Identifying the unknown

16th January 2017

Posted By Paul Boughton

Toxicology labs are seeing a host of new designer drugs appearing in their casework
The Selectra PFPP HPLC column plays a key role in the fight to identify new designer drugs
UCT’s Clean Screen XCEL I column can be used for the extraction of designer benzodiazepines and novel opiate-like compounds

Danielle Mackowsky reveals the in-lab battle to analyse emerging designer drugs

Toxicology labs are fighting to remain on top of the emerging designer drugs that are appearing in their casework.

Constantly evolving compounds, meant to mimic a variety of traditional drug classes such as cannabinoids, opiates and benzodiazepines, are found on the streets in unknown preparations and then subsequently on the lab bench.

On account of this, the accurate and rapid extraction methods for these drug classes in a wide variety of biological matrices are critical. These extraction methods need to not only be specific for the current generations of designer drugs, but also amendable to future compounds that emerge as drugs of abuse. The use of UCT’s solid phase extraction (SPE) methodologies meet the above criteria, while also providing scientists with clean, concentrated samples that can be analysed on either GC-MS or LC-MS/MS.

Synthetic cannabinoids first appeared as a legal high in the USA in the early 2000s. Advertised as being derived from naturally occurring herbal products, these compounds were boasted to be a safe and effective way to get high without testing positive for traditional drugs of abuse. In reality, these ‘spice’ sachets often contained plant material sprayed with chemicals that were once researched in legitimate academic settings for their endocannabinoid receptor properties. Early generations of these synthetic cannabinoids were named after John W. Huffman (JWH), the primary investigator who pioneered the synthesis of many of these compounds in the 1980s.

A polymeric-based sorbent

For analysis of the abovementioned synthetic cannabinoids, UCT has employed the use of its Styre Screen HLD polymeric-based sorbent. This approach allows for toxicologist to skip the preconditioning steps found in a traditional solid phase extraction protocol and also saves on overall solvent usage. The neutral chemical nature of the sorbent itself proves very effective for retention of both parent compounds and subsequent metabolites. Following sample loading, the column is washed with 100 mM acetate buffer (pH 5) and a 100 mM acetate buffer/methanol solution (75/25 v/v) to ensure unwanted matrix components are removed. Several combinations of buffer/ methanol washes were evaluated to ensure optimal cleanliness and analyte recovery ranging from 75% buffer/25% methanol to 50% buffer/50% methanol.

Although adequate recovery can be achieved for most synthetic cannabinoids under all conditions, it was noted that going above 25% methanol caused the metabolites of JWH compounds to be lost in the wash. For analyte elution, 100% ethyl acetate was determined to be the most effective solvent, selectively isolating analytes of interest while leaving unwanted matrix bound the sorbent.

In addition to synthetic cannabinoids, opiates have become a dominant player in the drug scene, specifically on account of the recent heroin epidemic plaguing the USA. Although strides have been made to not only prevent the over-prescribing of legal opiate medications, but also curb doctor-shopping amongst patients, they have been offset by heroin’s re-emergence as a popular drug of abuse. Cheaper than prescription medications obtained illegally in addition to providing a better euphoria to users, heroin continues to contribute to the surge in cases of opiate overdose in emergency rooms. While the treatment of a classically presenting heroin overdose and subsequent toxicological analysis has become routine, more recently many emerging novel analgesics can be found in addition to or in place of heroin itself, adding substantial complexity to the testing. Fentanyl, first identified in overdose cases in the 1980s, is now joined by compounds such as W-18, W-15, furanyl fentanyl, carfentanil and U-47700 in seized preparations and/or toxicology screens and confirmations.

Designer benzodiazepines have also created a considerable challenge for toxicologists, specifically in European countries. Many of the compounds being found in patient samples were once investigated to be used for pharmaceutical purposes. Black- market chemists have started to synthesise these drugs and often attempt to disguise them in pill form. These tablets can then either be wrongly marketed to customers as prescription-grade benzodiazepines or also be laced with other drugs of abuse.

For the extraction of both designer benzodiazepines and novel opiate-like compounds, UCT’s Clean Screen XCEL I column can be put to use. This sorbent also eliminates the column preconditioning step while providing laboratories with a sample preparation solution for a wide variety of forensic matrices. Each method requires only two wash steps prior to elution: D.I. H2O and 100 mM acetic acid for novel analgesics and acetate buffer (pH 5) and methylene chloride for designer benzodiazepines. Previous extraction optimisation for traditional benzodiazepines identified basified ethyl acetate as the elution solvent of choice for this class of compounds; thus, it was utilised in this extraction protocol. On the contrary, the novel analgesics studied did not exhibit the expected recovery rates as their traditional opiate counterparts when a basified solution of methylene chloride and isopropyl alcohol was used as an extraction solvent. Methanol with 2% ammonium hydroxide was ultimately selected due to its ability to elute both classic and emerging opiate-like drugs of abuse without compromising the recoveries for either category.

Smarter extraction

The aforementioned SPE methods allow for toxicologists to extract and identify designer and novel substances without compromising the recoveries of traditional drugs of abuse within the same drug classes. Although there is no way to predict what modifications will be made to these base structures and what will subsequently be abused in the drug market, it is still possible for forensic labs to be as prepared as possible for both extraction and instrumental methodologies by implementing universal SPE approaches as described above.

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Danielle Mackowsky is with UCT





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