ENZYMATIC ASSAY OF D-MANNOSE IN SERUM

We describe a new and improved enzymatic assay for determining the con centration of D-mannose in sera. Serum D-glucose is selectively conver ted to glucose-6 phosphate with the highly specific thermostable gluco kinase (EC 2.7.1.2) from Bacillus stearothermophilus. The anionic reac tion products and excess substrates are removed by a rapid and simple anion-exchange chromatography step in microcentrifuge spin columns. D- Mannose in the glucose-depleted sample is then assayed spectrophotomet rically by using coupled enzymatic reactions. The quantitative elimina tion of glucose from the serum samples allowed the accurate and reprod ucible assay of serum mannose in,the 0-200 mu mol/L range. Recovery of mannose added to serum (5-200 mu mol/L) was 94% +/- 4.4%. The intraas say CV was 6.7% at 40 mu mol/L mannose (n = 5; 39.6 +/- 1.6 mu mol/L) and 4.4% at 80 mu mol/L (n = 11; 75.0 +/- 1.8 mu mol/L); the interassa y CV at these concentrations was 12.2% (n = 7; 36.9 +/- 2.1 mu mol/L) and 9.8% (n = 7; 74.2 +/- 2.7 mu mol/L), respectively. Sera from 11 he althy human volunteers contained an average of 54.1 +/- 11.9 mu mol/L mannose (range 36-81 mu mol/L).