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Electrospray tandem mass spectrometry was used to determine steady-state serum and urinary inorganic sulfate and sulfate ester kinetic profiles of nine normal men after intravenous injection of the stable isotope sodium [[sup.34]S]sulfate. Sulfate ester appearance was traced by eliminating inorganic sulfate from samples, followed by hydrolysis of sulfate esters to inorganic sulfate for analysis. Whole body inorganic sulfate turnover in steady state was calculated using standard tracer techniques. Rate of appearance and disappearance of inorganic sulfate was 841 [+ or -] 49 [micro]mol/h. Average urinary inorganic sulfate excretion was 609 [+ or -] 41 [micro]mol/h, and the whole body sulfation rate (total rate of disappearance minus rate of urinary excretion) was 232 [+ or -] 36 [micro]mol/h. Tracer-labeled sulfate esters appeared in serum and urine within 1 h of tracer injection. The kinetics of inorganic sulfate and sulfate esters were linked by means of a compartmental model. The appearance and excretion of sulfate esters accounted for ~50% of the total sulfation rate. These results indicate that human whole body sulfation accounts for ~27% of inorganic sulfate turnover and that extracellular inorganic sulfate is an important pool for intracellular sulfation. A substantial fraction of newly synthesized sulfate esters promptly enters the extracellular space for excretion in the urine. inorganic sulfate; stable isotopes; sulfate esters; sulfation

Abstract Background: End-stage renal disease (ESRD) is associated with moderately severe hyperhomocysteinemia that is incompletely normalized by oral folic acid therapy and vitamin [B.sub.12]. Method: We administered 1 mg hydroxocobalamin [...]

We evaluated the use of urinary sulfur (S) excretion as a measure of sulfur amino acid (SAA) catabolism and the nitrogen/sulfur (N/S) molar balance ratio as an indicator of nonprotein SAA storage in growing piglets. After confirming that an intravenous dose of sulfate is fully recovered in urinary sulfate, we measured urinary S recovery after an intravenous dose of methionine in 6 piglets fed an adequate protein (AP) diet and 6 piglets fed a low protein (LP) diet with normal energy provision. As measured over 48 h, recoveries of the methionine load as urinary total S was 106% in the AP group but only 69% in the LP group (P < 0.05). On the baseline diets the N/S balance ratio in the AP group was 36, whereas that in the LP group was 30 (P < 0.05); immediately after the methionine load, this ratio remained constant in the AP group but decreased further, to 26 (P < 0.05) in the LP group. These results indicate that protein-deficient piglets accumulate relatively more S than N from their diet, and under these conditions a significant portion of the S derived from a methionine load is retained in nonprotein compounds. Urinary S excretion, a simple nontracer measurement, can provide an accurate measure of SAA catabolism, and the N/S balance ratio is a potentially useful indicator of changes in nonprotein SAA stores of growing piglets. KEY WORDS: * glutathione * methionine * protein restriction * sulfate * taurine

Persons with conventionally treated insulin-dependent diabetes mellitus (IDDM) appear to be impaired in their ability to reduce fed-state urea production appropriately in response to dietary protein restriction (Hoffer LJ, Taveroff A, and Schiffrin A. Am J Physiol 272: E59-E67, 1997). To determine whether these conclusions apply to whole body sulfur amino acid (SAA) catabolism, we used samples from this protocol to measure daily urinary sulfate excretion and fed-state sulfate production after a high-protein test meal before and after dietary protein restriction. Eight normal subjects and six IDDM subjects treated with twice-daily intermediate- and short-acting insulin consumed a mixed test meal containing 0.50 g protein/kg after adaptation to 4 days of high protein intake (1.28 g protein/kg body wt) and again after 5 days of dietary protein restriction (0.044 g/kg). Adaptation to protein restriction decreased daily urinary sulfate and urea-N excretion by ~80%. Over the first 24 h of protein restriction, urinary sulfate excretion decreased more than urea-N excretion for both the normal and IDDM subjects. Under conditions of a high prior protein intake, fed-state sulfate production was normal for the IDDM subjects; protein restriction reduced fed-state sulfate production by 51% (normal subjects) and 59% (IDDM subjects; not significant). We conclude that whole body SAA metabolism is normal in conventionally treated IDDM before and after dietary protein restriction. SAA catabolism, as measured by fed-state sulfate production, may be a convenient and useful method to determine the extent of whole body protein dysregulation in IDDM. adaptation; fed state; humans; sulfate; urea

The volume of human extracellular water (ECW) may be estimated from the sulfate space (SS). Although it may better approximate ECW volume than the bromide space, a common alternative, SS measurement is limited by the need to administer a radioactive substance, sodium [35S]sulfate. In this paper, we demonstrate the measurement of the SS using the stable isotope, sodium [34S]sulfate. Eight healthy nonobese men ingested 0.50-0.78 mg (3.47-5.42 [Mu]mol) [Na.sub.2]34S[O.sub.4]/kg body weight and 30 mg NaBr/kg body weight. Sulfate concentrations and 34S[O.sub.4] enrichments were measured by electrospray tandem mass spectrometry before and during the 5 h after tracer administration. SS was calculated by linear extrapolation of the natural logarithm of serum 34S[O.sub.4] concentrations obtained at h 2, 3 and 4 compared with h 3, 4 and 5. The SS obtained using values between h 3 and 5 (187 [+ or -] 17 mL/kg) was similar to published determinations using intravenous or oral radiosulfate, and was 80% of the simultaneously measured corrected bromide space (234 [+ or -] 10 mL/kg, P = 0.01). Oral sodium [34S]sulfate administration is a suitable technique for measuring ECW and avoids radiation exposure. KEY WORDS: * humans * extracellular water volume * corrected bromide space * sulfate space * body composition * stable isotope

