Is Xylazine Just the Beginning? —Two More Veterinary Drugs Found in Illicit Drug Supply

In my previous post, I wrote a blog about the veterinary tranquilizer xylazine and the dangers it poses as a toxic adulterant in the illicit drug market (Why is Everyone Talking About Xylazine?). In the time since, two more veterinary drugs—phenylbutazone and medetomidine—have also been identified in illicit drug materials. While these substances have not yet proliferated the illicit drug supply as xylazine has, we could see them continue to increase in popularity in a similar manner. 

Phenylbutazone is a non-steroidal anti-inflammatory drug (NSAID) that has analgesic and anti-inflammatory properties commonly used for the short-term treatment of pain and fevers in animals¹. It was originally used in humans to treat rheumatoid arthritis and gout but is no longer approved for any human use after harmful side effects were discovered¹. Adverse effects of phenylbutazone use can range from dizziness, headaches, and nausea to gastrointestinal bleeding, liver and kidney damage, and blood disorders which can lead to death². Medetomidine is a drug also used for veterinary purposes, primarily as an anesthetic. Medetomidine is a racemic mixture of dexmedetomidine and levomedetomidine, with dexmedetomidine being the active component.³ The effects of medetomidine in humans are not well studied.

Figure 1: Phenylbutazone	Figure 2: Medetomidine

Phenylbutazone and medetomidine have been detected alongside a number of illicit drugs, including heroin, fentanyl, xylazine, acetyl fentanyl, para-fluorofentanyl, diphenhydramine, and other adulterants^2,3. Much like I discussed in my xylazine blog, incorporating these two compounds into laboratory testing scopes is going to be a growing need. In the xylazine blog, we looked at adding the compound to an already developed, high-throughput drugs of abuse assay as a method of testing for xylazine. With phenylbutazone and medetomidine, I wanted to develop a new, rapid LC-MS/MS method that included these two analytes and the other drugs they are commonly found with.

To start developing a new method, normally I would head into the lab and start experimenting with columns, mobile phases, and gradients until I had a working chromatography method. Instead of this, I decided to use Restek’s Pro EZLC Chromatogram Modeler (Pro EZLC Online Tools) to do all the hard work for me. Pro EZLC is a virtual liquid chromatography modeler that can be used to develop and optimize LC methods without any hands-on time in the lab. I selected my compound list, made some changes based on the column I was using, and optimized the chromatography gradient until I was satisfied with the separation. Pro EZLC produced the method shown below. 

Figure 3: LC-MS/MS Method Generated by Pro EZLC Chromatogram Modeler

Using the method EZLC supplied me with, I ran a sample containing all the analytes on an LC-MS/MS. The following chromatogram was generated.

Figure 4: Chromatogram Generated Using Pro EZLC Chromatogram Modeler Method on LC-MS/MS System

Table 1: Experimental Analyte Retention Times

Peak #	Analyte	TR (min)
1	Xylazine	0.94
2	Heroin	1.09
3	Medetomidine	1.29
4	Acetyl fentanyl	1.61
5	Diphenhydramine	1.72
6	para-Fluorofentanyl	1.87
7	Fentanyl	1.94
8	Phenylbutazone	3. 46

The experimental results match up very well with what EZLC predicted, with an average difference in retention time of 6.7 seconds between the observed and expected values. This method which analyzes for newly trending toxic adulterants and common drugs of abuse demonstrates how Pro EZLC can be used to develop methods in a fraction of the time it would usually require. Pro EZLC is an especially valuable tool when working with compounds like medetomidine and phenylbutazone, which may not be routinely tested for by many laboratories at this point. 

References

1. Lees, Peter, and Pierre-Louis Toutain. “Pharmacokinetics, Pharmacodynamics, Metabolism, Toxicology and Residues of Phenylbutazone in Humans and Horses.” Veterinary Journal, vol. 196, no. 3, Elsevier BV, June 2013, pp. 294–303. https://doi.org/10.1016/j.tvjl.2013.04.019.
2. Shuda, Sarah A., M. S., et al. “Phenylbutazone: A Toxic Adulterant Found in Illicit Street Drugs.” The Center for Forensic Science Research and Education, Mar. 2023.
3. “Announcement of an Emerging Compound: Medetomidine.” National Forensic Laboratory Information System, 2023.