Determination of protein-bound urinary gamma-carboxyglutamic acid in calcium nephrolithiasis

Nephrocalcin is a urinary gamma-carboxyglutamic acid (Gla) containing protein that may be a physiological inhibitor of calcium oxalate nephrolithiasis. Nephrocalcin isolated from urine of stone formers seems to be abnormal in lacking Gla that is required for inhibitory activity. In order to study this hypothesis, we compared the protein-bound urinary Gla contents in 32 calcium oxalate stone formers and in 24 controls. Protein-bound Gla was resolved by reversed-phase high-performance liquid chromatography after elimination of free Gla, alkaline hydrolysis and precolumn derivatization with o-phthalaldehyde and mercaptoethanol. Protein-bound urinary Gla concentrations were similar in stone formers (0.83 +/- 0.38 mumol/l, mean +/- SD) and controls (0.81 +/- 0.27) and were less than 5% of free urinary Gla. However, excretion rates of free and protein-bound Gla (nmol/min) were higher in stone formers (P = 0.006 and P = 0.002). Positive correlations (P = 0.000) were observed between free and protein-bound Gla both in controls and in stone formers. These results do not support the hypothesis of a lacking Gla nephrocalcin in stone formers.