Orotic Acid Excretion Research Articles

Clinical findings of patients with hyperammonemia affected by urea cycle disorders with hepatic encephalopathy.

Urea cycle disorders (UCD) are a group of genetic diseases caused by deficiencies in the enzymes and transporters involved in the urea cycle. The impairment of the cycle results in ammonia accumulation, leading to neurological dysfunctions and poor outcomes to affected patients. The aim of this study is to investigate and describe UCD patients' principal clinical and biochemical presentations to support professionals on urgent diagnosis and quick management, aiming better outcomes for patients. We explored medical records of 30 patients diagnosed in a referral center from Brazil to delineate UCD clinical and biochemical profile. Patients demonstrated a range of signs and symptoms, such as altered levels of consciousness, acute encephalopathy, seizures, progressive loss of appetite, vomiting, coma, and respiratory distress, in most cases combined with high levels of ammonia, which is an immediate biomarker, leading to a UCD suspicion. The most prevalent UCD detected were ornithine transcarbamylase deficiency, followed by citrullinemia type 1, hyperargininemia, carbamoyl phosphate synthase 1 deficiency, and argininosuccinic aciduria. Clinical symptoms were highly severe, being the majority developmental and neurological disabilities, with 20% of death rate. Laboratory analysis revealed high levels of ammonia (mean ± SD: 860 ± 470 μmol/L; reference value: ≤80 μmol/L), hypoglycemia, metabolic acidosis, and high excretion of orotic acid in the urine (except in carbamoyl phosphate synthetase 1 [CPS1] deficiency). We emphasize the need of urgent identification of UCD clinical and biochemical conditions, and immediate measurement of ammonia, to enable the correct diagnosis and increase the chances of patients' survival, minimizing neurological and psychomotor damage caused by hepatic encephalopathy.

Orotic acid production from crude glycerol by engineered Ashbya gossypii

Orotic acid is an intermediate of the de novo pyrimidine biosynthesis that plays a crucial role in several nutraceutical supplements. In some microorganisms, blockage of this pathway downstream the dihydroorotate dehydrogenase (DHOD) leads to orotic acid accumulation. Here, the Ashbya gossypii Agura3 pyrimidine auxotroph was shown to accumulate and excrete orotic acid (~1.5 g/L), and this trait was further explored for the production of orotic acid from raw substrates. Metabolic engineering of this strain combined with culture conditions optimization led to a 3.6-fold increase in orotic acid production from crude glycerol. The mitochondrial DHOD encoded by AgURA9 was shown to be determinant for this phenotype. Heterologous expression of AgURA9 in engineered S. cerevisiae enabled the accumulation and excretion of orotic acid (~0.5 g/L). This study demonstrates the potential of A. gossypii for the valorization of crude glycerol to orotic acid and discloses the molecular determinants for its biosynthesis in fungi.

Ion chromatographic method for the determination of orotic acid in urine

Excess urinary orotic acid excretion occurs in patients with some inborn errors of metabolic pathways such as pyrimidine synthesis and urea cycle. Thus, rapid diagnosis of orotic aciduria has a vital importance for patients. In this paper, a novel method for determination of orotic acid in urine samples by ion chromatography with suppressed conductivity detection was investigated. The separation of orotic acid from urine matrix was accomplished by using an anion exchange column with optimized isocratic eluent program which utilized 50 mM NaOH. The other chromatographic conditions were as follows: the suppressor current was 31 mA; the flow rate of mobile phase was 0.25 mL min−1; the column temperature was 30 °C; sample loop volume was 10 μL. Under optimized conditions, the limit of detection was 0.2 μmol L‾1. Dramatically elevated orotic acid concentration was observed on pathological urine samples comparing to healthy urines, as expected. There are a good many advantages of the proposed method, but using an environmentally friendly reagent free system, no organic solvent employment and its quick nature, and being a sensitive and reliable method are the most obvious ones. The proposed method, therefore, may be utilized as an alternative technique for clinical laboratories.

