Author(s):
Atul S. Sayare, Varad M. Deth, Prashant D. Ghode, Ujjwala Kandekar, Dhanashree M. Alange, Shweta P. Ghode, Sayali M. Deth
Email(s):
atulsayare@gmail.com
DOI:
10.52711/0974-360X.2026.00236 
Address:
Atul S. Sayare1*, Varad M. Deth1, Prashant D. Ghode1, Ujjwala Kandekar1, Dhanashree M. Alange1, Shweta P. Ghode2, Sayali M. Deth2
1JSPM’s Rajarshi Shahu College of Pharmacy and Research, Pune, Maharashtra, India – 411033.
2Rasiklal M. Dhariwal Institute of Pharmaceutical Education and Research, Pune, (M.S) India.
*Corresponding Author
Published In:
Volume - 19,
Issue - 4,
Year - 2026
ABSTRACT:
The present study deals with development of reversed phase HPLC method for the quantitation of Ofloxacin in ophthalmic solutions utilizing as an internal standard (Ornidazole). The approach is easy, sensitive, accurate, reliable, and precise. The stationary phase used was a Qualisil-5 BDS C8 column (250cm × 4.6mm, 5µm size of particle), chromatographic separation was carried out. Mobile phase consisted of phosphate buffer (pH 3) and acetonitrile (78:22 %v/v). For the chromatographic analysis the volume of injection was 20µL and flow rate was set at 1.0mL/min with UV detection at 280nm. It was found that developed method was linear for Ofloxacin (3-15µg/mL, r²=0.999). The method validation was carried out as per ICH Q2 (R1) guidelines. In the accuracy assessment, recovery rates at 80%, 100%, and 120% levels ranged from 98.14% to 100.13%. The values of detection limit (LOD) and quantification limit (LOQ) were 2.02µg/mL and 6.14µg/mL, respectively. All validation criteria met the required acceptance limits, confirming that the method is fit for analysis of Ofloxacin in ophthalmic preparations.
Cite this article:
Atul S. Sayare, Varad M. Deth, Prashant D. Ghode, Ujjwala Kandekar, Dhanashree M. Alange, Shweta P. Ghode, Sayali M. Deth. RP-HPLC Method Development and Validation for the Estimation of Ofloxacin in Eye Drop. Research Journal of Pharmacy and Technology. 2026;19(4):1646-0. doi: 10.52711/0974-360X.2026.00236
Cite(Electronic):
Atul S. Sayare, Varad M. Deth, Prashant D. Ghode, Ujjwala Kandekar, Dhanashree M. Alange, Shweta P. Ghode, Sayali M. Deth. RP-HPLC Method Development and Validation for the Estimation of Ofloxacin in Eye Drop. Research Journal of Pharmacy and Technology. 2026;19(4):1646-0. doi: 10.52711/0974-360X.2026.00236 Available on: https://www.rjptonline.org/AbstractView.aspx?PID=2026-19-4-25
REFERENCES:
1. Rajendran SS. Santhi N. Kumar Nallasivan P. Sam Solomon WD. Venkata Narayanan R. Simultaneous Estimation of Cefixime and Ofloxacin in Bulk and Tablet Dosage Form. Asian Journal of Pharmaceutical Analysis. 2011; 1(2): 36-38. doi: 10.5958/2231–5675
2. Vichare V. Tambe V. Joshi SV. Dhole SN. New Simultaneous UV-Visible Spectrophotometric Methods for Estimation of Ofloxacin and Ketorolac Tromethamine in Ophthalmic Dosage Form. Asian Journal of Pharmaceutical Analysis. 2013, 3(2): 53-57. doi: 10.5958/2231–5675
3. Pasupuleti C. Mohammad AS. Rasheed N. Umadevi K. Adilakshmi D. Simultaneous Formulation, Estimation and Evaluation of Ofloxacin Microbeads. Asian Journal of Pharmaceutical Analysis. 2016; 6(3): 167-182. doi: 10.5958/2231-5675.2016.00026.0
4. Mali A. Jadhav S. Gorad R. Quazi A. Bath R. Patil M. Tamboli A. Simultaneous UV Spectrophotometric Methods for Estimation of Cefixime Trihydrate and Ofloxacin in Bulk and Tablet Dosage Form. Asian Journal of Pharmaceutical Research. 2016; 6(2): 100-106. doi: 10.5958/2231-5691.2016.00017.4
5. Bhusari KP, Chaple DR. Simultaneous Spectrophotometric Estimation of Ofloxacin and Ornidazole in Tablet Dosage Form. Asian Journal of Research in Chemistry. 2009; 2(1): 60-62. DOI: 10.5958/0974-4150
6. Wankhede SB, Prakash A, Chitlange SS. Simultaneous Spectrophotometric Estimation of Ofloxacin and Satranidazole inTablet Dosage Form. Asian Journal of Research in Chemistry. 2008; 1(1):9-11. DOI: 10.5958/0974-4150
7. Siva Kumar R. Nallasivan PK. Saravanakumar S. Kandasamy CS. Venkatnarayanan R. Simultaneous RP-HPLC Estimation of Nitazoxanide and Ofloxacin in Tablet Dosage Forms. Asian Journal of Research in Chemistry. 2009; 2(1): 43-45. DOI: 10.5958/0974-4150
8. Venkatachalam T. Narendiran S. Selvi PK. Kumar PD. RP-HPLC Method for Simultaneous Determination of Ofloxacin and Satranidazole in Tablet Dosage Form. Asian Journal of Research in Chemistry. 2009; 2(4): 469-471. DOI: 10.5958/0974-4150
