Fluispotter, a novel automated and wearable device for accurate volume serial dried blood spot sampling.

Aim: A novel automated serial dried blood spot (DBS) sampler, 'Fluispotter', was tested for its sampling performance. Materials & methods: An LC-MS/MS method was developed for the analysis of cortisol in DBS samples serially spotted by Fluispotter. The cortisol concentrations in 148 paired DBS and plasma samples were compared across a hematocrit (HCT) range of 22-55%. Results: The interassay accuracy and precision were <10%. Overall assay bias was negligible across the HCTs tested when analyzing the whole-spot DBS samples. The accuracy and precision of the blood volume in 10 µl DBS samples spotted by Fluispotters and micropipettes were within 3%. Deming regression and Bland-Altman analysis showed a good agreement of DBS-predicted and measured plasma cortisol. Conclusion: The Fluispotter performed serial sampling with high accuracy and precision of the sample blood volume.

Feedback from the European Bioanalysis Forum liquid microsampling consortium: capillary liquid microsampling and assessment of homogeneity of the resultant samples.

Following the completion of a detailed experimental protocol into the potential inhomogeneity of capillary liquid microsamples, which was performed at seven European Bioanalysis Forum member companies, the summary and conclusion on the data are reported here. It has been demonstrated that it is possible to generate homogeneous samples using these microsampling techniques; that the resultant microsamples can be accurate and precise and that capillary liquid microsampling data can be consistent with conventional larger volume plasma samples. However, the data contain some variability which is contributed to by the different range of experiences that each investigating site had with these techniques. Therefore, knowledge of the compounds, well-designed experiments and experience with these techniques are essential for the delivery of high quality data.

Feedback from the European Bioanalysis Forum liquid microsampling consortium: microsampling: assessing accuracy and precision of handheld pipettes and capillaries.

Aim: Microsampling in preclinical pharmacokinetics (PK) studies is currently widely adopted across the pharmaceutical industry. Materials & methods: The European Bioanalysis Forum liquid microsampling consortium member companies assessed the accuracy and precision of handheld pipettes and microcapillaries at volumes of less than 10 µl. The following key factors on pipetting performance were also evaluated: Pipette type (positive displacement, air displacement and microcapillary), experience of user and the liquid type. Water was selected as a best-case scenario for accuracy and precision determination and blood plasma as a 'real world' bioanalysis sample type. Conclusion: Accuracy and precision on the pipetted volume decreased at lower volumes and experienced laboratory technicians performed better compared with the infrequent users. With respect to the pipetting devices used, microcapillaries showed better or equivalent accuracy and precision compared with handheld pipettes across the volume range 1-8 µl independent of the matrix used.

Quantitative determination of the antidepressant vortioxetine and its major human metabolite in plasma.

BACKGROUND: Vortioxetine is a novel antidepressant that has been developed in a joint partnership between H. Lundbeck A/S and the Takeda Pharmaceutical Company, Ltd. RESULTS: A number of bioanalytical methods have been developed in order to support the nonclinical and clinical development of the drug. Method performance, long-term stability, urine analysis, unspecific binding and metabolites analysis are presented and discussed. CONCLUSION: Two different method applications for the quantification of vortioxetine and its major human metabolite in human plasma, an isocratic cation exchange HPLC-MS/MS method utilizing C8-SPE sample extracts and a reversed-phase UPLC-MS/MS method with gradient elution of protein precipitated sample extracts, have been validated according to current regulatory standards and applied in support to a large number of nonclinical as well as clinical studies.

Biological conversion of aripiprazole lauroxil - An N-acyloxymethyl aripiprazole prodrug.

