Balcke, P.

Haubenstock, A.

Kopsa, H.

Schmidt, P.

Zazgornik, J.

Clinical Chemistry and Laboratory Medicine (CCLM)

Undetermined

Dokumenten-Publikationsserver der Humboldt-Universität zu Berlin

Balcke et al.: Oxalic acid levels in haemodialysis patients 261J. CJin. Chem. Clin. Biochem.Vol. 22, 1984, pp. 261-263In Vitro Synthesis of Oxalic Acid Has No Relevant Influenceon Plasma Oxalic Acid Levels in Haemodialysis PatientsBy P. Balcke, P. Schmidt, J. Zazgornik, A. Haubenstock and H. KopsaL Medizinische Universit tsklinik (Vorstand: Prof. Dr. Dr. h.c. E. Deutsch) Wien(Received July 27/November 23, 1983/January 6, 1984)Summary: Plasma oxalic aeid concentrations were measured in 13 chronic haemodialysis patients. The meanplasma oxalic acid concentration was 128.0 ± 48.6 μπιοΐ/ΐ, being approximately 8 times higher than theplasma concentration of 14 volunteers (x = 16.8 ±5.2 μηιοΐ/ΐ). Ultrafiltrates obtained in vivo from thesepatients showed a mean oxalic acid concentration of 138.2 ± 56.5 μιηοΐ/ΐ. Since in vivo ultrafiltrates are freeof erythrocytes and plasma enzymes, an in vitro synthesis of oxalic acid from precursors by erythrocytes andplasma enzymes can be excluded. As the oxalic acid concentration of plasma corresponded to that of in vivoultrafiltrates, it is concluded that any in vitro formation of oxalic acid in haemodialysis patients must benegligibly small, and is irrelevant for the measurement of plasma oxalic acid levels in patients receiving regul rhaemodialysis.In vitro-Synthese von Oxals ure ist ohne Bedeutung f r die Bestimmung von Oxals ure im Plasmabei H modialysepatientenZusammenfassung: Bei 13 chronischen H modialysepatienten wurde die Konzentration von Oxals ure imPlasma bestimmt. Die mittlere Oxals urekonzentration im Plasma lag bei 128,0 ± 48,6 μπιοΐ/ΐ und war damitetwa 8 mal h her als der Wert, der bei 14 Referenzpersonen gemessen wurde (x = 16,8 ± 5,2 μιηοΐ/ΐ).Ultrafiltrate, die in vivo von diesen Dialysepatienten gewonnen worden waren, wiesen eine mittlere Oxals u-rekonzentration von 138,2 ± 56,5 μπιοΐ/ϊ auf. Da in in vivo-UItrafiltraten Erythrocyten und Plasmaenzymenicht enthalten sind, kann eine in vitro-Synthese von Oxals ure durch Erythrocyten und Plasmaenzyme ausVorstufen ausgeschlossen werden. Durch den Nachweis ann hernd gleich hoher Oxals urekonzentrationen inPlasma und Ultrafiltraten kann angenommen werden, da eine in vitro-Synthese von Oxals ure bei H mo-dialysepatienten vernachl ssigbar klein und unbedeutend f r die Bestimmung der Konzentration von Oxal-s ure im Plasma bei Dialysep tienten ist.IntroductionIncreased plasma oxalic acid concentrations havebeen described in patients undergoing chronic hae-modialysis (l, 2, 3). However the normal r nge ofplasma oxalic acid is still a matter of dispute. Usingchemical, fluorometric or enzymic methods the nor*mal r nge of plasma oxalic acid is about 10—20μπιοΐ/ΐ, being approximately 10 times higher thanthe concentration obtained by human in vivo isotop-ic investigations (r nge of plasma oxalic acid 1—3μιηοΐ/ΐ (4—7)). This discrepancy was recently re-J. Clin. Chem. Clin. Biochem. / Vol. 22, 1984 / No. 3solved by Akcay & Rose, who demonstrated thatoxalic acid is synthesized from precursors in vitro byerythrocytes when samples are allowed to stand, thusresulting in erroneously high plasma levels in nor-mals (8). An accumulation in the plasma of precur-sors of oxalic acid can be expected in uraemia. Thequestion therefore arises of whether the increasedvalues found in haemodialysis are due to a genuinephysiological increase of oxalic acid, or