New enzymatic method with tryptophanase for determining potassium in serum

Area of Science:

• Biochemistry
• Analytical Chemistry
• Enzymology

Background:

• Accurate serum potassium measurement is crucial for diagnosing and managing various medical conditions.
• Existing methods like flame photometry may have limitations in speed or specificity.
• Enzymatic assays offer potential for high specificity and sensitivity in ion determination.

Purpose of the Study:

• To develop a simple, rapid, and accurate enzymatic method for quantifying potassium ion concentration in human serum.
• To utilize tryptophanase and glutamate dehydrogenase for a novel potassium assay.
• To validate the performance of the developed enzymatic assay against a standard method.

Main Methods:

• Purification of tryptophanase (EC 4.1.99.1) from Escherichia coli K12.
• Enzymatic reaction involving tryptophanase, pyridoxal 5-phosphate, and potassium/ammonium ions.
• Elimination of endogenous ammonium using glutamate dehydrogenase (GLDH; EC 1.4.1.4).
• Quantification of ammonium produced, proportional to potassium concentration, via GLDH-catalyzed NADP+ formation.
• Spectrophotometric measurement of absorbance change at 340 nm.

Main Results:

• The assay demonstrated linearity for potassium concentrations up to 7.00 mmol/L.
• High precision was observed with within-assay coefficients of variation (CV) as low as 0.89% and day-to-day CVs of 0.99%.
• Excellent analytical recovery (98.7%–108.9%) and a high correlation coefficient (0.995) with flame photometry were achieved.
• The method showed minimal interference from hemoglobin, bilirubin, and other cations.

Conclusions:

• A simple, rapid, and precise enzymatic method for serum potassium determination has been successfully established.
• The assay utilizes readily available enzymes and standard spectrophotometric techniques.
• This enzymatic method provides a reliable alternative to traditional methods for clinical potassium analysis.