Douglas N. Buchanan

Effect of chemically induced propionic acidemia on neurobehavioral development of rats.

High levels of propionic acid (PPA) comparable to those of human propionic acidemia were achieved in blood (1-5 mmol/l) and brain (1 micromol/g) of rats by administering saline-buffered propionate (pH 7.4) subcutaneously twice a day from the 6th to the 28th day of life. PPA doses ranged from 1.44 to 1.92 micromol/g body weight as a function of animal age. Control rats were treated with saline in the same volumes. Growth and development of physical landmarks were assessed by monitoring the following parameters daily: body weight, upper incisor eruption, eye opening, and hair coat. Development of some reflexes was also monitored, and a specific subset of motor skills was evaluated at days 14 and 21 of life by the free-fall righting test and the spontaneous alternation test. Chronic PPA administration had no effect on body weight, cerebral cortex weight, or cerebellum weight, but caused slight but significant delays in the day of appearance of hair coat and eye opening, indicating an effect of PPA on the development of physical parameters. Free-fall righting was impaired in PPA-treated animals. On the other hand, PPA administration had no effect on the performance of the animals in the spontaneous alternation tests. Long-term effects of early PPA administration were investigated by assessing animal performance in an aversive task (two-way shuttle avoidance task) and in a nonaversive (open-field task) behavioral task at 60 days of age. PPA-treated rats did not habituate to the open field, and presented a lack of retention of the shuttle-avoidance task. Our results suggest that early postnatal PPA administration to rats alters normal development and induces long-term behavioral deficits in aversive and nonaversive tasks.

3-Hydroxy-3-methylglutaric aciduria: Response to carnitine therapy and fat and leucine restriction

A female infant, born to first cousin parents, lapsed into coma with severe metabolic acidosis on day three of life. The gas chromatographic/mass spectrometric urinary organic acid profile showed marked elevation of the leucine metabolites 3-hydroxy-3-methylglutaric, 3-methylglutaconic, 3-methylglutaric and 3-hydroxy-isovaleric acids. Less than 5% of the normal activity of the enzyme 3-hydroxy-3-methylglutaryl CoA lyase was detected in cultured skin fibroblasts. The patients total and free carnitine was initially low but rose to normal levels after placing her ondl-carnitine (100 mg kg−1 d−1). On a diet providing 87 mg kg−1 d−1 of leucine and only 25% of total calories as fat and 2 g kg−1 d−1 protein, the concentration of the urinary organic acids fell markedly. She is now 15 months old with normal growth and development. This regimen appears effective in the early treatment of 3-hydroxy-3-methylglutaric aciduria.

Analysis of α-Ketocarboxylic Acids by Ion Exchange Hplc With UV and Amperometric Detection

Abstract Cation exchange high performance liquid chromatography with tandem 200nm spectrophotometric and +1.15V amperometric detection was used for the analysis of severalOt α-ketocarboxylic acids. Derivatization of the α-ketocarboxyiic acids was not necessary. This technique was operable at 2.0 ug/ml concentrations of α-ketocarboxylic acids. The sensitivity limit was not investigated.

5-oxoprolinuria in an adolescent with chronic metabolic acidosis, mental retardation, and psychosis

5-Oxoprolinuria (pyroglutamic aciduria) is an inborn error of g luta thione metabol ism resulting from ei ther a generalized deficiency of g luta thione synthetase or a deficiency of 5-oxoprolinase; 5-oxoproline accumulates in body fluids. 5Oxoprol inuria was first reported in a 19-year-old Norwegian man with metabolic acidosis, re tardat ion, and spastic quadriparesis. 1 Metabol ic acidosis is a prominent feature when the deficient enzyme is g luta thione synthetase, but not 5-oxoprolinase. We report the clinical and biochemical data from a 12-year-old pat ient with 5-oxoprolinuria caused by gluta th ione synthetase deficiency. Her clinical history demonstrates the necessity of a complete diagnostic evaluation in pat ients with acidosis and in those with psychiatr ic symptoms.