to an in-creased conversion of accumulated precursors byerythrocytes and plasma enzymes in vitro. Ultrafil-262 Balcke et al.: Oxalic acid levels in haemodialysis patientstration before the haemodialysis, using dialysis ma-chines adapted for Ultrafiltration, offers the oppor-tunity to obtain ultrafiltrates free of erythrocytes andenzymes in vivo. Oxalic acid has a low molecularweight (Mr = 90) and shows negligible protein bind-ing; it therefore passes freely across the dialysismembrane, so that its concentration in the ultrafil-trate reflects its plasma level. In the present studypxalic acid concentrations were compared in in-vivoultrafiltrates and in contemporaneous plasma sam-ples of 13 patients undergoing chronic haemodialy-sis.Materials and MethodsUltrafiltrates were obtained in vivo from 13 chronic haemodialy-sis patients by 10 minutes Ultrafiltration before the beginning ofthe haemodialysis Session, s described by Schmidt and coworkers(9). No patient suffered from primary hyperoxalaemia. Capillarykidneys (ASAHI AM, 20) were used for Ultrafiltration. The trans-membrane pressure was 300 mm Hg. Blood samples were drawnsimultaneously and centrifuged within l hour (fig. 1). Former stu-dies showed that ultrafiltrate concentrations of low molecularweight substances like creatinine and potassium correspond toplasma levels after 10 minutes ultrafiltration (creatinine r = 0.88,potassium r = 0.92) (9). The behaviour of oxalic acid can be ex-pected to be similar to that of creatinine s the molecular weightsof these two substances differ only moderately (Mr creatinine 113,oxalic acid 90) and since protein binding of oxalic acid is minimal.Oxalic acid determinations were performed according to the pho-tometric method described by Krugers Dagneaux et al. (4). Sinceplasma proteins can influence the accuracy of the oxalic acid de-termination, ultrafiltrates of the plasma samples were used. Ultra-filtration of diluted plasma samples (1:10) was performed usingAmicon Filters Diaflo PM 10 with pore sizes corresponding to10000 Dalton.Cdpiliory kidneyBlood in ·Arterioll ineBlood ouiVenous lineResultsThe mean plasma oxalic acid concentration of the 13chronic haemodialysis patients under investigationwas 128.0 ± 48.6 μπιοΐ/ΐ, being approximately 8times higher than the mean plasma xjxalic acid levelof 14 normal volunteers (x = 16.8 ± 5.2 μπιο1/1).The mean oxalic acid level in the ifi-vivo ultrafil-trates of these patients was 138.2 ± 56.5 μηαοΐΛThus plasma oxalic acid concentrations were 8.7 percent lower than the mean oxalic acid concentrationmeasured in the in-vivo ultrafiltrates (Studenfs t-testf r paired samples: t = 2.26, p < 0.05). The lowermean oxalic acid level in plasma compared with thatof the in-vivo ultrafiltrates is probably due to smalllosses of oxalic acid during the plasma determinationprocedure. The correlation between the oxalic acidlevels in plasma and in the in-vivo ultrafiltrates washighly significant (r = 0.96, p < 0.001) (fig. 2).o 80Pump for negative pressureUUrafiltrate out40 80 120 160 200 240 28ΪΓOxalic ocid (p l a sma) [μπιοΐ/ΐ]Fig. 2. Correlation between oxalic acid concentration in plasmaand in-vivo ultrafiltrates.Regression line *Ideal regression lineFig. K Model of ultrafiltration procedure in vivo.T T T Transmembrane pressure (300 mmHg)[ j Points for drawing blood or ultrafiltrateUltrafiltrates obtained in this way and in-vivo ultrafiltrates wereused for oxalic acid determination.Since protein binding of oxalic acid is minimal, concentrations inin-vivo and in-vitro ultrafiltrates can be assumed to be