungal, cardiovascular, antiplatelet and antitumor effects. br It was reported that acridone alkaloids isolated -#102;-#114;-#111;-#109; many species related to genus Citrus, posses antiviral, antimicrobial, antifungal, mutagenic, antineoplastic effects, anti-herpes and antimalarial activities. br Citrus essential oils were found to exhibit activity against human tumor cell lines (MCF-7 and Hela). Also they have antifungal and antioxidant activities. br Reviewing the current literature, few reports were found dealing with the chemical constituents of our Citrus species, while the constituents of the parent plants C. medica, C. reticulata and C.limon were the subject of much more studies. br In the view of the previously mentioned points it was of great value to carry out phytochemical investigation and biological activities of some Citrus species growing in Egypt. br Citrus species under investigation: br • Citrus jambhiri Lush. (rough lemon) a hybrid of (C. medica X C. reticulata) br • Citrus pyriformis Hassk (ponderosa lemon) a hybrid of (C.limon X C. medica). br This study included two parts as follow; br Part I: Phytochemical investigation of Citrus jambhiri Lush. and Citrus pyriformis Hassk using different methods. br This part included seven chapters as follow; br Chapter I: Phytochemical investigation of Citrus jambhiri Lush. peel. br The petroleum ether, chloroform and ethyl acetate soluble fractions of the alcoholic extract were chromatographed over silica gel column followed by purification using preparative TLC and / or fractional crystallization to give 7, 6 and 3 compounds -#102;-#114;-#111;-#109; petroleum ether, chloroform and ethyl acetate fractions, respectively. The identification of these compounds was depending on spectral data (Ms: ESI, EI, FAB and HR/ESI, 1H-, 13C-NMR and 2D) and comparison with the authentic standard when available. These compounds were identified as follow; br A- Petroleum ether fraction: br 1. RLP1: mixture of β-sitosterol and stigmasterol. br 2. RLP2: Palmitic acid. br 3. RLP3: 5-Hydroxy-3, 6, 7, 8, 3’, 4’-hexamethoxyflavone. br 4. RLP4: 5-Demethylnobiletin*. br 5. RLP5 BI: 3, 5, 6, 7, 8, 3’, 4’-Heptamethoxyflavone. br 6. RLP5 BII: Tangeretin. br 7. RLP6: Nobiletin. br B- Chloroform fraction: br 1. RLC1: 5-Demethylnobiletin. br 2. RLC2: 5, 4’-Dihydroxy-6, 7, 8, 3’-tetramethoxyflavone. br 3. RLC3: Nobiletin. br 4. RLC4: Nomilin. br 5. RLC5: Limonin. br 6. RLC6: 6-Demethoxyflavone. br C- Ethyl acetate fraction: br 1. RLEt1: Naringin. br 2. RLEt2: Hesperidin. br 3. RLEt3: Neohesperidin. br All the isolated compounds in this study except hesperidin, neohesperidin and tangeretin, represent the first time for isolation -#102;-#114;-#111;-#109; C. jambhiri. br Chapter IV: Phytochemical investigation of Citrus pyriformis Hassk peel. br Dichloromethane and ethyl acetate soluble fractions obtained -#102;-#114;-#111;-#109; the alcoholic extract of the peel were chromatographed using silica gel column followed by purification through preparative TLC and / or fractional crystallization to give 7 and 3 compounds -#102;-#114;-#111;-#109; dichloromethane and ethyl acetate, respectively. Identification of the isolated compounds was carried out using physical, chemical and spectral means as (Ms: ESI, EI, FAB and HR/ESI, 1H-, 13C-NMR and 2D) in direct comparison with the authentic samples. These compounds were identified as follow; br A- Dichloromethane fraction: br 1. K-1: Citropten. br 2. K-2: Stigmasterol. br 3. K-3’: Ergosterol. br 4. K-3: Sitosteryl 6’-O-acyl -3 β –D-glucoside. br 5. K-4: Limonin. br 6. K-4: Deacetylnomilin. br 7. K-5: Sitosteryl β –D-glucoside. br B- Ethyl acetate fraction: br 1. P Et-1 is Naringin. br 2. P Et-2 is Hesperidin. br 3. P