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Evolution of Analytical Methods for the Determination of Ethanol in Blood and Breath for Clinical and Forensic Purposes

A. W. Jones

Published 2025 in WIREs Forensic Science

ABSTRACT

Laboratory procedures for measuring the concentration of alcohol (ethanol) in blood and other body fluids have been available for more than 100 years. The person's blood alcohol concentration (BAC) provides important information to confirm clinical signs and symptoms of drunkenness. Analysis of blood samples required separation of ethanol from the biological matrix by diffusion or distillation before oxidation with chromic acid and titrimetric analysis. By the 1950s enzymatic oxidation became possible after alcohol dehydrogenase (ADH) was extracted and purified from horse liver or yeast. The ADH method furnished a more sensitive and specific method of analysis and laboratory procedures could also be automated. The Breathalyzer instrument was developed in the 1950s and used as a surrogate for measuring the BAC directly. By the 1960s, gas–liquid chromatography (GLC) with flame ionization detector (FID) became the method of choice for analysis of ethanol and other volatiles for forensic purposes. After dilution of the blood sample with internal standard, direct injection and/or headspace (HS) sampling techniques were used. More recently, GC separations are combined with mass spectrometric (MS) detection, which permits more unequivocal identification of ethanol from its characteristic mass fragmentation pattern. Modern technology for breath‐alcohol analysis uses infrared spectrometry and/or electrochemical oxidation for the determination of ethanol. This article gives a retrospective account of major developments in methodology for the determination of ethanol in blood and breath for clinical and forensic purposes and pays homage to some of the pioneers in this domain of analytical toxicology. This article is categorized under: Toxicology > Alcohol Toxicology > Analytical Toxicology > Drug‐Impaired Driving

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