geographical-variation-of-antioxidant-constituent-in-garhwal-region-of

Views:
 
Category: Entertainment
     
 

Presentation Description

Numerous kind of antioxidants or protecting agent are present in the living body like Glutathione is the master antioxidant produce by the liver and uses free radicals to purify the body. Natural plant consuming antioxidants content is growing the interest for scientific research as well as industrial purposes. Pteridophytes (fern and fern allies) have drawn attention of plants seekers and horticultures since ancient period. Plant of Pteris vittata were collected from different geographical region of Uttarakhand Garhwal. The maximum yield of plant extract was found via the ethanol solvent i.e. 5.07-9.67%.

Comments

Presentation Transcript

slide 1:

Research Article Open Access Dobhal et al. J Appl Pharm 2018 10:3 DOI: 10.4172/1920-4159.1000266 Research Article Open Access Journal of Applied Pharmacy Journal of Applied Pharmacy ISSN: 1920-4159 Volume 10 • Issue 3 • 1000266 J Appl Pharm an open access journal ISSN: 1920-4159 Geographical Variation of Antioxidant Constituent in Garhwal Region of Uttarakhand: Chinese Brake Fern Pteris vittata Dobhal K Semwal A and Negi A Uttaranchal Institutes of Pharmaceutical Sciences Dehradun Uttarakhand India Abstract Numerous kind of antioxidants or protecting agent are present in the living body like Glutathione is the master antioxidant produce by the liver and uses free radicals to purify the body. Natural plant consuming antioxidants content is growing the interest for scientifc research as well as industrial purposes. Pteridophytes fern and fern allies have drawn attention of plants seekers and horticultures since ancient period. Plant of Pteris vittata were collected from different geographical region of Uttarakhand Garhwal. The maximum yield of plant extract was found via the ethanol solvent i.e. 5.07-9.67. DPV extract exhibited a maximum inhibition of 89.32 relatively closed to 91.96 inhibition of Ascorbic acid at the concentration of 0.1 mg/ml by DPPH radical scavenging assay method. The IC 50 of the DPV extract and Ascorbic acid was found to be 0.543 and 0.495 mg/ml by same. DPV extract showed a maximum inhibition of 72.33 relatively close to 77.42 inhibition of BHA at the concentration of 0.8 mg/ml by Hydrogen peroxide radical scavenging method. The IC 50 value of the DPV extract BHA was found to be 0.279 ± 0.005 mg/ml 0.257 ± 0.002 mg/ml by same. DPV extract showed a maximum inhibition of 84.32 relatively close to 87.96 inhibition of ascorbic acid at the concentration of 0.8 mg/ml by Nitrogen oxide scavenging method. The IC 50 value of the DPV extract BHA was found to be 0.233 ± 0.002 mg/ml to 0.218 ± 0.006 mg/ml by same. Keywords: Chinese fern Antioxidant Scavenging Phenolic compound Inhibition Introduction Natural antioxidants are required either prevent or cure the disorders caused by free radicals. As folk medicine the Pteridophytes which constitute fern and ferns allies have been known to man for more than 2000 years and also been mentioned in ancient literature 1. Pteridophytes are original vascular cryptogams which fourish well in worldly environment. In the world fora of pteridophytes 12000 species has been identifed among which 1000 species into 70 families and 191 genera are occur in India. Pteris vittata was also entitled