Volatile carboxylic acid profiling in physiological fluids

Many inborn errors of metabolism are characterized by an increase in the concentrations of various carboxylic acids in blood and urine. For example, the metabolic disorders propionic acidemia [ 11, methylmalonic aciduria [ 2,3], maple syrup urine disease [4], isovaleric acidemia [ 41 and glutaric aciduria type II [ 5,6] show increased amounts of volatile carboxylic acids in blood or urine. Procedures for the profiling and/or quantitation of these carboxylic acids require their isolation from blood or urine by vacuum distillation [6], extraction [ 7,8] , silicic acid absorption [ 91 or ion-exchange chromatography [lo] prior to gas chromatographic (GC) or gas chromatographic-mass spectrometric (GC-MS) analysis. These procedures are time-consuming and may result in losses of the more volatile acids. We have developed a rapid procedure for the profiling of volatile carboxylic acids in blood and urine that involves neither extraction or derivatization of the carboxylic acids prior to GC analysis.

Positive-ion thermospray liquid chromatography-mass spectrometry: detection of organic acidurias.

Positive-ion thermospray liquid chromatography-mass spectrometry (TSP-LC-MS) is used to detect organic acids via the direct injection of untreated urine from newborns and infants. Two methods are reported for the separation of organic acids. The separation of urinary organic acids is effected in either an acidic, pH 2.5 sulfuric acid, or a non-acidic, 0.05 M ammonium acetate, pH 6.8, mobile phase. Use of pH 2.5 sulfuric acid and an HPX-87H organic acid column produces better separation but has less sensitivity than the use of 0.05 M ammonium acetate, pH 6.8 and a C18 column. Positive ion TSP-LC-MS has been used to detect methylmalonic aciduria, 3-hydroxy-3-methylglutaric aciduria, propionic aciduria, isovaleric aciduria and argininosuccinic aciduria.

Volatile carboxylic acid profiling in physiological fluids: direct injection into a gas chromatograph/mass spectrometer

We present a procedure for the profiling of the volatile carboxylic acids and neutral compounds in blood or urine using the direct injection of the acidified sample into a gas chromatograph interfaced with a mass spectrometer by a jet separator. The non-volatile components remain at the head of the SP-1000 column while the volatile components move through the column. Up to sixty physiological samples can be analyzed before any degradation in mass spectrometer operating parameters is observed.

Photodiode array ultraviolet spectrophotometric profiling of carboxylic acids in physiological fluids

In this paper the use of a computer-controlled photodiode array spectrophotometric detector for the high-performance liquid chromatographic (HPLC) profiling of carboxylic acids in physiological fluids is reported. The ultraviolet spectrum of the flowing eluent is obtained at 6-sec intervals and is displayed as the absorbance at 190 nm. A three-dimensional (time versus wavelength versus absorbance) presentation of the HPLC profile facilitates peak identification through ultraviolet spectrum matching and relative retention time comparison with carboxylic acid standards. Several examples of HPLC urinary carboxylic acid profiles from infants with various inborn errors of metabolism are shown.

Metabolic studies in a mouse model of hepatorenal tyrosinemia: absence of perinatal abnormalities.

Radiation induced chromosomal deletions at the albino locus in the mouse, lethal when homozygous, cause abnormalities of expression of several unlinked liver specific genes. Recently, the gene encoding FAH was shown to be included in the deletions. Since in humans FAH mutations cause tyrosinemia type I, deletion homozygous mice were suspected of having tyrosinemia. Studies of plasma amino acids did not confirm this suspicion. Also, succinylacetone levels were normal in fetal and newborn livers of deletion homozygotes. The present evidence, therefore, does not support the assumption that the earlier described ultrastructural and enzyme abnormalities in deletion homozygotes are secondary effects of tyrosinemia caused by the deletion of FAH.