Urinary Organic Acid Excretion Research Articles

3-Hydroxy-3-Methylglutaric Acid Impairs Redox and Energy Homeostasis, Mitochondrial Dynamics, and Endoplasmic Reticulum-Mitochondria Crosstalk in Rat Brain.

3-Hydroxy-3-methylglutaryl-CoA lyase (HL) deficiency is a neurometabolic disorder characterized by predominant accumulation of 3-hydroxy-3-methylglutaric acid (HMG) in tissues and biological fluids. Patients often present in the first year of life with metabolic acidosis, non-ketotic hypoglycemia, hypotonia, lethargy, and coma. Since neurological symptoms may be triggered or worsened during episodes of metabolic decompensation, which are characterized by high urinary excretion of organic acids, this study investigated the effects of HMG intracerebroventricular administration on redox homeostasis, citric acid cycle enzyme activities, dynamics (mitochondrial fusion and fission), and endoplasmic reticulum (ER)-mitochondria crosstalk in the brain of neonatal rats euthanized 1 (short term) or 20 days (long term) after injection. HMG induced lipid peroxidation and decreased the activities of glutathione peroxidase (GPx) and citric acid cycle enzymes, suggesting bioenergetic and redox disruption, 1 day after administration. Levels of VDAC1, Grp75, and mitofusin-1, proteins involved in ER-mitochondria crosstalk and mitochondrial fusion, were increased by HMG. Furthermore, HMG diminished synaptophysin levels and tau phosphorylation, and increased active caspase-3 content, indicative of cell damage. Finally, HMG decreased GPx activity and synaptophysin levels, and changed MAPK phosphorylation 20 days after injection, suggesting that long-term toxicity is further induced by this organic acid. Taken together, these data show that HMG induces oxidative stress and disrupts bioenergetics, dynamics, ER-mitochondria communication, and signaling pathways in the brain of rats soon after birth. It may be presumed that these mechanisms underlie the onset and progression of symptoms during decompensation occurring in HL-deficient patients during the neonatal period.

Selective Intestinal Cobalamin Malabsorption with Proteinuria (Imerslund‐Gräsbeck Syndrome) in Juvenile Beagles

BackgroundSelective intestinal cobalamin malabsorption with mild proteinuria (Imerslund‐Gräsbeck syndrome; I‐GS), is an autosomal recessive disorder of dogs caused by mutations in AMN or CUBN that disrupt cubam function and which can present as a medical emergency.ObjectivesTo describe the clinical, metabolic, and genetic bases of I‐GS in Beagles.AnimalsFour cobalamin‐deficient and 43 clinically normal Beagles and 5 dogs of other breeds.MethodsClinical description and candidate gene genetic study. Urinary organic acid and protein excretion were determined by gas‐chromatography and SDS‐PAGE, respectively. Renal cubilin protein expression was assessed on immunoblots. Mutation discovery was carried out by PCR amplification and DNA sequencing of exons with flanking splice sites and cDNA of CUBN and AMN. Genotyping was performed by restriction enzyme digestion of PCR amplicons.ResultsJuvenile‐affected Beagles exhibited failure to thrive, dyshematopoiesis with neutropenia, serum cobalamin deficiency, methylmalonic aciduria, hyperammonemia, and proteinuria. Affected dogs' kidneys lacked detectable cubilin protein. All affected dogs were homozygous for a single‐base deletion in CUBN exon 8 (CUBN c.786delC), predicting a translational frameshift, and the 2 parents tested were heterozygous.ConclusionsThe CUBN mutation in juvenile I‐GS Beagles causes a more severe cobalamin malabsorption than in Border Collies with a different CUBN defect, but is similar to I‐GS caused by AMN mutations in Giant Schnauzers and Australian Shepherds. Awareness of the disorder and breed predispositions to I‐GS is crucial to precisely diagnose and promptly treat hereditary cobalamin malabsorption and to prevent disease in those dogs at risk in future generations.