as the Chinese brake Chinese ladder brake or simply ladder brake 23. Pteridophytes is predictable for the rich diversity of valuable antioxidants. Various works was accomplished towards the medicinal importance of pteridophytes by the researchers. Antioxidants are the chemical which are derived from the plant sources generally afect health but are not yet established nutrients. Phenolic rich content plant material simultaneously increasing the interest of market as antioxidant used in the food industry. Tese natural antioxidants improve the quality and nutritional value of food. Flavonoid related derivatives have remarkable antibacterial antiviral anti-infammatory anticancer and anti-allergic activities due to its scavenging activity 4. Te investigation was designed to explore the variation of the antioxidant content of whole plant of Chinese Brake in the diferent region of Uttarakhand. Material and Methods Plant material Te investigation was conducted out in the month of August- September 2018 at Uttaranchal Institute of Pharmaceutical Sciences Premnagar Dehradun Uttarakhand. Te fresh whole plant was collected through habitat of Uttarakhand i.e. Chamoli Dehradun Pauri Tehri and Uttarkashi. Te herbariums of plant were submitted to the Forest Research Institute Dehradun Uttarakhand. Te plant was sundried for 15-20 days in the laboratory and transformed into powder. Corresponding author: Dobhal K Uttaranchal Institutes of Pharma- ceutical Sciences Dehradun Uttarakhand India Tel: 7579082858 E-mail: kiran.dobhal2728gmail.com Received: October 31 2018 Accepted: November 09 2018 Published: November 17 2018 Citation: Dobhal K Semwal A Negi A 2018 Geographical Variation of Antioxidant Constituent in Garhwal Region of Uttarakhand: Chinese Brake Fern Pteris vittata. J Appl Pharm 10: 266. doi: 10.4172/1920-4159.1000266 Copyright: © 2018 Dobhal K et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License which permits unrestricted use distribution and reproduction in any medium provided the original author and source are credited. Preparation of plant extract Air dried plant powder was sofened in petroleum ether stored in tightly closed container. It was rotated on a rotary shaker at 190- 220 rpm for one day the clear fltrate was discarded. Petroleum ether was fully evaporated collects as powder. Further the powder was extracted out by the diferent solvent acetone benzene ethanol ethyl acetate and water respectively. By the centrifugation techniques the resulting extract was obtained as dry residues 5. All chemical solvent used during the experiments were analytical grade. Te plant extract were entitled consequently as their localities i.e Chamoli P. vittata CPV Dehradun P. vittata DPV Pauri P. vittata PPV Tehri P. vittata TPV and Uttarkashi P. vittata UPV. Qualitative phytochemical analysis Highest yield of extract was extracted in the ethanol solvent relatively to others. Terefore ethanol extract was subjected to preliminary qualitative phytochemical investigation for alkaloids proteins carbohydrates favonoids cardiac glycosides saponins catechins sugars steroids and triterpenoids tannins and phenols etc. 6. Quantitative phytochemical analysis Te phytochemicals constituent was enumerated by the standard procedures. Determination of total alkaloids TAS 1 gm of extract was liquefed in 40 ml of 10 acetic acid in ethanol

