Whole-body protein turnover in nephrotic subjects: a comparison between different degrees of proteinuria

In contrast to the increased hepatic albumin synthesis in subjects with proteinuria, little is known about the corresponding whole-body protein turnover rates (WPTR). The WPTR and the reutilisation rates (R) were investigated in 20 patients divided in three groups of different degrees of proteinuria (groups I-III: < 1, 1-3, > 3 g/m2/day, respectively). [15N]glycine was administered as a single oral pulse. Urine samples were taken over 2 days. After removing urinary proteins by trichloracetic acid, 15N-enrichment in the supernatant was measured by isotope ratio mass spectrometry. A three-compartment model was used to calculate WPTR and R, which showed a statistically significant difference between groups I and III (2.64 vs. 4.63 g/kg/day, and 70.4% vs. 80.8%, respectively, P < 0.01), whereas the net protein gain remained unchanged (0.13 vs. 0.22 g/kg/day). The higher the protein loss the higher the WPTR and the corresponding R. The severe protein loss provokes increased WPTR and R as well.     

Development and application of 15N-tracer substances for measuring the whole-body protein turnover rates in the human,especially in neonates: a review

Our research group of the Children's Hospital of the University of Rostock (Rostock group) has long-time experience in ¹⁵N-labelling and in using yeast protein and its hydrolysates for tracer kinetic studies to evaluate parameters of the whole-body protein metabolism in premature infants. The particular advantage of applying an economically convenient, highly ¹⁵N-enriched, and completely labelled yeast protein for evaluating protein turnover rates is the fact that the ¹⁵N dose is spread among all proteinogenic amino acids. The absorption has been improved by hydrolysing [¹⁵N]yeast protein with thermitase into a mixture of amino acids, dipeptides and tripeptides so that faecal analysis becomes unnecessary when determining turnover rates. The review shows that, in contrast to the application of single ¹⁵N-labelled amino acids with resulting overestimation of protein turnover rates, the ¹⁵N-labelled yeast protein thermitase hydrolysate represents the amino acid metabolism more closely without causing amino acid imbalances. The ¹⁵N-labelled yeast protein thermitase hydrolysate leads to the estimation of reliable protein turnover rates, particularly in premature infants.     

The Measurement of Muscle Protein Synthesis in Broilers with a Flooding Dose Technique: Use of 15N-Labelled Phenylalanine,GC-MS and GC-C-IRMS

Abstract

An experiment was carried out to measure fractional muscle protein synthesis rates (k _s ) in broilers with injection of a flooding dose of phenylalanine (1 ml/100 g body weight of 150 mM phenylalanine; 38 atom percent excess (APE) [¹⁵N]phenylalanine). K _s was calculated from the [¹⁵N] enrichment in phenylalanine of tissue-free and protein-bound phenylalanine using both gas chromatography mass spectrometry (GC-MS) and gas chromatography combustion isotope ratio mass spectrometry (GC-C-IRMS) for measurements after a 10 min isotope incorporation period.

The tertiary-butyldimethylsilyl (t-BDMS) derivatives of phenylalanine were used for gas chromatographic separation in both systems. GC-MS and GC-C-IRMS were calibrated for a range of 7 to 37 [¹⁵N]APE and 0 to 0.62 [¹⁵N]APE, respectively, and for sample sizes of 0.45 to 4.5 nmol phenylalanine and 7 to 40 nmol phenylalanine, respectively. Reproducibility of standards as a measure of precision varied from 0.06 to 0.29 [¹⁵N]APE and from 0.0004 to 0.0018 [¹⁵N]APE in GC-MS and GC-C-IRMS, respectively.

K _s was measured in the m. pectoralis major of broilers fed rye based diets (56%) which were provided either unsupplemented (-) or supplemented (+) with an enzyme preparation containing xylanase. K _s in breast muscles was significantly increased from 21.8%/d to 23.9%/d due to enzyme supplementation.

It can be concluded from the study that the measurement of protein synthesis in broilers with the flooding dose technique can be carried out by using [¹⁵N]phenylalanine, GC-MS and GC-C-IRMS.