Identification and assessment of markers of biotin status in healthy adults

Human biotin requirements are unknown and the identification of reliable markers of biotin status is necessary to fill this knowledge gap. Here, we used an outpatient feeding protocol to create states of biotin deficiency, sufficiency and supplementation in sixteen healthy men and women. A total of twenty possible markers of biotin status were assessed, including the abundance of biotinylated carboxylases in lymphocytes, the expression of genes from biotin metabolism and the urinary excretion of biotin and organic acids. Only the abundance of biotinylated 3-methylcrotonyl-CoA carboxylase (holo-MCC) and propionyl-CoA carboxylase (holo-PCC) allowed for distinguishing biotin-deficient and biotin-sufficient individuals. The urinary excretion of biotin reliably identified biotin-supplemented subjects, but did not distinguish between biotin-depleted and biotin-sufficient individuals. The urinary excretion of 3-hydroxyisovaleric acid detected some biotin-deficient subjects, but produced a meaningful number of false-negative results and did not distinguish between biotin-sufficient and biotin-supplemented individuals. None of the other organic acids that were tested were useful markers of biotin status. Likewise, the abundance of mRNA coding for biotin transporters, holocarboxylase synthetase and biotin-dependent carboxylases in lymphocytes were not different among the treatment groups. Generally, datasets were characterised by variations that exceeded those seen in studies in cell cultures. We conclude that holo-MCC and holo-PCC are the most reliable, single markers of biotin status tested in the present study.

DIFFERENTIAL DIAGNOSIS OF ORGANIC ACIDEMIA: CLINICAL AND NEUROIMAGING FINDINGS

Clinical differential DiagnosisThe organic acidemias are important in the differential diagnosis of metabolic and neurologic derangement in the neonate and of new-onset neurologic signs in the older child. A-Organic aciduriaSeveral disorders, not classified as primary disorders of organic acid metabolism, have a characteristic urinary organic acid profile that suggests the appropriate diagnosis. • Mevalonicaciduria, a disorder of cholesterol biosynthesis, shows mevalonic acid in the urine. • Glutaricacidemia type II, a disorder of fatty acid oxidation, has multiple organic acids in abnormal concentration in urine. These organic acids include ethylmalonic acid, glutaric acid, dicarboxylic acids, and glycine conjugates of medium chain dicarboxylic acids. • The fatty acylCoA-glycine conjugates that signal incomplete fatty acid oxidation and serves as signals to the diagnosis of MCAD defeciency and other disorders of fatty acid oxidation and transport. • Biotinidase deficiency, a disorder of biotin recycling, results in the urinary excretion of several unusual organic acids, including 3-hydroxy-isovaleric, 3-hydroxypropionic, 3-hydroxybutyric acids, and acetoacetate. Propionyl glycine may also be seen. • Mitochondrial diseases with disordered oxidative phosphorylation often demonstrate the presence of abnormal organic acids in the urine. B-AcidosisNon-genetic conditions, such as shock, sepsis, DKA, liver and kidney failure, thiamine deficiency, RTA, some drug intoxication cause acidosis- genetic conditions are include: inherited metabolic disorders of lactate and pyruvate metabolism and oxidative phosphorylation, disorders of the Krebs cycle such as fumarase deficiency. C-HyperammonemiaDisorders of the urea cycle and the hyperammonemia-hypoglycemia syndrome.Neuroimaging• A variety of MRI abnormalities have been described in the organic academia, including distinctive basal ganglia lesions in glutaricacidemia type I (GA I), white matter changes in maple syrup urine disease (MSUD), and abnormalities of the globus pallidus in methylmalonic acidemia. Macrocephaly is common in GA I. • Some differential agnosis of MRI findings in organic academia is consist of: HIE, mucopolysacaridosis, middle fossa arachnoid cyst, leighdisease, hexachlorophene toxicity in neonates, myelin splitting disorders. • Some organic aciduria such as L-2-Hydroxyglutaricaciduria may suggest leukodystrophy in MRI.

