U. Langenbeck

Correlations between branched-chain amino acids and branched-chain α-keto acids in blood in maple syrup urine disease

In 62 blood samples from 3 patients with classical maple syrup urine disease and from one patient with a variant form, a close linear correlation was found between levels of branched chain amino acids and their corresponding alpha-keto acids. Keto acids were determined as O-trimethylsilyl quinoxalinols by gas chromatography with a nitrogen-selective detector.

Restriction fragment length polymorphisms at the human parathyroid hormone gene locus

SummaryTwo common Pst I and Taq I restriction enzyme fragment length polymorphisms (RFLPs) were detected at the human parathyroid hormone (PTH) gene locus. The allele frequencies in a Northern German population were 0.578/0.422 (Pst I) and 0.628/0.372 (Taq I). The allele distributions follow Hardy-Weinberg expectations of equilibrium in the population. The Mendelian nature of the polymorphisms were confirmed in family studies.

A familial progressive neurodegenerative disease with 2-oxoglutaric aciduria

A boy and a girl born to a consanguineous Tunisian couple are suffering from a slowly progressive nervous disorder. Initially they both had normal psychomotor development with acquisition of gait and speech. First symptoms in the boy were athetoid movements during the second year of life. He later lost all motor and language skills and developed muscular rigidity and intention tremor. At the age of five years, he was completely bedridden while he appeared mentally much less affected. His younger sister followed a similar course. The major specific abnormality detected was a strikingly elevated excretion of 2-oxoglutaric acid, which was identified by gas liquid chromatography, mass spectrometry, and enzymatic analysis. 2-oxoglutarate dehydrogenase activity in homogenates of cultured skin fibroblasts was reduced to about 25% of control values in both children. Although the pathogenetic mechanisms leading to brain damage remain obscure, the finding strongly suggest an autosomal recessive neurometabolic disease with predominant involvement of the extrapyramidal system.

Maple syrup urine disease-therapeutic use of insulin in catabolic states

High and neurotoxic blood levels of leucine and its ketoanalogue develop in catabolic patients with maple syrup urine disease. The use of relatively high doses of insulin and additional glucose had a more pronounced effect on lowering leucine (and α-ketoisocaproate) blood levels than dietary elimination of leucine alone. This is demonstrated in 2 neonates after blood exchange transfusion and in one 4-months old patient suffering from febrile diarrhea.

Mitochondrial myopathy with lactic acidosis and deficient activity of muscle succinate cytochrome-c-oxidoreductase.

A male infant had severe muscular hypotonia from birth. Recurrent vomiting with dehydration and severe metabolic acidosis complicated the course. Elevated lactate (up to 12.3 mmol/l; n<2), pyruvate (0.4 mmol/l; n<0.05) and alanine levels were found in serum with an abnormal lactate/pyruvate ratio (>30; n<15). In urine the concentrations of lactate, pyruvate, alanine and of several intermediates of the citric acid cycle were increased. In muscle, numerous disseminated “ragged red fibres” were found by light microscopy; muscle fibres were found to contain subsarcolemmal aggregates of mitochondria, lipid droplets and glycogen by electromicroscopical methods. More-over, mitochondria with a typical circular arrangement of cristae were noticed.In liver homogenates normal activities of pyruvate carboxylase and pyruvate dehydrogenase complex were found; in liver mitochondria also succinate-cytochrome-c-oxidoreductase activity was normal. However, in muscle no succinate-cytochrome-c-oxidoreductase activity was detectable.The patient became increasingly lethargic and died because of sepsis at 5 months of age.

Influence of insulin on blood levels of branched chain keto and amino acids in man

Branched chain keto acids, their corresponding amino acids, glucose, glucagon, growth hormone, C-peptide and gastric inhibitory polypeptide were determined in 8 healthy subjects after an intravenous bolus injection of 0.1 U/kg insulin. Branched chain keto acids declined within 60 min, the corresponding amino acids within 20 min or later. Amino acids tended to return towards normal earlier than their keto acids. Blood glucose levels were normal 2 hr after insulin injection while keto and amino acids remained diminished for more than 3 hr. In 8 healthy controls, given physiological saline instead of insulin, the branched chain keto acids did not decline throughout the test. It is suggested that insulin diminishes blood levels of branched chain keto acids, that the intraorgan flux of branched chain keto acids is different from the flux of branched chain amino acids and that branched chain keto acids may serve to correct for hypoglycemia.

Peritoneal dialysis in maple-syrup-urine disease: studies on branched-chain amino and keto acids.

We report biochemical data on a child with MSUD who underwent peritoneal dialysis for severe metabolic imbalance. In confirmation of earlier data, the BCKA/BCAA ratios in blood had been found to be fairly stable in this patient during long-term dietary therapy.The child became comatose at comparatively low levels of leucine and KICA (ca. 2 mM each). At this time the blood/cerebrospinal fluid ratio for BCAAs and BCKAs was markedly diminished. During peritoneal dialysis, peritoneal clearance was highest for KIVA, but less for MEVA and BCAAs (40–50% or urea clearance), and least for the allegedly most toxic metabolite, KICA. The differences for BCKAs may be due to their differential protein binding. Given these individual differences, 1.8 to 8.7 initial plasma volumes were cleared in 14h with 24.21 of dialysis fluid. In the same time, urinary excretion of BCAAs and BCKAs was much less efficient.The data are discussed with regard to the pathobiochemical significance of high tissue levels of branched chain acids. A quantitative comparison between peritoneal dialysis and exchange transfusion is not yet possible.

