الفهرس 
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Abstract
Marine algae or seaweeds are one of the natural resources essentially for producing various bioactive secondary metabolites with potential for use in the development of new pharmaceutical and industrial agents.
The present study includes the following:
Part I: Chemical composition and biological activities of selected macroalgae
I.1. characterization and scientific classification of four selected algae
Four macroalgae are collected from Quseir and Marsa-Alam areas of Egyptian Red Sea shores during (2016-2017). The algal samples were identified by Dr. Khaled NM Elsayed (Assistant Professor of Algal Biotechnology at Botany and Microbiology Department, Faculty of Science, Beni-Suef University, Egypt). These algae are Halimeda tuna (green alga), Sargassum platycarpum (brown alga), Padina gymnospora (brown alga) and Phacelocarpus tristichus (red alga).
I.2. Preliminary phytochemical screening of all algal samples
Phytochemical screening of four selected macroalgae revealed the following:
• Flavonoids, steroids, terpenoids, phenols and carbohydrates were detected in four selected algae.
• Alkaloids and tannins were found only in S. platycarpum and P. gymnospora.
• Saponins were present only in S. platycarpum, P. gymnospora and P. tristichus.
• These algae showed negative response when tested for anthraquinones.
I.3. Chemical composition of algal extracts by GC-MS analysis
The chemical components of 70% methanol extracts of H. tuna, S. platycarpum, P. gymnospora and P. tristichus were characterized by GC–MS analysis. This study led to the following conclusions:
• Obviously, there were various classes of chemical compounds in four algal extracts such as fatty acids, esters, sterols, terpenes, alkaloids, hydrocarbons, aldehydes, etc.
• Major chemical constituents present in H. tuna extract were lup-20(29)-en-3á-ol, acetate (36.45%), dibutyl phthalate (29.52%), betulinaldehyde (4.26%), cyclooctasiloxane, hexadecamethyl (3.36%) and lupeol (3.02%).
• However, dichloroiodomethane (13.06%), neophytadiene (6.43%), dibutyl phthalate (5.04%), cyclononasiloxane, octadecamethyl (2.78%) and 10-octadecenoic acid, methyl ester (2.10%) were the major compounds in S. platycarpum.
• While hexadecanoic acid, methyl ester (14.09%), hexadecanoic acid (13.09%), cis-13-octadecenoic acid, methyl ester (10.99%), propanoic acid, 2-hydroxy-, ethyl ester (10.42%), 1-pentanol (6.99%), 2-pentanone, 4-hydroxy-4-methyl (6.67%), tetradecanoic acid (4.69%) and di-(9-octadecenoyl)-glycerol (3.26%) were the major components in P. gymnospora.
• Moreover, P. tristichus contained the highest percentage of hexadecanoic acid, methyl ester (20.57%), followed by 5, 8, 11, 14-eicosatetraenoic acid, methyl ester, (all-Z) (11.73%), stigmasta-5,24(28)-dien-3á-ol, (Z) (10.01%), methyl-9-octadecenoate (9.31%), methyl stearidonate (8.81%), trans-9-octadecenoic acid (4.70%), hexadecanoic acid (4.41%) and cis-5,8,11,14,17-eicosapentaenoic acid (3.82%) as major components.
I.4. Antimicrobial activity of algal extracts
The antimicrobial activity of four algal extracts prepared by 70% methanol was tested against nine micro-organisms using agar diffusion assay. The results of antimicrobial investigation revealed that:
• All four algal extracts exhibited relatively significant antimicrobial activity.
• P. gymnospora and S. platycarpum exhibited the strongest antimicrobial activity followed by P. tristichus and finally H.tuna.
I.5. Cytotoxicity of algal extracts
The cytotoxic activity of the 70% methanol extracts of H. tuna, S. platycarpum, P. gymnospora and P. tristichus was tested against HepG-2 and MCF-7 cell lines using cell viability assay. The results of cytotoxic activity investigated that:
• The tested algal extracts especially S. platycarpum extract showed the maximum cytotoxicity, followed by P. gymnospora, P. tritichus and finally H. tuna against HepG-2.
• While P. gymnospora extract showed the maximum cytotoxicity, followed by S. platycarpum, P. tritichus and finally H. tuna against MCF-7 cells.
Part II: Phytochemistry and biological activity of Sargassum platycarpum
II.1. GC-MS analysis of S. platycarpum fractions
The total extract (70% MeOH) of S. platycarpum was fractionated into 5 fractions (n-hexane, dichloromethane, ethyl acetate, n-butanol and aqueous fractions). The major and minor components of five fractions of S. platycarpum were detected using GC-MS analysis and the results showed the following:
• Hexadecanoic acid methyl ester was present in four fractions, n-hexane, dichloromethane, ethyl acetate and n-butanol fraction.