Infantile nephropathic cystinosis

CASE PRESENTATION A Caucasian girl, the product of a non-consanguineous union, was delivered after a full-term uneventful pregnancy with a birth weight of 4.1 kg. At 15 months, she presented to the emergency room with signs of a respiratory illness and was found to weigh 6.14 kg, below the 3rd percentile for age. She had no history of fevers, dehydration, or photophobia and there was no family history of renal or other medical disease. The patient had one elder sister who had no medical complaints. Physical examination revealed a fair-haired blue-eyed child with pale complexion. Details of her ethnic ancestry were unknown. Blood pressure was normal and the remainder of the physical examination was also unremarkable. Laboratory investigations at 15 months revealed blood urea nitrogen 9 mg per 100 ml (normal range (N)1⁄4 8–25), serum creatinine 0.6 mg per 100 ml (estimated creatinine clearance o60 ml min ), serum sodium 133 mmol l 1 (N1⁄4 136–145), potassium 3 mmol l 1 (N1⁄4 3.5–5.5), chloride 105 mmol l 1 (N1⁄4 96–106 mEq l ), total carbon dioxide (CO2) 18.7 mEq l 1 (N1⁄4 22–30), serum calcium 10.6 mg per 100 ml (N1⁄4 8.5–10.6 mg per 100 ml), and phosphate 3.4 mg per 100 ml (N1⁄4 2.5–4.6). Arterial blood gas was not drawn. Urinalysis revealed specific gravity o1.005, no protein or cells, and trace glycosuria. However, there was no evidence of phosphaturia (24 h urinary phosphorous excretion 0.2 mmol, N1⁄4 0.4–1.3), hypercalciuria, or increased urinary organic acid excretion. Measurement of urinary amino acids was not performed. Hematologic parameters and thyroid function tests, including thyroid stimulating hormone, T4, and free T4, were within normal limits. Based on the finding of hypokalemia, an investigation for possible Bartter’s syndrome was undertaken. These tests revealed markedly elevated plasma levels of renin (119 853 ng per 100 ml per h, N1⁄4 171–115) and aldosterone (715 ng per 100 ml, N1⁄4 3–35) that, together with the findings of normal blood pressure, hypokalemia, and hyponatremia, suggested a diagnosis of acquired Bartter’s syndrome. The patient was treated for the next two and a half years with indomethacin and electrolyte replacement, including potassium chloride and calcium carbonate. During this time, she gained weight and normalization of blood electrolytes. However, at 4 years of age, she was noted to have worsening renal insufficiency (serum creatinine 1.2–1.4 mg per 100 ml) and 4þ proteinuria on dipstick. At this time, an elevated thyroid stimulating hormone was noted (17.27 IU ml , N1⁄4 0.3–5.0), indicating hypothyroidism, and thyroid replacement therapy was commenced. Indomethacin was discontinued but renal function and proteinuria continued to deteriorate over the following year. At age 5 years, serum creatinine was 2.9 mg per 100 ml, urinary protein excretion was 1.8 g per 24 h, serum albumin was 3.4 g per 100 ml (normal42.5 g per 100 ml), and blood pressure was 80/50 mm Hg. A renal biopsy was performed.

Hematologic and Urinary Excretion Anomalies in Patients with Chronic Fatigue Syndrome

Patients with chronic fatigue syndrome (CFS) have a broad and variable spectrum of signs and symptoms with variable onsets. This report outlines the results of a single-blind, cross-sectional research project that extensively investigated a large cohort of 100 CFS patients and 82 non fatigued control subjects with the aim of performing a case-control evaluation of alterations in standard blood parameters and urinary amino and organic acid excretion profiles. Blood biochemistry and full blood counts were unremarkable and fell within normal laboratory ranges. However, the case-control comparison of the blood cell data revealed that CFS patients had a significant decrease in red cell distribution width and increases in mean platelet volume, neutrophil counts, and the neutrophil-lymphocyte ratio. Evaluation of the urine excretion parameters also revealed a number of anomalies. The overnight urine output and rate of amino acid excretion were both reduced in the CFS group (P < 0.01). Significant decreases in the urinary excretion of asparagine (P < 0.0001), phenylalanine (P < 0.003), the branch chain amino acids (P < 0.005), and succinic acid (P < 0.0001), as well as increases in 3-methylhistidine (P < 0.05) and tyrosine (P < 0.05) were observed. It was concluded that the urinary excretion and blood parameters data supported the hypothesis that alterations in physiologic homeostasis exist in CFS patients.