slide 2:

Citation: Dobhal K Semwal A Negi A 2018 Geographical Variation of Antioxidant Constituent in Garhwal Region of Uttarakhand: Chinese Brake Fern Pteris vittata. J Appl Pharm 10: 266. doi: 10.4172/1920-4159.1000266 Page 2 of 4 Volume 10 • Issue 3 • 1000266 J Appl Pharm an open access journal ISSN: 1920-4159 strained. It was concentrated add conc. ammonium hydroxide drop wise for the precipitation. Te precipitated material was further liquefed with dilute ammonium hydroxide again concentrated. Te resulting residue was alkaloid 5. Determination of total saponin content TSC 2 g of extract liquefed in 20 of aqueous ethanol. Te samples was heated for 90 minute strained out. Tis procedure revised atleast three times. Te resulting solution dissolved with ether shaken vigorously. Te saponin content was recovered by aqueous layer carefully through the mixture of n-butanol 5 sodium chloride solution. Te saponin content was weighed 7. Estimation of total phenolic content TPC Te total phenolic content was estimated by standard procedures with slight modifcation 8. 2.5 mg of plant sample was dissolved in 10 ml of ethanol sonicated. Tere was incubated the solution of plant sample methanol and distilled water Folin-Ciocalteu reagent for 5 minute. Add the 10 sodium carbonate solution covered through aluminum foil. It was again incubated for 20 min. Te absorbance of the sample was determined using a UV visible spectrophotometer at 765 nm. Gallic acid is used as standard phenolic compound. 100 μg/ mL solution of gallic acid was prepared by dissolving 10 mg of Gallic acid monohydrate in 100 mL of ethanol. By diluting the stock solution obtained diferent concentration ranging from 10-80 μg/ml. Total phenolic content was stated as μg of gallic acid equivalents GAE per ml. A standard calibration curve was drawn by plotting absorbance against concentration. It was found to be linear over this concentration range. Estimation of total favonoid content TFC Aluminium chloride technique was slightly modifed for the favonoid estimation 910. Prepared the mixture of sample solution methanol 10 aluminium chloride 1 M potassium acetate solution and distilled water in test tube incubated for 30 minutes. Te reaction mixture was treated with 1M sodium hydroxide solution. Rutin is used as standard favonoid compound. 10 mg of rutin dissolved in 10 mL of ethanol to get 100 μg/mL. By diluting the stock solution obtained diferent concentration ranging from 10-80 μg/ml. Te total favonoid content was expressed in microgram of rutin equivalents RE per gram of samples. Te absorbance was measured at 415 nm against reagent blank. Te calibration curve was drawn by plotting absorbance against concentration. It was found to be linear over this concentration range. DPPH radical scavenging activity Te DPPH radical scavenging activity was performed by standard procedures with slight modifcations 1112. DPPH 1 1-Diphenyl –2-picrylhydrazyl is a kind of stable free radical. Stock solution was prepared the concentration of 0.43 mg/ml. Ascorbic acid was used as standard. Prepared the solution of DPPH with sample control separately. Te control contained 0.1 ml ethanol in place of the plant sample. Te absorbance was measured at 517 nm by UV spectrophotometer 8. Antioxidant activity was expressed as percentage Antioxidant activity Ac − As /Ac × 100 Ac and As are the absorbance of control and sample respectively. Te IC 50 was calculated for sample control by Figure 1. Hydrogen peroxide scavenging activity Te hydrogen peroxide scavenging assay was carried out by standard procedure with minor modifcation 13. Hydrogen peroxide is an example of oxidizing agent. H 2 O 2 6v/v was prepared in phosphate bufer 0.1M pH 7.4. Butylated hydroxyl anisole BHA was used as standard. Te diferent concentration was prepared in the phosphate bufer hydrogen peroxide solution. Te absorbance was recorded at 230 nm by UV spectrophotometer. Scavenging activity was expressed as percentage inhibition H 2 O 2 scavenging activity A0 – A1 /A0 ×100 A0 is the absorbance of the control and A1 is the absorbance of the sample. Te IC 50 was calculated for sample control by Figure 2. Nitric oxide scavenging activity Te NO scavenging assay was carried out by the use of Griess Illovay reaction 14. Griess reagent was mixture of 0.1 sulfanilamide 0.2 phosphoric acid 0.01 N-1-naphthyl ethylene diamine dihydrochloride. Aqueous solution of sodium nitroprusside generates nitrogen oxide NO at pH7 interact with oxygen molecule to produce stable products nitrates nitrite. Sodium nitroprusside in phosphate bufered saline was mixed with diferent concentration of sample then dissolved in methanol incubated at 40 ο C for 90 minutes. Control was prepared without sample. Afer the incubation period add 0.1 ml of griess reagent. Ascorbic acid was used as standard. Te absorbance was calculated at 540 nm. Scavenging activity was expressed as percentage inhibition NO scavenging activity A0 – A1 /A0 ×100 0 20 40 60 80 100 0 0.5 1 1.5 i n h i b i t i o n conc.mg/ml PP adical can i n Ac ii y AA CPV DPV PPV TPV UPV Figure 1: DPPH radical scavenging inhibition versus concentration mg/ mL of standard and extracts. 0 20 40 60 80 100 0 0.2 0.4 0.6 0.8 1 INHIBITION conc.mg/ml ydroe n peroi de c ae ni n aay AA CPV DPV PPV TPV UPV Figure 2: Hydrogen peroxide scavenging inhibition versus concentration mg/mL.

