Rapid and simple LC‐MS/MS screening of 64 novel psychoactive substances using dried blood spots

Abstract

The range of novel psychoactive substances (NPS) including phenethylamines, cathinones, piperazines, tryptamines, etc. is continuously growing. Therefore, fast and reliable screening methods for these compounds are essential and needed. The use of dried blood spots (DBS) for a fast straightforward approach helps to simplify and shorten sample preparation significantly. DBS were produced from 10 µl of whole blood and extracted offline with 500 µl methanol followed by evaporation and reconstitution in mobile phase. Reversed-phase chromatographic separation and mass spectrometric detection (RP-LC-MS/MS) was achieved within a run time of 10 min. The screening method was validated by evaluating the following parameters: limit of detection (LOD), matrix effect, selectivity and specificity, extraction efficiency, and short-term and long-term stability. Furthermore, the method was applied to authentic samples and results were compared with those obtained with a validated whole blood method used for routine analysis of NPS. LOD was between 1 and 10 ng/ml. No interference from matrix compounds was observed. The method was proven to be specific and selective for the analytes, although with limitations for 3-FMC/flephedrone and MDDMA/MDEA. Mean extraction efficiency was 84.6 %. All substances were stable in DBS for at least a week when cooled. Cooling was essential for the stability of cathinones. Prepared samples were stable for at least 3 days. Comparison to the validated whole blood method yielded similar results. DBS were shown to be useful in developing a rapid screening method for NPS with simplified sample preparation.

High-throughput screening for new psychoactive substances (NPS) in whole blood by DLLME extraction and UHPLC–MS/MS analysis

“…Despite the diffusion of NPS in the illicit market, to date few are the analytical methods dealing with the multianalyte screening of these compounds in biological fluids, probably due to the relative new occurrence of the phenomenon. Published methods are mainly LC-MS/MS applications and deal with NPS in different biological samples: oral fluid [24,41], hair [25,[42][43][44], urine [19,26,27,45,46], blood [28,47,48], serum [29,30]. The methods available in the literature generally deal with a single class of analytes, and are performed after extraction of samples by SPME, SPE, or liquid/liquid extraction.…”

“…Furthermore, also in case of a positive result, the identification of the actual substance that can cross-react with the immunoassay must be carried out by a chromatographic/mass spectrometric technique, able to distinguish among similar structures and isomers. To overcome this problem, some screening methods based on chromatography coupled with mass spectrometry have been proposed [19][20][21][22][23][24][25][26][27][28][29][30]. The use of sophisticated techniques such as LC-MS/MS or GC-MS requires a clean-up step before the instrument analysis, especially when it is performed on highly complex matrices like blood.…”

Applying Dried Blood Spot Sampling with LCMS Quantification in The Clinical Development Phase of Tasquinimod

“…It was demonstrated that tasquinimod in spiked blood was stable on DBS cards for at least 24 months when stored at room temperature, as well as in whole blood stored at room temperature for 24 h. Also, ISR of DBS samples was shown to fulfil the 2/3/20 acceptance criteria. Acceptable stability for various compounds on DBS has been reported [17,18] and it has even been reported that DBS can increase stability for compounds found to be unstable in plasma [4,19].…”

Hematocrit-Independent Quantitation of Stimulants in Dried Blood Spots: Pipet versus Microfluidic-Based Volumetric Sampling Coupled with Automated Flow-Through Desorption and Online Solid Phase Extraction-LC-MS/MS Bioanalysis

“…Previously, good stability was described for up to 2 weeks at both RT and 4°C for amphetamine, MDMA, and methamphetamine while mephedrone was only stable when the DBS card was stored cooled. 24 Storage for 6 months at 4°C resulted in 50−75% of the initial concentrations measured for amphetamine, MDMA, and methamphetamine versus values between 84% and 96% when stored at −20°C. 25 In this study, analyte stability was evaluated after 5, 15, 30, and 50 days of storage at five different conditions: in sealed plastic bags at RT and 4 and −20°C, in a manual vacuum box at RT, and under nitrogen flow at RT (Figure S-2).…”