Development, validation and application of an HPLC method for reduced vitamin C qualification in HBOCs solution

Area of Science:

• Analytical Chemistry
• Biochemistry

Background:

• Hemoglobin-based oxygen carriers (HBOCs) are crucial for oxygen delivery.
• Accurate determination of vitamin C (Vc) is essential for HBOCs stability and efficacy.
• Existing methods for Vc determination in complex matrices like HBOCs can be time-consuming.

Purpose of the Study:

• To develop a rapid and sensitive high-performance liquid chromatography (HPLC) method for quantifying vitamin C (Vc) in HBOC solutions.
• To validate the method's performance characteristics, including linearity, recovery, precision, and limits of detection/quantification.
• To explore the application of the developed HPLC method in studying Vc's role in methemoglobin reduction within HBOCs.

Main Methods:

• High-performance liquid chromatography (HPLC) was employed for Vc determination.
• A sample preparation protocol involving polymerized hemoglobin precipitation was established.
• Method validation included linearity assessment, recovery studies, precision analysis, and determination of detection and quantification limits.

Main Results:

• The developed HPLC method provides rapid Vc determination in HBOCs within 5 minutes post-sample preparation.
• The method demonstrated excellent linearity (0.00-3.00 μg Vc), high recovery (103.56-108.68%), and good precision (RSD < 1.00%).
• Low limits of detection (0.002 ppm) and quantification (0.01 ppm) were achieved, indicating high sensitivity.

Conclusions:

• The established HPLC method is rapid, sensitive, and accurate for Vc determination in HBOC solutions.
• The method's application in studying methemoglobin reduction suggests higher pH accelerates the process.
• This HPLC technique holds significant potential for quality control and research in HBOC development.