slide 3:

Citation: Dobhal K Semwal A Negi A 2018 Geographical Variation of Antioxidant Constituent in Garhwal Region of Uttarakhand: Chinese Brake Fern Pteris vittata. J Appl Pharm 10: 266. doi: 10.4172/1920-4159.1000266 Page 3 of 4 Volume 10 • Issue 3 • 1000266 J Appl Pharm an open access journal ISSN: 1920-4159 A0 is the absorbance of the control and A1 is the absorbance of the sample. Results Yield and phytochemical estimation of crude extracts Ethanol extracts of P. vittata was appeared as green to dark green. Phytoconstituent was found to be in range of 7.37-.9.67. Plant extract was exposed the diferent qualitative test revealed positive for certain phytochemical tests. Alkaloids phenolic compounds favonoids saponins and tannins were the secondary metabolites present in the extract. Tese compounds play signifcant role of antioxidant activity of natural resources. Variation in the amount of these constituents directly infuences the antioxidant activity of compound 15. Total alkaloid content and total saponin content Alkaloid content of CPV DPV PPV TPV UPV was found to be in the range of 0.819 ± 0.053 0.152 ± 0.40 0.919 ± 0.906 0.115 ± 0.3 0.831 ± 0.628 μg /g respectively. Saponins content of CPV DPV PPV TPV UPV was found to be in the range of 0.595 ± 0.71 0.119 ± 0.185 0.737 ± 0.503 0.817 ± 0.125 0.541 ± 0.528 μg/g respectively. Among all of extract DPV and TPV revealed the maximum content of alkaloid and saponins. Alkaloids are the widely available phytochemicals responsible for the anticancer and antimicrobial activities. Saponins are applicable as a suitable option for phytochemicals to defend plant against several pathogens 16. Total phenol content TPC TPC was calculated from the calibration graph of gallic acid verses sample. Multiple reading was recorded for each sample. Te TPC was revealed to be in the huge variation range of 0.411 ± 0.327 to 1.653 ± 0.479 mg GAE/g Figure 3. Linear regression analysis y0.0163x+ 0.0083 R 2 0.0997 was applied to calculate TPC in which y is absorbance at 765 nm and x is the amount of gallic acid equivalent g/ ml per 20 gm extract. DPV extract was found the highest amount of TPC i.e. 1.605317 ± 0.4794 mg GAE/g. Hydroxyl group of phenol play important role in the scavenging activity of plant 1718. Total favonoids content TFC TFC was calculated from the calibration graph of rutin verses sample. Multiple reading was recorded for each sample. Te TFC was revealed to be in the huge variation range of 0.346 ± 0.89 to 0.714 ± 0.006 μg RE/g Figure 4. Linear regression analysis equation y0.015x+0.026 R 2 0.9994 was applied to calculate TFC in which y is absorbance at 415 nm and x is the amount of rutin equivalent RE per 20 g extract. DPV extract was found the highest amount of TFC i.e. 0.7143 ± 0.0064 μg RE/ g. Like phenolic favonoids have positive efect on human health 19. DPPH radical scavenging activity Ascorbic acid antioxidant content present in the extract directly react with DPPH radical produced а yellow diphenyl- β-picryl hydrazine complex. Deviation of discoloration directly proportional to phenolic favonoid content Figure 1. DPV extract was showed a maximum inhibition of 89.32 relatively close to of 91.96 inhibition of ascorbic acid at the concentration of 0.1 mg/ml. IC 50 of the DPV extract ascorbic acid was found to be 0.543 ± 0.002 mg/ml 0.495 ± 0.0005 mg/ml. Percentage inhibition of other was found to be in range of 51.44 to 74.33. IC 50 of other was found to be in the range of 0.871 ± 0.001 to 1.006 ± 0.007 mg/ml 2021. Hydrogen peroxide scavenging activity Hydrogen peroxide is oxygen containing species involved in the certain cellular activities like phagocytosis cell growth synthesis of biological compound. Hydrogen peroxide can cross cell membranes rapidly 22. Hydrogen peroxide scavenging activity of the extract are present in Figure 2. DPV extract showed a maximum inhibition of 72.33 relatively close to 77.42 inhibition of BHA at the concentration of 0.8 mg/ml. Te IC 50 value of the DPV extract BHA was found to be 0.279 ± 0.005 mg/ml 0.257 ± 0.002 mg/ml. Percentage inhibitions of others were found to be in the range of 39.44 to 68.33. IC 50 of others were found to be in range of 0.301 ± 0.001-0.553 ± 0.001 mg/ml. Nitric oxide scavenging activity Nitric oxide is potent chemical mediators which regulate the certain y 0.0163x + 0.0083 R² 0.997 0 0.2 0.4 0.6 0.8 1 1.2 0 0.2 0.4 0.6 0.8 Absorbance Concmicrogram/ml T ABS Linear ABS Figure 3: Standard curve between concentration of Gallic acid µg/ml and absorbance. y 0.0154x + 0.026 R² 0.9994 0 0.2 0.4 0.6 0.8 1 1.2 0 20 40 60 80 Absorbance Conc.microgram/ml ABS Linear ABS Figure 4: Standard curve between concentration of Rutin µg/ml and absorbance.