Riboflavin-responsive glutaryl CoA dehydrogenase deficiency

We report here riboflavin responsiveness in a patient with glutaryl CoA dehydrogenase (GCDH) deficiency, compound heterozygous for the S139L and P248L mutations and with 20% residual GCDH enzyme activity in vitro. Our results suggest the mitochondrial GCDH homotetramer remains intact with one of these mutations associated with the binding site of the single FAD cofactor and that pharmacological doses of the cofactor precursor may be sufficient to induce an increase in activity in the mutant GCDH enzyme, although not sufficient to normalise urinary organic acid excretion. Serine 139 is one of nine conserved amino acid residues that line the binding site of the protein and is in close proximity to both substrate and FAD cofactor. It is possible that steric alterations caused by substitution of serine with leucine at this position may be overcome with high cofactor concentrations. P248L is also associated with some residual GCDH activity in other patients and the unique combination of S139L with P248L may also explain the results in our patient. Responsiveness to riboflavin in our patient has been compared with two other patients with glutaric aciduria type 1 and minimal residual GCDH activity, one with homozygosity for the R257Q mutation and one with heterozygosity for the G354S mutation and a novel G156V mutation. A low lysine diet reduced glutaric acid excretion in our riboflavin-responsive GCDH-deficient patient almost to control values. She is now 21 years of age and clinically and neurologically normal.

Pharmacokinetics and Renal Excretion of Diphenhydramine and its Metabolites, Diphenylmethoxyacetic Acid and Diphenhydramine‐N‐Oxide, in Developing Lambs

The developmental disposition of diphenhydramine (DPHM) and its metabolites, diphenylmethoxyacetic acid (DPMA) and DPHM‐N‐oxide (DPHMNO), was investigated in postnatal lambs. Lambs received a DPHM intravenous (iv) bolus 15 days (Group A; n = 5) or 2 months (Group B; n = 6) after birth. Total body clearance of DPHM in postnatal lambs (Group A = 138.7 ± 80.5 mL/min/kg; Group B = 165.7 ± 51.3 mL/min/kg) was similar to the nonplacental clearance values (i.e., the component of fetal total body clearance that is not due to elimination via the placenta) estimated for fetal lamb (116.3 ± 49.6 mL/min/kg), and significantly greater than estimates in adult sheep (38.5 ± 12.3 mL/min/kg). In addition, Group A DPHM renal clearance (CLr, >1.80 ± 1.24 mL/min/kg) was similar to that of the fetus (2.06 ± 0.24 mL/min/kg), and significantly greater than that for Group B (0.26 ± 0.17 mL/min/kg) and the adult (0.012 ± 0.005 mL/min/kg). In contrast, similar to the fetal situation, postnatal DPMA CLr (Group A = 0.02 ± 0.02 mL/min/kg; Group B = 0.05 ± 0.01 mL/min/kg) was significantly less than adult values (0.53 ± 0.19 mL/min/kg). Because DPMA is not sequentially metabolized in sheep, the lower CLr in postnatal lambs results in longer apparent elimination half‐lives of this metabolite (Group A = 90.4 ± 32.2 h; Group B = 13.13 ± 11.0 h) compared with that in the adult (2.9 ± 1.6 h). No age‐related difference in DPHMNO CLR was observed. Alterations in the CLr of DPHM and DPMA are likely related to differences in the rate of development of mechanisms (i.e., tubular secretion and reabsorption and glomerular filtration rate) involved in the urinary drug excretion of organic acids and bases. © 2000 Wiley‐Liss, Inc. and the American Pharmaceutical Association J Pharm Sci 89: 1362–1370, 2000

Formation of Propionate After Short-Term Ethanol Treatment and Its Interaction with the Carnitine Pool in Rat

