To better understand the diagnosis and therapy of diseases of oxalate metabolism, we set out to measure plasma levels of sulfate, oxalate and citrate from single plasma samples, using a modification of our urinary ion-chromatography method. Previously determination of plasma oxalate was difficult und unreliable because of very low plasma levels (normal adults: 1 - 6 μM/l). Method: Blood was drawn in Li-heparin tubes and placed directly on ice and preserved immediately with 40 μL 1 M HCL / mL plasma in two ultracentrifugation steps. We found that these preparatory steps allow stability of plasma levels of oxalate and citrate for up to 21 days, if samples are stored at - 20° C. For measurement, plasma was diluted with 0.3 mM H3BO3 (1:3 or 1:5) and injected onto an ion chromatography system (DX 500, Dionex), equipped with an analytical column (AS11A) and a guard-column(AG11A) as the stationary phase. The mobile phase was NaOH, continuously degassed with nitrogen, and run as a linear gradient from 5 mM - 52.5 mM over 21 minutes. The eluent background conductivity was suppressed with an anion self regenerating suppressor (ASRS-I, Dionex) on the conductivity detector(300 μS). Peaknet software (Dionex) was used to calculate the concentration of the measured anion peaks. Standards (0.625 - 10 μM for oxalate and citrate and 1.25 - 20 μM for sulfate) were run daily to calibrate the program. Controls (2.5 and 7.5 μM) were run before and after the analysis of the plasma samples; high reliability and reproducibility (CV 1.95 - 4.7%) was obtained. Preliminary analysis of seven normal children with normal renal function yielded measurable oxalate (4.9 ± 2.6 μM/L), citrate (80.9± 26.9 μM/L) and sulfate (251.2 ± 64.1 μM/L). This suggests a true refinement in the method which allows a least detectable plasma oxalate level of 0.3 μM. Further work with primary defects of oxalate metabolism is ongoing. In conclusion, we have established a reproducible, precise method for determination of relevant plasma anions involved in mineral metabolism, which heretofore, have not been easily measurable.
B. Hoppe is supported by a grant from the German Research Foundation (DFG Ho 1272/4-1)
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Hoppe, B., Sailer, D. & Langman, C. A NEW METHOD FOR SIMULTANEOUS DETERMINATION OF SULFATE, OXALATE AND CITRATE IN PLASMA WITH ION-CHROMATOGRAPHY † 1662. Pediatr Res 41, 279 (1997). https://doi.org/10.1203/00006450-199704001-01681