Measurement of urinary catecholamines in small samples for mice

Area of Science:

• Biochemistry
• Analytical Chemistry
• Physiology

Background:

• Analysis of catecholamines in small urine samples is challenging due to stress sensitivity.
• Existing methods require large sample volumes or expensive mass spectrometry.
• A novel extraction method for unconjugated catecholamines in 20μL urine samples has been developed.

Purpose of the Study:

• To develop and validate a sensitive method for catecholamine analysis in micro-urine samples.
• To assess the impact of stress on urinary catecholamine levels.
• To establish a reliable measure of sympathetic activity.

Main Methods:

• Alumina extraction of unconjugated catecholamines from 20μL urine samples.
• Separation using high-performance liquid chromatography (HPLC).
• Detection via electrochemical detection (ECD) with 2,3-dihydroxybenzoic acid internal standard.

Main Results:

• Achieved 48% sample and 60% standard recovery.
• Limit of detection (LOD) of 1.2pg/mL for standards, enabling attomole-level detection in urine.
• Significantly increased norepinephrine and epinephrine levels observed 15 minutes after novel cage stress.

Conclusions:

• The developed technique effectively measures sympathetic activity in micro-samples.
• The method offers attomole-level sensitivity for 20μL urine samples.
• This provides a valuable tool for studying physiological responses to stress.