Syed Madhi Haider

Phytochemistry of Flower: Phytochemical screening analysis of Rosa indica L. Atya Hassan, Syed Madhi Haider, Iqra, Syed Mujtaba Abbas, Federal Urdu university of Arts, Science and technology Karachi, Pakistan University of Gujrat, Gujrat, Pakistan Abstract Rosa indica belongs to Rosaceae family with more than 100 of species and over 2000 cultivars. Various part of this plant has medicinal properties due to presence of different phytochemicals. Purpose of this study to determined the secondary metabolites present in rose petal through phytochemical screening analysis. Sample were tested include water, methanol, ethanol, acetic acid and ethyl acetate extract with different phytochemical tests. Phytochemical screening showed tannin, flavonoid, phenolic compounds, carbohydrate, alkaloid and other bioactive compounds were present in all solvent extract. Percent yield of phenolic compound, anthraquinones and alkaloid were higher in aqueous medium extract. It reliably possessed large number of pharmacological properties such as anti-cancer, anti-viral, anti-bacterial, anti-inflammatory, ant-allergic, anti-fungal and antiabortifacient and being employed for the treatment of different ailment in advanced system of medicines. key word:- Phytochemicals, phenolic compounds, anthraquinones, alkaloids, bioactive compounds, collection of rose petal presence of tannin, flavonoid, carbohydrate etc percent yeild calcuation Washing of rose petals Observation of colour changes Conclusion Extration of sample in various solvents phytochemical screening Introduction wide range of illness treated through medicinal plants from ancient time and have become more importance in medical science (Rivera-Mondragón et al., 2019). Different parts of plant contain various chemicals known as phytochemicals. Medicinal properties comprise from phytochemicals which improve the physiological balance of human beings (Owolabi et al., 2018). Vast range of medicinal plants in global has not yet been investigated about their usefulness in diseases treatment. In developing countries up to 80% of population dependent upon plants for their health (Hostettmann & Marston, 2002). Alkaloid, Flavonoid, Terpenoid and tannins are the bioactive compounds in different parts of plant. Bio active compound are those which have ability to interact with components of living tissues (Huang et al., 2016). 10,000 known compounds of flavonoid in plant kingdom with different major bioactivities including anti-inflammatory, anti-genotoxic, anti-cancer and antiglycative properties (Zahid et al., 2018) make flavonoid the king of phytochemicals. Rosa indica is traditional medicinal plant showing greatest advantage for controlling cancer growth, constipation, inflammation, heart and eye diseases (Raj et al., 2018). Rosa indica is perennial flower shrub belongs to Rosaceae family with more than 100 of species and over 2000 cultivars (Leghari et al., 2016). Different parts of this plant is used for medicinal purpose due to presence of various phytochemicals. Some species of rosa produce essential oils, 5 hydroxy methyl furfural pyrogallol (21.92%) and levoglucusan (5.69%). Acetic acid, mercaptohexyl ester, butanoic acid, artemiseol, methyl santonilate, caryophyllene oxide, 1,5octadecadine and ethyl easter are the essential oils in rosa indica (Rasheed et al., 2015). Different forms of phytochemicals screening analysis, antioxidant and antimicrobial analysis performed on Rosa indica L. phytochemical and anti-microbial activity of Rosa indica L studied by Rumana Saeed et.al in which antibacterial and antifungal activities found via different solvents (Saeed et al., 2014). Crude extract of Rosa indica petals used for antibacterial study by Deepika et.al (Pathak et al., 2019). Afifah et.al studied antioxidant activity of rose leaves (Afifah et al., 2021). Extraction and evaluation of antioxidant activity and free radicle scavenging potential correlated with biochemical component of red rose petal (Pal et al., 2018). Antioxidant analysis of rose petal tea carried out by Vinokur et.al (Vinokur et al., 2006). Two acids of medicinal properties isolated by Saaby et.al (Saaby et al., 2011). Similarly chemical composition of some rose fruits species also found by Ercisli et.al (Ercisli, 2007). Anti cancer effect of phytochemicals studied by Zahra et.al (Bathaie et al., 2015). Anthocyanin content of red rose petal