Organic acidurias are genetic disorders of mitochondrial metabolism that lead to the accumulation in tissues and biological fluids of organic acids. It has been demonstrated that interaction of carnitine with the cellular CoA pool, through the production of acyl-carnitines, is potentially critical for maintaining normal cellular metabolism under conditions of impaired acyl-CoA use and that exposure of humans and other mammals to ethanol effects leads to impairment of mitochondrial function. The aim of the present study was to evaluate the role of ethanol on urinary excretion of short-chain organic acids and endogenous carnitines in rats. The data reported show that ethanol significantly increases urinary excretion of propionate, methylmalonate, as well as free acetate, butyrate, pyruvate, lactate, and β-hydroxybutyrate. Furthermore, the increased formation of propionate and methylmalonate was dependent on the dose of ethanol; did not require the metabolism of ethanol, as was shown in experiments with pyrazole treatment of ethanol rats; and appears to be mediated by beta-adrenergic mechanisms because propranolol almost completely suppresses propionate accumulation. Alcohol administration also increased excretion of specific acyl-carnitines, corresponding to the accumulating acyl groups, whereas excretion of free carnitine was significantly reduced, with respect to control values. The data presented indicate that the short-term ethanol administration is associated with increased excretion of selected organic acids. This study suggests that endogenous carnitine pool might play a role against the deleterious effects of accumulating short-chain organic acids.

Aminoaciduria resulting from vigabatrin administration in children with epilepsy

Vigabatrin (γ-vinyl gamma aminobutyric acid), a recently developed antiepileptic drug, has been extensively evaluated in the treatment of drug-resistant epilepsy. Several case reports demonstrated that vigabatrin affects urinary excretion of several amino and organic acids. Fourteen children were investigated for the presence of abnormal urinary amino acids before and after treatment with vigabatrin. All demonstrated increased urinary excretion of amino acids, particularly β-alanine, γ-aminobutyric acid, and β-amino-isobutyric acid while on vigabatrin, which were not detected when off medication. These results emphasize the importance of obtaining urine for metabolic evaluation before the administration of vigabatrin.

Riboflavin and riboflavin-derived cofactors in adolescent girls with anorexia nervosa

Thyroid hormones, riboflavin, riboflavin cofactors, and organic acids were assessed in girls with anorexia nervosa. The objective was to examine the effect of malnutrition and low thyroid hormone concentrations on erythrocyte and plasma riboflavin metabolism and their relation with urinary organic acid excretion. Seventeen adolescent girls with anorexia nervosa [body mass index (BMI; in kg/m2): 14.8 +/- 2.2] and 17 age-matched, healthy girls (control subjects; BMI: 20.5 +/- 2.2) took part in the feeding study. Erythrocyte and plasma riboflavin as well as riboflavin cofactors (flavin mononucleotide and flavin adenine dinucleotide) were assessed by HPLC, whereas urinary organic acids were assessed by gas chromatography-mass spectrometry. Anorectic patients who began a feeding program had higher erythrocyte riboflavin (3.5 +/- 2.2 compared with <0.1 nmol/mol hemoglobin; P < 0.001), lower plasma flavin adenine dinucleotide (57.8 +/- 18.5 compared with 78.5 +/- 54.3 nmol/L; P < 0.05), and higher urinary ethylmalonic acid (7.12 +/- 4.39 compared with 1.3 +/- 2.8 micromol/mmol creatinine; P < 0.001) and isovalerylglycine (7.65 +/- 4.78 compared with 3.8 +/- 0.9 micromol/mmol creatinine; P < 0.05) concentrations than did control subjects. Triiodothyronine concentrations were low and negatively correlated with plasma riboflavin concentrations (r = -0.69, P < 0.01). Not all patients showed improvements in these biochemical indexes after 30 d of refeeding. The low triiodothyronine concentrations observed in anorexia nervosa could alter the extent of riboflavin conversion into cofactors, thus leading to high erythrocyte riboflavin concentrations, low plasma flavin adenine dinucleotide concentrations, and high rates of ethylmalonic acid and isovalerylglycine excretion.

Urinary organic acids in infant malnutrition.