Hyperammonemia in a Child Presenting with Growth Delay, Short Stature, and Diarrhea

A 5-year-old male child was referred to the metabolism clinic with a history of failure to thrive, chronic intermittent emesis, diarrhea, seizures, hypotonia, and growth hormone deficiency. He was also noted to experience frequent falling episodes. Routine biochemistry testing revealed increased plasma ammonia concentration at 184 μmol/L (reference interval, 16–47) and lactate dehydrogenase concentration at 445 U/L (reference interval, 110–295). Plasma free and total carnitine concentrations were decreased at 5 μmol/L (reference interval, 26–60) and 17.1 μmol/L (reference interval, 32–84), respectively. Orotic acid excretion in urine was also markedly increased at >80 mmol/mol creatinine (reference interval, 0.2–1.5). Plasma insulin-like growth factor-I (IGF-I)4 concentration was also decreased at <25 ng/mL (reference interval, 50–286). Paired plasma and urine specimens were collected for amino acid analysis using ultraperformance liquid chromatography. The results are shown in Table 1. View this table: Table 1. Amino acid analysis of paired plasma and urine. This patient has lysinuric protein intolerance (LPI), an autosomal recessive disorder of amino acid transport, caused by mutations in the solute carrier family 7, member 7 ( SLC7A7 ) gene (1, 2, 3). Although the worldwide incidence is not known, LPI is estimated to occur in approximately 1 in 60000 newborns in Finland and Japan (2). The SLC7A7 gene product is the …

Mild orotic aciduria in UMPS heterozygotes: a metabolic finding without clinical consequences

BackgroundElevated urinary excretion of orotic acid is associated with treatable disorders of the urea cycle and pyrimidine metabolism. Establishing the correct and timely diagnosis in a patient with orotic aciduria is key to effective treatment. Uridine monophosphate synthase is involved in de novo pyrimidine synthesis. Uridine monophosphate synthase deficiency (or hereditary orotic aciduria), due to biallelic mutations in UMPS, is a rare condition presenting with megaloblastic anemia in the first months of life. If not treated with the pyrimidine precursor uridine, neutropenia, failure to thrive, growth retardation, developmental delay, and intellectual disability may ensue.Methods and resultsWe identified mild and isolated orotic aciduria in 11 unrelated individuals with diverse clinical signs and symptoms, the most common denominator being intellectual disability/developmental delay. Of note, none had blood count abnormalities, relevant hyperammonemia or altered plasma amino acid profile. All individuals were found to have heterozygous alterations in UMPS. Four of these variants were predicted to be null alleles with complete loss of function. The remaining variants were missense changes and predicted to be damaging to the normal encoded protein. Interestingly, family screening revealed heterozygous UMPS variants in combination with mild orotic aciduria in 19 clinically asymptomatic family members.ConclusionsWe therefore conclude that heterozygous UMPS-mutations can lead to mild and isolated orotic aciduria without clinical consequence. Partial UMPS-deficiency should be included in the differential diagnosis of mild orotic aciduria. The discovery of heterozygotes manifesting clinical symptoms such as hypotonia and developmental delay are likely due to ascertainment bias.

Protective Effects of Tinospora cordifolia on Hepatic and Gastrointestinal Toxicity Induced by Chronic and Moderate Alcoholism.

Heavy alcohol intake depletes the plasma vitamins due to hepatotoxicity and decreased intestinal absorption. However, moderate alcohol intake is often thought to be healthy. Therefore, effects of chronic moderate alcohol intake on liver and intestine were studied using urinary vitamin levels. Furthermore, effects of Tinospora cordifolia water extract (TCE) (hepatoprotective) on vitamin excretion and intestinal absorption were also studied. In the study, asymptomatic moderate alcoholics (n = 12) without chronic liver disease and healthy volunteers (n = 14) of mean age 39 ± 2.2 (mean ± SD) were selected and divided into three groups. TCE treatment was performed for 14 days. The blood and urine samples were collected on Day 0 and 14 after treatment with TCE and analyzed. In alcoholics samples, a significant increase in the levels of gamma-glutamyl transferase, aspartate transaminase, alanine transaminase, Triglyceride, Cholesterol, HDL and LDL (P < 0.05) was observed but their level get downregulated after TCE intervention. Multivariate analysis of metabolites without missing values showed an increased excretion of 7-dehydrocholesterol, orotic acid, pyridoxine, lipoamide and niacin and TCE intervention depleted their levels (P < 0.05). In contrast, excretion of biotin, xanthine, vitamin D2 and 2-O-p-coumaroyltartronic acid (CA, an internal marker of intestinal absorption) were observed to be decreased in alcoholic samples; however, TCE intervention restored the CA and biotin levels. Vitamin metabolism biomarkers, i.e. homocysteine and xanthurenic acid, were also normalized after TCE intervention. Overall data depict that moderate alcohol intake is also hepatotoxic and decreases intestinal absorption. However, TCE treatment effectively increased the intestinal absorption and retaining power of liver that regulated alcohol-induced multivitamin deficiency.

