LC-MS/MS method for quantification of baclofen in hair: A useful tool to assess compliance in alcohol dependent patients?

To evaluate adherence to treatment, we developed and validated a novel liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for baclofen quantification in hair.Twenty mg was washed twice with dichloromethane, incubated in phosphate buffer (pH 5) for 10 minutes at 95°C, then extracted by liquid-liquid extraction in alkaline condition. Baclofen-d4 was used as the internal standard. This method was applied to assess compliance in4 treated alcohol-dependent patients (3 dead and one living). Blood quantification of baclofen and ethanol were performed in the 4 cases. Hair ethylglucuronide (ethanol metabolite, EtG) measurement (2x3 cm) was associated in 1 patient. Baclofen quantification in hair was validated over the range 10-5000 pg/mg. The accuracy was within 96.0%-110.9% and the precision was less than 9.3%. Baclofen segmental (3x2cm) hair concentrations found in the living patient were 4420, 4260, and 4380 pg/mg, reflecting a regular exposure over the last 6 months and suggesting patient compliance. However, the high EtG level found in this patient in the analyzed segments (225 pg/mg and 215 pg/mg) showed excessive alcohol consumption during the same period, suggesting therapeutic failure. In the 3 deceased patients, the non-segmental analysis of hair showed baclofen concentrations of 15, 545, and 2475 pg/mg. The low concentrations in the 2 first cases are compatible either with a poor compliance or to a beginning of a treatment. This is the first measurement of baclofen in hair of alcohol dependent patients. It could be used as a monitoring biomarker to assess patient's compliance.

Validated Method for the Quantification of Baclofen in Human Plasma Using Solid-Phase Extraction and Liquid Chromatography-Tandem Mass Spectrometry.

A highly sensitive and fully validated method was developed for the quantification of baclofen in human plasma. After adjusting the pH of the plasma samples using a phosphate buffer solution (pH 4), baclofen was purified using mixed mode (C8/cation exchange) solid-phase extraction (SPE) cartridges. Endogenous water-soluble compounds and lipids were removed from the cartridges before the samples were eluted and concentrated. The samples were analyzed using triple-quadrupole liquid chromatography-tandem mass spectrometry (LC-MS-MS) with triggered dynamic multiple reaction monitoring mode for simultaneous quantification and confirmation. The assay was linear from 25 to 1,000 ng/mL (r(2) > 0.999; n = 6). Intraday (n = 6) and interday (n = 15) imprecisions (% relative standard deviation) were <5%, and the average recovery was 30%. The limit of detection of the method was 5 ng/mL, and the limit of quantification was 25 ng/mL. Plasma samples from healthy male volunteers (n = 9, median age: 22) given two single oral doses of baclofen (10 and 60 mg) on nonconsecutive days were analyzed to demonstrate method applicability.

A small structural change resulting in improved properties for product development.

AZD9343 is a water-soluble gamma amino butyric acid (GABAB) agonist intended for symptomatic relief in gastroesophageal reflux disease (GERD) patients. The compound has good chemical stability in aqueous solutions, as well as in the solid state. Only one crystal modification has been observed to date. This polymorph is slightly hygroscopic (1.5% water uptake at 80% relative humidity (RH)), which is an improvement compared to the structurally similar agonist lesogaberan (AZD3355) which liquefies at 65% RH. Since the substance is very polar and lacks a UV chromophore, conventional separation and detection techniques cannot be used to characterize the substance and its impurities. The analytical techniques are described, focusing on the capillary electrophoresis method with indirect UV detection for assay and purity, the liquid chromatographic method for enantiomeric separation with derivatization with UV chromophore and three complementary nuclear magnetic resonance (NMR) approaches ((31)P-NMR, (13)C-NMR and (1)H-NMR) for impurities. For oral solutions, it was important to select the right concentration of phosphate buffer for the specific drug concentration and routinely use small additions of EDTA. I.V. solutions containing physiological saline as tonicity modifier could not be stored frozen at -20 °C. Properties of AZD9343 will be discussed in light of experiences from the structurally similar lesogaberan and (2R)-(3-amino-2-fluoropropyl)sulphinic acid (AFPSiA).

Evaluation of two modified quick, easy, cheap, effective, rugged and safe (QuEChERS) sample preparation methods for the analysis of baclofen and gabapentin in feeds by liquid chromatography tandem mass spectrometry.

This paper presents a new analytical method for the simultaneous determination of baclofen and gabapentin in feeds based on two modified quick, easy, cheap, effective, rugged and safe (QuEChERS) sample preparation methods and liquid chromatography tandem mass spectrometry (LC-MS/MS). For the two modified QuEChERS methods, samples were first extracted with acidified acetonitrile (5.0% acetic acid, v/v) without using acetonitrile salting-out extraction. Then, the first modified QuEChERS method was established according to the original QuEChERS cleanup procedure. For the second modified QuEChERS method, the extract was evaporated to dryness and reconstituted in acetonitrile. Subsequently, the analytes in the reconstituted solution were retained by primary secondary amine (PSA) and released from PSA with 1.0% formic acid in methanol. Finally, the eluate was evaporated and dissolved in 0.1% formic acid solution/methanol (v/v, 80:20). All of the samples were analyzed by LC-MS/MS on a Waters Acquity BEH C18 column with 0.1% formic acid in water/methanol as the mobile phase with gradient elution. The matrix effect, recovery, and repeatability, within laboratory reproducibility, and the LODs and LOQs of the two modified QuEChERS sample preparation methods were investigated and compared. Comparative results showed that the second method was obviously superior to the first method.