A rapid, automated approach to optimisation of multiple reaction monitoring conditions for quantitative bioanalytical mass spectrometry.

An improvement to the procedure for the rapid optimisation of mass spectrometry (PROMS), for the development of multiple reaction methods (MRM) for quantitative bioanalytical liquid chromatography/tandem mass spectrometry (LC/MS/MS), is presented. PROMS is an automated protocol that uses flow-injection analysis (FIA) and AppleScripts to create methods and acquire the data for optimisation. The protocol determines the optimum orifice potential, the MRM conditions for each compound, and finally creates the MRM methods needed for sample analysis. The sensitivities of the MRM methods created by PROMS approach those created manually. MRM method development using PROMS currently takes less than three minutes per compound compared to at least fifteen minutes manually. To further enhance throughput, approaches to MRM optimisation using one injection per compound, two injections per pool of five compounds and one injection per pool of five compounds have been investigated. No significant difference in the optimised instrumental parameters for MRM methods were found between the original PROMS approach and these new methods, which are up to ten times faster. The time taken for an AppleScript to determine the optimum conditions and build the MRM methods is the same with all approaches.

Quantitative analysis of pharmaceuticals in biological fluids using high-performance liquid chromatography coupled to mass spectrometry: a review.

1. The development of bio-analysis of drug molecules over the last 10 years is reviewed, focusing on advances in sample preparation, liquid chromatography and detection. 2. Developments have led to improvements in detection sensitivity, enhancements in specificity and increased capacity. 3. Emerging technologies such as monolithic column chromatography and miniaturized chip-based systems are discussed.

Absorption and disposition of ranitidine hydrochloride in rat and dog.

1. The pharmacokinetics of ranitidine were studied in the male beagle dog at a dose level of 50 mg (intravenous) or 5 mg/kg (oral). 2. After intravenous administration, Clp was moderate (10.4 ml/min/kg) with Clr accounting for approximately 30% of total clearance. Vdarea was 3.5 l/kg, resulting in a t1/2 of approximately 4 h. 3. After oral administration, F was good (73%) with peak plasma concentrations of ranitidine (2 micrograms/ml) achieved within 0.5-1 h hour after dosing. t1/2 (4.1 h) was similar to that observed after intravenous administration. 4. The absorption, metabolism and excretion of [14C]-ranitidine were studied in rat and dog after oral administration at a dose level of 50 mg/kg. 5. Urinary excretion was the major elimination pathway for radioactive drug-related material in both species (62-75% of the dose). Unchanged ranitidine was the major radioactive component in both rat and dog urine (0-24 h), accounting for approximately 40% of the dose in each case. 6. In dog, ranitidine undergoes N-oxidation (approximately 30% of dose) whereas in rat, N-oxidation, S-oxidation, N-demethylation and oxidative deamination are all evident, with each metabolite accounting for < 6% of the dose. 7. Two previously unreported metabolites of ranitidine were identified in rat urine using newly developed hplc and lc/ms methods. These metabolites result from single and di-N-demethylation of ranitidine and accounted for 4 and 1% of the dose respectively.

Thermospray liquid chromatography-mass spectrometry for the characterisation of sulphate ester conjugates.

Formation of polar conjugates is a well documented metabolic pathway for xenobiotics containing phenolic hydroxyl groups. This paper describes the analysis of two sulphate ester conjugates by fast atom bombardment mass spectrometry and thermospray liquid chromatography-mass spectrometry. Thermospray liquid chromatography-mass spectrometry proved the more successful technique for obtaining the molecular weight of the intact conjugate, but only by removal of the buffer from the high-performance liquid chromatography eluent.