Date of Award

Spring 5-2026

Document Type

Thesis

Degree Name

Master of Science (MS)

Department/Program

Forensic Science

Language

English

First Advisor or Mentor

Marta Concheiro-Guisan

Second Reader

Ana Pego

Third Advisor

Michelle Carlin

Abstract

While the window of detection for direct ethanol measurement in the blood can be up to 8 hours, the alcohol metabolite phosphatidylethanol (PEth) can be measured anywhere from 7 days to 1 month later, depending on the level of consumption. PEth is a unique phospholipid formed from phosphatidylcholine present in the membranes of red blood cells. For accurate and reliable measurement of PEth, the analysis of the most abundant analogs, phosphatidylethanol 16:0/18:1 and 16:0/18:2, is recommended. Assessment of drinking history is useful in multiple forensic and clinical settings, such as determining prenatal alcohol exposure. This study aimed to improve on current methods for the analysis and detection of PEth 16:0/18:1 and PEth 16:0/18:2 using dried blood spots (DBS) and liquid chromatography–tandem mass spectrometry (LC–MS/MS). This method was applied to high–risk pregnant women across three visits to investigate alcohol use during pregnancy. One 10 mm whole punch from DBS cards was extracted in 1 mL of methanol for 90 minutes in the presence of the internal standards, PEth 16:0/18:1–d5 and PEth 16:0/18:2–d5 at 0.1 μg/mL, followed by centrifugation, evaporation, and reconstitution with 200 μL of acetonitrile: methanol (90:10 v/v). The chromatographic separation was performed in gradient mode on a C18 column. The analysis was achieved by LC–MS/MS in negative ESI mode. A total of 596 authentic samples were analyzed, with mean concentrations of 70 ng/mL for PEth 16:0/18:1 and 68 ng/mL for PEth 16:0/18:2. These primary findings demonstrate that DBS–PEth analysis can improve the detection of prenatal alcohol exposure.

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