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DOI: 10.1542/neo.2-8-e192 2001;2;192 NeoReviews
Gregory M. Enns and Seymour Packman Investigations
Diagnosing Inborn Errors of Metabolism in the Newborn: Laboratory
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The online version of this article, along with updated information and services, is
Online ISSN: 1526-9906. Illinois, 60007. Copyright © 2001 by the American Academy of Pediatrics. All rights reserved. by the American Academy of Pediatrics, 141 Northwest Point Boulevard, Elk Grove Village,it has been published continuously since 2000. NeoReviews is owned, published, and trademarked NeoReviews is the official journal of the American Academy of Pediatrics. A monthly publication,
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Diagnosing Inborn Errors ofMetabolism in the Newborn:Laboratory InvestigationsGregory M. Enns, MB, ChB*
and Seymour Packman, MD†Objectives After completing this article, readers should be able to:
1. List the initial laboratory tests used to assess the infant suspected of having an inbornerror of metabolism.
2. Delineate the most common cause of a decreased anion gap.3. List the specialized tests that should be undertaken in neonates suspected of having
inborn errors of metabolism.
IntroductionApproximately 4% of individuals born in the United States have a genetic or partly geneticdisorder. Inborn errors of metabolism contribute significantly to this total. Althoughindividually rare, the aggregate incidence of metabolic disease is relatively high and may begreater than 1 in 1,000 newborns. Newborn screening programs using tandem massspectrometry that can detect approximately 20 inborn errors of metabolism typically havereported an incidence of 1 in 5,000. Because there are hundreds of known metabolicconditions, the aggregate estimate seems reasonable.
Relatively few metabolic diseases produce symptoms in the neonate. Many disorders,such as the sphingolipidoses, mucopolysaccharidoses, purine and pyrimidine disorders,and neuronal ceroid lipofuscinoses, produce slowly progressive encephalopathies, al-though histologic abnormalities may be present in the fetal central nervous system by 4 to5 months of gestation. Because inborn errors of metabolism that present in the newbornperiod often have nonspecific features, appropriate laboratory investigations are requiredto avoid misdiagnoses such as sepsis or asphyxia.
Basic InvestigationsMany medical centers are not equipped to perform the specialized investigations needed toevaluate a patient in whom an inborn error of metabolism is suspected, such as detailedamino acid, organic acid, or acylcarnitine analyses. However, simple laboratory tests,including measurements of blood gases, electrolytes, glucose, lactate, and ammonia levelsand basic urinalysis, often provide the initial clues to a possible underlying metabolicdisease. Although the presence of a specific inborn error of metabolism cannot beconfirmed until biochemical genetic laboratory results are available, a diagnosis may besuspected and an inborn error disease category reasonably hypothesized on the basis offindings from simple laboratory studies and the clinical presentation. Such an initialhypothesis is important, because the physician must initiate appropriate therapy withoutdelay and without a final diagnosis to decrease the morbidity or mortality associated withthese conditions. Initial laboratory investigations for the assessment of the critically illneonate suspected of having an inborn error are shown in Table 1.
The Anion GapThe anion gap is an important diagnostic tool in evaluating metabolic acidosis. The termis a misnomer because it implies a “gap” between cation and anion concentration when, infact, the concentration of total serum cations equals the concentration of total anions. Theanion gap is commonly estimated as [Na1]2([Cl2]1[HCO3
22]), with the normal value
*Assistant Professor of Pediatrics; Director, Biochemical Genetics Program, Stanford University School of Medicine, Stanford,CA.†Professor of Pediatrics, Division of Medical Genetics, Department of Pediatrics, University of California, San Francisco, CA.