A Novel Description of Increased Urinary Orotic Acid Excretion in Pediatric Trauma

The diagnostic utility of orotic aciduria for suspected inborn errors of metabolism is well established. Orotic aciduria has been reported in traumatized adults, but not in children. A 2-year-old hyperammonemic girl with cardiac arrest and abdominal compartment syndrome secondary to non-accidental trauma with significant orotic aciduria is presented. To our knowledge, this is the first report highlighting the association of transiently elevated urinary orotic acid and mild uricosuria with severe trauma in children. Further investigations into the clinical use of orotic acid excretion as a biomarker of the catabolic state of pediatric trauma or other critical illness are warranted. J Endocrinol Metab. 2014;4(1-2):32-35 doi: http://dx.doi.org/10.14740/jem209w

A Novel Description of Increased Urinary Orotic Acid Excretion in Pediatric Trauma

The diagnostic utility of orotic aciduria for suspected inborn errors of metabolism is well established. Orotic aciduria has been reported in traumatized adults, but not in children. A 2-year-old hyperammonemic girl with cardiac arrest and abdominal compartment syndrome secondary to non-accidental trauma with significant orotic aciduria is presented. To our knowledge, this is the first report highlighting the association of transiently elevated urinary orotic acid and mild uricosuria with severe trauma in children. Further investigations into the clinical use of orotic acid excretion as a biomarker of the catabolic state of pediatric trauma or other critical illness are warranted. J Endocrinol Metab. 2014;4(1-2):32-35 doi: http://dx.doi.org/10.14740/jem209w

Megaloblastic Anemia—A Rare Cause

A 2- year- old boy presented with non responsive megaloblastic anemia, growth failure and developmental delay. Blood levels of B(12), folic acid and iron were normal. Tandem mass spectroscopy for common inborn errors of metabolism did not reveal any abnormality. There was an increased excretion of orotic acid in urine. The authors report this as a rare cause of megaloblastic anemia.

Dihydrofolate Reductase Deficiency Is Caused by a Homozygous DHFR Mutation and Leads to Congenital Megaloblastic Anemia and Cerebral Folate Deficiency