The metabolic derangements in severe protein-energy malnutrition (PEM) are only partially known, due to the limitations of blood collection in these patients. Urinary excretion of organic acids was studied by gas chromatography-mass spectrometry in 39 infants with four types of PEM: 1) upon hospital admission, as soon as eventual infections had been cleared, and salt and water deficits corrected, but before oral feeding was started; 2) after start of protein alimentation; 3) on the day of discharge. All of the patients showed an increased excretion of various organic acids at some point of their hospital stay, regardless of the clinical type of PEM. In nearly half of the malnourished children, results were suggestive of blocks in the pathways of propionate (15.4% with increased methylmalonate and 25.6% with 2-methylcitrate), of fatty acid beta-oxidation (30.8% with raised dicarboxylic acids with low or low normal 3-hydroxybutyrate), or of both pathways (12.8%). These abnormalities may have been caused by cofactor deficiencies (biotin, vitamin B12, riboflavin, carnitine, niacin). Dicarboxylic acids were excreted in high amounts since the initial sample, probably due to increased mobilization of fatty acids. Increased 2-methylcitrate and methylmalonate excretion was observed more frequently once patients started to be orally fed. The accumulation of potentially toxic acyl-CoA precursors of these compounds could contribute to the known clinical worsening of some malnourished infants after suddenly increased protein intake. Other less specific metabolites, such as 3-hydroxybutyrate, lactate, 4-hydroxyphenyllactate, fumarate, succinate, and 4-hydroxyphenylacetate, were also abnormally excreted in some patients. The analysis of urinary organic acids provides a new approach for the metabolic study of PEM and may have diagnostic and therapeutic implications.

Effects of hypoxia on urinary organic acid and hypoxanthine excretion in fetal sheep.

Severe birth asphyxia leads to a transient organic aciduria and increased hypoxanthine excretion. To investigate its origin and timing, we analyzed urine from 12 late gestation fetal sheep in utero subjected to moderately severe isocapnic hypoxia for 1 h. In six fetuses the carotid sinus nerves were cut to determine whether reflex peripheral vasoconstriction contributed to the changes in excretion. After a control period of 1 h, maternal inspired oxygen was reduced for 1 h so that fetal arterial oxygen tension fell significantly from 2.86 +/- 0.12 kPa (mean +/- SEM) to 1.55 +/- 0.04 kPa. The ewes were returned to normoxia, and monitoring was continued for 1 h. Fetal heart rate, arterial blood pressure, and femoral arterial blood flow (intact fetuses only) were recorded, and arterial pH, blood gases, and lactate were measured. Urine collected via a bladder catheter was analyzed for organic acids and hypoxanthine with gas chromatography-mass spectrometry. In intact fetuses, hypoxia increased excretion of hypoxanthine and several organic acids, notably lactic acid and intermediates of valine catabolism. Changes were apparent by 15 min, significant by 45 min, and maximal after reoxygenation. In denervated fetuses, there were small, significant, increases in organic acids and hypoxanthine by 45 min of hypoxia, but there was no surge in excretion posthypoxia. Hypoxia caused a large, significant, fall in femoral arterial blood flow in intact fetuses. We conclude that the extent of the reflex peripheral vasoconstriction, particularly in skeletal muscle, determines the amount of organic acid and hypoxanthine excretion and may explain similar biochemical disturbances after birth asphyxia. Urinary lactic acid measurement has a potential value for grading birth asphyxia.

