Accurate measurement of pyruvate is particularly important in the detection of mitochondrial enzyme disorders. We describe a simple high-performance liquid chromatography assay for pyruvate with derivatization and fluorescence detection. Between-run variation is below 4·5% and the method is linear between 1 and 1200μmol/L. The mean recovery is 101%. After sample pretreatment, the calculated pyruvate concentration proved to be stable for 15 days when samples were kept at −20°C.
References
1.
MunnichARötigAChretienDCormierVBourgeronTBonnefontJ-PClinical presentation of mitochondrial disorders in childhood. J Inherit Metab Dis1996;19: 521–7.
2.
Poggi-TravertFMartinDBillette de VillemeurTBonnefontJ-PVassaultARabierDMetabolic intermediates in lactic acidosis: compounds, samples and interpretation. J Inherit Metab Dis1996;19: 478–88.
3.
ChariotPRatineyRAmmi-SaidMHérigaultRAdnotSGherardiR.Optimal handling of blood samples for routine measurement of lactate and pyruvate. Arch Pathol Lab Med1994;118: 695–7.
4.
ChanAGoldRArpS.Pflughaupt KW, Toyka KV, Reichmann H. Standardized bicycle ergometry test in mitochondrial myopathies. Indications, interferences and clinical parameters. Nervenarzt1998;69: 472–84.
5.
HermannH-PPieskeBSchwarzmüllerEKeulJJustHHasenfussG.Haemodynamic effects of intracoronary pyruvate in patients with congestive heart failure: an open study. Lancet1999;353: 1321–3.
GlosterJAHarrisP.Observations on an enzymic method for the estimation of pyruvate in blood. Clin Chim Acta1962;7: 206–21.
8.
LiaoJCHoffmanNEBarboriakJJRothDA. High-performance liquid chromatography of pyruvic and α-ketoglutaric acids and its application to urine samples. Clin Chem1977;23: 802–5.
9.
TeradaHHayashiTKawaiSOhnoT.High-performance liquid chromatographic determination of pyruvic acid and α-ketoglutaric acid in serum. J Chromatogr1977;130: 281–6.
10.
PaillaKBlonde-CynoberFAusselCde BandtJ-PCynoberL.Branched-chain keto-acids and pyruvate in blood: measurement by HPLC with fluorimetric detection and changes in older subjects. Clin Chem2000;46: 848–53.
11.
LinS-SMiyawakiONakamuraK.Continuous production of L-alanine with NADH regeneration by a nanofiltration membrane reactor. Biosci Biotechnol Biochem1997;61: 2029–33.
12.
GallemannDEyerP.Formation of hydrogen peroxide during precipitation of red cells with perchloric acid. A cautionary note for precise determination of pyruvate, GSH, and NAD(P)H. Anal Biochem1990;191: 347–53.
13.
AbasoloMIGaozzaCHFernándezBM. Kinetic study on the anelation of heterocycles. I. Quinoxalinone derivatives synthesized by the Hinsberg reaction. J Heterocyclic Chem1987;24: 1771–5.
14.
LiRKenyonGL. A spectrophotometric determination of α-dicarbonyl compounds and its application to the enzymatic formation of α-ketobutyrate. Anal Biochem1995;230: 37–40.
15.
StijntjesGJte KoppeleJMVermeulenNPE. High-performance liquid chromatography-fluorescence assay of pyruvic acid to determine cysteine conjugate β-lyase activity: application to S-1,2-dichlorovinyl-1-cysteine and S-2-benzothiazolyl-1-cysteine. Anal Biochem1992;206: 334–43.
16.
YangWRothK.S.Defect in α-ketobutyrate metabolism: a new inborn error. Clin Chim Acta1985;145: 173–82.
17.
LandaasSPettersenJE. Clinical conditions associated with urinary excretion of 2-hydroxybutyric acid. Scand J Clin Lab Invest1975;35: 259–66.
18.
RosalkiSBWilkinsonJH. Reduction of α-ketobutyrate by human serum. Nature1960;188: 1110–1.
19.
HenryRJCannonDCWinkelmanJW, editors. Clinical Chemistry, 2nd edn.London: Harper and Row, 1974.
20.
LamersKJDoesburgWHGabreelsFJRomsomACRenierWOWeversRAReference values of blood components related to fuel metabolism in children after an overnight fast. Clin Chim Acta1985;145: 17–26.
21.
FosterKJAlbertiKGMMHinksLLloydBPostleASmythePBlood intermediary metabolite and insulin concentrations after an overnight fast: reference ranges for adults, and interrelations. Clin Chem1978;24: 1568–72.
22.
ArtuchRVilasecaMAFarréCRamonF.Determination of lactate, pyruvate, β-hydroxybutyrate and acetoacetate with a centrifugal analyser. Eur J Clin Chem Clin Biochem1995;33: 529–33.
23.
PanteghiniMPaganiF.Biological variation of lactate and pyruvate. Clin Chem1993;39: 908.