• Likewise, hexadecanoic acid was found in n-hexane, dichloromethane, and ethyl acetate fraction.
• Compounds such as oleic acid, methyl ester, oleic acid, eicosyl ester, octylphenol diethoxylate and octadecanoic acid were the major components that also found in n-hexane fraction.
• In addition, kjellmanianone and loliolide were the main compounds present in dichloromethane fraction.
• Furthermore, oleic acid, methyl ester, 1-butanol, 3-methyl acetate and Z-(13, 14-epoxy)tetradec-11-en-1-ol acetate were the major compounds found in ethyl acetate fraction.
• Also, 2(3H)-furanone, dihydro-5-propyl, formic acid, butyl ester and silane, dimethylisobutoxybutoxy were the main components present in n-butanol fraction.
• Beside, 2-allyl-5-t-butylhydroquinone and 7,9-di-tert-butyl-1-oxaspiro-(4,5)-deca-6,9-diene-2,8-dione were the major constituents reported in the aqueous fraction.
II. 2. Antimicrobial activity of S. platycarpum fractions
Antimicrobial activities of total extract (70% MeOH) and its five fractions of S. platycarpum were evaluated against nine pathogens using agar well diffusion method. The results indicated the following:
• All algal extracts were inhibited significantly the microbial growth of some tested pathogens with variable potency.
• The total extract (70% methanol) exhibited significantly inhibition of the growth of most tested pathogens.
• Upon fractionation, ethyl acetate fraction exhibited the highest antimicrobial activity followed by dichloromethane, n-hexane, n-butanol and finally aqueous fraction.
II.3. Cytotoxic activity of S. platycarpum fractions
Cytotoxicity of total extract (70% MeOH) and its five fractions was evaluated against human cancer cells (HepG-2 and MCF-7 cells) using cell viability assay. The results revealed the following:
• The total extract (70% methanol) showed significant cytotoxic activity against HepG-2 and MCF-7.
• The dichloromethane fraction showed the most potent cytotoxic activity against HepG-2 and MCF-7 cell lines followed by ethyl acetate, aqueous, n-butanol and finally n-hexane.
II.4. Isolation and structural elucidation of compounds from S. platycarpum
Phytochemical study results in nine compounds isolated from S. platycarpum for the first time. The isolated compounds were characterized as hexadecanoic acid (1), oleic acid (2), saringosterols (3 and 4, 1:1 mixture of C-24epimers), β-sitosterol (5), glycoglycerolipid (6), loliolide (7), kjellmanianone (8) and caulerpin (9) by spectroscopic techniques.
II.5. In vitro and in silico studies on the cytotoxic activity of eight compounds (1-8) from brown alga Sargassum platycarpum against HepG-2
• The cytotoxicity of all isolated compounds (1-8) was evaluated using cell viability assay against HepG-2 (liver cancer cell line). The results indicated that saringosterols (3 and 4, 1:1 mixture of C-24epimers) displayed the highest cytotoxicity, followed by kjellmanianone (8), hexadecanoic acid (1), loliolide(7), glycoglycerolipid(6), β-sitosterol(5) and finally oleic acid (2).
• The results obtained from in silico studies are in a good agreement with those obtained from experimental. where all isolated compounds (1-8) can be utilized as significant compounds against human liver cancer cells, especially compounds 3 and 4.
II.6. In vitro and in silico studies on biological activity of compound (9) isolated from a new source S. platycarpum
• Caulerpin, a bis-indole alkaloid is isolated from a new source Sargassum platycarpum, brown alga (family Sargassaceae) for the first time.
• The results of in vitro antifungal activity indicated that caulerpin moderatly inhibited the growth of C. albicans and C. neoformans. Also, this algal pigment did not exhibited antifungal activity against are A. fumigatus and P. expansum.
• The findings of cytotoxicity indicated that caulerpin exhibited potential cytotoxicity against new type of human cancer cells (HepG-2) using cell viability assay.
• The results of in silico study evaluated that caulerpin-T/U complexes are stable in the presences of water molecules. Thus, the findings of our study will be useful for giving an insight into the caulerpin/bases complexes that could be helpful in future experimental studies to develop the performance of caulerpin molecules as natural cancer candidate drug.
Part III: Phytochemical studies and biological effects of P. gymnospora
III.1. GC-MS analysis of P. gymnospora fractions
The total extract (70% MeOH) of P. gymnospora was fractionated into 3 fractions (n-hexane, ethyl acetate and n-butanol fractions). The major and minor components of three fractions of P. gymnospora were detected using GC-MS analysis and the results showed the following:
• Hexadecanoic acid, methyl ester was found in three fractions of P. gymnospora, n-hexane, ethyl acetate and n-butanol fraction.