AbstractAbstract 1006Folate deficiency can be associated with megaloblastic anemia, neurological and mental disorders, cardiovascular disease, embryonic defects, in particular neural tube defects, and, possibly, malignancies. The importance of an intact intracellular folate metabolite cycle is illustrated by the severity of symptoms in rare inborn disorders of folate metabolism.Dihydrofolate reductase (DHFR) catalyzes the reduction of dihydrofolate (DHF) to tetrahydrofolate (THF) and at a lower rate of folic acid (FA) to DHF. DHFR plays a key role in maintaining intracellular folate homeostasis. A distinct genetic defect associated with DHFR deficiency has not been described.We examined three children of healthy distantly related parents presenting with megaloblastic anemia and neurological symptoms of varying severity. Total serum folate and cobalamin, homocysteine, urinary excretion of methylmalonic acid, orotic acid, formiminoglutamic acid, and cobalamin binding capacity were normal. Red blood cell (RBC) folate was below the normal range in all patients. Cerebrospinal fluid (CSF) analysis revealed very low 5-methyl-tetrahydrofolate (5MTHF) in one and the absence of detectable CSF 5MTHF in two siblings. Treatment with folinic acid (5-formyl-tetrahydrofolate; 5FTHF) increased RBC folate in all patients associated with a normalization of RBC indices and bone marrow morphology and led to CSF 5MTHF normalization.According to a recessive inheritance model, we conducted a screen for homozygous chromosomal regions using genome-wide analysis of single nucleotide polymorphisms. This analysis revealed a short overlap on the long arm of chromosome 2 and a second common region on the long arm of chromosome 5. This region included the DHFR gene locus. Genomic sequence analysis of DHFR exons and neighboring nucleotides revealed a homozygous mutation DHFR c.458A>T in exon 5 (RefSeq NM_000791) leading to the amino acid change DHFR p.Asp153Val (p.D153V) in all siblings.Assuming DHFR deficiency we analyzed folate metabolites in RBCs and plasma by liquid chromatography tandem mass spectrometry (LC-MS/MS). The folate profile in RBCs and plasma was compatible with DHFR deficiency. To examine DHFR function we studied the formation of THF out of DHF in EBV-immortalized lymphoblastoid cells from patients, their heterozygous mother and healthy control individuals. DHFR activity was severely reduced in all patient cells (12, 88, 51 pmol THF per hour and mg protein, respectively) as compared to controls (median 3741, range 1441–6828 pmol/h*mg) whereas cells of the heterozygous mother exhibited DHFR activity close to the lower limit of the control range (1156 pmol/h*mg). A fluorescent Methotrexate (MTX) binding assay showed a markedly reduced MTX binding capacity in EBV-immortalized lymphoblastoid cells of all three patients. Cells of the heterozygous mother presented an intermediate binding capacity. To assess whether the reduced DHFR activity was caused by an altered mRNA and protein expression due to the DHFR p.Asp153Val mutation we performed RT-PCR and western blot analyses. RT-PCR revealed a single ∼450bp cDNA fragment without significant quantitative differences between DHFR wildtype, DHFR p.Asp153Val heterozygous, or homozygous cells. In contrast, DHFR protein expression was reduced in EBV-immortalized lymphoblastoid cells carrying the DHFR mutation as compared to wildtype cells. Semiquantitative assessment revealed a reduction of the amount of intact DHFR protein in heterozygous lymphoblastoid cells to about 70 percent, in homozygous cells from one patient to 50 percent and from the other patients to 20–30 percent as compared to wildtype cells.In conclusion, the homozygous mutation DHFR p.Asp153Val causes DHFR deficiency and leads to a complex hematological and neurological disease which can successfully be treated with 5FTHF. Disclosures:No relevant conflicts of interest to declare.

Les déficits du cycle de l’urée chez les patients adultes

Urea cycle disorders (UCD) usually present after 24 h to 48 h of life with failure to thrive, lethargy and coma leading to death, but milder forms may occur from infancy to adulthood. Survival of children with UCD has significantly improved and the need for transitional care to adulthood has emerged. Adult onset UCD present with chronic or acute neurological, psychiatric and digestive symptoms associated with protein avoidance. Ornithine transcarbamylase (OTC) deficiency, which is inherited as an X-linked disorder, is the most well-described UCD in adults. Acute decompensations associate the triad of encephalopathy, respiratory alkalosis and hyperammonemia. Acute encephalopathy is characterized by brain edema, which is life-threatening without treatment. Specific urea cycle enzyme deficiency can be suspected in the presence of abnormal plasma amino acids concentrations and urinary excretion of orotic acid. A measurement enzyme activity in appropriate tissue, or DNA analysis if available, is required for diagnosis. Treatment requires restriction of dietary protein intake and the use of alternative pathways of waste nitrogen excretion with sodium benzoate and sodium phenylbutyrate. Patients with acute forms may need hemodialysis or hemodiafiltration. Therapeutic goals for OTC deficiency are to maintain plasma ammonia<80 micromol/L, plasma glutamine<1,000 micromol/L, argininemia 80-150 micromol/L and branched chain amino acids within the normal range, in order to prevent episodes of potentially lethal acute hyperammonemia. Potentially fatal acute hyperammonemia may occur in male or female patients at any age. Ammonia should be measured promptly in case of acute neurological and psychiatric symptoms or coma.