9. Prabhu S, Vijay Amirtharaj R, Senthilkumar N. Simultaneous RP-HPLC Method Development and Validation of Cefixime and Ofloxacin in Tablet Dosage Form. Asian Journal of Research in Chemistry. 2010; 3(2): 367-369. DOI: 10.5958/0974-4150
10. Pattanayak P. Chaudury YP. Sharma R. Mohapatra P. Setty DK. Simultaneous Spectrophotometric Estimation of Nitazoxanide and Ofloxacin in Combained Tablet Dosage Form. Research Journal of Pharmacy and Technology. 2009; 2(2): 291-293. DOI: 10.52711/0974-360X
11. Pattan SR. Lateef SN. Kendre PN. Dighe NS. Godge RK. Simultaneous Estimation of Ofloxacin and Satranidazole in Bulk and Dosage form by Ratio Spectra Derivative Spectroscopy. Research Journal of Pharmacy and Technology. 2009; 2 (3): 531-533. DOI: 10.52711/0974-360X
12. Shingote V. Mankar SD. Dighe SB. A Review Article on Analytical Methods Development and Validation. Research Journal of Science and Technology. 2022; 14(1): 77-3. doi: 10.52711/2349-2988.2022.00012
13. Game MD. Sakarkar DM. Gabhane KB. Tapar KK . Deokate UA. Absorbance Correction and Vierodt’s Spectrophotometric Methods for the Simultaneous Determination of Ofloxacin and Nitazoxanide in Combined Tablet Dosage Form. Research Journal of Pharmacy and Technology. 2010; 3(1): 214-216. DOI: 10.52711/0974-360X
14. Patel, S.K.N., et al., Analytical Method Development and Validation of Ofloxacin Eye Drop by HPLC. Journal of Current Chemical and Pharmaceutical Sciences. 2011. 1(1): 59-64.
15. Davis R. and Bryson HM. Levofloxacin: A Review of its Antibacterial Activity, Pharmacokinetics and Therapeutic Efficacy. Drugs. 1994; 47(4): 677-700. DOI: 10.2165/00003495-199447040-00008
16. Drlica K. and Zhao X. DNA Gyrase, Topoisomerase IV, and the 4-Quinolones. Microbiology and Molecular Biology Reviews. 1997; 61(3): 377-392. DOI: 10.1128/mmbr.61.3.377-392.1997
17. Hussy P. et al Effect Of 4-Quinolones and Novobiocin on Calf Thymus DNA Polymerase Alpha Primase Complex, Topoisomerases I and II, and Growth of Mammalian Lymphoblasts. Antimicrobial Agents and Chemotherapy, 1986; 29(6): 1073-1078. DOI: 10.1128/AAC.29.6.1073
18. Hosomi J. et al Mutagenicity of Norfloxacin and AM-833 in Bacteria and Mammalian Cells. Reviews of Infectious Diseases, 10 (Suppl. 1). 1988; S148–S149.
19. Gootz T. Barrett J. Sutcliffe J. Inhibitory Effects of Quinolone Antibacterial Agents on Eucaryotic Topoisomerases and Related Test Systems. Antimicrobial Agents and Chemotherapy. 1990. 34(1): 8-12. doi: 10.1128/aac.34.1.8
20. Lawrence JW. et al 4‐Quinolones Cause a Selective Loss of Mitochondrial DNA from Mouse L1210 Leukemia Cells. Journal of Cellular Biochemistry. 1993; 51(2): 165-174. doi: 10.1002/jcb.240510208.
21. Al-Soud YA. Al-Masoudi NA. Ferwanah AS. Synthesis and Properties of New Substituted 1,2,4-Triazoles: Potential Antitumor Agents. Bioorganic and Medicinal Chemistry. 2003; 11 (8): 1701-1708. https://doi.org/10.1016/S0968-0896(03)00043-9
22. Zhou G. and Pan J. Polarographic and Voltammetric Behaviour of Ofloxacin and its Analytical Application. Analytica Chimica Acta. 1995; 307(1): 49-53. https://doi.org/10.1016/0003-2670(95)00028-X
23. Rizk M. et al Differential Pulse Polarographic Determination of Ofloxacin in Pharmaceuticals and Biological Fluids. Talanta. 1998; 46(1): 83-89. https://doi.org/10.1016/S0039-9140(97)00249-X
24. Tamer A. Adsorptive Stripping Voltammetric Determination of Ofloxacin. Analytica Chimica Acta. 1990; 231: 129-131. https://doi.org/10.1016/S0003-2670(00)86407-7
25. Okazaki O. Aoki H. and Hakusui H. High Performance Liquid Chromatographic Determination of (S)-(−)-Ofloxacin and its Metabolites in Serum and Urine Using a Solid-Phase Clean-Up. Journal of Chromatography B: Biomedical Sciences and Applications. 1991; 563(2): 313-322. https://doi.org/10.1016/0378-4347(91)80037-D
26. Ohkubo T. Kudo M. and Sugawara K. Determination of Ofloxacin in Human Serum by High-Performance Liquid Chromatography with Column Switching. Journal of Chromatography B: Biomedical Sciences and Applications. 1992; 573(2): 289-293. DOI: 10.1016/0378-4347(92)80131-9
27. Wright DH. et al Determination of Quinolone Antibiotics in Growth Media by Reversed-Phase High-Performance Liquid Chromatography. Journal of Chromatography B: Biomedical Sciences and Applications. 1998; 709(1): 97-104. DOI: 10.1016/s0378-4347(98)00006-1