Carnitine palmitoyltransferase 2 deficiency: the time-course of blood and urinary acylcarnitine levels during initial L-carnitine supplementation.

Carnitine palmitoyltransferase 2 (CPT2) deficiency is one of the most common mitochondrial beta-oxidation defects. A female patient with an infantile form of CPT2 deficiency first presented as having a Reye-like syndrome with hypoglycemic convulsions. Oral L-carnitine supplementation was administered since serum free carnitine level was very low (less than 10 micromol/L), indicating secondary carnitine deficiency. Her serum and urinary acylcarnitine profiles were analyzed successively to evaluate time-course effects of L-carnitine supplementation. After the first two days of L-carnitine supplementation, the serum level of free carnitine was elevated; however, the serum levels of acylcarnitines and the urinary excretion of both free carnitine and acylcarnitines remained low. A peak of the serum free carnitine level was detected on day 5, followed by a peak of acetylcarnitine on day 7, and peaks of long-chain acylcarnitines, such as C16, C18, C18:1 and C18:2 carnitines, on day 9. Thereafter free carnitine became predominant again. These peaks of the serum levels corresponded to urinary excretion peaks of free carnitine, acetylcarnitine, and medium-chain dicarboxylic carnitines, respectively. It took several days for oral L-carnitine administration to increase the serum carnitine levels, probably because the intracellular stores were depleted. Thereafter, the administration increased the excretion of abnormal acylcarnitines, some of which had accumulated within the tissues. The excretion of medium-chain dicarboxylic carnitines dramatically decreased on day 13, suggesting improvement of tissue acylcarnitine accumulation. These time-course changes in blood and urinary acylcarnitine levels after L-carnitine supplementation support the effectiveness of L-carnitine supplementation to CPT2-deficient patients.

The phenylpropionic acid load test: experience with 72 children at-risk for beta-oxidation disorders.

The urinary excretion of metabolites of orally administered phenylpropionic acid (PPA) in 72 children, aged 2 days to 16 years, thought to be at-risk of medium acyl CoA dehydrogenase deficiency has been studied. Forty had presented as Reye Syndrome, 9 as a Reye-like syndrome and 24 were sibs of decreased RS, sibs of RLS cases or sibs of infants who had died suddenly and without explanation where an autopsy revealed the presence of very heavy fatty infiltration of the liver. These studies demonstrated that PPA metabolites are maximally excreted during the 3 hours following the oral load and that this urine collection should be diagnostic. PPA loading is a relatively simple, safe test which is part of the investigation of a patient suspected of having an inborn error of metabolism.

Abnormal fatty acid metabolism in patients in hopantenate therapy during clinical episodes.

Calcium 4-(2,4-dihydroxy-3,3-dimethylbutyramido)butyrate hemihydrate (hopantenate), a cerebral metabolic enhancer used in Japan since 1978, is a homologue of pantothenic acid. Using mass spectrometry, we found urinary excretion of 4-hydroxydodecanedioic acid, 4-hydroxytetradecanedioic acid and a series of 2-hydroxydicarboxylic acids (C8-C14), in addition to a series of odd- and even-numbered dicarboxylic acids (C5-C12) and 3-hydroxydicarboxylic acids (C8-C14) in patients receiving hopantenate during episodes of Reye's-like syndrome. Our findings suggest that an acute intoxication associated with hopantenate occurs owing to pantothenic acid deficiency or the inhibition of CoA-requiring reactions during stress, i.e. infection, prolonged fasting, or malnutrition.

Recurrent, familial Reye-like syndrome with a new complex amino and organic aciduria.

Five of 13 siblings from a Jewish-Ashkenazi family suffered from recurrent Reye-like episodes. During attacks, these patients excreted alpha-keto-adipic, alpha-hydroxy-adipic, and alpha-aminoadipic acids, branched-chain keto acids and saccharopine in addition to lactic, pyruvic, and dicarboxylic acids characteristic of Reye syndrome. The serum concentrations of citrulline and alpha-amino-adipic acid were elevated and carnitine was at the upper limit of the normal range. Serum acetoacetate level was 4-5 times the beta-hydroxybutyrate level, but the pyruvate/lactate ratio was normal. Notably, plasma ketone bodies were lower than expected from the degree of catabolism. When the patients were symptom-free, no abnormal amino or organic acids in serum or urine were detected. These findings might be interpreted as a functional impairment at three different biochemical sites: fatty acid beta-oxidation, dehydrogenase complexes of the pyruvic, alpha-ketoglutaric, alpha-ketoadipic, and branched-chain keto acids, and pyruvate carboxylase. We suggest that in this hereditary disorder a toxic substance, exogenously or endogenously derived, interfered at multiple sites in different metabolic pathways.

