Gas chromatographic/mass spectrometric analysis of stable isotopes of 3-methylhistidine in biological fluids: Application to plasma kineticsin vivo

Abstract

A simple and rapid method for measuring 3-methylhistidine (3MH) in plasma and urine is described. Internal standard, 1-methylhistidine (1MH), was added to plasma, acidified and absorbed onto cation-exchange columns. It was then eluted from columns, dried, and derivatized for gas chromatography/mass spectrometry. A major fragment of 3MH was monitored at 238 u and 3-methyl-(methyl-2H3)histidine (d3-3MH) (used for in vivo kinetics) at 241 u, whereas 1MH was monitored at 340 u and eluted 0.5 min later than 3MH. Standard curves for plasma analysis were linear and nanamole amounts of 3MH in plasma were determined with a precision of 3.5%. 3MH was also quantitated in urine; however, because of substantial amounts of 1MH, (18O2)1MH was used as the internal standard. Nanamole amounts of 3MH were determined in urine with a precision of 2.7%. Application of the 3MH analytical method was used to develop a kinetic compartmental model by using the stable isotope of 3MH, d3-3MH. Cattle, like humans, quantitatively excrete 3MH in the urine. A young bovine was injected with d3-3MH and the enrichment curve in plasma was evaluated in order to obtain a steady-state production rate of 3MH. The decay curve was modeled through the use of NIH-SAAM modeling program. The kinetics of d3-3MH from plasma were adequately described by a three-pool compartmental model. The de novo production rate of 3MH estimated in the calf was 665 mumol per day. This corresponded to an estimated fractional turnover rate of 1.56% per day, which was similar to estimates obtained from urine collections.(ABSTRACT TRUNCATED AT 250 WORDS)

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