Methodological considerations in the development of HPLC-MS methods for the analysis of rodent plasma for metabonomic studies.

A study of the factors involved in obtaining valid global metabolite profiles from the HPLC-MS of rat or mouse plasma for the purposes of metabonomic analysis has been undertaken. Plasma proteins were precipitated with three volumes of either methanol or acetonitrile. Chromatographic separations were performed on a C18-bonded stationary phase using 3.5 and 5 mum particles packed into 2.1 and 4.6 mm i.d. formats, respectively, and on a C8 phase using 3.5 mum particles and a 2.1 mm i.d. column. Three reversed-phase gradient solvent systems, based on acidified water-acetonitrile, acidified water-methanol and acidified water-methanol-acetonitrile mixtures, were investigated. The column eluent was analysed with both positive and negative electrospray ionisation using a quadrupole-linear ion trap mass spectrometer. These studies revealed that while accurate classification of sample type can be made, there are a number of methodological problems associated with the analysis of plasma with respect to factors such as repeatability and column longevity. In particular, special care has to be taken to ensure that the analytical system is properly "conditioned" by the repeated injection of matrix samples. The use of biological quality control (QC) samples provided an important means of monitoring method performance. Finally, the source of the plasma (Zucker wild-type or (fa/fa) rat or mouse tumour model) also appeared to have an effect on the repeatability of the methodology.

Going the extra mile: beyond health teaching to political involvement.

TOPIC: Addressing community health problems through political involvement. PURPOSE AND SOURCES OF INFORMATION: This article describes how a group of RN-BSN students completing an assigned community-assessment and health-teaching project in a small, rural, southern county exceeded course requirements to address a significant community health problem. Specifically, after documenting a high rate of dental caries among local children and consulting with state officials and other experts, these students involved themselves in local politics in an effort to persuade county officials to implement community water fluoridation. CONCLUSIONS: These RN-BSN students successfully demonstrated their ability to move beyond a focus on individuals to embrace the concept of community as client. In the process, they honed their skills in advocacy, communication, and political involvement, and achieved all of their BSN program's objectives.

Instrumental and experimental effects in LC-MS-based metabolomics.

The experimental complexity of a metabolomics study can cause uncontrolled variance that is not related to the biological effect being studied and may distort or obscure the data analysis. While some sources can be controlled with good experimental techniques and careful sample handling, others are inherent in the analytical technique used and cannot easily be avoided. We discuss the sources and appearance of some of these artifacts and show ways in which they can be detected using visualization and statistical tools, allowing appropriate treatment prior to multivariate analysis (MVA).

Investigation of analytical variation in metabonomic analysis using liquid chromatography/mass spectrometry.

Sources of analytical variation in high-performance liquid chromatography/mass spectrometry (HPLC/MS), such as changes in retention, mass accuracy or signal intensity, have been investigated to assess their importance as a variable in the metabonomic analysis of human urine. In this study chromatographic retention and mass accuracy were found to be quite reproducible with the most significant source of analytical variation in the data sets obtained being the result of changes in detector response. Depending on the signal intensity threshold used to define the presence of a peak a sample component could be present in some replicate injections and absent in others within the same run. The implementation of a more sophisticated data software analysis package was found to greatly reduce the impact of detector response variability resulting in improved data analysis.

Fast mass spectrometry-based methodologies for pharmaceutical analyses.

Historically, most bioanalytical methods for drug analysis in pharmaceutical industry were developed using HPLC coupled with UV or fluorescence detection. However, there is a trend toward interfacing separation technologies with more sensitive tandem mass spectrometry (MS/MS)-based systems. MS/MS detection offers complete resolution of the parent compounds from their first pass metabolites to avoid extra efforts for separation and sample clean-up procedures resulting in shorter run times. With the increasing demand for ever faster screening, there is a continuing demand for bioanalytical methods possessing higher sample throughput for both in vitro and in vivo drug metabolism and pharmacokinetic evaluations to accelerate the discovery process. This review focuses on the current approaches for fast MS-based assays (cycle-time less than 5 min) of pharmaceuticals and their metabolites that have been reported in the peer-reviewed publications.