الفهرس 
Aim of the workOnly 14 pages are availabe for public view
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Abstract
Summary
This thesis entitled includes three chapters and ends with a list of references.
The first chapter covers the literature survey up to date of benzoxazinoenes, Quinazolinones and 1,3,5 triazine derivatives synthesis and application.
The second chapter consists of the equipment, chemicals and software used in the work. It also includes two parts: Chemistry part: preparation of 2-phenyl-4H-benzo[d][1,3]oxazin-4-one (170), 3-hydroxy-2-phenylquinazolin-4(3H)-one (172), 4-hydrazinyl-2-phenyl-3,4-dihydroquinazoline (174), 3-amino-2-phenylquinazolin-4(3H)-one (175), N-(2-(hydrazinecarbonyl)phenyl)benzamide (176), 3-((4,6-dichloro-1,3,5-triazin-2-yl)amino)-2-phenyl-2,3-dihydroquinazolin-4(4aH)-one (177), 6-chloro-2,4-diamino acid-1,3,5-triazine derivatives (178a,b), 2,2’-((6-((1-oxo-3-phenyl-3,8a-dihydroisoquinolin-2(1H)-yl)amino)-1,3,5-triazine-2,4-diyl)bis(azanediyl))diacetic acid (179a,b). ((4-chloro-6-((4-oxo-2-phenyl-4,4a-dihydroquinazolin-3(2H)-yl)amino)-1,3,5-triazin-2-yl)amino) acids (180a,b), 2-((4-hydroxy-6-((4-oxo-2-phenyl-4,4a-dihydroquinazolin-3(2H)-yl)amino)-1,3,5-triazin-2-yl)amino) acids (181a,b), 3-((4,6-dihydroxy-1,3,5-triazin-2-yl)amino)-2-phenyl-2,3-dihydroquinazolin-4(4aH)-one (182) and N-(2-(2-(4-sustituted benzylidene) hydrazinecarbonyl)phenyl)benzamide (183a-e).
Biology part: Cytotoxicity screening, morphological examination of the apoptosis induced by the most active compounds and flow cytometric analysis of apoptosis
The third chapter deals with the discussion and interpretation of the results obtained from the above-mentioned chapter.
Preparation of 2-phenyl-4H-benzo[d][1,3]oxazin-4-one 170 and 3-hydroxy-2 phenylquinazolin-4(3H)-one (172): 2-phenyl-4H-benzo[d][1,3]oxazin-4-one (170) was prepared from anthranilic acid and benzoyl chloride in pyridine as reported.120 The prepared 2-phenyl-benzoxazinone was used as starting material for the preparation of different nitrogen containing heterocyclic compounds as follows. First 2-substituted benzoxainones were prepared by the reaction of 2-phenyl-4H-benzo[d][1,3]oxazin-4-one (170) with different nitrogen nucleophiles as shown in (Scheme 51).
Scheme 51. Preparation of 2-phenylquinazolin-4(H)-one 171 and 3-hydroxy-2-phenylquinazolin-4(3H)-one 172
2-phenylquinazolin-4(H)-one (171) was prepared by the reaction of 2-phenyl-4H benzo[d][1,3]oxazin-4-one (170) with equimolar amount of formamide. The reaction proceeded by heating under reflux for 2 h. The reaction mixture was then poured into crushed ice and then filtered. The pure product is then obtained by recrystallization from dioxane. While 3-hydroxy-2-phenylquinazolin-4(3H)-one (172) was prepared by the reaction of benzoxazinone derivative (170) with 3 equivalents of hydroxylamine hydrochloride in ethanol. After the reaction was completed the reaction mixture cooled, poured into iced HCl. The crude product was filtered off and purified by recrystallized from ethanol.
The 4-hydrazinyl-2-phenyl-3,4-dihydroquinazoline (174) was prepared from 2-phenylquinazolin-4(H)-one (171) in two steps, first chlorination at position 4 of quinazoline ring occurred by the reaction of compound 171 with mixture of phosphorus oxychloride and phosphorous pentachloride on water bath for 2 h. After the reaction was completed the reaction mixture was cooled then poured slowly into crushed ice. The solid formed was filtered off, washed with water and purified by crystallization with ethanol. Hydrazinolysis of the pure 4-chloro derivative 173 in butanol yielded the corresponding 4-hydrazinyl-2-phenyl-3,4-dihydroquinazoline( 174) (Scheme 52). The structure of the chloro derivative 173 and the hydarzino derivative 174 were confirmed by spectroscopic methods.