O-trimethylsilylquinoxalinol derivatives of aromatic α-keto acids : Mass spectra and quantitative gas chromatography

As an extension of earlier work on aliphatic α-keto acids, a method is described for the quantitative gas chromatographic determination of urinary aromatic α-keto acids. The keto acids are derivatized with o-phenylenediamine to yield the quinoxalinols. These compounds are chromatographed after trimethylsilyation. The aromatic keto acids are stabilized by sodium dithionite (4 mg/ml urine) and storage below 0°. The final derivatives are stable for weeks at room temperature. Low resolution mass spectra are reported. The fragmentation mechanims are elucidated by analysis of O-trimethylsilyl-(TMS)-quinoxalinols, O-(TMS-d9)-quinoxalinols and O-TMS-6(7)-chloroquinoxalinols.

Serum branched chain amino and keto acid response to fasting in humans.

Eight healthy individuals were fasted for 72 hours. The concentrations of the branched chain keto acids (BCKA), branched chain amino acids (BCAA), C peptide, and glucagon were determined in peripheral venous blood. alpha-ketoisocaproic acid, alpha-keto-beta-methyl-n-valeric acid, and alpha-ketoisovaleric acid increased significantly within 36 hours along with the corresponding amino acids. After 60 hours of starvation, the concentrations of BCKA and BCAA declined despite the fact that the subjects were still in the fasting state. These changes were accompanied by a decrease in the concentrations of C peptide and an increase in glucagon levels. It is suggested that in starving man insulinopenia may contribute to the rise in BCKA concentrations and that the increase in BCKA may be a mechanism to reduce proteolysis.

Blood levels of branched-chain alpha-keto acids in uremia: Therapeutic implications.

ZusammenfassungDie Serumkonzentrationen verzweigtkettiger Ketosäuren (BCKAs), d.h. α-Ketoisocapronsäure (KICA), α-Keto-isovaleriansäure (KIVA) und α-Keto-β-methyl-n-valeriansäure (KMVA) sowie ihrer korrespondierenden Aminosäuren wurden bei Urämikern mit kompensierter chronischer Niereninsuffizienz sowie bei hämodialysierten Urämikern und bei nierengesunden Kontrollen bestimmt.Urämiker hatten signifikant niedriger BCKA-Konzentrationen im Serum als nierengesunde Kontrollen. Es bestand eine negative Korrelation zwischen der BCKA-Konzentration und derjenigen von Kreatinin bzw. von Harnstoff. BCKAs ließen sich im Hämofiltrat nachweisen. Die Konzentrationen von KICA und KMVA waren signifikant höher, diejenige von KIVA identisch verglichen mit den entsprechenden Konzentrationen im Hämofiltrat. Daraus läßt sich eine unterschiedliche Eiweißbindung der BCKAs vermuten. Orale Einnahme von 5 g Ca-KICA führte bei einer gesunden Versuchsperson zu einem vorübergehenden, deutlichen Anstieg von Serum-KICA und Leuzin. Der maximale Anstieg der KICA trat früher ein als das Leuzinmaximum.Die jeweils 28tägige Gabe eines Gemisches verzweigtkettiger Keto- und Aminosäuren (6–9 g/Tag) mit oder ohne Eiweißbeschränkung der Kost auf 0,5 g/kg/Tag führte bei Patienten mit kompensierter chronischer Niereninsuffizienz zu keiner Änderung der BCKA-Konzentrationen im Serum.Die Bestimmung verzweigtkettiger Ketosäuren im Blut bietet neue Möglichkeiten zur Erforschung des Stickstoffmetabolismus sowie zur Überprüfung des Stellenwerts von Ketosäuren bei der Behandlung von Patienten mit chronischer Niereninsuffizienz.SummarySerum levels of branched-chain keto acids (BCKAs), i.e., α-keto-isocaproic acid (KICA), α-keto-isovaleric acid (KIVA) and α-keto-β-methyl-n-valeric acid (KMVA) as well as their corresponding amino acids were determined in uremic patients with compensated chronic renal failure, patients on hemodialysis, and in subjects without renal insufficiency.Uremic patients had significantly lower BCKA levels than controls without renal insufficiency. There was a negative correlation between serum BCKAs and the levels of blood urea and creatinine. BCKAs were detectable in the hemofiltrate. The concentrations of KICA and KMVA were significantly higher, that of KIVA identical compared to the respective concentrations in the hemofiltrate. This suggests a different protein binding of BCKAs. Oral administration of 5 g Ca-KICA to a healthy subject resulted in a transient increase in serum KICA and leucine. The maximum increase in KICA preceded the leucine peak.Serum BCKA levels did not change significantly in patients with compensated renal failure, who were — for 28 days each — first on an unrestricted diet plus supplementation, then solely on an unrestricted diet, followed by a protein-restricted diet (0.5 g/kg/day) plus supplementation and finally on a protein-restricted diet alone. Supplementation was with 5 essential amino acids, 4 keto acids and 1 hydroxyacid (6–9 g/day).The determination of BCKAs in serum offers a promising investigatory tool to study nitrogen metabolism in healthy and uremic subjects and might help to further evaluate the role of keto acids in the treatment of chronic renal failure.