Development of a reference method for determining urinary oxalate by means of isotope dilution-mass spectrometry (ID-MS) and its usefulness in testing existing assays for urinary oxalate.

An accurate and reproducible GC-MS reference method for determining urinary oxalate is presented. With forty 24-h urines, comparisons were made between this reference method and four other existing procedures: a GC method, an HPLC method and two enzymatic assays. The results of the first two methods were in accordance with the GC-MS method. The performance of the enzymatic kits was less satisfactory. The use of a GC-MS reference method in evaluating existing and newly developed methods is recommended.

An improved high performance liquid chromatographic method for determining urinary oxalate, making use of an ID-MS reference method.

With the aid of a newly developed isotope dilution mass spectrometric (ID-MS) measurement of urinary oxalate, an existing HPLC method for assaying this substance was investigated. Results obtained with this method were too high, apparently due to a systematic error. Subsequently, this method was improved by additional prepurification in three different ways: (a) precipitation with CaSO4 to Ca-oxalate, (b) adsorption (and subsequent desorption) to columns supplied with a commercial kit for urinary oxalate and (c) the enzymatic destruction of urinary oxalate present. This work demonstrates the successful use of a reference method in improving a newly developed analytical procedure and presents a reliable and reproducible HPLC method for determining urinary oxalate.

Circadian variation of catecholamine excretion in rats: correlation with locomotor activity and effects of drugs.

Rats housed in metabolic cages were used to study the circadian variation in the urinary excretion of free catecholamines. Small samples of urine (25-100 microliter) were analyzed for adrenaline, noradrenaline and dopamine by high pressure liquid chromatography (HPLC) and electrochemical detection. Various ways in which the values for excretion of catecholamines can be expressed (per min; per ml; per mmol creatinine; as ratio over dopamine) were calculated and discussed. Correction for excretion of creatinine resulted in the lowest variations coefficient among the experimental data. The correction for creatinine removed the circadian rhythm present in the output of noradrenaline (NA) and dopamine (DA). Adrenaline corrected for creatinine still displayed a pronounced circadian variation which was related to the overall locomotor activity of the animals (as recorded by photocells). Collection of 1 hr samples instead of 3 hr samples resulted in a worsening of the relationship between the excretion of adrenaline and locomotor activity. Finally, the possibility that the DA antagonist haloperidol, the DA agonist dipropyl-5,6-2-amino-6,7-dihydroxytetrahydronaphtalene (dipropyl-5,6 ADTN) and the alpha-antagonist phentolamine, could modify the excretion of free urinary catecholamines was investigated. Haloperidol and 5,6-dipropyl-ADTN did not change the output of the catecholamines, but phentolamine induced a strong increase in the excretion of NA. The latter observation suggest that at least part of the excretion of NA may originate from peripheral noradrenergic neurotransmission.