Determination of plasma lactic acid concentration and specific activity using high-performance liquid chromatography.
Assessment of lactate metabolism is of particular interest during exercise and in disease states such as diabetes, shock, and absorptive abnormalities of short-chain fatty acids by the colon. We describe an analytical method that introduces radio-active tracers and high-performance liquid chromatography (HPLC) to simultaneously analyze concentrations and specific activities (SAs) of plasma lactate. The HPLC conditions included separation on a reversed-phase column (octadecylsilane) and an isocratic buffer (30% acetonitrile in water). [3H]Acetate served as an internal standard. Lactate and acetate were extracted from plasma samples with diethyl ether following a pH adjustment to less than 1.0 and back-extracted into a hydrophilic phase with sodium carbonate (2 mM, pH greater than 10.0). Lactate is detected in the ultraviolet range (242 and 320 nm) by derivatization with alpha-bromoacetophenone. Control plasma samples were studied after an overnight fast for precision and analytical recovery. Calibration curves were linear in the range 0.18-6.0 mM (r = 0.92). The precision was 3% and the analytical recovery was 87%. The detection limit of the method was 36 pmol. Determination of lactate metabolism was performed in a patient with chronic congestive heart failure who was administered primed-continuous L-[U-14C]lactate (10 microCi bolus and 0.3 microCi/min continuously) during a 60-min rest period. Mean arterial lactate concentration and SA were 1.69 +/- 0.2 mM and 253.8 +/- 22 dpm/mumol, respectively. Systemic lactate turnover was 25.65 mumol/kg per min. Lactic acid systemic turnover, organ uptake and release rates can be accurately determined by isocratic HPLC.
Bleiberg
B
B
Department of Medicine, Albert Einstein College of Medicine, Bronx, NY 10461.
Steinberg
J J
JJ
Katz
S D
SD
Wexler
J
J
LeJemtel
T
T
D020155:3-Hydroxybutyric Acid; D000085:Acetates / Q000097:blood; D002851:Chromatography, High Pressure Liquid; D006333:Heart Failure / Q000097:blood; D006801:Humans; D006885:Hydroxybutyrates / Q000097:blood; D007773:Lactates / Q000097:blood*; D019344:Lactic Acid; D012588:Scintillation Counting
D000085:Acetates; D006885:Hydroxybutyrates; D007773:Lactates; D019344:Lactic Acid; D020155:3-Hydroxybutyric Acid