Urolithiasis: HPTLC Method for Quantitative Detection of Rutin and Quercetin in an Herbal Plant
Keywords:Densitometry, <i>Ocimum basilicum</i>, Quercetin, High-performance Thin-layer Chromatography (HPTLC), Rutin, Validation
AbstractAnti-inflammatory, Hypoglycemic, Hepatoprotective, Antihyperlipidemic, Anti-Ulcerative, Cardioprotective Stimulant, Sedative, Hypnotic, Anticonvulsant Activity, Memory Retention, Stroke Preventive Activity, Antimicrobial Activity, Antimycobacterial, Antiviral, Larvicidal, Antiparasitic, Chemopreventative, Chemo-modulatory, Anticancer, Cytoprotective activities were found in a variety of herbal plants, including Ocimum basilicum. The purpose of this study was to design and develop a new HPTLC method that was accurate, precise and cost-effective for simultaneous measurement of rutin and quercetin in a hydroalcoholic extract of Ocimum basilicum seeds. The mobile phase was Toluene: Ethyl Acetate: Methanol: formic acid (6:4:3:1, v/v/v/v/v) and densitometric scanning was performed at 254 nm. Merck TLC aluminium sheets of silica gel 60 F254, (10 x 10 cm) with a thickness of 250 mm was used as stationary phase. At a wavelength of 254 nm, rutin and quercetin were detected. The constituents were resolved satisfactorily, with Rf values of 0.25 ± 2.01 for rutin and 0.80 ± 0.64 for quercetin, respectively. Linearity (300-1300 ng/spot for rutin and quercetin) was used to test the method’s accuracy and reproducibility. For both analytes, intra- and inter-day precision, as evaluated by coefficient of variation, was less than 3%. For rutin, the detection and quantification limits were 46.52 and 140.96 ng/spot, respectively, and for quercetin, they were 81.79 and 247.84 ng/spot. The proposed approach was found to be precise, accurate, repeatable, and specific, and it could be used to measure quercetin and rutin in samples at the same time.
Schreiner M, Huyskens-Keil S. Phytochemicals in fruit and vegetables: Health promotion and postharvest elicitors. Cri Rev Plant Sci. 2006; 25:267-78. https://doi.org/10.1080/07352680600671661
Soleas GJ, Grass L, Josephy PD, Goldberg DM, Diamandis EP. A comparison of the anticarcinogenic properties of four red wine polyphenols. Clin Biochem. 2002; 35:119-24. https://doi.org/10.1016/S0009-9120(02)00275-8
Middleton EM, Teramura AH. The role of flavonol glycosides and carotenoids in protecting soybean from ultraviolet-B damage. Plant Physiol. 1993; 103:741-52. https://doi.org/10.1104/pp.103.3.741. PMid:12231976. PMCid:PMC159044
Hein KE, Tagliaferro AR, Bobilya DJ. Flavonoid antioxidants: Chemistry, metabolism and structure-activity relationships. J Nutr Biochem. 2002; 13(10):572-84. https://doi.org/10.1016/S0955-2863(02)00208-5
Bors W, Heller W, Michel C, Saran M. Flavonoids as antioxidants: determination of radical-scavenging efficiencies. Method Enzymol. 1990; 186:343-55. https://doi.org/10.1016/0076-6879(90)86128-I
Dixon RA, Dey PM, Lamb CJ. Phytoalexins: enzymology and molecular biology. Adv Enzymol RAMB.1983; 55:1-136.
Rice-Evans CA, Miller NJ, Bolwell PG, Broamley PM, Pridham JB. The relative antioxidant activities of plantderived polyphenolic flavonoids. Free Radical Res. 1995; 22(4):375-83. https://doi.org/10.3109/10715769509145649. PMid:7633567
Agati G, Azzarello E, Pollastri S, Tattini M. Flavonoids as antioxidants in plants: location and functional significance. Plant Sci. 2012; 196:67-76. https://doi.org/10.1016/j.plantsci.2012.07.014. PMid:23017900
Du F, Zhang F, Chen F, Wang A. Advances in microbial heterologous production of flavonoids. Afr J Microbiology Res. 2011; 5(18):2566-74. https://doi.org/10.5897/AJMR11.394
Middleton EJ. Effect of plant flavonoids on immune and inflammatory cell function. Adv Exp Med Biol. 1998; 439:175-82. https://doi.org/10.1007/978-1-4615-5335-9_13. PMid:9781303
Kumar S, Pandey AK. Chemistry and biological activities of flavonoids: An overview. Scientific World J. 2013; 2013:1- 16. https://doi.org/10.1155/2013/162750. PMid:24327805. PMCid:PMC3845396
Nijveldt RJ, Nood EL, Van Horn DEC, Boelens PG, Van Norren K, van Leeuwen PAM. Flavonoids: A review of probable mechanisms of action and potential applications. Am J Clin Nutr. 2001; 74(4):418-25. https://doi.org/10.1093/ajcn/74.4.418. PMid:11566638
Bilal A, Jahan N, Ahmed A, Bilal SN, Habib S, Hajra S. Phytochemical and pharmacological studies on Ocimum basilicum linn — A review. Int J Curr Res. 2012; 4(23).
International Conference on Harmonization. Validation of Analytical Procedures Methodology Q2B; 1996.
Method for determination of forskolin in crude drug and pharmaceutical dosage form. Chromatographia. 2008; 67:441-47. https://doi.org/10.1365/s10337-008-0521-x
Alam P, Ali M, Ahmad SA. A validated HPLC method for estimation of cordifolioside A in Tinospora cardifolia, Miers and marketed formulations. J Chromatogr Sci. 2009; 47: 910-13. https://doi.org/10.1093/chromsci/47.10.910. PMid:19930804
Parveen R, Baboota S, Ahmad S, Ali J, Ahuja A. Stabilityindicating HPTLC method for quantitative estimation of silybin in bulk drug and pharmaceutical dosage form. Biomed Chromatogr. 2010; 24:639-47. https://doi.org/10.1002/bmc.1340. PMid:19816854
Rozylo JK, Janicka M. Different planar techniques for prediction of solute retention in column liquid chromatography. J Planar Chromatogr. 1996; 9:418-24.
Weins C, Hauck HE. Advances and developments in thin layer chromatography. LC-GC Int. 1996; 4:455-71.
Poole CF, Poole SK. Instrumental thin layer chromatography. Anal Chem. 1994; 66:27A-37A. https://doi.org/10.1021/ac00073a001
Sejal P, Niraj V. Validated spectrofluorimetric method for estimation of piperine in an ayurvedic formulation. Asian J Pharm Clin Res. 2012; 5(4):231-233.
Niraj V, Sangita P. Development and Validation of RP-HPLC Method for Simultaneous estimation of Nebivolol and Indapamide in Pharmaceutical Dosage Form. Asian J Pharm Anal. 2014; 4(3):98-102.