investigated through maceration and percolation method (Saptarini et al.). Effetc of Rosa damascna extract on primary dysmenorrhea (Bani et al., 2014). Citronally disaccharide glycoside aroma precursor isolated of rose flower also studied (Oka et al., 1998). LC-UV-MS/MS analysis of 71 species of rosa (Porter et al., 2012) performed. Luvenia et.al analysed the phytochemical, FTIR and NMR crude extract of Acacia planifrons seeds (Fernando et al., 2019). Manikandan et.al used ethanolic extract of rosa indica petals and biosynthesized siler nano particles (Manikandan et al., 2015). Amrita et.al synthesized ZnO nano particles from rosa indica leaves (Raj & Lawerence, 2018). Amit et.al performed comparative study on Rosa indica, Moringa olifera and Azadirachta indica for antibacterial , antioxidant and photosynthetic study (Charan & Gupta, 2013). Antibacterial and antifungal activity of Rosa damascene studied by Shohayeb et.al (Shohayeb et al., 2014). Main interest of this research paper to performed the different phytochemicals tests for Rosa indica flowers. Large number of phytochemicals with different functional groups are present in Rosa indica with health effects. Different biochemical such as vitamins especially vitamin A, B, K and E and potassium and iron present in different parts of Rosa indica (Nanda & Das, 2015). While the extract of rose plant act as anti-depressant, ant bacterial, anti-fungal, anti-septic, anti-inflammatory, and digestive stimulant. Similarly the extract of petal contain anthocyanins that have high antioxidant activity and anti-inflammatory activity (AKh et al., 1993). Material and Methods Present work is performed for the identification of phytochemicals of Rosa indica flower belongs to genus rosa . Collection and Drying the rosa indica flowers collected from Teen Hatti, local market of flower geographically situated in Karachi, Pakistan. Flowers petals were washed with tap water then with distilled water for the removal od surface dust and contaminant. Petals were dried in shadow at room temperature for couple of hours. After drying petals were ground in normal grinder for better extraction Extraction Powder was soaked into different solvents including distilled water, methanol, ethanol, acetic acid and ethyl acetate at room temperature for a week. The filtered extract then keeps in glass bottles at room temperature. Phytochemical screening Medicinal plants have bio active compound known as phytochemicals. For determination of phytochemical in different solvent of petals, reported tests (Belay & Sisay, 2014; Rakholiya et al., 2016; Seow et al., 2013; Sheel et al., 2014). Table 1 Described the following test for difierent phytochemicals. Table 1 Different phytochemical screening with procedure Phytochemical Procedure References Alkaloid (Wagner’s test) 3mL of sample + Wagner reagent (Tiwari et al., 2011) Saponins (Frothing test) Extract diluted with water + shaken (Pandey et al., 2014) Extract + 5mL CHCl3 + Shaken then filtered + add (Namadina & Sani, 2018) Anthraquinones (Bontrager’s test) 10% ammonium solution Steroid (Liebermann-Burchard test) 2mL of organic extract dissolved in 2mL CHCl3 + (kumar Bargah, 2015) H2SO4(c) and CH3COOH Carbohydrate (Molisch Test) Filtrate treated with 2 drops of alcoholic α naphthol (Yamini et al., 2019) + 2mL of H2SO4(c) Phenolic compounds (Ferric chloride test) Water extract + 2mL of 5% FeCl3 (Vimalkumar et al., 2014) Flavonoid (NaOH test) Extract + few drops of NaOH + yellow colour (Anza et al., 2015) appear + dilute acid yellow colour disappear Tannin ((Ferric chloride test) 5mL of sample + few drops of FeCl3 (Fitrilia et al., 2015) Triterpenes (Salkowski test) 5mL of extract + few drops of CHCl3 with H2SO4(c) (Tongco et al., 2014) Phlobatannins (HCl test) 2mL of extract boiled with 2mL of 10% HCl (Sharma & Patel) Terpenoids (Salkowski test) 1mL of extract + few drops of CHCl3 with H2SO4(c) (Kale, 2020) Result and Discussion . water and methanol were the most polar solvents among all solvents used in phytochemical screening. Both solvents were also recommended in literature (Kidaha et al., 2013; Seow et al., 2013). Different colour observation for various phytochemicals mentioned in table 1. All sought bioactive phytochemicals were detected in all solvent mentioned in table 2. According to results mentioned in table 3, phytochemical yield in aqueous extract was high than other