Comprehensive method to detect nitazene analogues and xylazine in wastewater
Open accessAbstract
A class of novel synthetic opioids, nitazenes, have infiltrated the illicit drug market worldwide. Their high potency leading to overdose and fatality is a public health concern. Xylazine is a non-opiate sedative typically used for veterinary applications. The drug has recently appeared as an adulterant in the informal drug market, particularly within opioid drugs such as fentanyl and heroin. Determining the extent of use of these drugs is challenging due to low levels consumed. Wastewater analysis has been applied to monitor novel psychoactive substance use at a population level. This study presents the validation of a sensitive analytical method for the detection and quantification of trace levels of nitazenes and xylazine in wastewater. A solid phase extraction method was employed, yielding a concentration factor of 1000-fold. The method was applied to wastewater samples collected in Australia. Five different nitazenes were found near the limit of detection and quantitation levels, with detection frequencies ranging from 3 to 6% of all samples analysed. Xylazine was detected at a higher detection frequency (26% of all samples analysed). The method presented in this study could be used as an early warning system to track the use of xylazine and nitazenes internationally.
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Older gas chromatography-mass spectrometry methods frequently lack the necessary sensitivity for nitazenes, often missing them in casework.
“Furthermore, older gas chromatography–mass spectrometry methods also frequently lack the necessary sensitivity for nitazenes, often missing them in casework [50,51,52].”
From Nitazenes: The Emergence of a Potent Synthetic Opioid Threat (2025) · cites this paper as [50]
The Results and discussion section says gas chromatography may lack the sensitivity or selectivity needed to detect nitazenes in wastewater. It does not describe older GC–MS methods, establish that this happens frequently, or report nitazenes being missed in forensic casework. (LLM, conf=0.88)
Experimental — accuracy not yet measuredForensic laboratories are adapting to analyse nitazenes in alternative matrices such as hair for retrospective exposure assessment, dried blood spots for micro-sampling in clinical settings, and wastewater for early warning and epidemiological monitoring.
“Forensic laboratories are also adapting to analyse nitazenes in alternative matrices such as hair [62] for retrospective exposure assessment, dried blood spots [54,63] for micro-sampling in clinical settings, and wastewater for early warning and epidemiological monitoring [50,64].”
From Nitazenes: The Emergence of a Potent Synthetic Opioid Threat (2025) · cites this paper as [50]
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- Authors
- Keller, E. L., Peake, B., Simpson, B. S., White, J. M., & Gerber, C.
- Affiliations
- University of South Australia (AU)
- Corresponding author
- Gerber, Cobus
- Venue
- Environmental Science and Pollution Research
- Published
- 2025-04-22
- Volume
- 32
- Issue
- 57
- Pages
- 30863-30870
- ISSN
- 0944-1344 (ISSN-L)
- 1614-7499
- PMC
- PMC12811310
- OpenAlex
- W4409635835
- Cited by
- 13
- References
- 20
- Citations by year
- 2026: 7 · 2025: 6
- FWCI
- 6.61
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Locations
- Environmental Science and Pollution Research (publishedVersion, cc-by)
- PubMed (publishedVersion)
- PubMed Central (publishedVersion, cc-by)
- Adelaide Research & Scholarship (AR&S) (University of Adelaide) (submittedVersion, cc-by)
- Funders
- Australian Criminal Intelligence Commission
- SA Health
- University of South Australia
- Topics
- Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Toxicology › Forensic Toxicology and Drug Analysis
- Physical Sciences › Environmental Science › Pollution › Pharmaceutical and Antibiotic Environmental Impacts
- Physical Sciences › Chemistry › Analytical Chemistry › Analytical chemistry methods development
- Keywords
- nitazenes, xylazine, synthetic opioids, wastewater-based epidemiology, new psychoactive substances, drug adulteration, solid-phase extraction, trace detection, early warning systems
- MeSH terms
- Environmental Monitoring, Water Pollutants, Chemical, Xylazine, Solid Phase Extraction, Wastewater
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