Micellar electrokinetic chromatographic screening of letrozole and its metabolite in human urine: Validation and robustness/ruggedness evaluation

Abstract

A simple, rapid, and sensitive method has been proposed and validated to directly quantify letrozole (LE) and its metabolite, bis-4-cyanophenylmethanol (ME) in urine samples (without any additional treatment) by micellar electrokinetic capillary chromatography (MEKC). In an effort to improve the selectivity and sensitivity of the method, the chemical and instrumental parameters were optimized. The best conditions were: 70 mM borate buffer (pH 9.2) and 40 mM SDS as BGE, 25 kV and 20 degrees C as working voltage and temperature, respectively, with hydrodynamic injection for 6 s. The reliability of the proposed method was also proved by means of a validation procedure based on precision, accuracy, linearity, LOD (15 microg/L for both of them) and LOQ studies. Moreover, an innovatory experimental and statistical design approach, upon a Plackett-Burman fractional factorial model, which involves the simultaneous evaluation of the global robustness and ruggedness effects, was applied. As it has been already stated, the proposed method has been successfully used to directly quantify both compounds in human urine samples, without any additional treatment, but the previously reached LOD and LOQ values can be improved by applying an SPE preconcentration procedure, also developed and optimized by the authors in this work. Real determinations of these analytes in clinical urines of a patient under LE treatment were performed, too.

Determination of letrozole, anastrozole and exemestane by capillary zone electrophoresis

“…At the beginning of determinations we have selected the type of background electrolyte (BGE) by checking the electrophoretic behavior of the analytes using an acidic BGE -phosphoric acid 25 mM and a basic BGE -25 mM borax. After our prelimary results we selected a 25 mM borax (pH 9.3) as BGE also taking in consideration previous electrophoretic studies [17][18][19][20][21].…”

“…Several capillary electrophoretic methods have been reported for determination of AIs in combination with other compounds, mainly micellar electrokinetic capillary chromatographic (MEKC) [17][18][19][20]. So far, there is no published electrophoretic method for the simultaneous determination of all three AIs selected in this research.…”

Letrozole Determination by Capillary Zone Electrophoresis and UV Spectrophotometry Methods

“…Initially, we choose to start with a 25 mM phosphate buffer, an acidic background electrolyte (BGE), but the electrophoretic signal of the substance was difficult to detect (low value of area and height). A standard BGE containing 25 mM borax (pH 9.3) was than selected, taking in consideration also previous electrophoretic studies, however the disadvantage in a basic environment was that LET migrated very close to the electroosmotic flow (EOF) [16][17][18][19]. Adding sodium dodecyl sulfate (SDS) in the 25 mM borax BGE did not resolved the problem.…”

“…Until now, on a base of detailed literature survey, it was found that LET could be estimated along with other compounds only by MEKC [16][17][18][19].…”

“…Similar data has been previously obtained at the administation of a single dose of letrozole (2.5 mg) with 14C-labeled within two weeks when the recovered radioactivity from urine was 88.2 ± 7.6% (5.0 ± 2.4% unmetabolized letrozole, 63.2 ± 11.2% carbinole glucuronide and about 8% other metabolites) [15]. A few capillary electrophoretic (CE) methods for LET determination have been already reported in the literature, mainly micellar electrokinetic capillary chromatographic (MEKC) [16][17][18][19]. The electrophoretic methods are more advantageous through accessibility than other expensive analytical methods as HPLC and RP-HPLC, LC-MS/MS or SPE-UPLC-MS/MS [20][21][22][23][24].…”