Scheme 52. Preparation of 4-hydrazinyl-2-phenyl-3,4-dihydroquinazoline (174)
Preparation of 3-amino-2-phenylquinazolin-4(3H)-one (175) and N-(2 (hydrazinecarbonyl) phenyl)benzamide (176): Hydrazinolysis of 2-phenyl-4H-benzo[d][1,3]oxazin-4-one (170) gives two different products depending on the reaction conditions. 121, 122 Depending on the duration of the reaction time and the strength of the given wave irradiation the lack of microwave irradiated power and a relatively short reaction time leads to compounds 176. 122 The proposed mechanism is shown in Scheme 3. The nucleophile attack of hydrazine hydrate on C carbonyl lactone ring causes opening of lactone ring and forms amide groups to obtain N-(2-(hydrazinecarbonyl)phenyl)benzamide derivative (176). While excessive microwave irradiation and relatively long reaction time or heating under reflux in presence of EtOH 121 resulted in intramolecular attack of -NH at C=O amide followed by H2O release. The nucleophile -NH attack causes the cyclization of the quinazolinone ring and afforded compound 3-amino-2-phenylquinazolin-4(3H)-one derivative (175), Scheme 53.
Scheme 53. Preparation of 3-amino-2-phenylquinazolin-4(3H)-one 175 and N-(2-(hydrazinecarbonyl)phenyl)benzamide (176).
Preparation of 3-((4,6-dichloro-1,3,5-triazin-2-yl)amino)-2-phenyl-2,3-dihydroquinazolin-4(4aH)-one (177), 6-chloro-2,4-diamino acid-1,3,5-triazine derivatives (178 a,b) and 2,2’-((6-((1-oxo-3-phenyl-3,8a-dihydroisoquinolin-2(1H)-yl)amino)-1,3,5-triazine-2,4-diyl)bis(azanediyl))diacetic acid (179a,b): The monosubstituted s-triazine derivative( 177) was prepared via nucleophilic aromatic substitution of one chlorine atom of cyanuric chloride at 0-5C with the prepared 3-amino-2-phenylquinazolin-4(3H)-one (175). The reaction occurred by stirring for 2 h at 0oC using dioxane / water as solvent. Sodium carbonate was used to capture the liberated HCl. (Scheme 4). The monochlorinated s-triazine derivatives (178a,b) were prepared via nucleophilic aromatic substitution of two chlorine atoms from cyanuric chloride by two molecules of glycine and β-alanine. The reaction was carried out in acetone/water solvent mixture and in the presence of sodium carbonate as a base to scavenge the liberated HCl. Then neutralization with dil HCl to obtain the desired product (Scheme 54).
Scheme 54. Preparation of 3-((4,6-dichloro-1,3,5-triazin-2-yl)amino)-2-phenyl-2,3-dihydroquinazolin-4(4aH)-one( 177), 6-chloro-2,4-diamino acid-1,3,5-triazine derivatives (178a,b) and 2,2’-((6-((1-oxo-3-phenyl-3,8a-dihydroisoquinolin-2(1H)-yl)amino)-1,3,5-triazine-2,4-diyl)bis(azanediyl))diacetic acid( 179a,b).
The C3-symmetrical s-triazine derivatives (179 a, b) were prepared by two different methods depending on the order of replacing chlorine atoms of cyanuric chloride with the corresponding N-Nucleophiles. First C3-symmetrical s-triazine derivatives (179 a, b) were prepared by the reaction of the solution of 3-amino-2-phenylquinazolin-4(3H)-one (175) in dioxane with the aqueous solution of diamino acid-1,3,5-triazine derivatives (178a,b) in the presence of Na2CO3. The reaction mixture was heated under reflux for 24 h. The desired pure product was filtered off, washed well with water and dried. Second, C3-symmetrical s-triazine derivatives (179 a, b) were prepared by the reaction of a solution of the previously prepared 3-((4,6-dichloro-1,3,5-triazin-2-yl)amino)-2-phenyl-2,3-dihydroquinazolin-4(4aH)-one (177) in dioxane with the aqueous solution of glycine or β-alanine in presence of Na2CO3. The reaction mixture heated under reflux for 12 h. The desired pure product filtered off, washed well with water and then dried.
Preparation of ((4-chloro-6-((4-oxo-2-phenyl-4,4a-dihydroquinazolin-3(2H)-yl)amino)-1,3,5-triazin-2-yl)amino) acids (180a,b), 2-((4-hydroxy-6-((4-oxo-2-phenyl-4,4a-dihydroquinazolin-3(2H)-yl)amino)-1,3,5-triazin-2-yl)amino) acids (181a,b) and 3-((4,6-dihydroxy-1,3,5-triazin-2-yl)amino)-2-phenyl-2,3-dihydroquinazolin-4(4aH)-one( 182).