Et-3 is Neohesperidin. br All the isolated compounds in this study except hesperidin, neohesperidin and ergosterol, represent the first time for isolation -#102;-#114;-#111;-#109; C. pyriformis. Sitosteryl 6’-O-acyl -3 β –D-glucoside represents the first time of isolation -#102;-#114;-#111;-#109; genus Citrus. br Chapter V: Identification and quantitative determination of flavonoid glycosides in both Citrus species. br br Flavonoidal content obtained -#102;-#114;-#111;-#109; the different organs of Citrus jambhiri Lush. and Citrus pyriformis Hassk peels was determined using LC-MS technique and 7 flavonoid glucosides have been identified in the ethyl acetate fractions. Identification was carried out using authentic samples, whenever possible in addition to chromatographic and spectroscopic means. br Hesperidin is detected in all ethyl acetate fractions but diosmetin 6-C-β-D-glucoside has not been found in Citrus pyriformis Hassk. br Also, hesperidin represents the main peak in the chromatogram of Citrus jambhiri peel while neohesperidin is the major peak in Citrus pyriformis rind chromatogram. br Chapter VI: Phytochemical investigation of limonoids in the seeds of Citrus jambhiri Lush. and Citrus pyriformis Hassk. br Sun-dried Citrus seeds of C. jambhiri Lush.(290 gm) and C. pyriformis Hassk (100 gm) were cleaned, powdered and defatted with petroleum ether followed by soaking overnight with MeOH at room temperature. br The MeOH extract was diluted with H2O, extracted 3 times with CH2Cl2 and the solvent evaporated under vacuum to yield 9 g and 3 g -#102;-#114;-#111;-#109; C. jambhiri Lush. and C. pyriformis, respectively. br Dichloromethane fractions -#102;-#114;-#111;-#109; both seeds were subjected to HPLC-MS to identify the limonoid aglycones, -#119;-#104;-#101;-#114;-#101; 11 compounds were identified -#102;-#114;-#111;-#109; C. pyriformis and 7 compounds were detected only in C. jambhiri Lush. br Crystallization of CH2Cl2 fraction obtained -#102;-#114;-#111;-#109; C. jambhiri Lush. afforded the isolation of limonin and nomilin in pure form. While, column chromatography of the CH2Cl2 fraction obtained -#102;-#114;-#111;-#109; C. pyriformis resulted in the isolation of the following compounds; br 1. PS1: Cholesterol. br 2. PS2: Stigmasterol. br 3. PS3: Nomilin. br 4. PS4: Sitosteryl 6’-O-acyl -3 β –D-glucoside. br 5. PS5: Limonin. br 6. PS6: Ichangin. br 7. PS7: Deacetylnomilin. br 8. PS8: Sitosteryl-β-D-glucoside. br All the isolated compounds in this study represent the first time for isolation -#102;-#114;-#111;-#109; C. pyriformis Hassk seeds. br Chapter VII: Comparative analyses of the flavors -#102;-#114;-#111;-#109; Citrus jambhiri Lush. and Citrus pyriformis Hassk with GLC-MS. br The essential oils were prepared -#102;-#114;-#111;-#109; the peel and juice of Citrus jambhiri Lush. and Citrus pyriformis by steam distillation and solvent extraction, respectively. The yield percentage and the physical characters of the obtained oils were determined. The oils were analyzed using capillary GlC and GlC-MS. The study of the essential oils revealed that, altogether 94 compounds were identified. In both species, the major component was found to be limonene. In C. jambhiri, a total of 84 components were identified -#102;-#114;-#111;-#109; which 70 in fruit rind hydrodistilled oil. On the other hand, Solvent extracted rind and juice were found to contain 71 and 44 components, respectively. Sixty five compounds were identified in the hydrodistilled oil of C. pyriformis. A total of 77 and 31 compounds were identified in the rind and juice extract, respectively. Solvent extraction has the advantage of recovering the higher molecular weight natural compounds. -#102;-#114;-#111;-#109; these