Solid-phase extraction of phospholipids from hemoglobin solutions using Empore styrene-divinylbenzene disks

Area of Science:

• Biochemistry
• Analytical Chemistry
• Biomedical Engineering

Background:

• Red blood cells are a source for hemoglobin-based oxygen carriers.
• Phospholipids in red blood cells can be toxic if present in hemoglobin therapeutics.
• Accurate detection of phospholipids is crucial for ensuring the safety of hemoglobin-based oxygen carriers.

Purpose of the Study:

• To develop a rapid and efficient method for extracting phospholipids from hemoglobin solutions.
• To reduce the time and solvent consumption in sample preparation for hemoglobin-based oxygen carriers.
• To ensure the safety of hemoglobin-based oxygen carriers by quantifying phospholipid levels.

Main Methods:

• Styrene-divinylbenzene Empore disks were used for phospholipid extraction.
• The method was applied to stroma-free hemoglobin preparations.
• Phospholipid recovery and disk capacity were evaluated.

Main Results:

• Empore disks efficiently extracted phospholipids from hemoglobin solutions, with average yields of 92% for four types and 65% for phosphatidylserine.
• Sample preparation time and solvent use were reduced by an average of 80% compared to liquid-liquid extraction.
• The Empore disk method demonstrated capacity for over 0.3 mg of total phospholipids and was successfully used for quantitation.

Conclusions:

• Empore disk extraction is a highly effective method for removing phospholipids from hemoglobin solutions.
• This technique significantly reduces sample preparation time and solvent requirements.
• The method is suitable for routine analysis and quality control of hemoglobin-based oxygen carriers.