Article inborn errors of metabolism
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being approximately 8 to 16 mEq/L (8 to 16 mmol/L).However, the actual concentration of unmeasured an-ions (all anions other than chloride and bicarbonate)is about 23 mEq/L (23 mmol/L), not 12 mEq/L(12 mmol/L) (Table 2). This difference between actual
and calculated unmeasured anions may be explainedwhen the equation used to calculate the anion gap isrewritten to include the concept of electroneutrality.The total cations, Na1 and the unmeasured cations(UC), must equal the total anions of Cl2, HCO3
22,and the unmeasured anions (UA): Na11UC5(Cl21HCO3
22)1UA. Therefore, Na12(Cl21HCO322)5
UA2UC. When unmeasured cations are subtractedfrom unmeasured anions, a value of 12 mEq/L(12 mmol/L) is obtained (Table 2). Therefore, theclinically used term of “anion gap” actually represents thedifference in concentration between unmeasured anionsand unmeasured cations, and is equal to Na12(Cl21HCO3
22). It is this last subtraction that is used forestimation in clinical circumstances.
An increased anion gap is caused by either an increasein the concentration of unmeasured anions or a decreasein the concentration of unmeasured cations (Table 3).A decreased anion gap is rarer, but it may be seen if theunmeasured anion concentration is decreased or if theunmeasured cation concentration is increased. In prac-tice, the most common cause of a decreased anion gap ishypoalbuminemia, although an increased or decreasedanion gap also may result from laboratory errors.
Metabolic acidosis frequently is accompanied by anincreased anion gap, usually caused by the accumulationof organic acids that titrate bicarbonate. Organic acidstypically responsible for causing an increase in the aniongap include lactic acid, ketone bodies (acetoacetate andbeta-hydroxybutyrate), and unusual organic acids such asmethylmalonic acid, propionic acid, and their derivatives(Table 4). On the other hand, when metabolic acidosisoccurs as a result of bicarbonate loss, the anion gap is
Table 1. Initial LaboratoryInvestigation of a SuspectedInborn Error of Metabolism inthe NewbornBlood Tests
Complete blood countBlood gasesElectrolytes, calcium, magnesiumGlucoseLactate, pyruvateAmmoniaLiver enzymes, prothrombin time, and partial
thromboplastin timeQuantitative amino acid analysisCarnitine levels (total, free, esterified)Acylcarnitine profilePlasma for storage at 220°C
Urine Tests
Routine urinalysisOrganic acidsFerric chloride test, DNPH, AcetestT,
reducing substances (see text)Urine for storage at 220°C
DNPH 5 dinitrophenylhydrazine
Table 2. Normal SerumConcentrations of“Unmeasured Cations” and“Unmeasured Anions”
Unmeasured Cations(mEq/L [mmol/L])
Unmeasured Anions(mEq/L [mmol/L])
K1 4.5 Protein 15Ca21 5 PO4
22 2Mg21 1.5 SO4
22 1Organic acids 5
Total 11 23
Adapted with permission from Oh MS, Carroll HJ. The anion gap. NEngl J Med. 1977;297:814–817. Copyright © 1977 MassachusettsMedical Society. All rights reserved.
Table 3. Causes of IncreasedAnion GapDecreased Unmeasured Cation
Hypokalemia, hyopcalcemia, hypomagnesemia
Increased Unmeasured Anion (MUDPILES)
Methanol, metforminUremiaDiabetic ketoacidosisParaldehyde, phenforminInborn error of metabolism, inorganic anions
(phosphate, sulfate), iron, isoniazidLactic acidemiaEthanol, ethylene glycolSalicylates, solvents, strychnine
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normal because of a concomitant increase in Cl2. Hyper-chloremic metabolic acidosis usually is caused by exces-sive gastrointestinal bicarbonate loss (eg, diarrhea) orrenal tubular acidosis. Metabolic diseases associated withrenal tubular acidosis are shown in Table 5. A basicapproach to metabolic acidosis is illustrated in Figure 1.
Lactic AcidosisAbnormal accumulation of lactic acid is a common causeof metabolic acidosis with increased anion gap in theneonate. Most often, lactic acidosis is caused by tissuehypoxia due to inadequate circulation or diminishedoxygen supply. Sepsis, heart failure (congenital heart
disease, cardiomyopathy), pulmonary hypertension, andsevere anemia may cause lactic acidosis. However, oncethe underlying cause is treated, the lactic acidosis resolvesrelatively quickly. Inborn errors of metabolism that areassociated with lactic acidosis include disorders of pyru-vate metabolism, mitochondrial respiratory chain func-tion, gluconeogenesis, and fatty acid oxidation and someorganic acidemias (Table 6).
