Determination of xylazine, xylazine-4-hydroxy, and 2,6-dimethylaniline in canine urine: veterinary and forensic applications in two suspected poisoning cases

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Arslan Z., Bulut C., Bilgiç B., Öztürk Y. E., Or M. E., Türkmen Z.

FORENSIC TOXICOLOGY, cilt.44, sa.3, ss.1-7, 2026 (SCI-Expanded, Scopus)

  • Yayın Türü: Makale / Tam Makale
  • Cilt numarası: 44 Sayı: 3
  • Basım Tarihi: 2026
  • Doi Numarası: 10.1007/s11419-026-00784-1
  • Dergi Adı: FORENSIC TOXICOLOGY
  • Derginin Tarandığı İndeksler: Health Research Premium Collection (ProQuest), Scopus, Science Citation Index Expanded (SCI-EXPANDED), EMBASE, MEDLINE
  • Sayfa Sayıları: ss.1-7
  • İstanbul Üniversitesi-Cerrahpaşa Adresli: Evet

Özet

Abstract

Purpose Xylazine, an α2-adrenergic agonist, is widely used in veterinary medicine for anesthesia, sedation, and analgesia.

However, non-therapeutic use, overdose, and deliberate administration may pose serious health risks to animals, necessitating

reliable detection methods for both clinical and forensic purposes.

Methods In this study, a liquid chromatography-high-resolution mass spectrometry (LC-HRMS) method was developed and

fully validated for the determination of xylazine and its metabolites in canine urine.

Results The method exhibited excellent selectivity and linearity over the range of 0.05–100 ng/mL, with coefficients of

determination (R²) ≥ 0.9989. Recovery values ranged from 70.3% to 93.4%, with matrix effect values between 78.6% and

95.6%, and process efficiency values ranging from 74.9% to 94.3%. Precision and accuracy values were within ± 15% at all

quality control levels, and the analytes were stable under all tested conditions (+ 8 °C for 3 days, + 4 °C and − 20 °C for 5

days). In two forensic cases, urine samples collected from canines presenting with unconsciousness contained xylazine and

its metabolites, while no other sedative or toxic agents were detected.

Conclusion The presence of both 2,6-dimethylaniline (DMA) and 4-hydroxyxylazine supports the necessity for including

metabolite analysis in exposure assessment. This study highlights the applicability of LC-HRMS for the detection of

xylazine-related compounds in canine urine and its potential use in veterinary and forensic toxicology contexts.

Highlights

● Validated LC-HRMS method for detecting xylazine, DMA, and 4-hydroxyxylazine in canine urine.

● Xylazine concentrations exceeded metabolite levels in both cases.

● DMA levels were higher than 4-hydroxyxylazine.

● The method showed low LOD/LOQ and acceptable validation performance.

● Applicability was confirmed in two forensic cases involving unconscious dogs.