• Stigmasta-5,24(28)-dien-3-ol,(3á,24), trans-13-octadecenoic acid, methyl ester, tricyclo[4.3.1.13,8]undecane,3-methyl and 6-hydroxy-4,4,7a-trimethyl-5,6,7,7a-tetrahydrobenzofuran-2(4H)-one were the major components in n-hexane fraction.
• However, the ethyl acetate fraction contained high percentage of oleic acid, tetradecanoic acid, n-hexadecanoic acid and loliolide.
• While 17-octadecenoic acid, methyl ester and butyl 9,12-octadecadienoate were the major compounds in n-butanol fraction.
III.2. Antimicrobial activity of P. gymnospora fractions
Antimicrobial activities of total extract (70% MeOH) and its three fractions of P. gymnospora were evaluated against the same nine pathogens that are mentioned previously using agar well diffusion method. The results indicated the following:
• The total extract (70% methanol) exhibited significantly inhibition of the growth of most tested pathogens.
• Upon fractionation, the n-hexane fraction exhibited the highest antimicrobial activity followed by ethyl acetate and finally n-butanol.
III.3.Cytotoxic activity of P. gymnospora fractions
Cytotoxicity of total extract (70% MeOH) and its three fractions was evaluated against human cancer cells (HepG-2 and MCF-7 cells) using cell viability assay. The results revealed the following:
• The highest cytotoxicity was recorded by total extract (70% methanol) against HepG-2 and MCF-7 compared with its three fractions.
• Upon fractionation, the ethyl acetate fraction had the most potent cytotoxic activity against HepG-2 and MCF-7 followed by n-hexane and finally n-butanol.
III.4. Isolation, characterization, in vitro cytotoxic activity of four compounds (1-4) from brown alga Padina gymnospora
Phytochemical investigation of P. gymnospora results in four compounds isolated for the first time. These isolated phytoconstituents were identified as saringosterols (1 and 2, 1:1 mixture of C-24epimers), oleic acid (3) and hexadecanoic acid (4). These isolated compounds are the same compounds that already isolated previously from S. platycarpum and their cytotoxicity against HepG-2 cells was already reported in this work.
Part IV: Evaluation of antiviral activity of all isolated compounds from S. platycarpum and P. gymnospora against COVID-19 by in silico approaches
IV.1.The inhibitory effect of all isolated compounds from S. platycarpum and P. gymnospora against SARS-CoV-2 main protease insights from molecular docking analysis and molecular dynamic simulation
In the current study, we isolated nine compounds from brown alga S. platycarpum and four compounds from P. gymnospora for the first time. All these isolated compounds exhibited cytotoxicity against HepG-2 cell line according to our results in this study. In addition, these compounds are already abundant and isolated from other different species of plants or marine algae and showed a broad spectrum of other biological activities like antimicrobial, antioxidant, anticancer, antiviral, anti-inflammatory, etc. that making them an attractive target to evaluate their potential to become significant candidate inhibitors against SARS-CoV-2 receptors.
In this study we aimed to the inhibitory effect of all isolated compounds in our study as potential inhibitors of SARS-CoV-2-3CL main protease (PDB ID: 6LU7) and SARS-CoV main proteases (PDB IDs: 2GTB and 3TNT) by in silico approaches. The results showed the following:
• Briefly, caulerpin isolated from S. platycarpum is highly effective against SARS-CoV-2 main protease and can be explored further for drug repurposing against the successful inhibition of COVID-19.
IV.2. Destabilizing the structural integrity of COVID-19 by caulerpin and its derivatives along with some antiviral drugs: An in silico approache for a combination therapy
In this study we aimed to evaluate the inhibitory effect of caulerpin and its analogs against the SARS-CoV-2 main protease Mpro (PDB ID: 6LU7) and SARS-CoV-2 spike protein Sp (PDB ID: 6VYB) by in silico approaches. Also, study the effect of side chain replacement of caulerpin on the inhibitory effect compared with some used drugs which currently are used as SARS-CoV-2 inhibitors such as lopinavir, simeprevir, hydroxychloroquine, chloroquine and amprenavir. The results reported the following:
• Breifly, the combination of the studied compounds and simeprevir are highly effective against SARS-CoV-2 protease, and these drugs can be explored further for drug repurposing against the successful inhibition of COVID-19.
• Finally, the antiviral activity of lopinavir, simeprevir, hydroxychloroquine, chloroquine and amprenavir could be increased against SARS-CoV-2 by using caulerpin and its derivatives as combination therapy.