compounds: tangeretin, heptamethoxyflavone, hexamethoxyflavone and nobiletin were identified in detectable amount in the rind extract of C. jambhiri while, osthol, meranzin, isomeranzin and 5-isopentenyloxy-7-methoxycoumarin were identified (as coumarins) in C. pyriformis. Moreover, citropten, bergapten, α-tocopherol, stigmasterol and β-sitosterol were found in both species. br Part II: Biological activities of Citrus jambhiri Lush. and Citrus pyriformis Hassk. br This part includs five chapters and all the biological tests represent the first report form both species. br Chapter I: DPPH free radical scavenging activity. br This study revealed that the total extracts, fractions, pure compounds and essential oils obtained -#102;-#114;-#111;-#109; both species possessed antioxidant activity by scavenging DPPH free radical. br Neohesperidin which was isolated -#102;-#114;-#111;-#109; the peel of C. pyriformis Hassk and C. jambhiri Lush. demonstrated stronger activity than hesperidin. Polymethoxylated flavonoids which isolated -#102;-#114;-#111;-#109; petroleum ether fraction of peel of rough lemon showed higher IC50 but these compounds together in the fraction showed synergistic effect. br It was found that, the essential oil -#102;-#114;-#111;-#109; C. pyriformis Hassk showed strong antioxidant in comparison with the other oil -#102;-#114;-#111;-#109; C. jambhiri Lush. br Chapter II: Inhibition of 5-lipoxygenase by essential oils, extracts and fractions. br br The 5-LOX assay of essential oils, total extracts and different fractions of C. jambhiri Lush. and C. pyriformis Hassk was studied. br Citrus essential oils demonstrated a very good activity against 5-LOX. There is a good correlation between the activities of d-limonene with those of citrus oils evaluated. This should not be surprising since d-limonene is the main component of C. jambhiri Lush. and C. pyriformis oils. Moreover, the petroleum ether fraction obtained -#102;-#114;-#111;-#109; both species showed higher activity compared with the other fractions. br Chapter III: In vitro antitrypanosomal activity using resazurin dye. br This study was applied on the total methanol extracts of the peel of C. jambhiri and C. pyriformis, hesperidin -#102;-#114;-#111;-#109; ethyl acetate of C. jambhiri and neohesperidin -#102;-#114;-#111;-#109; C. pyriformis and essential oils -#102;-#114;-#111;-#109; both species. It was found that, the essential oils of both species showed substantial activity against Trypanosoma brucei brucei compared with the other tested samples. br Chapter IV: Antimicrobial Activity. br The essential oils of both species showed antimicrobial activity against all tested Gram (+) bacteria and antifungal activity against all tested strains. The oils possess no activity against Gram (-) strains except Pseudomonas fluorescens. br Chapter V: MDR Reversal and cytotoxicity by secondary metabolites -#102;-#114;-#111;-#109; C. jambhiri and C. pyriformis. br br It is confirmed during this study that Citrus compounds can inhibit p-gp dependent efflux and lead to accumulation of antitumor drug inside tumor cell. br Most of the compounds especially polymethoxyflavones exhibited cytotoxicity against different cell lines. br In conclusion: br The biological studies in this work (part II) represent the first time for C. jambhiri and C. pyriformis. br Concerning the MDR reversal, this study demonstrates that some Citrus compounds are good substrates of p-gp. Since they show a low cytotoxicity they could be considered as a promising lead compounds for the design of more efficient MDR chemosensitizers or reversal agents. br Recommendation, daily intake of Citrus fruits protects human body -#102;-#114;-#111;-#109; chronic disease.