Heterogeneity of symptomatology in two male siblings with thymine uraciluria

We report studies in two brothers found to have thymine uraciluria (McKusick 274270). The diagnosis was initially made during the neonatal period of the younger boy (Y.R.). He was born at term plus 10 days to first-cousin Asian parents, weighing 3.90kg (on the 10th centile). Twenty-one hours after delivery the baby was lethargic and feeding poorly. Tube feeding was necessary for the first three days of life due to poor suck. He had bilateral hydroceles and a large anterior fontanelle. Head circumference was 32.5 cm (below the 3rd centle) and length 52.5 cm (on the 90th centile). He was fel~ to have coarse facial features with a bulbous nose, held his neck in extension and had a feeble cry. Analysis of urinary organic acid excretion led to the discovery of thymine uraciluria. Y.R. has two older siblings, a 6-year-old sister (N.R.) and a 5-yearold brother (H.R.). Both are healthy and attend normal school. H.R., however, has been found to have the same biochemical defect as Y.R. He does not share any of the dysmorphisms and had a normal neonatal history. His head circumference of 53 cm lies between the 50th and 98th centiles for age. His language development appears normal, but he does have problems with phonology for which he is receiving speech therapy. Urinary purines from the siblings have subsequently been quantified by highpressure liquid chromatography. The proband had uracil and thymine excretions of 582 and 519/zmol/mmol creatinine, respectively. His older brother was found to have similarly elevated excretions at 313 and 170/.tmol/mmol creatinine, but the same metabolites were not increased in his 6-year-old sister (15 and 1/.tmol/mmol creatinine). The enzyme dihydropyrimidine dehydrogenase has been measured in leukocytes from Y.R. and his parents. There was no detectable activity in cells from the index case, whereas the parents had activities approximately half those of normal controls: father 2.76, mother 2.45, control 5.0 nmol/h per mg protein. Thymine uraciluria was first described by van Gennip (1981) in a 4-year-old boy with transient seizures, speech retardation and behavioural problems. Since that time, 19 other affected children have been reported, 13 of whom have been from Holland. There has been considerable clinical heterogeneity but with a predominance of neurological abnormality. The more severely affected children often have microcephaly. A number of affected adults have been described who are quite healthy and are detected either during family studies or following a severe neurotoxic reaction to the drug 5-fluorouracil. The enzyme dihydropyrimidine dehydrogenase is necessary for the metabolism of this chemotherapeutic drug. The observations in the family described here clearly illustrate that there can be considerable phenotypic variabilty, even within a family. This is also the first report of this condition among people of Asian origin, and the first case in the United Kingdom.

Impaired ketogenesis in patients with adult-type citrullinemia

Background/Aims: To clarify the mechanism causing fatty liver in adult-type citrullinemia, the effect of fasting on blood levels of free fatty acids, triglycerides, and ketone bodies was investigated in two cases. Methods: Blood and urine samples were collected from two patients and healthy volunteers 12, 15, 17, 21.5, and 24 hours after their last meal. Results: During 24-hour fasting free fatty acid concentrations increased in both cases to the concentrations found in the healthy volunteers. The levels of blood ketone bodies (β-hydoxybutyrate and acetoacetate) were markedly suppressed throughout the fasting test without any increase in urinary excretion of ketone bodies or organic acids in both cases when plasma citrulline concentrations were more than 10-fold higher than in controls. Serum triglyceride concentrations in case 1 paradoxically increased from 185 mg/dL to 294 mg/dL during 24-hour fasting when the citrulline concentration was extremely high. When hemodialysis was performed and plasma citrulline consequently decreased to near the normal level in case 1, levels of both serum triglycerides and blood ketone bodies responded normally to 24-hour fasting. Conclusions: These data suggest that ketogenesis was impaired in adult-type citrullinemia.

Biochemical investigation of a Brazilian patient with a defect in mitochondrial acetoacetyl‐coenzyme‐A thiolase

A case report of 3-ketothiolase deficiency due to a defect of mitochondrial acetoacetyl-CoA thiolase protein in a Brazilian boy and its biochemical investigation is presented. The child had moderate generalized hypotonia, EEG alterations and crises of metabolic acidosis following infections. Hypotonia and EEG abnormalities disappeared with a low protein diet, and physical and mental development are normal. Urinary organic acid excretion was typical of 3-ketothiolase deficiency, showing consistently high levels of 2-methyl-3-hydroxybutyric acid and tiglylglycine. Activation of acetoacetyl-CoA thiolase activity by potassium (K) ion in cultured fibroblasts was not observed, demonstrating the lack of activity of mitochondrial acetoacetyl-CoA thiolase. In addition, the signal for the mitochondrial acetoacetyl-CoA thiolase protein was undetectable in the immunoblot analysis. In the pulse-chase experiments, the signal for mitochondrial acetoacetyl-CoA thiolase was detected after a 1-h pulse but not after a 24-h chase. These results indicate that the deficiency was caused by an unstable mitochondrial acetoacetyl-CoA thiolase protein.

Effects of birth asphyxia on urinary organic acid excretion.