Orotic Acid Excretion and Arginine Metabolism

The urinary excretion of orotic acid, an intermediate in the pyrimidine biosynthetic pathway, is markedly increased in many inborn errors of the urea cycle and in a number of other disorders involving arginine metabolism. Carbamoyl phosphate, which accumulates within hepatic mitochondria in patients with ornithine transcarbamoylase deficiency, can diffuse to the cytosol and enter the pyrimidine pathway, resulting in greatly increased orotic acid production and excretion. This orotic aciduria also occurs in inborn errors of the mitochondrial ornithine/citrulline transporter, arginase, argininosuccinate synthetase, and argininosuccinate lyase. Increased orotic acid excretion is also found in a number of hypoargininemic states, such as lysinuric protein intolerance. However, orotic aciduria should not be used uncritically as an index of arginine deficiency because it is found in patients with arginase deficiency who exhibit hyperargininemia. Increased orotic acid excretion can also arise as a result of impairments of pyrimidine synthesis, whether brought about by a genetic defect (e.g., in UMP synthase) or by drugs that inhibit the terminal part of the pathway (e.g., allopurinol or 6-azauridine). When used appropriately, measurement of urinary orotic acid is a valuable tool for the study of many derangements of arginine metabolism, including arginine depletion, and to assess the efficacy of therapies used to replete this amino acid.

Purine and pyrimidine metabolites in children's urine

Various enzyme defects in the metabolic pathways of purines and pyrimidines are known, which result in different diseases occurring in children. They mainly affect kidney function, central nervous system, immunological and blood system. For example, complete deficiency of HPRT (hypoxanthine-guanine-phosphoribosyl-transferase) causes the Lesch Nyhan syndrome, which is characterized by hyperuricemia, mental retardation, choreoathetosis and compulsive self-mutilation. XDH deficiency (xanthine-dehydrogenase) causes in arthropathia and myopathia. For screening for these and other enzyme defects, urinary purine and pyrimidine excretion is considered a simple diagnostic tool. The purpose of the present study was to establish a reverse phase HPLC screening method for urinary purines and pyrimidines and to establish age related reference ranges in children for the urinary excretion of orotic acid, uracile, pseudouridine, uric acid, hypoxanthine, xanthine, thymine, 7-methylguanine, inosine, guanosine and adenosine.

Influence of dietary arginine on sexual dimorphism of arginine metabolism in mice

We have studied the influence of dietary arginine on tissue arginine content, and arginine metabolism in CD1 mice. Dietary arginine restriction produced by feeding mice with a low arginine diet (0.06%) produced a marked decrease in arginine concentrations in the plasma, skeletal muscle and kidney of female mice (72%, 67% and 54%, respectively) while in male mice the decreases were smaller (58% in blood and 18% in the skeletal muscle). This diet abolished not only the sexual dimorphism in arginine content observed in mice fed with the diet containing 1% arginine, but also reduced renal activities of arginase and nitric oxide synthase in the female mice and ornithine decarboxylase and the decarboxylation of arginine in the male mice. Urinary putrescine excretion was dramatically reduced by arginine restriction in the male mice whereas orotic acid excretion increased about 30 fold in both sexes; urea and creatinine excretion did not change. Taken together our results indicate that dietary arginine plays a relevant role in the maintenance of the sexual dimorphism in arginine content and arginine metabolism in CD1 mice, and that this may have physiological significance because of the important effects that arginine-derived products exert on a variety of cellular processes.

