Simultaneous determination of L- and D-methotrexate using a sequential injection analysis/amperometric biosensors...

Area of Science:

• Analytical Chemistry
• Biotechnology
• Biomedical Engineering

Background:

• Methotrexate (Mtx) is a crucial chemotherapy drug with stereoisomers L-Mtx and D-Mtx.
• Accurate and simultaneous determination of both isomers is vital for therapeutic efficacy and minimizing toxicity.
• Existing analytical methods may lack the speed, precision, or on-line capabilities required for synthesis monitoring.

Purpose of the Study:

• To develop and validate a sequential injection analysis (SIA) system for the simultaneous determination of L-methotrexate and D-methotrexate.
• To integrate amperometric biosensors as sensitive detectors within the SIA system.
• To assess the system's suitability for on-line process control during Mtx synthesis.

Main Methods:

• Sequential Injection Analysis (SIA) system implementation.
• Amperometric biosensors utilizing L-amino acid oxidase (L-AAOD), L-glutamate oxidase (L-Glox), D-amino acid oxidase (D-AAOD), and horseradish peroxidase (HRP).
• Simultaneous detection of L-Mtx and D-Mtx at the pmol/L to nmol/L concentration range.

Main Results:

• The SIA/biosensor system demonstrated high precision and accuracy.
• Achieved very low limits of detection for both L-Mtx and D-Mtx.
• The system enabled simultaneous quantification of L- and D-Mtx with a sample throughput of 34 samples per hour.
• The method proved reliable for on-line synthesis process control.

Conclusions:

• The developed SIA system with amperometric biosensors provides a robust and efficient platform for simultaneous L-Mtx and D-Mtx determination.
• This method offers significant advantages in terms of precision, accuracy, and reduced sample/buffer consumption.
• The system is well-suited for real-time monitoring and control in pharmaceutical synthesis processes.