solvents. Primary and secondary metabolites of plant was numerous functions. Alkaloid detected in all solvents with different yield. Percent yield of alkaloid in aqueous extract was about 8.77%. Alkaloid used as hallucinogenic agent and also stimulant, anticancer drug, analgesic, anti-bacterial and cholinomeimerics (Wansi et al., 2013). But in some cases alkaloid from plants can cause serious illness even death. Pyrrolizidine alkaloids known as plant toxins which cause severe illness in human(Ibanez, 2005). They are the large group of hepatotoxin, pneumotoxin ang genotoxin with pyrrolizidine in their structure (Diaz, 2015) Table 1 Different phytochemical with positive results S. No. Active Compounds Plant Extracts Aq. Ex. Eth. Ex. Mth. Ex. A.A. Ex. E.A. Ex. 1 Alkaloids + + + + + 2 Anthraquinones + + + + + 3 Flavonoids + + + + + 4 Phlobtanins + + + + + 5 Saponins + + + + + 6 Steroids + + + + + 7 Tannins + + + + + 8 Terpenoids + + + + + 9 Carbohydrates + + + + + 10 Triterpenes + + + + + 11 Coumarins + + + + + 12 Phenolic compounds + + + + + 13 Quinines + + + + + Table 2 Different phytochemical tests with observations Phytochemical Procedure Observation Alkaloid (Wagner’s test) 3mL of sample + Wagner reagent Reddish/Brown/whitish ppt formed Saponins (Frothing test) Extract diluted with water + shaken Honeycomb froth Anthraquinones Extract + 5mL CHCl3 + Shaken then filtered + add 10% Pink colour in upper layer (Bontrager’s test) ammonium solution Steroid (Liebermann- 2mL of organic extract dissolved in 2mL CHCl3 + H2SO4(c) Burchard test) and CH3COOH Carbohydrate (Molisch Test) Filtrate treated with 2 drops of alcoholic α naphthol + 2mL of Green colour Brown ring formation H2SO4(c) Phenolic compounds (Ferric Water extract + 2mL of 5% FeCl3 Violate/Green colour Extract + few drops of NaOH + yellow colour appear + dilute Dark yellow colour chloride test) Flavonoid (NaOH test) acid yellow colour disappear Tannin ((Ferric chloride 5mL of sample + few drops of FeCl3 Brownish green 5mL of extract + few drops of CHCl3 with H2SO4(c) Yellowish to golden test) Triterpenes (Salkowski test) yellow Phlobatannins (HCl test) 2mL of extract boiled with 2mL of 10% HCl Red ppt Terpenoids (Salkowski test) 1mL of extract + few drops of CHCl3 with H2SO4(c) Reddish brown interface Table 3 Percent yield of active compounds (w/w) S. No. Active compounds 1 Phenolic Aq.Ex Eth.Ex Mth. Ex. E.A. Ex. A.A. Ex. 0.4 0.1 0.28 0.78 - compounds 2 Coumarins 2.49 0.53 0.16 0.53 - 3 Quinines 0.12 0.43 0.99 0.84 1.48 4 Anthraquinones 1.94 0.47 1.42 1.22 2.9 5 Flavonoids 0.66 0.54 0.19 0.27 - 6 Alkaloids 8.77 9.51 8.51 8.37 8.25 Saponins related to glycosidic compounds present in plant kingdom (Furuya, 1988). Saponin was also detected in all solvents. Saponins related to plants metabolites with pancreatic lipase inhibitory effect (Xu et al., 2005). On other hand saponin may be toxic. These compounds have haemolytic activity on human erythrocytes. This was due to interaction with sterol present in erythrocyte membrane, which leads the membrane permeability and finally loss of haemoglobin (Baumann et al., 2000). Similarly some saponin containing plants are toxic for ruminants, cause gastrointestinal, liver and kidney degeneration (Wina et al., 2005). Anthraquinones were also found in all solvent. The percent yield of anthraquinones in aqueous solution and methanolic solution was 1.94 % and 1.42% respectively. These structurally related to anthracene and sometime referred as 9,10-dioxoanthracene. These present in plants as glycoside (Bruneton, 2012). These compound impart colour to plants and also used as natural dye substance (Simpson & Amos, 2017). Anthraquinones used as semiconductor and having anti-fungal, anti-viral, anti-bacterial and parasitic activity (Yadav et al., 2019). Tannins belongs to phytochemical found in many plants. All extract gave positive result for tannin. This phytochemical responsible for various pharmaco-therapeutic effect (Ferreira et al., 2008). In the form of proanthocynidin beneficial effect on vascular health. Rose petal have antiinflammatory, antioxidant, antibacterial and antiviral activities due to presence of tannin. Phlobatannins also present in extract. Phlobatannins belongs to C-ring isomerized oligomeric form (Ferreira et al., 1999). . Flavonoid and phenolic compounds were also present in petal extract of Rosa indica. Percent yield of flavonoid and phenolic extract in aqueous extract were about to 0.66% and 0.4% respectively. Phenolic compounds classified into various range of groups according to structure. Most phenolic compounds synthesized in the endoplasmic reticulum of plants and stored in vacuoles (Gan et al., 2019). Rose contained high level of antioxidant activity due to presence of high concentration of phenolic compounds. (Choi et al., 2015). Polyphenolic content in red rose higher than the pink, white rose, reportedly attributed to influenced of redcolour anthocyanin pigment.