Plasma urea-cycle-related amino acids, ammonium levels, and urinary orotic acid excretion in short-bowel patients managed with an oral diet

Background and Aims: The small intestine contains several enzymes involved in arginine synthesis and converts glutamine to citrulline, the major compound for endogenous arginine synthesis. This study was conducted to assess the plasma status of urea-cycle intermediates and orotic urinary excretion in short-bowel patients. Methods:Thirteen stable short-bowel syndrome patients (7 men; 60.2±15.2 years) were studied. Patients were divided into moderately resected (Group A; n=6) and severely resected (Group B; n=7) according to their remnant bowel length (Group A: 61–150 cm; Group B: ≤60 cm). All subjects were consuming an oral diet plus dietetic supplements. Plasma urea-cycle amino acids, ammonium and urinary orotic acid were determined. Results: Plasma glutamine levels were significantly higher in both patient groups than in the control group ( P<0.001). Regarding citrulline, Group B levels were significantly lower vs. controls ( P<0.001). Comparisons between patient groups showed higher arginine in Group A ( P<0.05) and non-statistically lower citrulline in Group B. Blood ammonium and orotic urinary excretion were normal. Conclusions: Although plasma citrulline and glutamine alterations were found, patients showed no hyperammonemia or orotic aciduria, which suggests a certain degree of adaptation in arginine and related amino acid metabolism, when an adequate dietary supply of arginine is provided.

ガスクロマトグラフィー質量分析による発育不良黒毛和種牛における尿の代謝プロフィル解析

To detect metabolic abnormalities, we performed gas chromatography/mass spectrometry (GC/MS) on growth-retarded Japanese Black cattle. Many organic acids, amino acids, sugars, nucleic-acid bases, and glycine conjugates were detected in urine from 18 normal, cattle. Increased excretion of orotic acid was observed in one growth-retarded calf. In 3 calves with renal tubular dysplasia, concentrations of hippurate, suberate, and phenols tended to be higher and concentrations of propionylglycine lower than in normal calves. In 14 other growth-retarded calves, concentration of hippurate was higher and concentration of propionylglycine tended to be lower than in normal calves. Concentrations of organic acids (4-hydroxyphenylacetate, methylcitrate, cis-aconite, and glycolate) and amino acids tended to be lower.

Ornithine aminotransferase, a potential target for the treatment of hyperammonemias.

Ornithine-delta-aminotransferase (OAT) (EC 2.6.1.13) is a pyridoxal-5' phosphate dependent mitochondrial matrix enzyme. It controls the L-ornithine (Orn) level in tissues by catalysing the transfer of the delta-amino group of Orn to 2-oxoglutarate. The products of this reaction are L-glutamate-gamma-semialdehyde and L-glutamate. Among the compounds known to inhibit (or inactivate) OAT, only L-canaline and (SS)-5-(fluoromethyl)ornithine [(SS)-5FMOrn] are selective for OAT. Treatment of laboratory animals with 5FMOrn causes a dramatic accumulation of Orn in most tissues and organs, and the enhanced formation of urea due to saturation of ornithine:carbamoyltransferase with its substrate. The enhancement of urea formation by increased endogenous levels of Orn is comparable with that produced by large doses of Orn and arginine, a treatment known to enhance the detoxification of ammonia. However, protection to lethal doses of ammonium salts by exogenous Orn is rapidly fading. In contrast, inactivation of OAT by a small dose of 5FMOrn renders a long-lasting protective effect against various forms of hyperammonemic states. Among these the reduction of ammonia concentrations in blood and tissues, and the reduction of the pathologic excretion of orotic acid to normal levels in mice with hereditary defects of the urea cycle, were most impressive. In human hereditary OAT deficiency the elevated intraocular concentrations of Orn are considered to be a cause of gyrate atrophy. This is presumably the reason, why OAT has not been considered as a therapeutically useful target. Chronic inactivation of OAT by repeated administration of 5FMOrn, caused elevated intraocular Orn concentrations, but this treatment had no effect on the function and histology of the visual system, or the behaviour of adult mice. The confirmation of this and related observations in higher species will show, whether OAT inactivation has potentials in the treatment of hyperammonemic states.