N-acyloxyalkylation of NH-acidic compounds can be a prodrug approach for e.g. tertiary or some N-heterocyclic amines and secondary amides and have the potential to modify the properties of the parent drug for specific uses, for example its physicochemical, pharmacokinetic or biopharmaceutical properties. Aripiprazole lauroxil was prepared as a model compound for such prodrugs and its bioconversion was investigated both in vitro and in vivo. Theoretically, N-acyloxyalkyl derivates of NH-acid compounds undergo a two-step bioconversion into the parent NH-acidic drug through an N-hydroxyalkyl intermediate. However, to our knowledge no published studies have investigated the formation of an intermediate in vivo. In the present study, it was demonstrated that the assumed N-hydroxymethyl intermediate was readily observed both in vitro and in vivo. In vivo, the observed plasma concentration of the intermediate was at the same level as the drug (aripiprazole). When prodrug intermediates are formed, it is important to make a proper pharmacological, pharmacokinetic and toxicological evaluation of the intermediates to ensure patient safety; however, several challenges were identified when testing an N-acyloxyalkyl prodrug. These included the development of a suitable bioanalytical method, the accurate prediction of prodrug bioconversion and thereby the related pharmacokinetics in humans and the toxicological potential of the intermediate.

IS addition in bioanalysis of DBS: results from the EBF DBS-microsampling consortium.

BACKGROUND: In the framework of wider exploration of the application of dried blood spots (DBS) in bioanalysis, by the DBS consortium of the European Bioanalytical Forum, one team of five laboratories investigated the merits of the various ways of IS addition prior to LC-MS/MS analysis. A set of 22 pharmaceutical compounds with log P in the range of 0-10 was selected for this purpose. Assessments were made of precision, recovery, and of the effects of prolonged storage. RESULTS: Assay precision was not significantly different for 3 month-aged samples as compared with 'fresh' samples stored for 7-22 days. Extraction recovery from 3 month-aged spots decreased for some of the analytes; the most widely employed addition of IS in the extraction solvent does not compensate for recovery in such cases. CONCLUSION: From the overall results, it is clear that there is no 'one size fits all' approach to IS addition in DBS bioanalysis.

Evaluation of an isochronic study design for long-term frozen stability investigation of drugs in biological matrices.

Long-term stability is a basic parameter in bioanalytical method validation; however, no criteria for conducting long-term stability studies are specified in current guidelines. We present an evaluation of a modified statistical approach applied to a study design utilizing an isochronic analysis (collection of samples to be analyzed at one time point) to determine the long-term stability and, further, a comparison with the most widely used continuous design. The presented approach has been used in regulated bioanalysis at Lundbeck for the past 7 years and has, in this period, been applied to 121 studies; all providing conclusive data. The isochronic approach eliminates day-to-day variation, reduces labor and adds to the flexibility in the laboratory. The statistical evaluation used is based on the relative difference between baseline samples and stability test samples as well as 90% confidence intervals for the mean concentration for each of the stability test points.

In vivo evaluation of tablets and capsules containing spray-dried o/w-emulsions for oral delivery of poorly soluble drugs.

It is recognised that poorly soluble drugs may show an increased oral bioavailability when incorporated in o/w-emulsions. Encapsulating the emulsion lipid droplets in hydroxypropyl methylcellulose (HPMC) by spray drying has been demonstrated to preserve an improved bioavailability releasing lipid droplets from the powder in vivo. However, the spray-dried powder is cohesive and bulky requiring additional processing to improve handling. This was resolved in previous work where a directly compressible dry emulsion formulation was described. The purpose of the present study is to investigate the oral bioavailability resulting from administration of a directly compressible dry emulsion as a tablet and compare it with a HPMC dry emulsion powder and a simple lipid solution. Four female Beagle dogs received a single dose of each formulation containing the same amount of medium-chain triglycerides (MCT) and a model drug, Lu 28-179. Cyclodextrin solutions administered orally and intravenously were used as references. The absolute bioavailability decreased in the order cyclodextrin solution (0.14), HPMC dry emulsion (0.11), technically improved dry emulsion (0.10) and MCT solution (0.06). The directly compressible dry emulsion tablets were concluded to be comparable to a HPMC dry emulsion powder in terms of bioavailability. The lack of statistically significant differences relative to a MCT solution was ascribed to a low and variable absolute oral bioavailability of the model drug.