When evaluating a neonate who has an anion gapmetabolic acidosis, it is helpful to determine if the gap is“explained” by lactate. In other words, if the anion gapbecomes normal when lactate is taken into account—Na12(Cl21HCO3
221lactate)58 to 16 mEq/L (8 to16 mmol/L)—the acidosis is accounted for by the in-crease in blood lactate concentration. Conversely, if thegap remains substantially elevated after the lactate con-centration has been considered, other unmeasured an-ions must be contributing to the increased anion gap. Inthe latter case, an organic acidemia or cause of significantketosis (diabetic ketoacidosis) is a diagnostic possibility.The lactate-to-pyruvate (L/P) ratio may help in diagno-
Figure 1. Approach to the investigation of metabolic acidosis.Adapted from Clarke JTR, 1996, p. 78 with permission ofCambridge University Press.
Table 4. Inborn Errors ofMetabolism With IncreasedAnion GapOrganic Acidemias
Propionic acidemiaIsovaleric acidemiaMethylmalonic acidemiaHolocarboxylase synthetase deficiencyMultiple acyl-CoA dehydrogenase deficiency3-Hydroxyisobutyric acidemia3-Hydroxy-3-methylglutaryl-CoA (HMG-CoA) lyase
deficiency
Fatty Acid Oxidation Defects
Short-chain acyl-CoA dehydrogenase (SCAD)deficiency
Medium-chain acyl-CoA dehydrogenase (MCAD)deficiency
Long-chain 3-hydroxyacyl-CoA dehydrogenase(LCHAD) deficiency
Trifunctional protein deficiencyVery long-chain acyl-CoA dehydrogenase (VLCAD)
deficiencyCarnitine uptake deficiencyCarnitine-acylcarnitine translocase (CAT) deficiencyCarnitine palmitoyltransferase 2 (CPT-2) deficiency
Congenital Lactic Acidosis
Pyruvate dehydrogenase deficiencyPyruvate carboxylase deficiencyMitochondrial respiratory chain disorders
Tricarboxylic Acid Cycle Defects
Fumaric aciduriaAlpha-ketoglutarate dehydrogenase deficiency
Disorders of Gluconeogenesis
Phosphoenolpyruvate carboxykinase deficiencyFructose-1,6-bisphosphatase deficiency
Table 5. Inborn Errors ofMetabolism Without IncreasedAnion Gap (Renal TubularAcidosis)GalactosemiaHereditary fructose intoleranceGlycogen storage disease, types I and IIIPhosphoenolpyruvate carboxykinase deficiencyTyrosinemia, type ICystinosisCarnitine palmitoyltransferase 1 deficiencyMitochondrial respiratory chain disordersLowe syndrome*Carbonic anhydrase II deficiency†
*Phosphatidylinositol-4,5-bisphosphate 5-phosphatase deficiency.†Osteopetrosis with renal tubular acidosis.
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sis. An elevated L/P ratio (.25) may be present inmitochondrial disorders or pyruvate carboxylase defi-ciency type A, and the ratio is normal in pyruvate dehy-drogenase deficiency or gluconeogenesis disorders.(Note that the L/P ratio also may be elevated in hypox-emic states.) It may be necessary to repeat the lactatemeasurement several times because spurious elevationsmay result from a difficult blood draw and local tissuehypoxia. An approach to the evaluation of a child whohas persistently elevated lactate is shown in Figure 2.
HypoglycemiaThe sick neonate commonly exhibits hypoglycemia re-gardless of the etiology of the illness. Hypoglycemiaassociated with severe systemic illness caused by sepsis orasphyxia is usually relatively easy to control by the admin-istration of glucose at, or slightly above, the basal neona-tal glucose oxidation rate (4 to 6 mg/kg per minute). Incontrast, hypoglycemia due to inborn errors of metabo-lism may be somewhat recalcitrant to therapy; intrave-nous glucose supplementation may be required at highlevels or it may be difficult to wean the infusion.