Impaired oxidative metabolism of salicylate in Reye's syndrome.

Administration of salicylates during prodromal viral illness has been associated with the development of Reye's syndrome (RS). We studied salicylate biotransformation in RS patients and compared it with those on chronic salicylate therapy for juvenile rheumatoid arthritis (JRA). Urine of RS patients contained significantly more salicylic acid and less gentisic acid than that of JRA patients while the conjugated metabolites were not different between the two groups. These results suggest decreased salicylate microsomal oxidation in RS. The role of altered salicylate metabolism in the pathogenesis of RS is unclear.

Urinary C6-C12 dicarboxylic acylcarnitines in Reye's syndrome.

C6-C12 dicarboxylic acylcarnitines have been identified for the first time in urine from a 2-year-old girl presenting with Reye's syndrome. The acylcarnitines were extracted by ion-exchange chromatography and analysed, both underivatised and as methyl esters using high-resolution fast-atom-bombardment mass spectrometry and B/E-linked scanning. The acylcarnitines were quantified by capillary gas chromatography of the acids extracted after hydrolysis of the acylcarnitine esters. Dodecandioylcarnitine was present in the highest concentration (35.9 mmol/mol creatinine) which exceeded the urinary free dodecandioic acid concentration. The adipic, suberic and sebacic acylcarnitine concentrations were less than 10% of the respective free acid concentrations. It is possible that beta-oxidation of dicarboxylic acids is partially inhibited in Reye's syndrome leading to accumulation of precursor dodecandioyl CoA which is metabolised to dodecandioylcarnitine. The accumulation of these metabolic intermediates may be significant in the pathogenesis of Reye's syndrome.

Serum and urinary carnitine and organic acids in Reye syndrome and Reye-like syndrome.

Free and acyl-carnitine in serum and urine, and urinary organic acids were measured in 6 patients with Reye syndrome and Reye-like syndrome. The free and total carnitine concentrations were significantly reduced in serum during the acute phases of the diseases. Thus, the ratio of acylcarnitine to free carnitine was significantly increased. Urinary excretion of acylcarnitine was greatly increased, and the acylcarnitine to total carnitine ratio was therefore greater than in controls. The urinary organic acids comprised large amounts of lactic acid, dicarboxylic acids and ketone bodies. It is suggested that carnitine deficiency is induced as more carnitine is consumed to buffer the increased amount of toxic acyl-CoA compounds metabolized from free fatty acids and the many organic acids. These results indicate that administration of L-carnitine should generally be considered in patients with Reye syndrome and Reye-like syndrome.

Urinary dicarboxylic acids in Reye syndrome.

Urine from 12 patients with Reye syndrome was examined by gas-liquid chromatography for identification of organic acids. Large amounts of lactic acid, dicarboxylic acids (adipic, suberic, and sebacic), and 3-OH butyric acid were noted. The mean (+/- SD) total dicarboxylic acid concentration was 0.98 +/- 0.24 mg/mg creatinine, compared with 0.006 +/- 0.010 mg/mg creatinine in controls, n = 140; the mean in patients with Reye syndrome was higher (1.40 +/- 0.26 mg/mg creatinine, n = 8) when the samples were obtained prior to initiation of therapy, but declined rapidly after administration of hypertonic glucose, exchange transfusion, and osmotic diuretics. The total urine excretion of dicarboxylic acids plus urine ketones at the time of presentation correlated well with the plasma lactate (r2 = 0.9676) and peak blood ammonia (r2 = 0.9216) levels. Our results document the occurrence of significant dicarboxylic aciduria in Reye syndrome and indicate that fatty acid metabolism is more impaired in this disorder than previously appreciated.

Application of high resolution fast atom bombardment and constant B/E ratio linked scanning to the identification and analysis of acylcarnitines in metabolic disease.

Acylcarnitines, a biologically important group of metabolites which have thus far eluded characterization by mass spectrometry, exhibit very intense fast atom bombardment mass spectra dominated by parent cations. The combination of high resolution selected ion detection and daughter ion analysis using the linked scan at constant B/E ratio has enabled the unequivocal identification of the acylcarnitines in the urine of children with propionic acidemia, methylmalonic aciduria and Reye's Syndrome. Quantitative analysis of acetylcarnitine and propionylcarnitine in selected samples was accomplished by isotope dilution, utilizing (2H3)acetyl- and (2H5)propionylcarnitines as internal standards. These techniques will allow assessment of the therapeutic use of L-carnitine in these disorders.

Mass spectrometric identification of abnormal aromatic compounds in the urine of a child with Reye's like syndrome.

