Bioluminescent flow sensor for L-phenylalanine determination in serum

Area of Science:

• Biotechnology
• Analytical Chemistry
• Enzyme Assays

Background:

• L-phenylalanine (Phe) is an essential amino acid with clinical significance.
• Accurate quantification of Phe in serum is crucial for diagnosing and managing certain metabolic disorders.
• Existing methods for Phe determination can be time-consuming or require complex instrumentation.

Purpose of the Study:

• To develop a highly sensitive and rapid bioluminescent flow sensor for L-phenylalanine determination in serum.
• To investigate and compare the performance of different immobilized phenylalanine dehydrogenase (PheDH) enzymes.
• To validate the sensor's accuracy and reliability against established methods.

Main Methods:

• Development of a bioluminescent flow sensor system.
• Immobilization of phenylalanine dehydrogenase (PheDH) from Bacillus badius, Bacillus sphaericus, and Rhodococcus sp. M 4.
• Monitoring of reduced nicotinamide adenine dinucleotide (NADH) production using bacterial bioluminescent enzymes.
• Analysis of sensor performance, including sensitivity, linearity, and precision.

Main Results:

• The sensor demonstrated high sensitivity and rapid response for L-phenylalanine detection.
• The immobilized PheDH from Bacillus badius exhibited superior transformation rates and sensitivity.
• Linear response was observed from 1 to 100 microM, with a detection limit of 0.5 microM (10 pmoles).
• Intra- and inter-assay coefficients of variation were below 5%, with recoveries between 90-101%.

Conclusions:

• The developed bioluminescent flow sensor provides a sensitive, rapid, and reliable method for serum L-phenylalanine quantification.
• The Bacillus badius-based PheDH reactor offers optimal performance for this application.
• The sensor's results showed excellent agreement with chromatographic methods and normal reference values, indicating its clinical utility.