Using capillary gas chromatography-mass spectrometry, the effects of birth asphyxia on the urinary organic acid profile of term babies was investigated. Random urine samples were collected on days 1 and 8 from 19 babies with fetal distress, 19 with moderate birth asphyxia and 12 with severe asphyxia causing encephalopathy. Controls were 27 well neonates. Statistically significant abnormalities were found only for the severely asphyxiated group: increased concentrations of lactic, pyruvic, 3-hydroxybutyric, 4-hydroxyphenyllactic and 4-hydroxy-3-methoxymandelic acids, and excretion of four abnormal metabolites, 2-hydroxybutyric, 2-oxoisocaproic, 2-hydroxyisovaleric and 2-oxo-3-methylvaleric acids. Six other babies had increased lactic acid excretion, associated in four with transient 'jitterness' or hypotonia. Organic acid studies may help to grade the severity of perinatal asphyxia in the outcome or intervention studies.

Ethylmalonic/Adipic Aciduria: Effects of Oral Medium-Chain Triglycerides, Carnitine, and Glycine on Urinary Excretion of Organic Acids, Acylcarnitines, and Acylglycines

A 9-y-old girl with ethylmalonic/adipic aciduria was hospitalized to determine the possible therapeutic efficacy of oral carnitine and glycine supplementation. To provoke a mild metabolic stress, her diet was supplemented with 440 mg/kg/d of medium-chain triglycerides. She was treated successively with carnitine (100 mg/kg/d) for 5 d, neither carnitine nor glycine for 2 d, and then glycine (250 mg/kg/d) for 6 d. Consecutive 12-h urine collections were obtained throughout the entire period. The urinary excretion of eight organic acids, four acylglycines, and four acylcarnitines, which accumulate as a result of a metabolic block of five mitochondrial acyl-CoA dehydrogenases, were quantitatively determined by capillary gas chromatography, stable isotope dilution gas chromatography/mass spectrometry, and radioisotopic exchange HPLC, respectively. The excretion of each group of metabolites was calculated as the mean percentage of total output (mumol/24 h) during the four phases of the protocol (organic acids/acylglycines/acylcarnitines = 100.0%): 1) regular diet (3 d); 88.1/10.8/1.1; 2) medium-chain triglyceride supplementation (4); 82.5/15.6/1.9; 3) medium-chain triglycerides plus carnitine (5); 79.2/8.2/12.6; and 4) medium-chain triglycerides plus glycine (6); 81.0/18.7/0.3. Comparison between total and individual excretion of acylglycines and acylcarnitines indicates that oral glycine supplementation enhanced the conjugation and excretion of fatty acyl-CoA intermediates as efficiently as carnitine. We propose that oral glycine supplementation should be considered in the treatment of other inborn errors of metabolism associated with abnormal urinary excretion of acylglycines.

Urinary organic acid excretion during feeding of medium-chain or long-chain triglyceride diets in patients with non-insulin-dependent diabetes mellitus

Medium-chain triglycerides (MCTs) are absorbed and metabolized differently from long-chain triglycerides (LCTs). Recent data indicate that MCTs may be useful as a dietary substitute in a variety of clinical disorders. The current studies were undertaken to characterize urinary organic acid excretion in patients with non-insulin-dependent diabetes mellitus during 4 d of an LCT or MCT diet. Urinary excretion of the dicarboxylic acids adipic, suberic, and 3-hydroxysebacic and the (omega-1) hydroxylation products 5-hydroxyhexanoic acid and 7-hydroxyoctanoic acid, was increased during MCT feeding as compared with LCT feeding. Urinary suberic and 7-hydroxyoctanoic acid excretions were increased 55- and 30-fold, respectively, during the MCT-substituted diet. Urinary organic acid profiles provide information on the fate of lipids during MCT feeding and may also be useful in assessing complicance during clinical trials employing MCT-substituted diets.

Carnitine Metabolism and Urinary Organic Acid Excretion in Reye's Syndrome and Salicylate Intoxication.

Carnitine Metabolism and Urinary Organic Acid Excretion in Reye's Syndrome and Salicylate Intoxication.