equal toplasma levels.DiscussionThe mean plasma oxalic acid concentration in chron-ic haemodialysis patients was about 8 times higherthan that in normal volunteefs. The mean oxalic acidlevel in plasma was not higher than the mean oxalicacidoconcentration of in-vivo ultrafiltrates. Further-more oxalic acid concentrations ιη* plasma and inplasma ultrafiltrates showed a highly significant cor-J. Clin. Chem. Clin. Biochem. / Vol. 22, 1984 / No. 3Balcke et al.: Oxalic acid levels in haemodialysis patients · 263relation. Since an in vitro synthesis of oxalic acid canbe excluded in in-vivo ultrafütrate, we assume thatspontaneous generation of oxalic acid in blood sam-ples is of negligible significance in the determinationof plasma oxalic acid levels in uraemic patients.However, our results do not necessarily disprove therelevance of in vitro synthesis of oxalic acid for themeasurement of plasma oxalic acid concentrations innormals. It was suggested that the estimated normalränge for plasma oxalic acid is erroneously high, ow-ing to the in vitro conversion of glyoxylic acid (a pre-cursor) to oxalic acid by erythrocytes in blood sam-ples on Standing (8). Akcay & Rose showed that gly-oxylic acid added to blood samples is converted tooxalic acid immediately by erythrocytes (8). Rehmertand coworkers demonstrated the in-vitro synthesisof oxalic acid in plasma of normals. The formation ofoxalic acid could be prevented by addition of enzymeinhibitors (3). In addition to glyoxylic acid, otherprecursors, like glycolic acid, ascorbic acid or gly-cine, could contribute to in vitro synthesis of oxalicacid (10).. In uraemia, the accumulation of thesecompounds, resulting in elevated plasma levels, canbe expected or has even been proved. Therefore anincreased in vitro formation of oxalic acid from pre-cursors might be expected in uraemia. Our results,however, demonstrate that the in vitro conversion ofthese precursors could make only a negligibly smallcontribution to the elevated plasma oxalic acid con-centrations in uraemic patients. In fact, such a con-version cannot be demonstrated by the method used.It can therefore be assumed that the increased plas-ma oxalic acid concentrations measured in uraemiagenuinely reflect the accumulation of oxalic acid inrenal failure.References1. Zarembsky, P. M. & Hodgkinson, A. (1966) Nature 272,511-512.2. Balcke, P., Schmidt, P., Zazgornik, J., Kopsa, H. & Deutsch,E. (1980) N. Engl. J. Med. 303, 944.3. Rehmert, U., Wicher, K., Rüge, W. & Bahlmann, J. (1983)Lab. Med. 7, 29-32.4. Krugers Dagneaux, P. G. L. C, Klein Elhorst, J. T. & Ol-thuis, F. M. F. G. (1976) Clin. Chim. Acta 77, 319-325.5. Hatch, M., Bourke, E. & Costello, J. (1977) Clin. Chem. 23,76-78.6. Hodgkinson, A. & Wilkinson, R. (1974) Clin. Sei. Mol. Med.46, 61-73.7. Kohlbecker, G. & Butz, M. (1981) J. Clin. Chem. Clin. Bio-chem. 79, 1103-1106.8. Akcay, T. & Rose, G. A. (1980) Clin. Chim. Acta 707, 305-311.9. Schmidt, P., Endler, A. T., Kopsa, H., Zazgornik, J. &Balcke, P. (1980) 5. Donausymposion für Nephrologie, Linz.10. Schröder, R. (1980) Dtsch. Med. Wochenschr. 105. 997-1001.Dr. P. BalckeI. Medizinische Universitätsklinik WienLazarettgasse 14A-1090 WienJ. Clin. Chem. Clin. Bipchem, / Vol. 22, 1984 / No. 3

In Vitro Synthesis of Oxalic Acid Has No Relevant Influence on Plasma Oxalic Acid Levels in Haemodialysis Patients

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In Vitro Synthesis of Oxalic Acid Has No Relevant Influence on Plasma Oxalic Acid Levels in Haemodialysis Patients

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