A manual spectrophotometric method for the measurement of serum sodium and potassium by enzyme activation.

Manual procedures suitable for use on standard benchtop spectrophotometers have been developed for the enzymatic determination of Na+ and K+ in serum. Both assays require only minimal modification of reagents already available for BM/Hitachi analyzers and are performed in an endpoint mode, allowing up to 20 assays per run. The addition of a stop reagent is required--dipotassium EDTA for the Na+ assay and sodium dodecyl sulphate for the K+ assay. The most important criterion for achieving good assay performance is the precise pipetting of sample and reagent. Within-run imprecision is < 1% for Na+ and K+, and between-run imprecision < 1.5%, for both assays at all but the lowest concentrations of K+. Enzymatic electrolyte results compare well with flame photometry, however the assays are more prone to interference by very high concentrations of bilirubin or triacylglycerols than those performed on automated, dual-wavelength kinetic analyzers. It is possible to correct for most interferences by inclusion of appropriate sample and reagent blanks.

Effect of fatty acids on rat liver nuclear T3-receptor binding.

The effects of selected fatty acids (linoleic, oleic, and palmitic) on triiodothyronine (T3)-receptor binding were compared in isolated rat hepatocytes, rat liver nuclei, and receptor protein. Scatchard analysis indicated that the inhibition of T3-receptor binding by fatty acids was characterized by an increase in Kd and no change in maximum binding capacity (MBC). In isolated receptors, the rank order of potency for inhibition was linoleic acid greater than oleic acid greater than palmitic acid. The Ki for oleic acid in isolated receptors was the same as that for whole nuclei (15.4 +/- 1.3 v 16.3 +/- 1.9 mumol/L, respectively), indicating that the inhibition of nuclear T3 binding is probably at the level of the receptor protein itself. In isolated hepatocytes, linoleic acid was more potent than oleic acid in inhibiting T3 binding to nuclear receptors. Cell-associated T3 was not affected by the presence of fatty acids, implying that cellular uptake of T3 was not inhibited. High concentrations of fatty acids were necessary for inhibition of T3-receptor binding in isolated hepatocytes, with linoleic acid being one to two orders of magnitude less potent in isolated hepatocytes compared with isolated receptors (Ki, 179 +/- 12 v 4.4 +/- 0.5 mumol/L, respectively). It is concluded that the inhibitory effect of fatty acids on T3-receptor binding in isolated rat hepatocytes probably occurs at the level of the nuclear receptor, and does not involve an inhibition of the access of T3 to the receptor. However, in vivo it seems unlikely that fatty acids will have access to the nuclear receptors in sufficiently high concentrations to affect T3-receptor binding in liver cells.

Urinary risk factors in calcium oxalate stone disease: comparison of men and women.

The daily excretion of calcium, oxalate, uric acid and glycosaminoglycans, the 24-h urinary pH and volume, and the inhibitory effects of the urines on calcium oxalate crystal growth and aggregation, were measured in 44 normal women, 41 normal men, 32 female stone formers and 63 male stone formers. No significant differences could be found between the normal men and women, the male and female stone formers, or between the patients and their normal controls with regard to the excretion of oxalate and glycosaminoglycans, and the urinary pH. The normal women exhibited significantly lower urinary volumes and excreted less calcium per day than did the other subject groups. The excretion of calcium by the female stone formers was indistinguishable from that of both groups of men. The male and female stone formers did not differ from their corresponding control groups with regard to the excretion of urate, but both groups of male subjects had significantly higher daily urate excretions than did either female category. This was attributed to the greater body weights of the men. There were no discernible differences between any of the subject groups with regard to the inhibitory effects of their urines on calcium oxalate crystal growth, but urines from both groups of female subjects demonstrated a significantly greater inhibitory influence on crystal aggregation than did those of the men. It would appear that the relatively low incidence of uninfected calcium oxalate urolithiasis in women compared with men may be attributable to (a) a lower daily calcium excretion and (b) a higher inhibitory activity of their urines towards crystal aggregation.

Inhibitory activity of whole urine: a comparison of urines from stone formers and healthy subjects.

The inhibitory activity of whole urines from 32 healthy subjects and 50 calcium oxalate renal stone formers was assessed in terms of their ability to withstand increasing quantities of oxalate before undergoing spontaneous nucleation of calcium oxalate, and their response to a standard 30-mumol challenge of oxalate above their measured metastable limits. The concentrations of calcium (p less than 0.05), oxalate (p less than 0.05), urate (p less than 0.01) and glycosaminoglycans (p less than 0.005) were significantly lower in the stone formers than in the controls and were associated with a significantly higher 24-h urinary volume (p less than 0.001). The majority of urine samples precipitated envelope crystals of calcium oxalate dihydrate, while the remainder precipitated the monohydrate. A significantly (p less than 0.02) greater proportion of the urines from stone formers than from controls deposited calcium oxalate monohydrate, and this was attributed to a lower concentration of calcium in these urines. The minimum amounts of oxalate necessary to induce crystal nucleation did not differ between the two groups, but when the measured metastable limits were expressed as the product of the total (i.e. endogenous + that added to induce nucleation) concentrations of oxalate and calcium at which precipitation occurred, then these limits were significantly lower (p less than 0.05) in the stone formers than in the healthy subjects. However, when the metastable limits of a subgroup of stone formers and controls matched for 24-h urinary volume and calcium and urate concentrations were compared, no differences between the groups could be discerned.(ABSTRACT TRUNCATED AT 250 WORDS)

Factors affecting measurement of urinary oxalate.

Using a gas-chromatographic method, we examined the effects of phosphate concentration, added calcium chloride, and pH on precipitation of oxalate from urine. All three factors are important, but the pH of precipitation is particularly so, especially in the presence of even normal concentrations of ascorbic acid. At pH 8, increases in measured oxalate ranged from 20% at an ascorbic acid concentration of 1 mmol/L to more than 300% at 15 mmol/L. Ascorbic acid is rapidly converted to oxalate at alkaline pH. We also investigated the stability of both untreated and acidified urine containing ascorbic acid during storage for up to one month at -70, -20, and 4 degrees C, and room temperature. After one month, untreated collections were stable at -70 degrees C and acidified collections at -20 and -70 degrees C. We recommend conditions for assay and storage of urine specimens that are to be assayed for oxalate under which positive interference by ascorbic acid is minimized.