Rajasekaran R., Suresh P. K.
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Rajasekaran R.1, Suresh P. K.2*
1Research Scholar, School of Biosciences and Technology, VIT, Vellore, India - 632014.
2School of Biosciences and Technology, VIT, Vellore, India - 632014.
Volume - 14,
Issue - 6,
Year - 2021
Identification and isolation of active principles from Lepidium sativum L. garden cress seeds and their chemical/biological characterization would aid in drug development (templates for synthesizing derivatives or as a value-added product). Initially, this study dealt with the phytochemical, qualitative and quantitative analysis for phenolic and flavonoid compounds. Subsequently, different physical and chemical extraction techniques -Simple Crude extraction (CRU), Ultra Sonication-Assisted Extraction (UAE), Microwave-Assisted Extraction (MAE) and classical Soxhlet extraction were adopted to select the best Lepidium sativum L. seed extract based on their activity in certain classical, in vitro antioxidant assay systems. The selected best extract was further analyzed for characterization using UPLC, GC-MS. CRU MeOH extracts showed a higher Total phenolic content of 4464.1±349.7mg GAE/100g. The UAE method showed a higher total flavonoid content of 1520.6±182.2mg QUE/100g. CRU-MeOH showed good antioxidant activity based on the free radical scavenging DPPH assay (IC50: 50.61µg/mL). A 100µg/mL concentration of CRU-MeOH was found to be 217.82±12.82 FRAP value. At 400 µg/mL, the NO scavenging assay was reported to be 62.11±4.84%. The present study indicates that extracts from the polar solvents shows better antioxidant scavenging potential than that of the non-polar extracts. Based on the relatively superior performance of the CRU-MeOH extract, UPLC-PDA data was generated. The validity of our chromatography conditions was shown by the presence of quercetin at a retention time 8.555 (validated by our internal standard), apart from the presence of this important bioactive molecule in our extract. As another strand of evidence for the presence of important bioactive molecules, p-coumaric acid, and ferulic acid were also present in our extracts, following correlation of their UPLC-PDA data with published findings. Also, GC-MS analysis of the CRU MeOH extract showed that 24 compounds (8 major and 16 minor peaks). One of the compounds detected (cis-vaccenic acid) is in consonance with published data. The presence of bioactive components, with known antioxidant and cell death potential, validates our experimental flow for the development and/or refinement of crude extract-based drugs. However, the variability perforce warrants an inter-laboratory harmonization of protocols for making meaningful comparisons.
Cite this article:
Rajasekaran R., Suresh P. K. Physical and Chemical methods of extraction of Bioactive Molecules from Lepidium sativum Linn. and Antioxidant Activity-based screening and selection of extracts-Probable Phytochemical, Chromatography and mass spectroscopy analysis-based correlates. Research Journal of Pharmacy and Technology. 2021; 14(6):3082-2. doi: 10.52711/0974-360X.2021.00539
Rajasekaran R., Suresh P. K. Physical and Chemical methods of extraction of Bioactive Molecules from Lepidium sativum Linn. and Antioxidant Activity-based screening and selection of extracts-Probable Phytochemical, Chromatography and mass spectroscopy analysis-based correlates. Research Journal of Pharmacy and Technology. 2021; 14(6):3082-2. doi: 10.52711/0974-360X.2021.00539 Available on: https://rjptonline.org/AbstractView.aspx?PID=2021-14-6-29
1. Jadhav Sameer S., Salunkhe Vijay R, Magdum Chandrakant S.. Daily Consumption of Antioxidants:-Prevention of Disease is better than Cure. Asian Journal of Pharmaceutical Research. 2013. 3(1): 33-39.
2. Arya B, Krishnaveni K, Sambathkumar R. Review on Antioxidant Supplements use in Cancer Chemotherapy. Research Journal of Pharmacology and Pharmacodynamics.2020; 12(1):21-24.
3. Charde RM., Dhongade HJ., Charde MS, Joshi SB. Evaluation of Wound Healing, Anti-Inflammatory and Antioxidant Activity of Rhizomes of Curcuma longa. Research Journal of Pharmacology and Pharmacodynamics. 2010; 2(1):42-47.
4. Attia ES, Amer AH, Hasanein MA. The hypoglycemic and antioxidant activities of garden cress (Lepidium sativum L.) seed on alloxan-induced diabetic male rats. Natural Product Research. 2019; 33(6): 901-905.
5. Mahassni SH, Al-Reemi RM. Apoptosis and necrosis of human breast cancer cells by an aqueous extract of garden cress (Lepidium sativum) seeds. Saudi Journal of Biological Sciences. 2013; 20(2): 131-139.