(Yang & Shin, 2017). Flavonoid associated with health promoting effect and indispensable component in pharmaceutical, medicinal and cosmetic industries. Their anti-cancer, anti-inflammatory, anti- mutagenic activities coupled with their capacity to modulate the enzyme activity functions (Panche et al., 2016). In most angiosperm family many flavonoids recognized as flower pigments. Number of flavonoid also reported for anti- cholinesterase activity. Inhibitory studies done on varios flavonoid such as quercetin, rutin, kaempferol 3-O-β-D-galactoside showed that concentration dependent inhibition ability against butyrylcholinesterase (Khan et al., 2009). Salkowski test performed on sample for identification of terpenoid and triterpene. Both compounds are detected in all extracts. Terpene are the largest class of phytochemicals with 40,000 reported structural compounds so far (Gershenzon & Dudareva, 2007). The basic chemical structure of terpene dependent upon the isoprene (C5H8) units. On number of isoprene unit’s terpene further classified as mono-, sesqui-, di and tri terpene. Terpene and terpenoid were slightly differed from each other on basis of no.of isoprene units(Cox-Georgian et al., 2019) . Terpenoid found to be useful in therapy of different diseases and also anti-bacterial, anti-viral, anti-allergic and anti-parasitic activity (Paduch, 2007). Liebermann-Burchard test performed for steroid detection in all extracts. It was observed that extract of Rosa indica petal showed positive results in all solvents for steroid. Steroid present in all plants as secondary metabolite. They are derived from cholesterol and all steroid molecules possessed basic perhydroxyl cyclopentanophenanthrene skeleton. Steroid widely used in drug designing (Ur Rahman et al., 2017). Beside this carbohydrate also detected in sample. Carbohydrate known as polyhydroxy carbonyl compounds. this basic building block of cell wall classified as mon-, di, and poly saccharides. Mono and di saccharides known as true sugar. Sugar reduced the water potential of petals and promoting the influx of water for cell enlargement and flower opening (Naveen et al., 2007).In plant defence system sugars play key role for the regulation of defence gene (Ehness et al., 1997; Roitsch et al., 2003). Role of some sugar regarding immunity known as ‘’sweet immunity’’ and sugar ‘’enhanced defenced’’ concept (Bolouri Moghaddam & Van den Ende, 2013). Conclusion it was concluded from the phytochemical screening analysis of Rosa indica that all phytochemicals such as carbohydrates, flavonoid, tannins, steroid, saponin and terpenoids were present in petal of rose. Presence of phytochemical additionally dependent on solvent. Red colour of rose was due to presence of high amount of anthocyanin. Presence of different phytochemical in rose petal indicated that the medicinal paramountcy of rose. Anti-viral, anti-inflammatory, anti-bacterial, anti-allergic, anti-fungal and anti-carcinogenic activities of rose petal comprised the pharmacological importance of this plant. Due to different medicinal properties of Rosa indica plant, different form of medicines can prepare from rose petal for cancer, allergies treatment in future. Rosa indica plant may hold potential in providing benefits for human nutrition and health. Acknowledgment Author would like to thanks the supervisor for proving the all necessary facilities and helped for writing the research paper. Conflict of interest Author hereby that there was no conflict of interest regarding research performance and paper writing and publication. References Afifah, D. N., Yessy, Y., Rahmani, F. N., Ningsih, A. L., Sari, C. D. P., Yudapraja, R. R., Praptiwi, P., & Ridwan, R. (2021). ANTIOXIDANT ACTIVITY OF RED ROSE LEAVES (Rosa chinensis Jacq.) EXTRACT. Acta Biochimica Indonesiana, 3(2), 81-88. 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