Analytical characterization and pharmacological evaluation of the new psychoactive substance 4‐fluoromethylphenidate (4F‐MPH) and differentiation between the (±)‐threo and (±)‐erythro diastereomers

Abstract

Misuse of (±)-threo-methylphenidate (methyl-2-phenyl-2-(piperidin-2-yl)acetate; Ritalin®, MPH) has long been acknowledged, but the appearance of MPH analogs in the form of ‘research chemicals’ has only emerged in more recent years. 4-Fluoromethylphenidate (4F-MPH) is one of these recent examples and this study presents the identification and analytical characterization of two powdered 4F-MPH products that were obtained from an online vendor in 2015. Interestingly, the products appeared to have originated from two distinct batches given that one product consisted of (±)-threo-4F-MPH isomers whereas the second sample consisted of a mixture of (±)-threo and (±)-erythro 4F-MPH. Monoamine transporter studies using rat brain synaptosomes revealed that the biological activity of the 4F-MPH mixture resided with the (±)-threo- and not the (±)-erythro isomers based on higher potencies determined for blockage of dopamine uptake (IC50 4F-MPHmixture = 66 nM vs. IC50 (±)-threo = 61 nM vs. IC50 (±)-erythro = 8,528 nM) and norepinephrine uptake (IC50 4F-MPHmixture = 45 nM vs. (±)-threo = 31 nM vs. IC50 (±)-erythro = 3,779 nM). In comparison, MPH was three times less potent than (±)-threo-4F-MPH at the dopamine transporter (IC50 = 131 nM) and around 2.5-times less potent at the norepinephrine transporter (IC50 = 83 nM). Both substances were catecholamine selective with IC50 values of 8,805 nM and >10,000 nM for (±)-threo-4F-MPH and MPH at the serotonin transporter. These findings suggest that the psychostimulant properties of (±)-threo-4F-MPH might be more potent in humans than MPH.

Analysis of six ‘neuro‐enhancing’ phenidate analogs

“…[11,13] Since methylphenidate is a widely regulated medicine, structurally diverse and often more potent NPS of the phenidate class have entered the drug market. [14,15] Molecular structures and the respective activities at DAT have been reported for many of these compounds in the scientific literature, making the targeted synthesis of potent phenidate analogues for the drug market likely. [16] Phenidates were recently seized by German authorities, such as 4methylmethylphenidate (2), 3,4-dichloromethylphenidate (3), ethylphenidate (4), 3,4-dichloroethylphenidate (5), and N-benzylethylphenidate (7), whereas ethylnaphthidate (6) was obtained from an Internet retailer ( Figure 2).…”

“…Numbers in parentheses correspond to the values for the known parent methyl-ester compounds. [15,25] effect rather than its chemical structure. [27] Procedure (1) can be difficult to implement and pharmaceutical legislation in, for example, Germany is not applicable to 'substances that […] are only consumed to intoxicate while being detrimental to health' according to a recent ruling by the European Court of Justice (ECJ).…”

Addressing the challenges in forensic drug chemistry

“…Interestingly, it was revealed that one powdered product consisted of a diastereomeric mixture represented by racemic (±)‐ threo and (±)‐ erythro 4F‐MPH whereas the other product consisted of (±)‐ threo ‐4F‐MPH only, thus, reflecting distinct batches prepared during synthesis and incomplete product clean‐up. Furthermore, after chromatographic separation, both forms were subjected to in vitro pharmacological characterization using rat brain synaptosomes, which revealed that the biological activity, i.e., catecholamine selective blockade of monoamine reuptake, was associated with (±)‐ threo form …”

Challenges in the identification process of phenidate analogues in LC‐ESI‐MS/MS analysis: Information enhancement by derivatization with isobutyl chloroformate

“…MPH is rapidly absorbed and hydrolyzed by carboxylesterase 1 liver enzymes to the inactive metabolite ritalinic acid (RA), which is excreted primarily in urine . Because MPH is a widely regulated medicine, several closely related unregulated substances, which are often more potent, have been launched on the NPS market . Multiple cases of misuse of phenidate analogues have been observed among recreational drug users, presumably due to their psychostimulant properties.…”

“…Several scientific articles have been published in recent years regarding analytical characterization of phenidate analogues by nuclear magnetic resonance (NMR), infrared (IR) spectroscopy, gas chromatography‐mass spectrometry (GC‐MS), and liquid chromatography‐electrospray ionization‐tandem mass spectrometry (LC‐ESI‐MS/MS) . The confirmation of such chemical identities by MS techniques represents a challenging task because these analogues present information‐poor mass spectra in LC‐ESI‐MS/MS and in GC‐EI‐MS analysis, with a fragment/product ion representing mainly or only the secondary amine‐containing residue.…”