Zopiclone behaved differently from most benzodiazepines in a six-matrix autopsy study: its vitreous-to-peripheral-blood ratio had a median close to one. That result supports vitreous as a potentially useful specimen for this drug. It does not make vitreous and blood interchangeable in an individual death investigation; the 16 paired ratios ranged from 0.13 to 3.7.
The study selected cases through blood findings
Øiestad and colleagues collected specimens during medicolegal autopsies from June 2013 to November 2016. Of 183 project cases, 109 had one or more benzodiazepines or z-hypnotics detected and were included. The 109 were not all zopiclone cases: 16 had zopiclone findings in peripheral blood and vitreous.
Routine peripheral-blood screening triggered analysis of the project matrices. This design did not establish the frequency with which vitreous would detect zopiclone when blood screening was negative. It also did not measure population prevalence or diagnostic sensitivity across every autopsy.
How were the specimens collected and handled?
Autopsy generally occurred within one week after death. Peripheral blood was obtained by transecting the iliac vein, allowing the initial flow to stop, and collecting femoral blood expressed by thigh and groin compression. Collection from both sides was often needed to obtain 40–50 mL. The procedure was designed to avoid collecting blood from the superior vessels.
Cardiac blood, pericardial fluid, psoas and lateral vastus muscle, and vitreous were also collected. After laboratory receipt, specimens were refrigerated at 4°C, typically for one to two weeks while routine screening was completed. Matrices from the same case were analyzed together as far as possible.
Most included bodies had no or slight putrefaction: 77 and 24 cases, respectively. Four had moderate putrefaction and four lacked that information. The study did not establish equivalent performance in severe decomposition.
What did the 16 zopiclone pairs show?
Peripheral-blood zopiclone had a median of 0.097 mg/L and range of 0.0038–8.1 mg/L. Vitreous had a median of 0.16 mg/L and range of 0.011–1.73 mg/L. The median of individual vitreous/blood ratios was 0.97, with a range of 0.13–3.7.
The ratio of the two group medians is a different calculation from the median of individual paired ratios. Consequently, 0.16 divided by 0.097 should not be substituted for the reported 0.97 paired-ratio median.
The vitreous–blood Spearman correlation was 0.90 and significant at the 0.01 level. Correlation indicates ordering across the available observations, not equality or a narrow prediction interval for a new case.
Why the finding should not be generalized to every sedative
For diazepam, the study reported a median vitreous/blood ratio of 0.084 in 14 pairs; for alprazolam it was 0.35 in 15 pairs. The authors considered zopiclone’s lower protein binding a relevant explanation for its different pattern. This was an interpretation of the observed drug-specific distributions, rather than proof that protein binding alone determined every ratio.
The authors also considered greater zopiclone stability in alternative matrices a possible explanation for some higher concentrations. The casework comparison was not a controlled stability time course and could not quantify how much storage changed each result.
Screening, confirmation and reporting limits
Routine screening used UHPLC–MS/MS, followed by separate confirmatory methods. The confirmation-method quantitation limit for zopiclone was 0.00029 mg/L. Values below quantitation limits were excluded from evaluation. The findings therefore concern reported quantifiable results, not every possible low-level detection.
All matrices used whole-blood calibration, a limitation acknowledged by the authors. They separately evaluated recovery, matrix effects, accuracy and precision in the alternative matrices. Limited vitreous volume also prevented some analyses for other drugs. This was not a guarantee that every available vitreous specimen would support every assay.
No antemortem concentrations were available to identify which postmortem matrix best represented the concentration during life. A positive zopiclone result did not establish impairment or causal contribution. For the broader specimen question, see vitreous versus blood for postmortem drug screening.
Primary source
Øiestad EL, Øiestad ÅML, Middelkoop G, Brochmann GW, Thaulow CH, Vindenes V. Comparative Study of Postmortem Concentrations of Benzodiazepines and Z-Hypnotics in Several Different Matrices. Journal of Analytical Toxicology. 2023;47:287–298. doi:10.1093/jat/bkac106.
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