Hypoglycemia normally stimulates ketone body for-mation by increased mitochondrial beta-oxidation offatty acids. Disorders of fatty acid oxidation and hyper-insulinism are associated with low plasma and urineketone body concentrations (hypoketotic hypoglyce-mia). Some disorders of gluconeogenesis, including gly-cogen storage disease type I, phosphoenolpyruvate car-boxykinase deficiency, and fructose-1,6-bisphosphatasedeficiency, may cause secondary inhibition of ketoneformation and, therefore, also may be associated withhypoketotic hypoglycemia. On the other hand, the pres-ence of urinary ketones in the neonate is unusual. If aprominent ketosis is observed in association with hypo-glycemia and an anion gap metabolic acidosis, an organicacidemia should be suspected. Ketosis and hypoglycemia
Figure 3. Approach to the investigation of hypoglycemia.HFI5hereditary fructose intolerance, FAOD5fatty acid oxida-tion disorders, HMG53-hydroxy-3-methylglutaryl-CoA lyasedeficiency, CDG5congenital disorders of glycosylation (for-merly carbohydrate-deficient glycoprotein syndrome),FBP5fructose-1,6-bisphosphatase deficiency, PEPCK5phos-phoenolpyruvate carboxykinasae deficiency, SCOT5succinyl-CoA:3-oxoacid-CoA transferase deficiency, SCHAD5short-chain 3-hydroxyacyl-CoA dehydrogenase deficiency [SCHADdeficiency also may be associated with hypoketotic hypogly-cemia, and abnormal urine organic acids suggestive of a FAODmay be present].
Table 6. Inborn Errors ofMetabolism Associated WithLactic AcidosisPrimary Lactic Acidosis
Defects of pyruvate metabolismPyruvate dehydrogenase deficiencyPyruvate carboxylase deficiency
Mitochondrial respiratory chain defects
Secondary Lactic Acidosis
Gluconeogenesis disordersPhosphoenolpyruvate carboxykinase (PEPCK)
deficiencyFructose-1,6-bisphosphatase deficiency
Carbohydrate disordersGlycogen storage disease, type IHereditary fructose intolerance
Fatty acid oxidation defectsOrganic acidemias
Holocarboxylase synthetase deficiencyBiotinidase deficiency*Propionic, methylmalonic, isovaleric acidemias3-Hydroxy-3-methylglutaryl-CoA (HMG-CoA)
lyase deficiencyMultiple acyl-CoA dehydrogenase deficiency
*Tends to present later in infancy.
Figure 2. Approach to the investigation of lactic acidosis.L/P5lactate/pyruvate ratio, TCA5tricarboxylic acid cycle,PC5pyruvate carboxylase, GSD5glycogen storage disease,PDH5pyruvate dehydrogenase. Adapted with permission fromScriver et al, eds, 2001.
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also may be present in maple syrup urine disease(MSUD), but metabolic acidosis is not a typical findingearly in the course of disease. An approach to the evalu-ation of hypoglycemia is shown in Figure 3.
HyperammonemiaHyperammonemia is a life-threatening condition thatmay occur in the absence of other obvious metabolicderangements. Respiratory alkalosis due to central stim-ulation of ventilation by ammonium is often present. Theammonia level should be evaluated immediately in allinfants who have altered consciousness or a child whohas encephalopathy may be misdiagnosed as havingsepsis or another more common disorder. Significantneonatal hyperammonemia (.200 mcmol/L and often$2,000 mcmol/L) most frequently is associated with adefect of the urea cycle. However, infants who havetransient hyperammonemia of the newborn (THAN),organic acidemias (eg, propionic, isovaleric, and methyl-malonic acidemias), and some fatty acid oxidation defects(eg, carnitine uptake defect, carnitine palmitoyl trans-ferase 1 deficiency, and carnitine-acylcarnitine translo-case deficiency) may have similar ammonia elevationscaused by a secondary inhibition of the urea cycle bytoxic metabolites.
