Shift in pH of biological fluids during storage and processing: effect on bioanalysis

Area of Science:

• Biochemistry
• Analytical Chemistry
• Pharmacology

Background:

• Storage and processing of biological samples can alter their pH.
• Changes in pH can significantly impact the stability of pH-labile compounds.
• Accurate interpretation of in vivo study results relies on stable sample conditions.

Purpose of the Study:

• To monitor pH changes in ex vivo plasma, bile, and urine.
• To investigate the impact of storage and processing on sample pH.
• To evaluate methods for stabilizing sample pH and preserving compound integrity.

Main Methods:

• Monitoring pH of plasma, bile, and urine under various storage conditions (time, temperature).
• Assessing pH changes after sample processing (ultrafiltration, centrifugation, precipitation, evaporation).
• Illustrating pH shift effects using model compounds and drug stability studies.

Main Results:

• Ex vivo plasma, bile, and urine pH increased upon storage and processing.
• Increased pH led to significant degradation of pH-labile compounds.
• Specific examples showed substantial drug loss (e.g., compound I, II) due to pH shifts.
• Methanol precipitation and evaporation resulted in a high pH (9.5), causing degradation.

Conclusions:

• Plasma pH shifts during processing can yield misleading in vivo study results.
• Degradation of pH-labile compounds can be prevented by stabilizing sample pH.
• A 10% CO2 atmosphere is recommended for stabilizing plasma pH for stability/protein binding studies.
• Citrate or phosphate buffers are suitable for stabilizing pH in bile and urine, and for extensive plasma sample preparation.