6. Kadam D, Palamthodi S, Lele SS. LC-ESI-Q-TOF-MS/MS profiling and antioxidant activity of phenolics from L. sativum seedcake. Journal of food science and technology. 2018; 55(3): 1154-1163.
7. Ait-Yahia O, Bouzroura SA, Belkebir A, Kaci S, Aouichat AB. Cytotoxic activity of flavonoid extracts from Lepidium Sativum (Brassicaceae) seeds and leaves International Journal of Pharmacognosy and Phytochemical Research. 2015; 7(6): 1231-1235
8. Zamzami MA, Baothman OA, Samy F, Abo-Golayel MK. Amelioration of CCl4-Induced Hepatotoxicity in Rabbits by Lepidium sativum Seeds. Evidence-Based Complementary and Alternative Medicine. 2019; 2019.
9. Basaiyye SS, Kashyap S, Krishnamurthi K, Sivanesan S. Induction of apoptosis in leukemic cells by the alkaloid extract of garden cress (Lepidium sativum L.). Journal of Integrative Medicine. 2019 ;17(3): 221-228.
10. Al-Asmari AK, Athar MT, Al-Shahrani HM, Al-Dakheel SI, Al-Ghamdi MA. Efficacy of Lepidium sativum against carbon tetra chloride induced hepatotoxicity and determination of its bioactive compounds by GC-MS. Toxicology reports. 2015; 2: 1319-1326.
11. Alqahtani FY, Aleanizy FS, Mahmoud AZ, Farshori NN, Alfaraj R, Al-sheddi ES, Alsarra IA. Chemical composition and antimicrobial, antioxidant, and anti-inflammatory activities of Lepidium sativum seed oil. Saudi Journal of Biological Sciences. 2019; 26(5): 1089-1092.
12. Zia-Ul-Haq M, Ahmad S, Calani L, Mazzeo T, Del Rio D, Pellegrini N, De Feo V. Compositional study and antioxidant potential of Ipomoea hederacea Jacq. and Lepidium sativum L. seeds. Molecules. 2012; 17(9): 10306-10321.
13. Ghasemzadeh A, Jaafar HZ, Rahmat A. Effects of solvent type on phenolics and flavonoids content and antioxidant activities in two varieties of young ginger (Zingiber officinale Roscoe) extracts. Journal of Medicinal Plants Research. 2011; 5(7): 1147-1154.
14. Singh M, Jha A, Kumar A, Hettiarachchy N, Rai AK, Sharma D. Influence of the solvents on the extraction of major phenolic compounds (punicalagin, ellagic acid and gallic acid) and their antioxidant activities in pomegranate aril. Journal of Food Science and Technology. 2014; 51(9): 2070-2077.
15. Teh SS, Bekhit AE, Birch J. Antioxidative polyphenols from defatted oilseed cakes: effect of solvents. Antioxidants. 2014; 3(1): 67-80.
16. Samaram S, Mirhosseini H, Tan CP, Ghazali HM. Ultrasound-assisted extraction (UAE) and solvent extraction of papaya seed oil: Yield, fatty acid composition and triacylglycerol profile. Molecules. 2013; 18(10): 12474-12487.
17. Zhang ZS, Wang LJ, Li D, Jiao SS, Chen XD, Mao ZH. Ultrasound-assisted extraction of oil from flaxseed. Separation and Purification Technology. 2008; 62(1): 192-198.
18. Farhat A, Fabiano-Tixier AS, Visinoni F, Romdhane M, Chemat F. A surprising method for green extraction of essential oil from dry spices: microwave dry-diffusion and gravity. Journal of Chromatography A. 2010; 1217(47): 7345-7350.
19. Malar J, Chairman K, Singh AR, Vanmathi JS, Balasubramanian A, Vasanthi K. Antioxidative activity of different parts of the plant Lepidium sativum Linn. Biotechnology Reports. 2014;3:95-98.
20. Doke S, Guha M. Identification of extraction conditions for determination of phenolic contents of garden cress seed (Lepidium sativum L.) and its milled fractions. Food analytical methods. 2015; 8(4): 1053-7.
21. Umesha SS, Naidu KA. Antioxidants and antioxidant enzymes status of rats fed on n-3 PUFA rich Garden cress (Lepidium sativum L) seed oil and its blended oils. Journal of Food Science and Technology. 2015; 52(4): 1993-2002.
22. Tayel AA, Salem MF, El‐Tras WF, Brimer L. Exploration of Islamic medicine plant extracts as powerful antifungals for the prevention of mycotoxigenic Aspergilli growth in organic silage. Journal of the Science of Food and Agriculture. 2011; 91(12): 2160-2165.
23. Muthukumaran P, Shanmuganathan P, Malathi, C. In Vitro Antioxidant Evaluation of Mimosa pudica. Asian Journal of Pharmaceutical Research. 2011; 1(2): 44-46.