Measurement of blood gases, electrolytes, and urineketones may help to distinguish hyperammonemia dueto a urea cycle defect from that due to an organicacidemia. Unlike organic acidemias, urea cycle defectsusually are not associated with significant metabolic aci-dosis or ketosis. However, tissue hypoxia may supervenein the critically ill neonate, making a distinction difficultin practice. Precise diagnosis of urea cycle disorders re-quires serum amino acid quantitation, urine organicacid analyses (especially for orotic acid), and fibroblastor hepatocyte measurement of enzyme activity. Othercauses of neonatal hyperammonemia include thehyperammonemia-hyperornithinemia-homocitrullinuriasyndrome, the hyperinsulinism-hyperammonemia syn-drome, and lysinuric protein intolerance. A guide to thedifferential diagnosis of neonatal hyperammonemia isshown in Figure 4.
Simple Urine TestsNeonates have a mild physiologic hyperketonemia thatdoes not result in ketonuria significant enough to bedetected by dipstick methods. This normal ketogenesismay be attenuated in small-for-gestational-age and pre-term infants. Routine urinalysis or dipstick test for ke-tones may offer critical information. The presence ofsignificant ketonuria in a neonate who has an anion gap
metabolic acidosis may occur in organic acidemias orlater stages of MSUD. Alternatively, hypoketosis in theface of severe distress may point to an underlying fattyacid oxidation disorder.
Urine color and odor also may yield diagnostic cluesto an underlying genetic defect. Other simple urinescreening tests include the Clinitestt (reducing sub-
Figure 4. Approach to the investigation of hyperammonemia.ASA5argininosuccinic acid, THAN5transient hyperammone-mia of the newborn, LPI5lysinuric protein intolerance, HHH5hyperammonemia-hyperornithinemia-homocitrullinuria syn-drome, OTC5ornithine transcarbamylase, CPS5carbamyl phos-phate synthetase, NAGS5N-acetylglutamate synthetase. Urineorotic acid also tends to be elevated in LPI and HHH, butcitrulline concentration is normal.
Table 7. Causes of Urine ReducingSubstances
Compound Disorder/Source
Glucose Diabetes mellitusRenal Fanconi syndrome
Galactose GalactosemiaSevere liver disease
Fructose Hereditary fructoseintolerance
Essential fructosuriaXylose Pentosuria4-Hydroxyphenylpyruvic
acidTyrosinemia
Homogentisic acid AlkaptonuriaOxalic acid HyperoxaluriaUric acid HyperuricosuriaAscorbic acid Exogenous administrationSalicylates Exogenous administration
Adapted with permission from Blau et al, 1996.
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stances), ferric chloride test (oxoacids), dinitrophenylhy-drazine (DNPH) test (2-oxoacids), Acetestt (ketones),and nitroprusside test (sulfur-containing acids). Com-pounds screened for and the corresponding disordersdetected are listed in Tables 7 through 10.
Specialized InvestigationsThese investigations are not available in many centersand require specialized equipment and expert interpreta-tion. Nevertheless, all children suspected of having in-born errors of metabolism should have plasma and urineobtained for these analyses and sent to a reference labo-ratory that has expertise in biochemical genetic diagno-sis.
Plasma Amino Acid AnalysisPlasma amino acid analysis is indicated for any infantsuspected of having an inborn error of metabolism.Characteristic patterns of amino acid elevation or de-
crease in conjunction with resultsfrom urine organic acid analysislead to a diagnosis of metabolic dis-ease in many cases. For example,the branched-chain amino acidsleucine, isoleucine, and valine areincreased in MSUD; glycine is ele-vated in many organic acidemias;and methionine is often high in ty-rosinemia. Characteristic organicacid findings confirm the diagnosisin each case. The citrulline concen-tration is helpful in differentiatingurea cycle defects. Very high levelsare present in argininosuccinic acid
synthetase deficiency (citrullinemia); high levels inargininosuccinic acid lyase deficiency; and low or unde-tectable levels in N-acetylglutamate synthetase, carbamylphosphate synthetase, and ornithine transcarbamylasedeficiencies. An elevated glutamine concentration fromtransamination of glutamic acid is also a clue to underly-ing hyperammonemia. Similarly, elevated alanine levelsmay reflect lactic acidosis; an increased lactate concentra-tion leads to the formation of pyruvate via lactate dehy-drogenase, and pyruvate can be converted to alanine byalanine aminotransferase. A CSF-to-plasma glycine ratiogreater than 0.08 is diagnostic of nonketotic hypergly-cinemia, a disorder in which other metabolic laboratoryfindings are typically normal. Urine amino acids areanalyzed less frequently, but such analysis may detectpatterns of dibasic or neutral amino acid elevations char-acteristic of specific amino acid transport defects (eg,Hartnup disease, cystinuria). Generalized aminoaciduriais seen in the renal Fanconi syndrome, but it also may be
present in normal neonates due toimmaturity of tubular transport.