slide 4:

Citation: Dobhal K Semwal A Negi A 2018 Geographical Variation of Antioxidant Constituent in Garhwal Region of Uttarakhand: Chinese Brake Fern Pteris vittata. J Appl Pharm 10: 266. doi: 10.4172/1920-4159.1000266 Page 4 of 4 Volume 10 • Issue 3 • 1000266 J Appl Pharm an open access journal ISSN: 1920-4159 activities like smooth muscle relaxation neuronal signaling inhibition of platelet aggregation and regulation of cell mediated toxicity etc. It is a difusible free radical 22. Te scavenging of nitrogen oxide is present in Figure 5. DPV extract showed a maximum inhibition of 84.32 relatively close to 87.96 inhibition of ascorbic acid at the concentration of 0.8 mg/ml. Te IC 50 value of the DPV extract BHA was found to be in the range of 0.233 ± 0.002 mg/ml to 0.218 ± 0.006 mg/ml. Percentage inhibitions of others were found to be in the range of 41.44-76.33. IC 50 of others were found to be in the range of 0.259 ± 0.0052 to 0.499 ± 0.001 mg/ml. Conclusion Uttarakhand is rich source of natural sources. Alkaloid saponins phenolic and favonoid constituent are possessing efcient antioxidant activity in natural resources. It is unfortunate that these forms were discounted group of plants in biodiversity in spite of their economic value is familiar in worldwide. We summarized the outcomes that antioxidant activity of chinese brake fern was strongly revealed by ethanol extract of Dehradun Tehri district. It is very much supportive in the era of new drug investigation for the cancer tumors disease. Terefore cultivation should be supported as a medicinal plant for the development of health care products for aging and chronic disease. Te plant extract was obtained from diferent graphical region showed the strong presence of natural antioxidant. It is concluded that Dehradun and Tehri District possessing more favorable condition for the cultivation of Chinese bracken fern. Tis work will provide a foundation for further isolation of natural antioxidant constituent especially Dehradun and Tehri district of Uttarakhand. Acknowledgment We the authors are grateful to Chancellor Vice-Chancellor Director of Uttaranchal University for provide the kind knowledge and support. Conficts of Interest The authors declare there is no confict of interest. References 1. Kirtikar KR Basu BD Basu LM 1935 Indian Medicinal Plants. Allahabad. 2. Semwal A Kunwar A Prasad A Kumar A 2016 Azolla- An Environment Eco- Friendly Pteridophytic Species. Eur J Biomed Pharma Sci 3: 210-213. 3. Dixit RD Bhatt GK 1975 Ferns-a much-neglected group of medicinal plants. J Res Indian Med 10: 68-76. 4. Dhar ML Dhar MM Dhaman BN Mehrotra BN Roy C 1968 Screening various Indian ferns for biological activity. Indian J Experi Biol 6: 232-247. 5. Harbone JB 1998 Phytochemical methods. 3 rd Edn. Chapman and Hall London. 