DEVELOPMENT AND VALIDATION OF A FAST AND SENSITIVE UHPLC-PDA METHOD FOR THE QUANTIFICATION OF URSOLIC ACID IN POLY(L-LACTIC ACID) NANOCAPSULES
Objective: The aim of the present study is to develop and validation of a ultra-high performance liquid chromatography (UHPLC) method to determine the ursolic acid content and its encapsulation efficiency (EE) in lipid-core nanocapsules prepared from poly (L-lactic acid).
Methods: A simple UHPLC-PDA method was developed and validated for the quantitative determination of ursolic acid in poly(L-lactic acid) nanocapsules. The chromatographic conditions used were: RP-C18 column, isocratic mobile phase containing acetonitrile:water (92:8, v/v), flow rate of 0.8 ml/min, column temperature of 50°C, and detection at 203 nm. The following parameters were evaluated: Specificity, linearity, limits of detection and quantification, precision, accuracy, and robustness.
Results: The method was specific to the ursolic acid and linear (r=0.9998) in the range of 10–100 μg/ml. The limits of detection and quantification were 1.35 and 4.10 μg/ml, respectively. The precision was demonstrated by a relative standard deviation less than 2%. Adequate accuracy (98.35%±0.82) was obtained. Changes in flow rate, mobile phase, and column temperature did not significantly alter the peak area and the retention time of the ursolic acid. The mean EE was 99.89%.
Conclusion: The method proved to be fast, sensitive, and simple for quantifying ursolic acid in nanocapsules and was successfully used for determining the EE.
2. Mlala S, Oyedeji AO, Gondwe M, Oyedeji OO. Ursolic acid and its derivatives as bioactive agents. Molecules 2019;24:2751.
3. Wo?niak ?, Sk?pska S, Marsza?ek K. Ursolic acid-a pentacyclic triterpenoid with a wide spectrum of pharmacological activities. Molecules 2015;20:20614-41.
4. López-Hortas L, Pérez-Larrán P, González-Muñoz M, Falqué E, Domínguez H. Recent developments on the extraction and application of ursolic acid. A review. Food Res Int 2018;103:130-49.
5. Sundaresan A, Harini R, Pugalendi KV. Ursolic acid and rosiglitazone combination alleviates metabolic syndrome in high fat diet fed C57BL/6J mice. Gen Physiol Biophys 2012;31:323-33.
6. Gutiérrez-Rebolledo GA, Siordia-Reyes AG, Meckes-Fischer M, Jiménez-Arellanes A. Hepatoprotective properties of oleanolic and ursolic acids in antitubercular drug-induced liver damage. Asian Pac J Trop Med 2016;9:644-51.
7. Singh AK, Pandey H, Ramteke P, Mishra SB. Nano-suspension of ursolic acid for improving oral bioavailability and attenuation of Type II diabetes: A histopathological investigation. Biocatal Agric Biotechnol 2019;22:101433-43.
8. Ramos-Hryb AB, Pazini FL, Kaster MP, Rodrigues AL. Therapeutic potential of ursolic acid to manage neurodegenerative and psychiatric diseases. CNS Drugs 2017;31:1029-41.
9. Checker R, Sandur SK, Sharma D, Patwardhan RS, Jayakumar S, Kohli V, et al. Potent anti-inflammatory activity of ursolic acid, a triterpenoid antioxidant, is mediated through suppression of NF-?B, AP-1 and NF-AT. PLoS One 2012;7:e31318.
10. Gai W, Yu D, Wang X, Wang P. Anti-cancer effect of ursolic acid activates apoptosis through ROCK/PTEN mediated mitochondrial translocation of cofilin-1 in prostate cancer. Oncol Lett 2016;12:2880-5.
11. Chan EW, Soon CY, Tan JB, Wong SK, Hui YW. Ursolic acid: An overview on its cytotoxic activities against breast and colorectal cancer cells. J Integr Med 2019;17:155-60.
12. Zhang C, Wang C, Li W, Wu R, Guo Y, Cheng D, et al. Pharmacokinetics and pharmacodynamics of the triterpenoid ursolic acid in regulating the antioxidant, anti-inflammatory, and epigenetic gene responses in rat leukocytes. Mol Pharm 2017;14:3709-17.
13. Nahak P, Karmakar G, Chettri P, Roy B, Guha P, Besra SE, et al. Influence of lipid core material on physicochemical characteristics of an ursolic acid-loaded nanostructured lipid carrier: An attempt to enhance anticancer activity. Langmuir 2016;32:9816-25.
14. Biswas S, Mukherjee PK, Harwansh RK, Bannerjee S, Bhattacharjee P. Enhanced bioavailability and hepatoprotectivity of optimized ursolic acid-phospholipid complex. Drug Dev Ind Pharm 2019;45:946-58.
15. Asai T. Nanoparticle-mediated delivery of anticancer agents to tumor angiogenic vessels. Biol Pharm Bull 2012;35:1855-61.
16. Yamamoto E, Miyazaki S, Aoyama C, Kato M. A simple and rapid measurement method of encapsulation efficiency of doxorubicin loaded liposomes by direct injection of the liposomal suspension to liquid chromatography. Int J Pharm 2018;536:21-8.
17. Nadal JM, Toledo MG, Pupo YM, Paula JP, Farago PV, Zanin SM. A stability-indicating HPLC-DAD method for determination of ferulic acid into microparticles: Development, validation, forced degradation, and encapsulation efficiency. J Anal Methods Chem 2015;2015:1-10.