Urine Organic Acid AnalysisThe analysis of urine organic acidsby gas chromatography-mass spec-trometry is often key to diagnosingor excluding the presence of an in-born error of metabolism. If an or-ganic acidemia is severe enough topresent in a neonate, characteristiccompounds are likely to be de-tected as long as the urine samplewas collected while the infant was ina catabolic state. Diagnostic metab-olites may be absent or difficult todetect once therapy has been initi-
Table 8. The Urine Ferric Chloride TestColor Compound Disorder
Blue-green Phenylpyruvic acidImidazolepyruvic acidHomogentisic acid
PhenylketonuriaHistidinemiaAlkaptonuria
Greenish-gray Branched-chain oxoacids MSUDGreen Hydroxyphenylpyruvate TyrosinemiaCherry red Acetoacetic acid 3-Oxothiolase deficiency
Diabetic ketoacidosisPurple Ketones 3-Oxothiolase deficiency
The ferric chloride test detects oxo-acids, and differently colored complexes may form depending on thecompound. MSUD5maple syrup urine disease. Adapted with permission from Blau et al, 1996.
Table 9. The Urine Dinitrophenylhydrazine (DNPH)and AcetestT
DNPH Acetest Positive Compound Disorder/Source
1 2 Phenylpyruvic acid Phenylketonuria1 2 2-Oxoisovaleric acid MSUD1 2 2-Oxoisocaproic acid1 2 2-Oxo-3-methylvaleric acid1 2 Imidazolepyruvic acid Histidinemia1 1 Acetone Organic acidemias*2 1 2-Methylacetoacetate2 1 Butanone1 2 4-Hydroxyphenylpyruvate Tyrosinemia, liver disease1 1 Pyruvate Lactic acidosis
*Propionic acidemia, methylmalonic acidemia, 3-oxothiolase deficiency. MSUD5maple syrup urinedisease. Adapted with permission from Blau et al, 1996.
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ated and metabolic balance has been restored. Diagnosisis based on pattern recognition of specific organic acidelevations. For example, elevations of methylmalonate,3-hydroxypropionate, and methylcitrate are characteris-tic of methylmalonic acidemia. Dicarboxylic acids arepresent in fatty acid oxidation defects, but also may beseen after valproate administration or in patients receiv-ing supplemental feedings containing medium-chaintriglycerides (MCT). However, the pattern of dicarboxy-lic acid elevation tends to be different in MCT supple-mentation (sebacic.suberic.adipic) than in fatty acidoxidation disorders (adipic.suberic.sebacic). Disordersof pyruvate metabolism may show only high lactate andpyruvate peaks, whereas mitochondrial respiratory chaindisorders, in addition to increased lactate and pyruvate,often show multiple nonspecific elevations, includingethylmalonic acid and tricarboxylic acid cycle intermedi-ates (eg, 2-oxoglutaric, fumaric, and succinic acids).
Carnitine LevelsCarnitine (hydroxytrimethylaminobutyric acid) trans-ports long-chain fatty acids across the inner mitochon-drial membrane and, therefore, is essential for the properfunction of the fatty acid oxidation cycle. Carnitine issynthesized by the liver and kidney and present in thediet, but secondary deficiency is relatively common. Lowcarnitine levels are common in preterm infants and neo-nates receiving total parenteral nutrition without ade-quate carnitine supplementation for long periods. Somemetabolic disorders also may result in secondary carni-tine deficiency. Because carnitine has an additional role asa “metabolic scavenger,” acylcarnitine esters accumulatein times of decompensation in fatty acid oxidation de-fects or inborn errors of organic acid metabolism and
then are excreted in the urine. Under normal circum-stances, the concentration of acylcarnitine esters is low,with most of the plasma carnitine being in a free, unes-terified form. An elevation of carnitine esters (anesterified-to-free carnitine ratio .0.30) may be seen infatty acid oxidation defects, organic acidemias, and keto-sis. Such an elevation in the esterified fraction may be aclue to diagnosing an underlying inborn error.