Effect of lysine infusion on urea cycle in lysinuric protein intolerance

Poor intestinal absorption and excessive renal loss of dibasic amino acids result in low plasma concentrations in patients with lysinuric protein intolerance (LPI). Arginine and ornithine deficiency impair the function of the urea cycle and cause hyperammonemia after protein intake, while chronic lysine deficiency may cause growth failure and lead to reduced bone density in such patients. Since high lysine concentrations inhibit several enzymes of the urea cycle in the liver, lysine supplementation may induce hyperammonemia in LPI. We thus studied how LPI patients tolerate high plasma lysine by intravenous (IV) infusion of 3.3 mmol/kg lysine hydrochloride over 90 minutes in 6 adult patients and 4 healthy controls. The plasma lysine concentration (mean +/- SD, range) peaked in the patients (9,114 +/- 1,864, 7,156 to 12,044 micromol/L) and controls (10,185 +/- 2,253, 7,714to 13,122 micromol/L) at 90 minutes. Urinary lysine excretion peaked in the second 2-hour urine collection in the patients (4,582 +/- 1,276, 3,018 to 6,315 micromol/m2 body surface area per hour) and in the first 2-hour collection in the controls (5,373 +/- 1,766, 3,551 to 7,286 micromol/m2/h). Two patients had mild nausea but no hyperammonemia and one patient had moderate hyperammonemia (peak, 112 micromol/L) at the end of the infusion. Orotic acid excretion increased in 2 subjects with a peak excretion rate of 33 and 251 micromol/m2/h in the third 2-hour collection after starting the load. All other subjects remained asymptomatic and showed no change in plasma ammonia or urinary orotic acid excretion. We thus conclude that an acute increase in plasma lysine caused minimal clinical or biochemical untoward effects in patients with LPI. Moderate increases in plasma lysine after low-dose oral supplementation with lysine or well-absorbed lysine derivatives are probably well tolerated in LPI.

Determination of urinary orotic acid and uracil by capillary zone electrophoresis

We describe a simple method for measuring orotic acid and uracil concentration in urine by capillary zone electrophoresis in 20 m M Na-borate buffer, pH 9.2. The method was applied for studying a patient with HHH (hyperornithinemia, hyperammonemia and homocitrullinuria) syndrome. A high value of uracil excretion was found during periods of relatively low orotic acid excretion and normal ammonemia. The orotic acid level in urine was increased by increasing protein intake.

Aberrations of ammonia metabolism in ornithine carbamoyltransferase-deficient spf-ash mice and their prevention by treatment with urea cycle intermediate amino acids and an ornithine aminotransferase inactivator

Sparse fur with abnormal skin and hair (spf-ash) mice are deficient in ornithine carbamoyltransferase (OCT) activity, but their OCT protein is kinetically normal. We administered ammonium chloride to spf-ash mice, in order to analyze ammonia metabolism and to find a rationale for the therapy of OCT deficiency. Ammonia concentration in the liver of spf-ash mice increased to a level much higher than in the control. Ammonium chloride injection caused an increase in ornithine (Orn) 5 min after injection and an increase in the sum of Orn, citrulline (Cit) and arginine (Arg) for at least 15 min in the liver of control mice, but no increase in Orn, Cit and Arg in the liver of spf-ash mice. Treatment of spf-ash mice with Arg 5–20 min prior to the injection of ammonium chloride kept the hepatic ammonia concentration at a level comparable to that without the load. A significant reciprocal relationship between ammonia and Orn concentrations in the liver of spf-ash mice 5 min after an ammonium chloride load with or without Arg strongly suggests that ammonia disposal is dependent on the supply of Orn. In spf-ash mice loaded with tryptone as a nitrogen source, Arg supplementation showed a dramatic decrease in urinary orotic acid excretion in a dose-dependent manner. Similar effects were observed with Cit and Orn at the same dose, and a long-lasting effect with an ornithine aminotransferase inactivator, 5-(fluoromethyl)ornithine, at a much lower dose. The rate of urea formation in liver perfused with ammonium chloride was lower in spf-ash mice than in controls, but with the addition of Orn to the medium it increased to a similar level in control and spf-ash mice. These results indicate that OCT is not saturated with Orn in vivo under physiological conditions and that the administration or enrichment of the urea cycle intermediate amino acids enhances the OCT reaction so that the ammonia metabolism of OCT-deficient spf-ash mice is at least partially normalized.