Aims: RP-HPLC-UV method was developed and validated for estimation of Carvacrol release rom
Carvacrol loaded nanobeads through study of interaction between method parameters on method responses by
Response Surface Methodology (RSM).
Background: Literature review reveals that there is no validated analytical method for determining drug release of
Carvacrol loaded nanobeads.
Objective: The main objective of the research is to develop a novel robust method for estimation of carvacrol in drug
release sample by applying RSM. Objective is to study the effect /interaction of method parameters like mobile phase
ratio, pH and flowrate to the method output/response such as retention time, peak area, tailing factor, theoretical
plates. To develop the design space and set the limit for all the method parameter is another objective. Finally the
optimized method would be validated as per ICH Q2(R1)guideline for parameters like accuracy, specificity,
precision, robustness, limit of detection, limit of quantitation.
Methods: HPLC method was optimized by RSM approach using Design of Expert software. Box behnken design
was applied using three factors and four responses.17 trials as per RSM were performed and final optimised LC
method was selected based on design space. LC-UV method was validated as per ICH Q2 (R1) guideline. Drug
release study was performed at pH 7.2 and pH 8.
Results: Mobile phase finalised was Tetrahydrofuran: Acetonitrile: Water: Triethylamine (70:18:12:1 %v/v) having
pH 6.5 adjusted with glacial acetic acid. Chromatographic column used was C18, 5µm particle size. Carvacrol was
detected at 275nm and 1 mL/min flow rate. In-vitro drug release of Carvacrol from topical formulation is performed
in a Franz diffusion cell. Drug release at pH 7.2 was found to be 82.8689 which is less than 90.1664 in phosphate
buffer pH 8 at 24 hours.
Conclusion: RSM was useful to minimize the experimental trials, study the interaction of method variation with
method responses and give design space for robust method. Method was proved to be precise, specific, accurate and
robust and suitable for Carvacrol release estimation.