Acylcarnitine ProfileThe plasma acylcarnitine profile is determined by fast-atom bombardment or electrospray tandem mass spec-trometry (MS/MS). Whereas free and esterified carnitinelevels may offer a clue to the presence of a fatty acidoxidation defect or organic acidemia, the acylcarnitineprofile allows determination of the biochemical compo-sition of the elevated esterified fraction. For example,elevated isovalerylcarnitine is seen in isovaleric acidemia,and long-chain fatty acylcarnitines (C14, C14:1, C16, andC18 acylcarnitines) are characteristic of very long-chainacyl-CoA dehydrogenase (VLCAD) or carnitine palmi-toyl transferase 2 (CPT-2) deficiency. MS/MS is beingused increasingly in newborn screening programs todetect an additional 15 to 30 metabolic disorders. SuchMS/MS newborn screening could provide critical diag-nostic information.
Additional AnalysesOther studies may be required based on the clinicalpresentation. Examples include plasma very long-chainfatty acids, phytanic acid, and erythrocyte plasmalogenanalyses for the diagnosis of peroxisomal disorders; gly-cosylated transferrin analysis for congenital disorders ofglycosylation; and urine bile acids for disorders of bileacid synthesis. Final confirmation of the presence of aninborn error of metabolism may require enzymology onfibroblast, liver, or muscle or molecular genetic (DNA)analysis.
Postmortem DiagnosisAppropriate specimens should be collected from all in-fants who die unexpectedly from unknown causes. Pre-and perimortem blood and urine samples should be sentfor metabolic investigations, as outlined in Table 1. Theneonatal blood spot also may be retrieved for MS/MSanalysis. If consent is given for an autopsy, portions ofmuscle, heart, and liver should be snap-frozen immedi-ately for possible future analysis. A skin, diaphragm,and/or lung biopsy should be obtained for establishing afibroblast cell line. Only by making a diagnosis can thefamily be provided with accurate genetic counseling for
Table 10. The Urine NitroprussideTest
Compound Disorder/Source
Cystine CystinuriaGeneralized aminoaciduriaHyperargininemia
Homocystine HomocystinuriaCobalamin C and D diseaseMTHFR deficiencyVitamin B12 deficiency
Glutathione GlutathionuriaKetones 1 high creatinine Dehydration
Nitroprusside reacts with sulfur acids to form a pink to purple coloredcomplex. MTHFR5methylene tetrahydrofolate reductase. Adaptedwith permission from Blau et al, 1996.
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future pregnancy and for evaluations of additional familymembers at risk.
SummarySimple laboratory tests can yield valuable informationand lead the clinician to a preliminary diagnosis of anunderlying inborn error of metabolism in the acutely illneonate. For example, an infant who has isolated hyper-ammonemia can be presumptively considered to have aurea cycle disorder or THAN and treated accordingly.A baby who has some combination of hyperammonemia,hypoglycemia, and metabolic ketoacidosis, with or with-out lactic acidosis, may be suspected of having an organicacidemia. If hypoketotic hypoglycemia is present, espe-cially if associated with liver failure (a Reye syndrome-likepresentation) or cardiomyopathy, a disorder of fatty acidoxidation should be strongly considered. Finally, variouscombinations of mellituria (positive reducing substanc-
es), hypoglycemia, metabolic acidosis, and lactate andpyruvate elevations suggest a defect of carbohydrate me-tabolism (Fig. 5). Confirmation of the presence of aninborn error of metabolism, however, requires specificbiochemical analysis by specialized centers and oftenadditional enzymology or DNA studies. The physiciancaring for an acutely ill neonate must consider metabolicdisorders upon initial presentation. In many instances,only rapid diagnosis and management can prevent deathor significant morbidity.
