Analysis of glucocorticoids in DBS: profiles of dexamethasone, methylprednisolone and deflazacort after oral administration, correlation with plasmatic concentrations and possibilities to establish reporting levels to detect oral administration

Code: DBS20GC01RV

Increasing interest has been dedicated to DBS in sports drug testing, due to the advantages regarding sample collection and transportation.

Few data is published in the literature regarding analysis of GCs in DBS and concentrations of GCs after administrations of GCs, and it deserve to be studied.

The objective of the project will be to study the detection of oral administration of GCs in DBS. The research will be focused on the following specific objectives:

- To collect DBS samples in excretion studies of DEX, MP and DEF with administration of the compounds in one single oral dose. For DEX, multiple oral doses will be also studied.

- To develop and validate methods to detect DEX, MP and DEF metabolite in DBS.

- To measure concentrations of the compounds in the DBS samples collected after oral administration of GC.

- To evaluate correlation between DBS and plasma concentrations

Main findings

Dried Blood Spots (DBS) have increasing interest in sports drug testing due to easier sample collection compared to urine and blood, and advantages regarding sample transportation and storage. The objective of the project was to study the detection of oral administration of glucocorticoids (GCs) in DBS samples. The concentrations of dexamethasone (DEX), methylprednisolone (MP) and deflazacort (DEF) and respective metabolites were evaluated after single-dose oral treatments or multiple-dose oral treatments (for DEX). For that purpose, analytical methods for the quantitation of these GCs and metabolites, and cortisol (CORT) in DBS samples were developed and validated.

After single oral dose, GCs were detected in all DBS samples collected during the first 8 h post-administration, showing maximum concentrations in the samples collected 1 to 2 h post-administration, and decreasing over time. In the multiple-dose oral treatment, both DEX and 6βOH-DEX showed an increase after each dose followed by a decrease over time. For DEF, only the metabolites 21DES and 6βOH-21DES were detected. The suppression of CORT due to the administration of the GCs was observed in all studies.

The correlation between DBS and plasma was good and was established by calculating the DBS-to-plasma ratios for each compound. Based on the data of the study minimum reporting levels in DBS samples were proposed for DEX, MP and 21DES.

Dried Plasma Spots (DPS) were also collected in the administration studies and methods to quantify DEX, 21DES and CORT in these samples were also developed and validated. The methods showed higher variabilities compared to DBS analysis. They were applied to the analysis of DPS samples collected after the administration of DEX or DEF. The correlation between DPS and plasma samples was evaluated and no significant differences were obtained, in general, for the analytes evaluated.

Compared to plasma, both DPS and DBS present similar advantages regarding sample collection, stability, transportation and storage, however an important limitation is the larger volumes of blood required to collect DPS samples. As a general conclusion, DBS analysis is preferred to DPS for antidoping testing.