Although mild abnormalities of amino acid metabolism frequently exist in conventionally treated insulin-dependent diabetes mellitus (IDDM), their physiologic and nutritional importance is uncertain. We tested whether a tendency toward body N loss can be either masked or revealed in insulin-treated IDDM by changing the level of protein in the diet. After adaptation to a protein-free diet adequate in all other nutrients, obligatory urinary N excretion of intensively treated IDDM subjects was significantly greater than normal, indicating an impaired ability to recycle endogenous amino acids during protein restriction. When the preceding diet was high in protein, urea N production after consumption of a mixed test meal matched the amount of N consumed for both normal and diabetic subjects. However, when the test meal was preceded by 5 d of protein restriction, conventionally treated IDDM subjects failed to adaptively reduce postprandial urea production as effectively as normal or intensively treated IDDM subjects. Thus, even during insulin treatment, the ability to maximally recycle endogenous amino acids is impaired in IDDM, as is the ability to adaptively increase dietary amino acid retention in response to protein restriction. KEY WORDS: nutrition; malnutrition; urea; amino acids; fed state

The kinetic and metabolic effects of leucine was analyzed in normal human subjects that were infused with the amino acid isotope L-[5,5,5-H3]leucine, L-[1-13C]valine and L-[1-13C]threonine. Amino acid isotope infusions in normal human volunteers reduced plasma leucine enrichment and leucine [alpha-ketoisocaproic acid]. Furthermore, the difference between plasma and intracellular leucine enrichments decreased with the expansion of the amino acid pool.

The effects of diabetes on dietary amino acid retention were analyzed in normal and diabetic subjects after a high protein diet followed by a mixed test meal (MTM) and after a protein-deficient diet. Postprandial urea production was reduced in normal subjects and insulin-dependent diabetes mellitus patients under extensive insulin treatment after a five day regimen of protein-deficient diet preceded by MTM. Furthermore, diabetic subjects exhibited higher rates of protein catabolism due to energy deficiency and suboptimal insulin provision.

The relationship between dietary protein content and glutathione (GSH) metabolism in rats was evaluated. Rats fed high protein diets did not have increased liver GSH content, plasma GSH turnover, or plasma methionine and cysteine contents as compared to controls acclimated to 20% casein-based purified diets. A sigmoidal relationship between total GSH flux and liver GSH content was shown. Noninvasive blood sampling of the various parameters showed that the best indirect predictor of liver GSH content was plasma cysteine levels.

One feature of the adaptation to dietary protein restriction is reduced urea production over the hours after consumption of a test meal of fixed composition. This adaptation is impaired in conventionally treated insulin-dependent diabetes mellitus (Hoffer LJ, Taveroff A, and Schiffrin A. Am J Physiol Endocrinol Metab 272: E59-E67, 1997). We have now tested the response to a test meal containing less protein and included as a main outcome variable the production of sulfate, a specific indicator of sulfur amino acid catabolism. Six normal men consumed a mixed test meal containing 0.25 g protein/kg and 10 kcal/kg while adapted to high (1.5 g [multiplied by] [kg.sup.-1] [multiplied by] [day.sup.-1]) and low (0.3 g [multiplied by] [kg.sup.-1] [multiplied by] [day.sup.-1]) protein intakes. They followed the identical protocol twice. Six subjects with insulin-dependent diabetes consumed the test meal while adapted to their customary high-protein diet. Adaptation to protein restriction reproducibly reduced 9-h cumulative postmeal urea N and S production by 22-29% and 49-52%, respectively (both P [is less than] 0.05). Similar results were obtained for a postmeal collection period of 6 h. The response of the diabetic subjects was normal. We conclude that reductions in postmeal urea and sulfate production after protein restriction are reproducible and are evident using a postmeal collection period as short as 6 h. Sulfate production effectively depicts fed-state adaptation to protein restriction. urea; stable isotope; fed state; amino acid oxidation

Six normal men consumed a mixed test meal while adapted to high (1.5 g [multiplied by] [kg.sup.-1] [multiplied by] [day.sup.-1]) and low (0.3 g [multiplied by] [kg.sup.-1] [multiplied by] [day.sup.-1]) protein intakes. They completed this protocol twice: when the test meals included 3 mg/kg of [sup.15N]alanine ([[sup.15N]]Ala) and when they included 30 mg/kg of intrinsically labeled [[sup.15N]]Spirulina platensis ([[sup.15N]]SPI). Six subjects with insulin-dependent diabetes mellitus (IDDM) receiving conventional insulin therapy consumed the test meal with added [[sup.15N]]Ala while adapted to their customary high-protein diet. Protein restriction increased serum alanine, glycine, glutamine, and methionine concentrations and reduced those of leucine. Whether the previous diet was high or low in protein, there was a similar increase in serum alanine, methionine, and branched-chain amino acid concentrations after the test meal and a similar pattern of [sup.15N] enrichment in serum amino acids for a given tracer. When [[sup.15N]]Ala was included in the test meal, [sup.15N] appeared rapidly in serum alanine and glutamine, to a minor degree in leucine and isoleucine, and not at all in other circulating amino acids. With [[sup.15N]]SPI, there was a slow appearance of the label in all serum amino acids analyzed. Despite the different serum amino acid labeling, protein restriction reduced the postmeal transfer of dietary [sup.15N] in [sup.15N]Ala or [[sup.15N]]SPI into [[sup.15N]]urea by similar amounts (38 and 43%, respectively, not significant). The response of the subjects with IDDM was similar to that of the normal subjects. Information about adaptive reductions in dietary amino acid catabolism obtained by adding [[sup.15N]]Ala to a test meal appears to be equivalent to that obtained using an intrinsically labeled protein tracer. humans; stable isotope; fed state; amino acid oxidation