Background: Letrozole (LTZ) is a potent aromatase inhibitor which blocks estrogen synthesis
in post-menopausal women. Suitable analytical methods are required to quantify letrozole in bulk
and its nanoformulations. Hence a sensitive and robust reverse-phase high performance liquid chromatographic
method (RP-HPLC) was developed for the estimation of LTZ.
Methods: The method was developed using Kinetex C18 column (250 mm×4.6 mm; 5µ) and mobile
phase comprising acetonitrile (ACN): acetate buffer (pH 4.5) mixture (50:50% v/v) was used to affect
the chromatographic separation at 0.8 mL/min flow rate. The responses were measured at 240 nm.
Force degradation studies were done by exposing LTZ to acid- and alkali-induced hydrolysis, oxidation
(H2O2), thermal and photolysis. Two-level factorial design was used to validate the method as per ICH
Q2 (R1) using Design-Expert® software. The effects of independent variables on flow rate, pH, acetonitrile
content and column temperature were recorded as responses.
Results: Method was linear over the concentration range of 5-2500 ng/mL with a correlation coefficient
(R2) of 1.000. Force degradation studies revealed that LTZ was stable to all stress agents except the
basic conditions. The inter-day precision results were reproducible with relative standard deviation (%
RSD) of 0.078. The peak area RSD and mean recovery of LTZ was found to be <1.0% and 99-102% in
drug solution and <1.0% and 98-106% in nanoformulations respectively. Deliberate changes in independent
chromatographic parameters analyzed using analysis of variance (ANOVA) indicated that the
model was significant (p<0.0001).
Conclusion: The developed analytical method was successfully utilized to quantify LTZ in bulk and its