Validation of a Liquid Chromatographic/Tandem Mass Spectrometric Method for the Determination of Scopolamine Butylbromide in Human Plasma: Application of the Method to a Bioequivalence Study

Abstract

A sensitive and specific LC/MS/MS method was developed and validated for the determination of scopolamine butylbromide in human plasma. Scopolamine butylbromide and propanolol (internal standard) were extracted from the plasma by liquidliquid extraction with dichloromethane as the extraction solvent and separated on a C18 analytical column (50 4.6 mm id) maintained at 40C. The analytes were eluted at a constant flow rate of 0.45 mL/min; the mobile phase consisted of acetonitrile and a buffer of 5 mM ammonium acetate and 0.1 formic acid (60 + 40, v/v). The mass spectrometer, equipped with an electrospray source in the positive ionization mode, was set up in the multiple-reaction monitoring mode to monitor the transitions m/z 360.6 > 102.5 (scopolamine butylbromide) and m/z 259.7 > 115.6 (propanolol). The chromatographic separation was obtained within 2.0 min, and the responses were linear over the concentration range of 0.1040.00 ng/mL. The mean extraction recoveries of scopolamine butylbromide and propanolol from plasma were 69.00 and 80.76, respectively. Method validation parameters, such as specificity, linearity, precision, accuracy, and stability, were within the acceptable range. Moreover, when the proposed method was successfully applied to a pharmacokinetic study of healthy human volunteers, the results showed that the two scopolamine butylbromide formulations tested are not bioequivalent in rate and extent of absorption.

Development and validation of a UPLC‐ESI‐MS/MS method for the determination of N‐butylscopolamine in human plasma: Application to a bioequivalence study

“…Manfio et al proposed an LC-MS/MS method and prior liquid/ liquid extraction for dichloromethane, linear in the range of 0.1-40 ng/ml, for N-butylscopolamine determination in serum, the recovery in their study was 69% for N-butylscopolamine, lower than comparing to this present study. [12] The results of protein precipitation of the developed method, using acetonitrile as a precipitation agent, allowed mean recoveries of N-butylscopolamine (94%) and IS (84%) at the specified concentration levels, confirming the suitability of the method for the plasma samples (Table 4). For the extraction, different organic solvents and mixtures were also evaluated, including ethyl acetate, diethyl ether, dichloromethane, and hexane; however, the recovery was worse than achieved with acetonitrile.…”

“…[6,8] A pharmacokinetic study has been based on gas chromatography-tandem mass spectrometry (GC-MS/MS) and liquid chromatography-tandem mass spectrometry (LC-MS/ MS). [9][10][11][12] In the GC-MS/MS method, the scopolamine and the internal standard (IS) mexiletine were extracted from serum by using a single step liquid/liquid extraction; however, after that, a derivatization step was necessary. [9] In the method reported by Oertel et al, the scopolamine and atropine (IS) were extracted and cleaned up by using solid-phase extraction (SPE).…”

A fast and sensitive UHPLC–MS/MS method for the determination ofN‐butylscopolamine in human plasma: application in a bioequivalence study

“…Following oral administration, the maximum concentration (C max ) of N-butylscopolamine is between 0.25 and 1.5 h. However, no pharmacokinetic data about the single-dose administration of Buscopan ® (10 mg) in patients or healthy volunteers exist. Manfio, dos Santos, Favreto, Hoffman, andMertin (2009) andFavreto et al (2012) obtained C max values of 1.61 ± 1.22 ng/mL and 0.29 ± 0.23 ng/mL at 2:85 ± 1:35 h and 4:26 ± 1:48 h, respectively, after the administration of two tablets of Buscopan ® (10 mg) in 24 healthy volunteers. Prior to these studies, pharmacokinetics data were collected following the oral administration of 500 mg of this drug in clinical research conducted by the patent holder (Tytgat, 2007).…”

Simultaneous determination of atropine, scopolamine, and anisodamine fromHyoscyamus nigerL. in rat plasma by high-performance liquid chromatography with tandem mass spectrometry and its application to a pharmacokinetics study

“…So far, numerous analytical methods including TLC , CCC , GC , HPLC coupled with UV , fluorescence , or MS detection , CE , and gas‐liquid chromatography (GLC) with MS have been developed for the determination of these three alkaloids, most of which were developed for quantitative determination of them in plants, foods, and medical preparations, not in biological samples. Only a few procedures for the detection of the tropane alkaloids and their metabolites in biological matrices have been reported in the literature so far. The methods determining the three tropanes in biological samples are listed in Table .…”

“…The methods determining the three tropanes in biological samples are listed in Table . Some of them detected only one or two compounds , while some could simultaneously determine the three alkaloids, but using LLE or SPE sample preparation, which need a larger sample volume so that they are not suitable for the determination in rat plasma . Tian et al.…”