24. Muthukumaran P, Abirami C, Priyatharsini. Antioxidant and Free Radical Scavenging Activity Azima tetracantha Lam. Research Journal of Pharmaceutical Dosage Forms and Technology. 2013; 5(6): 315-319.
25. Umesha SS, Naidu KA. Antioxidants and antioxidant enzymes status of rats fed on n-3 PUFA rich Garden cress (Lepidium sativum L) seed oil and its blended oils. Journal of Food Science and Technology. 2015; 52(4): 1993-2002.
26. Jose BE, Panneer Selvam P. Identification of Phytochemical Constituents in the Leaf Extracts of Azima tetracantha Lam using Gas Chromatography-Mass Spectrometry (GC-MS) analysis and Antioxidant Activity. Asian Journal of Research in Chemistry. 2018; 11(6): 857-862.
27. Eswarudu MM, Eswaraiah MC, Kumar KP, Sudhakar K. Ultra Performance Liquid Chromatography (UPLC): A Preeminent Technique in Pharmaceutical Analysis.. Research Journal of Pharmacy and Technology. 2012. 5(12): 1484-1489.
28. Sanganna B., Kulkarni AR. Antioxidant and Anti-colon cancer activity of fruit peel of Citrus reticulate essential oil on HT-29 cell line. Research Journal of Pharmacy and Technology. 2013.6(2): 216-219.
29. George VC, Kumar DR, Rajkumar V, Suresh PK, Kumar RA. Quantitative assessment of the relative antineoplastic potential of the n-butanolic leaf extract of Annona muricata Linn. in normal and immortalized human cell lines. Asian Pacific Journal of Cancer Prevention. 2012;13(2):699-704.
30. Rispail, N, Morris, P, Webb, Kj. Phenolic compounds: extraction and analysis. Lotus japonicus. Handbook.Springer, Dordrecht.2005; 349-354.
31. Chemat S, Aït-Amar H, Lagha A, Esveld DC. Microwave-assisted extraction kinetics of terpenes from caraway seeds. Chemical Engineering and Processing: Process Intensification. 2005; 44(12): 1320-6.
32. Spigno G, De Faveri DM. Microwave-assisted extraction of tea phenols: a phenomenological study. Journal of Food Engineering. 2009; 93(2): 210-217.
33. Verma SC, Jain CL, Padhi MM, Devalla RB. Microwave extraction and rapid isolation of arjunic acid from Terminalia arjuna (Roxb. ex DC.) stem bark and quantification of arjunic acid and arjunolic acid using HPLC‐PDA technique. Journal of Separation Science. 2012; 35(13): 1627-33.
34. Palma M, Barroso CG. Ultrasound-assisted extraction and determination of tartaric and malic acids from grapes and winemaking by-products. Analytica Chimica Acta. 2002; 458(1): 119-130.
35. Derakhshan Z, Ferrante M, Tadi M, Ansari F, Heydari A, Hosseini MS, Conti GO, Sadrabad EK. Antioxidant activity and total phenolic content of ethanolic extract of pomegranate peels, juice and seeds. Food and Chemical Toxicology. 2018; 114:108-111.
36. Tiwari P, Patel RK. Estimation of Total Phenolics and Flavonoids and Antioxidant Potential of Ashwagandharishta Prepared by Traditional and Modern Methods. Asian Journal of Pharmaceutical Analysis. 2013. 3(4): 147-152.
37. Sinha A, Suresh PK. Enhanced induction of apoptosis in HaCaT cells by luteolin encapsulated in PEGylated liposomes-Role of Caspase-3/Caspase-14. Applied Biochemistry and Biotechnology. 2019; 188(1): 147-64.
38. Marcocci L, Packer L, Droy-Lefaix MT, Sekaki A, Gardès-Albert M.  Antioxidant action of Ginkgo biloba extract EGb 761. InMethods in enzymology 1994; 234: 462-475.
39. Etim OE, Ekanem SE, Sam SM. In Vitro Antioxidant Activity and Nitric Oxide Scavenging Activity of Citrullus lanatus Seeds. In Vitro. 2013; 3(12).
40. Sakakibara H, Honda Y, Nakagawa S, Ashida H, Kanazawa K. Simultaneous determination of all polyphenols in vegetables, fruits, and teas. Journal of Agricultural and Food Chemistry. 2003; 51(3): 571-581.
41. Agarwal J, Verma D. Antioxidant activity- guided fractionation of aqueous extracts from Lepidium sativum and identification of active flavonol glycosides. Academia Arena 2011; 3 (12); 10–13.
42. Diwakar BT, Dutta PK, Lokesh BR, Naidu KA. Physicochemical properties of garden cress (Lepidium sativum L.) seed oil. Journal of the American Oil Chemists' Society. 2010; 87(5): 539-548.