Newly prepared C3-symmetrical s-triazine derivatives 180a,b, 181a,b and 182 were prepared from the dichloro 1,3,5-triazine derivative (177) as shown in (Scheme 5). First, a solution of 3-((4,6-dichloro-1,3,5-triazin-2-yl)amino)-2-phenyl-2,3-dihydroquinazolin-4(4aH)-one (177) in dioxane was added dropwise with stirring to a stirred mixture of the amino acids and Na2CO3 in water. The reaction mixture was stirred overnight at room temperature, neutralized with hydrochloric acid till complete precipitation, filtered off and washed well with acetone and dried to obtain the desired monochloro triazine derivatives 180a, b which were used for preparation of the tri substituted triazine by stirring overnight with solution of aqueous NaOH, followed by reflux for 4 h. The reaction mixture was then cooled till complete precipitation. The desired product 181 a,b filtered off, wash well with water and acetone and dried. On the other hand, the chlorinated s-triazine derivative 177 was converted to the hydroxy s-triazine derivative 182 reaction with aqueous sodium hydroxide solution at room temperature for 24 h under stirring, followed by reflux for 4 h. (Scheme 55).
Scheme 55. Preparation of ((4-chloro-6-((4-oxo-2-phenyl-4,4a-dihydroquinazolin-3(2H)-yl)amino)-1,3,5-triazin-2-yl)amino) acids (180a,b), 2-((4-hydroxy-6-((4-oxo-2-phenyl-4,4a-dihydroquinazolin-3(2H)-yl)amino)-1,3,5-triazin-2-yl)amino) acids( 181a,b) and 3-((4,6-dihydroxy-1,3,5-triazin-2-yl)amino)-2-phenyl-2,3-dihydroquinazolin-4(4aH)-one (182).
Preparation of N-(2-(2-(4-sustituted benzylidene) hydrazinecarbonyl)phenyl)benzamide (183a-e): Different hydrazones 183a-e were prepared by the reaction of N-(2-(hydrazinecarbonyl)phenyl)benzamide (176) with different aromatic aldehydes by reflux in ethanol in presence of 2 drops of acetic acid for about 6 h. Hydrazones precipitated during the reaction, collected by filtration, then recrystallized from ethanol to afford the pure product (Scheme 56).
Scheme 56 Preparation of N-(2-(2-(4-sustituted benzylidene) hydrazinecarbonyl)phenyl) benzamide (183a-e).
The structure of all prepared compounds was determined by spectroscopic methods (IR, 1H NMR and 13C-NMR).
Biological evaluation: Cytotoxicity screening: Based on the detected EC100 values on normal human cells all the evaluated compounds showed better safety profiles compared to doxorubicin except 177, 178a, 179a, 179b, and 180a. Among the investigated newly prepared compounds 180b was the safest recording the highest EC100 value, followed by 183a, 183d, 180a. Promising safety profiles were also observed for Schiff’s bases 183b, 183c, and 183e as well as the beta alanine-substituted one 181b. Schiff’s bases 183a, 183b and 183d exhibited outstanding anticancer activities with single-digit nanomolar IC50 values against triple negative breast cancer cells; MDA-MB 231 being more potent than doxorubicin as well as all the evaluated new compounds and precursors in the current study. Besides, this observation its safety profile was promising. ((4-chloro-6-((4-oxo-2-phenyl-4,4a-dihydroquinazolin-3(2H)-yl)amino)-1,3,5-triazin-2-yl)amino) acids (180a) and (180b) showed slightly less potent anticancer activities as evidenced by the IC50 values against MDA-MB 231. On the other hand, compounds 180a, 181a, 181b, and 183 a-d were more active than doxorubicin against liver cancer cells; HepG-2b.
The two cancer cell lines (MDA-MB 231 and HepG-2) were examined for morphological changes when treated with the most active compounds in comparison with the untreated cancer cells and cells treated with the reference doxorubicin. All the treated cells obviously lost their normal shapes. Additionally, their characteristic severe shrinkage indicated potent anticancer activities of the tested compounds, especially Schiff’s base 183a, in comparison to doxorubicin.
Schiff’s base 183a as well as the Schiff’s bases 183b and 183d possessed higher capability to induce apoptosis (>61%) in the tested human cancer cells than doxorubicicn (<32%). Schiff’s base 183a allowed higher potential to induce apoptosis (>65.59%) in MDA-MB 231 and HepG-2). These results were consistent with MTT assay results.