6. Khandelwal KR Sethi V 2016 Practical Pharmacognosy. 26 th Edn. Nirali Prakashan. 7. Obdoni BO Ochuko PO 2001 Phytochemical studies and comparative effcacy of the crude extracts of some homostatic plants in Edo and Delta States of Nigeria. Global J Pure Appl Sci 8: 203-208. 8. Slinkard K Singleton VL 1977 Total phenol analysis: Automation and comparison with manual methods. Am J Enol Vitic 28: 49-55. 9. Kumaran A Karunakaran R 2006 Anti-oxidant and free radical scavenging activity of an aqueous extracts of Coleus aromaticus. Food Chem 97: 109-114. 10. Alothman M Bhat R Karim AA 2009 Antioxidant Capacity and Phenolic Content of Selected Tropical Fruits from Malaysia Extracted with Different Solvents. Food Chem 115: 785-788. 11. Blois MS 1958 Antioxidant determinations by the use of a stable free radical. Nature 181: 1199-1200. 12. Lee SM Na MK An RB Min BS Lee HK 2003 Antioxidant activity of two phloroglucinol derivatives from Dryopteris crassirhizomaI. Biol Pharm Bull 26: 1354-1356. 13. Keser S Celik S Turkoglu S Yilmaz O Turkoglu J 2012 Hydrogen Peroxide Radical Scavenging and Total Antioxidant Activity of Haworthn. Chem J 02: 9-12. 14. Garrat DC 1964 The Quantitative analysis of Drugs. Chapman and Hall Ltd Japan. 15. Muraleedharannair M Johnson M Mony M Zachariah M Solomon J 2012 Inter-specifc variation studies on the phyto-constituents of Christella and Adiantum using phytochemical methods. As Paci J Tro Biomed 2: S40-S45. 16. Hassan SB Gullbo J Hu K Berenjian S Morein B et al. 2013The Nanoparticulate Quillaja Saponin BBE is selectively active towards renal cell carcinoma Anticancer Res 33: 143-151. 17. Ismail A Marjam ZM Foong CW 2004 Total antioxidant activity and Phenolic content in selected vegetables. Food Chem 87: 581-586. 18. Kahkonen MP Hopia AI Vuorela HJ 1999 Antioxidant activity of plant extracts containing phenolic compounds. J Agric Food Chem 47: 3954-3962. 19. Montoro P Braca A Pizza C De TN 2005 Structure-antioxidant activity relationships of favonoids isolated from different plant species. Food Chem 92: 349-55. 20. Silva CG Herdeiro RS Mathias CJ Panek A Silveira C et al. 2005 Evaluation of antioxidant activity of Brazilian plants Pharmacol Res 52: 229-233. 21. Chang HC Huang GJ Agrawal DC Kuo CL Wu CR et al. 2007 Antioxidant activities and polyphenol contents of six folk medicinal ferns used as “Gusuibu”. Bot Stud 48: 397-406. 22. Nishaa S Vishnupriyam AM Sasikumar JM Hephzibah PC Gopalakrishnan VK 2012 Antioxidant activity of ethanolic extract of Maranta arundinacea .L tuberous rhizomes. Asian J Pharm Clin Res 25: 85-88. 0 10 20 30 40 50 60 70 80 90 100 0 0.2 0.4 0.6 0.8 1 INHIBITION conc.mg/ml i roe n oi de c ae ni n aay CPV DPV PPV TPV UPV AA Figure 5: Nitric oxide scavenging inhibition versus concentration mg/ml.

authorStream Live Help