Suggested ReadingBlau N, Duran M, Blaskovics ME, eds. Physician’s Guide to the
Laboratory Diagnosis of Metabolic Diseases. London, England:Chapman and Hall; 1996
Burlina AB, Bonafe L, Zacchello. Clinical and biochemical ap-proach to the neonate with a suspected inborn error of aminoacid and organic acid metabolism. Semin Perinatol. 1999;23:162–173
Clarke JTR. Acute metabolic illness in the newborn. In: A ClinicalGuide to Inherited Metabolic Diseases. New York, NY: Cam-bridge University Press; 1996:176–204
Greene CL, Blitzer MG, Shapira E. Inborn errors of metabolismand Reye syndrome: differential diagnosis. J Pediatr. 1988;113:156–159
Greene CL, Goodman SI. Catastrophic metabolic encephalopathiesin the newborn period: evaluation and management. Clin Peri-natol. 1997;24:773–786
Oh MS, Carroll HJ. The anion gap. N Engl J Med. 1977;297:814–817
Packman S. Approach to inherited metabolic disorders that presentin the newborn period and infancy. In: Rudolph AM, HoffmanJIE, Rudolph CD eds: Rudolph’s Pediatrics. Stamford, Conn:Appleton and Lange; 1996:291–294
Packman S. Metabolic encephalopathies. In: Berg B, ed. ChildNeurology: A Clinical Manual. Philadelphia, Pa: JB LippincottCo; 1994:51–59
Rinaldo P, Yoon H-R, Yu C, Raymond K, Tiozzo C, Giordano G.Sudden and unexpected neonatal death: a protocol for thepostmortem diagnosis of fatty acid oxidation disorders. SeminPerinatol. 1999;23:204–210
Saudubray J-M, Charpentier C. Clinical phenotypes: diagnosis/algorithms. In: Scriver CR, Beaudet AL, Sly WS, Valle D, eds.The Metabolic and Molecular Bases of Inherited Disease. NewYork, NY: McGraw-Hill; 2001:1327–1403
Sue CM, Hirano M, DiMauro S, De Vivo DC. Neonatal presenta-tions of mitochondrial metabolic disorders. Semin Perinatol.1999;23:113–124
Ward JC. Inborn errors of metabolism of acute onset in infancy.Pediatr Rev. 1990;11:205–216
Figure 5. Diagnostic approach to inherited metabolic disor-ders during infancy. Reprinted with permission from PackmanS, 1996.
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NeoReviews Quiz
5. Anion gap, estimated as the difference between the sum of concentrations of cations (sodium, potassium,and other unmeasured cations) and the sum of concentrations of anions (chloride, bicarbonate, and otherunmeasured anions), is useful in evaluating a newborn who is suspected of having a metabolic disorder. Ofthe following, a decreased anion gap is most consistent with:
A. Hyperchloremic metabolic acidosis.B. Hypoalbuminemia.C. Lactic acidosis.D. Methylmalonic acidemia.E. Renal tubular acidosis.
6. Hypoglycemia normally stimulates ketone body formation by increased mitochondrial beta-oxidation offatty acids. Failure of this compensatory response results in hypoketotic hypoglycemia. Of the following,the most likely cause of hypoketotic hypoglycemia is:
A. Hyperinsulinism.B. Lactic acidosis.C. Maple syrup urine disease.D. Methylmalonic acidemia.E. Propionic acidemia.
7. Plasma amino acid analysis is indicated in any infant suspected of having an inborn error of metabolism.The plasma concentration of citrulline is particularly helpful in differentiating urea cycle defects. Of thefollowing, the deficient enzyme most likely to yield a very high plasma concentration of citrulline is:
A. Argininosuccinic acid synthetase.B. Carbamyl phosphate synthetase.C. N-acetylglutamate synthetase.D. Ornithine transcarbamylase.E. Pyruvate dehydrogenase.
inborn errors of metabolism laboratory findings
e200 NeoReviews Vol.2 No.8 August 2001 by Amanda Sandford on November 1, 2009 http://neoreviews.aappublications.orgDownloaded from
DOI: 10.1542/neo.2-8-e192 2001;2;192 NeoReviews
Gregory M. Enns and Seymour Packman Investigations
Diagnosing Inborn Errors of Metabolism in the Newborn: Laboratory
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