18. Lü H, Lee R, Huang J, Chen J, Go VW, Li Z, et al. A new HPLC-UV method for the quantification of terpenoids and antioxidant activity of commercial loquat leaf tea and preparation. J Food Meas Charact 2020;14:1085-91.
19. Owczarek A, Ku?ma ?, Wysoki?ska H, Olszewska M. Application of response surface methodology for optimisation of simultaneous UHPLC-PDA determination of oleanolic and ursolic acids and standardisation of ericaceae medicinal plants. Appl Sci 2016;6:244-61.
20. Stebounova L, Ebert SM, Murry LT, Adams CM, Murry DJ. Rapid and sensitive quantification of ursolic acid and oleanolic acid in human plasma using ultra-performance liquid chromatography-mass spectrometry. J Chromatogr Sci 2018;56:644-9.
21. Zheng Z, Zhao X, Zhu S, Dang J, Qiao X, Qiu Z, et al. Simultaneous determination of oleanolic acid and ursolic acid by in vivo microdialysis via UHPLC-MS/MS using magnetic dispersive solid phase extraction coupling with microwave-assisted derivatization and its application to a pharmacokinetic study of Arctium lappa L. Root extract in rats. J Agric Food Chem 2018;66:3975-82.
22. Antonio EA, Junior OR, Khalil NM, Mainardes RM. A validated HPLC-PDA method for the analysis of ursolic acid content in poly (lactic acid) nanoparticles. Curr Chromatogr 2016;3:117-22.
23. Almeida MM, Bou-Chacra NA, Tavares VF, Baby AR, Kaneko TM, Velasco MV. Determination of ursolic acid from polymeric nanoparticles using high-performance liquid chromatography method. Lat Am J Pharm 2012;8:1187-91.
24. Baishya R, Nayak DK, Kumar D, Sinha S, Gupta A, Ganguly S, et al. Ursolic acid loaded PLGA nanoparticles: In vitro and in vivo evaluation to explore tumor targeting ability on b16f10 melanoma cell lines. Pharm Res 2016;33:2691-703.
25. Chesnut S, Salisbury JJ. The role of UHPLC in pharmaceutical development. J Sep Sci 2007;30:1183-90.
26. The International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use. Validation of Analytical Procedures: Text and Methodology Q2 (R1); 2005.
27. Chen G, Kim H, Kim E, Yoon J. Synthesis of high-molecular-weight poly (L-lactic acid) through the direct condensation polymerization of L-lactic acid in bulk state. Eur Polym J 2006;42:468-72.
28. Hyon S, Jamshidi K, Ikada Y. Synthesis of polylactides with different molecular weights. Biomaterials 1997;18:1503-8.
29. Horváth T, Szabó TJ, Marossy K. Molar mass determination of microwave initiated polycondensation produced PLLA by capillary viscometry method. J Phys 2020;1527:12-9.
30. Fessi H, Puisieux F, Devissaguet JP, Ammoury N, Benita S. Nanocapsule formation by interfacial polymer deposition following solvent displacement. Int J Pharm 1989;55:R1-4.
31. Rudnik LA, Farago PV, Budel JM, Lyra A, Barboza FM, Klein T, et al. Co-loaded curcumin and methotrexate nanocapsules enhance cytotoxicity against non-small-cell lung cancer cells. Molecules 2020;25:1913.
32. Justus B, Kanunfre CC, Budel JM, de Faria MF, Raman V, de Paula JP. New insights into the mechanisms of French lavender essential oil on nos-small-cell lung cancer cell growth. Ind Crops Prod 2019;136:28-36.
33. Almeida MA, Nadal JM, Klein T, de Paula JP, Budel JM, Novatiski A, et al. Innovative phytoformulation containing capsaicinoids: Microparticles development, analytical method validation, and anti-ulcer effect. Pharmacogn Mag 2018;14:290-6.
34. Gaspar MD, Lyra AM, Camargo GA, Nadal JM, Teitelbaum AP, dos Santos FA, et al. Development and validation of a reversed-phase UHPLC-PDA method for determination of chlorhexidine in meth (acrylic) nanocapsules. Lat Am J Pharm 2020;39:792-8.
35. Lopes CE, Langoski G, Klein T, Ferrari PC, Farago PV. A simple HPLC method for the determination of halcinonide in lipid nanoparticles: Development, validation, encapsulation efficiency, and in vitro drug permeation. Braz J Pharm Sci 2017;53:1-9.
36. Ghalia MA, Dahman Y. Biodegradable poly (lactic acid)-based scaffolds: Synthesis and biomedical applications. J Polym Res 2017;24:1-22.
37. Moazzen N, Khanmohammadi M, Garmarudi AB, Kazemipour M, Dogaheh MA. Optimization and infrared spectrometric evaluation of the mechanical properties of PLA-based bio-composites. J Macromol Sci A 2019;56:17-25.
38. Carletto B, Berton J, Ferreira TN, Dalmolin LF, Paludo KS, Mainardes RM, et al. Resveratrol-loaded nanocapsules inhibit murine melanoma tumor growth. Colloids Surf B Biointerfaces 2016;144:65-72.
This work is licensed under a Creative Commons Attribution 4.0 International License.
The publication is licensed under CC By and is open access. Copyright is with author and allowed to retain publishing rights without restrictions.