We are currently examining the effects of the administration of lithium salts to humans on the urinary excretion of organic acids. It is already established that there is an increased excretion of a-oxoglutarate during therapy with lithium salts (Bond et a/., 1972; Jenner, 1973), and an appreciation of the range of acids affected might assist in the interpretation of this phenomenon. Complete 24h urine collections were obtained from four patients maintained on controlled diets both before and after the start of treatment with lithium salts. We also obtained specimens from a patient discontinuing therapy with lithium salts. The organic acids were isolated either by ion-exchange chromatography on Dowex I (formate form), with elution with 12~-formic acid, o r by extraction from acidified urine with ether or ethyl acetate. The acids were converted into the trimethylsilyl derivatives by using bistrimetliylsilyltrifluoroacetamide containing 1 % (v/v) of chlorotrimethylsilane, either directly or after conversion of ketone groups into the methoximes (Dalgliesh etal., 1966). The resulting mixtures were then examined by g.1.c. on OV-101 and OV-17 columns. Mass spectra were obtained with a Perkin-Elmer 270 gas chromatographmass spectrometer. The ‘metabolic profiles’ obtained by g.1.c. of urinary acids are very complex, a single run revealing at least 70 distinct peaks. The true complexity is much greater, since most of the peaks contain more than one component. A number of studies on humans involving g.1.c. profiles of this type have been reported (see, e.g., Jellum et al., 1971), but these have been concerned with rather gross changes in metabolism. To utilize the full potential of the profiles it is necessary to eliminate random variation. This entails attention, not only to chromatographic detail, but also to the control of diet and other variables. Even so, under practical conditions with patients, some day-to-day variations are seen in the profiles, and careful comparison is necessary to distinguish random from significant changes. The major components can be estimated roughly from the g.1.c. trace. The direct quantitative determination of minor components is not possible except in very favourable cases, and, once a substance is suspected of changing, a more specific method of determination must be sought. The simplest ad hoc solution in many cases is to synthesize the 2H-iabelled compound and to use it as an internal standard. The isotope ratio is determined either by multiple ion monitoring or by the method of repetitive scans (Lee & Pollitt, 1972). We have so far used the method of repetitive scans in this study to determine methylmalonate, succinate, glutarate, adipate, pinielate and suberate. For each patient urine collections made on three separate days before treatment with lithium salts were compared with collections made on three separate days during treatment with lithium salts. The results from one of these patients are shown in Table 1. The rises in succinate, glutarate and adipate were seen in all patients studied and were statistically highly significant. The change in glutarate had roughly the same magnitude as the change in a-oxoglutarate. Examination of the g.1.c. traces also showed three other peaks that increased sharply on treatment with lithium salts. These were identified as being the trimethylsilyl derivatives of fumarate, a-hydroxyglutarate and 2-hydroxy-4oxoglutarate. Malate excretion also rises. The fumarate content of the urine before and after treatment with lithium salts was compared by measuring the m/e 259 peak in the OV-17 repetitive scans (where fiimarate overlaps succinate) and comparing these with the m/e 265 peak of the deuterated succinate standard. In the example shown in Table 1 the fumarate increased fourfold on treatment with lithium salts. Very similar changes, in the revcrse direction, were seen in the patient discontinuing treatment with lithium salts.
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Jun 1, 1990
G A Mills + 1
Open Access
