Background and aim: We have received special request for nitrous oxide analysis on traffic case samples since the beginning of 2020. The gas can cause dizziness, disorientation and fainting spells that might affect the ability to drive, but it is not yet covered in the Danish traffic regulations regarding driving under the influence of drugs. A surge of national and international traffic incidents have heightened attention and the need for analysis of nitrous oxide in blood samples. We present a retrospective analysis of nitrous oxide analysis results to gain insight into the scope and characteristics of traffic cases involving living drivers suspected of driving under the influence of nitrous oxide.

Methods: To accommodate police requests for testing, we have developed a semiquantitative headspace GC-MS analysis method for nitrous oxide in blood. Estimation of nitrous oxide concentrations is done by comparing sample results to freshly made calibration curves and additions of Xenon gas as internal standard. Positive samples have been defined as having estimated concentrations greater than 0.1 mL nitrous oxide per L blood. Nitrous oxide test results from traffic cases in Greater Copenhagen and Eastern Denmark have been assessed through a three-year (2020-2022) retrospective analysis. The prevalence of nitrous oxide has been compared to other commonly found drugs of abuse in traffic cases in Greater Copenhagen during the same period.

Results: The case load for nitrous oxide analysis has been steadily increasing over the three-year period, with a total of 62 traffic case blood samples tested for the presence of nitrous oxide. In total, 52 of the 62 cases were positive and most offenders were young males. Nitrous oxide was the seventh most detected drug in blood samples from traffic cases in Greater Copenhagen in 2022.

Conclusion: We present a simple and effective method for the semiquantitative forensic determination of driving under the influence of nitrous oxide. It has been possible to detect nitrous oxide abuse in the majority of cases as requested by the police. The frequency compared to other drugs warrents more analytical development and investigation into the effects of nitrous oxide on the ability to drive safely.