The urine of a young child with hypoglycemia and a Reye's like syndrome contained an excess of unusual aromatic products with a three carbon chain, phenylpropionylglycine and 3- and 4-(hydroxyphenyl)propionic and 3-(3-methoxy-4-hydroxyphenyl)propionic acids, as well as of organic acids usually found in fatty acid beta-oxidation defects: the mono- and dicarboxylic acids derived from the respective (omega-1) and omega-oxidation of C6 to C10 fatty acids.

Urea nitrogen excretion in critically ill children.

Urinary urea nitrogen (UUN) excretion as an index of both total nitrogen excretion and protein catabolism was assayed in 32 children (aged 2 months to 15 years, median 6 years) (50% mechanically ventilated) during an intensive care unit course of one to ten days (median three days). The daily UUN excretion was 4.38 +/- 2.22 gm/sq m (171 +/- 89 mg/kg) (N = 121 patient days). The average daily UUN excretion (N = 32 children) was well described by a linear regression equation for square meters of body surface area (BSA) (milligrams of UUN = 4,421.5 x BSA; r2 = .903). This linear relationship permitted the valid comparison of both individuals and subgroups despite wide age differences. Excretion data in the mechanically ventilated vs the spontaneously breathing children, and in four diagnostic subgroups (Reye syndrome, seven; sepsis, six; elective surgery, seven; and miscellaneous, 12) were evenly distributed about the regression line for body surface area. Variability in average daily UUN excretion was on individual basis, and was independent of diagnostic or therapeutic subgroup.

Aflatoxin and Reye's syndrome: a case control study.

Aflatoxin levels were determined in serum and urine of 17 patients with Reye's syndrome and in control subjects. No significant difference in aflatoxin levels was found for the two groups. However, 23% of all persons studied had levels of aflatoxin indicative of recent exposure. Aflatoxin levels were associated with ingestion of cornmeal and corn bread but not peanut-containing products. This prevalence of aflatoxin may be of public health importance.

Identification of 5-hydroxyhexanoic acid in the urine of twin siblings with a Reye's-like syndrome associated with dicarboxylic aciduria and hypoglycaemia and with similarities to Jamaican vomiting sickness.

Twin male infant siblings who presented in Harrow, UK, with a Reye's-like syndrome associated with profound hypoglycaemia, vomiting, diarrhoea, coma and death in one child, with dicarboxylic aciduria, and similarities to Jamacian vomiting sickness (hypoglycin toxicity) have been shown to excrete large amounts of a previously unrecorded urinary organic acid. This has been identified as 5-hydroxyhexanoic acid by gas chromatography mass spectrometry using a synthesized standard. Concentrations observed were 340 and 330 mg g-1 creatinine in the two patients. The metabolic precursor of the urinary acid is suggested to be hex-4-enoic acid, a probable chemical toxin closely related to the active organic acid metabolite of hypoglycin. The possibility of omega - 1 oxidation of hexanoic acid to 5-hydroxyhexanoic acid in these and other patients with dicarbocylic aciduris is also discussed.

Evidence for hypertyraminemia in Reye's syndrome.

Utilizing a specific and sensitive radioimmunoassay, palsma and urine tyramine were measured in 14 consecutive patients with liver biopsy-proven Reye's syndrome. Plasma tyrosine was measured in 11 of these patients. The results revealed significant (P less than .003) elevation in plasma (3.4 +/- .52 ng/ml) (mean +/- SEM) and urine (1.00 +/- .26 mg/24 hr) tyramine as well as plasma tyrosine (204 +/- 52.5 mumole/liter) at the onset of the disease when compared to the levels of tyramine and tyrosine in a group of hospitalized patients without hepatic disorders. Furthermore, there was a positive correlation between plasma tyramine and days in coma (r = .86; P less than .001), and between plasma tyramine and tyrosine (r = 0.80; P less than .001). These data suggest that there is s substantial disturbance of tyrosine metabolism in Reye's syndrome and that the accumulation of this amino acid and its metabolite, tyramine, may contribute to the encephalopathy of this disease.

Dicarboxylic aciduria: the response to fasting.

The urine of a child who presented with an episode of a disease resembling Reye's syndrome was found to contain large quantities of the dicarboxylic acids adipic and suberic acids, as well as the glycine conjugate of suberic acid, suberyl glycine. A variety of other dicarboxylic acids, both saturated and unsaturated, were also found in the urine at the time of the attack. It was found that the excretion of these unusual metabolites could be markedly increased by fasting for periods of greater than 10 h. These results indicate that the patient may have a defect in fatty acid oxidation which becomes clinically significant during periods of prolonged fasting.