pKa determinations by using a HPLC equipped with DAD as a flow injection apparatus

Area of Science:

• Analytical Chemistry
• Physical Chemistry

Background:

• Accurate determination of pKa values is crucial for understanding chemical and biological processes.
• Traditional spectrophotometric methods can be time-consuming and require larger sample volumes.

Purpose of the Study:

• To develop and validate a semi-automated, spectrophotometric method for pKa determination.
• To leverage High-Performance Liquid Chromatography with a Diode Array Detector (HPLC-DAD) for enhanced analysis.
• To assess the method's performance regarding sample consumption, speed, sensitivity, and precision.

Main Methods:

• A semi-automated flow injection analysis system was configured using an HPLC-DAD.
• Spectrophotometric measurements were performed across a wide pH range while maintaining constant ionic strength.
• pKa values were determined for benzoic acid and 2-aminopyridine.
• The method was also applied to samples dissolved in non-aqueous solvents.

Main Results:

• The developed method demonstrated low sample consumption, rapid sample throughput, high sensitivity, and excellent precision.
• Experimental pKa values for benzoic acid (~4.0) and 2-aminopyridine (~6.8) closely matched literature values.
• The impact of non-aqueous solvents on pKa values was minimal (0.04–0.40 pH units) compared to conventional techniques.

Conclusions:

• The semi-automated HPLC-DAD spectrophotometric method provides an efficient and accurate approach for pKa determination.
• This technique is suitable for analyzing compounds in both aqueous and non-aqueous media.
• The method offers significant advantages in terms of speed, sample efficiency, and precision over traditional approaches.