43. Ullah MA, Tungmunnithum D, Garros L, Hano C, Abbasi BH. Monochromatic lights-induced trends in antioxidant and antidiabetic polyphenol accumulation in in vitro callus cultures of Lepidium sativum L. Journal of Photochemistry and Photobiology B: Biology. 2019; 196: 111505.
44. Indumathy R, Aruna A. Free radical scavenging activities, total phenolic and flavonoid content of Lepidium sativum (Linn.). International Journal of Pharmacy and Pharmaceutical Sciences. 2013;5:634-637.
45. Inglett GE, Rose DJ, Chen D, Stevenson DG, Biswas A. Phenolic content and antioxidant activity of extracts from whole buckwheat (Fagopyrum esculentum Möench) with or without microwave irradiation. Food Chemistry. 2010; 119(3): 1216-1219.
46. Krishnan RY, Rajan KS. Microwave assisted extraction of flavonoids from Terminalia bellerica: study of kinetics and thermodynamics. Separation and Purification Technology. 2016; 157: 169-178.
47. Ait-yahia O, Perreau F, Bouzroura SA, Benmalek Y, Dob T, Belkebir A. Chemical composition and biological activities of n-butanol extract of Lepidium sativum L (Brassicaceae) seed. Tropical Journal of Pharmaceutical Research. 2018; 17(5): 891-896.
48. Bimakr M, Rahman RA, Saleena Taip F, Adzahan NM, Islam Sarker Z. Ultrasound-assisted extraction of valuable compounds from winter melon (Benincasa hispida) seeds. International Food Research Journal. 2013; 20(1): 331–338.
49. Seidi K, Jahanban-Esfahlan R, Abasi M, Abbasi MM. Anti tumoral properties of Punica granatum (Pomegranate) seed extract in different human cancer cells. Asian Pacific Journal of Cancer Prevention. 2016; 17(3): 1119-1122
50. Gallo M, Ferracane R, Graziani G, Ritieni A, Fogliano V. Microwave assisted extraction of phenolic compounds from four different spices. Molecules. 2010;15(9): 6365-6374.
51. Fan QL, Zhu YD, Huang WH, Qi Y, Guo BL. Two new acylated flavonol glycosides from the seeds of Lepidium sativum. Molecules. 2014;19(8): 11341-11349.
52. Fernandes de Oliveira AM, Sousa Pinheiro L, Souto Pereira CK, Neves Matias W, Albuquerque Gomes R, Souza Chaves O, de Souza V, De Fátima M, Nóbrega de Almeida R, Simões de Assis T. Total phenolic content and antioxidant activity of some Malvaceae family species. Antioxidants. 2012; 1(1): 33-43
53. Piluzza G, Bullitta S. Correlations between phenolic content and antioxidant properties in twenty-four plant species of traditional ethnoveterinary use in the Mediterranean area. Pharmaceutical Biology. 2011; 49(3): 240-7.
54. Buitrago D, Buitrago-Villanueva I, Barbosa-Cornelio R, Coy-Barrera E. Comparative Examination of Antioxidant Capacity and Fingerprinting of Unfractionated Extracts from Different Plant Parts of Quinoa (Chenopodium quinoa) Grown under Greenhouse Conditions. Antioxidants. 2019; 8(8): 238.
55. Hussein HJ, Hameed IH, Hadi MY. Using Gas Chromatography-Mass Spectrometry (GC-MS) Technique for Analysis of Bioactive Compounds of Methanolic Leaves extract of Lepidium sativum. Research Journal of Pharmacy and Technology. 2017;10(11):3981-3989.
56. Chemat S, Aït-Amar H, Lagha A, Esveld DC. Microwave-assisted extraction kinetics of terpenes from caraway seeds. Chemical Engineering and Processing: Process Intensification. 2005; 44(12): 1320-1326
57. Hamid Khan, Mushir Ali, Alka Ahuja, Javed Ali. Validated UPLC/Q-TOF-MS Method for Simultaneous Determination of Metformin, Glimepiride and Pioglitazone in Human Plasma and its Application to Pharmacokinetic Study. Asian Journal of Pharmacy and Technology. 2017; 7(1): 27-32.
58. Goh E, Gledhill A. Analysis of polyphenols in fruit juices using acquity UPLC H-class with UV and MS detection. 2011; 1–9.
59. Orlovskaya TV, Chelombit’Ko VA. Phenolic compounds from Lepidium sativum. Chemistry of Natural Compound. 2007; 43 (3): 323
60. Taniguchi M, Lindsey JS. Database of absorption and fluorescence spectra of> 300 common compounds for use in photochem CAD. Photochemistry and Photobiology. 2018; 94(2): 290-327.