Diagnosing Inborn Errors of Metabolism in the Newborn: Laboratory

Investigations
Gregory M. Enns and Seymour Packman
NeoReviews 2001;2;192
DOI: 10.1542/neo.2-8-e192

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Article

inborn errors of metabolism

Diagnosing Inborn Errors of

Metabolism in the Newborn:
Laboratory Investigations
Gregory 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 inborn
error 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.

Introduction
Approximately 4% of individuals born in the United States have a genetic or partly genetic
disorder. Inborn errors of metabolism contribute significantly to this total. Although
individually rare, the aggregate incidence of metabolic disease is relatively high and may be
greater than 1 in 1,000 newborns. Newborn screening programs using tandem mass
spectrometry that can detect approximately 20 inborn errors of metabolism typically have
reported an incidence of 1 in 5,000. Because there are hundreds of known metabolic
conditions, 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, although histologic abnormalities may be present in the fetal central nervous system by 4 to
5 months of gestation. Because inborn errors of metabolism that present in the newborn
period often have nonspecific features, appropriate laboratory investigations are required
to avoid misdiagnoses such as sepsis or asphyxia.

Basic Investigations
Many medical centers are not equipped to perform the specialized investigations needed to
evaluate a patient in whom an inborn error of metabolism is suspected, such as detailed
amino acid, organic acid, or acylcarnitine analyses. However, simple laboratory tests,
including measurements of blood gases, electrolytes, glucose, lactate, and ammonia levels
and basic urinalysis, often provide the initial clues to a possible underlying metabolic
disease. Although the presence of a specific inborn error of metabolism cannot be
confirmed until biochemical genetic laboratory results are available, a diagnosis may be
suspected and an inborn error disease category reasonably hypothesized on the basis of
findings from simple laboratory studies and the clinical presentation. Such an initial
hypothesis is important, because the physician must initiate appropriate therapy without
delay and without a final diagnosis to decrease the morbidity or mortality associated with
these conditions. Initial laboratory investigations for the assessment of the critically ill
neonate suspected of having an inborn error are shown in Table 1.

The Anion Gap

The anion gap is an important diagnostic tool in evaluating metabolic acidosis. The term
is a misnomer because it implies a gap between cation and anion concentration when, in
fact, the concentration of total serum cations equals the concentration of total anions. The
anion gap is commonly estimated as [Na]([Cl][HCO32]), 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.
e192 NeoReviews Vol.2 No.8 August 2001

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inborn errors of metabolism laboratory findings

Initial Laboratory
Investigation of a Suspected
Inborn Error of Metabolism in
the Newborn
Table 1.

Hypokalemia, hyopcalcemia, hypomagnesemia

Methanol, metformin
Uremia
Diabetic ketoacidosis
Paraldehyde, phenformin
Inborn error of metabolism, inorganic anions
(phosphate, sulfate), iron, isoniazid
Lactic acidemia
Ethanol, ethylene glycol
Salicylates, solvents, strychnine

Complete blood count

Blood gases
Electrolytes, calcium, magnesium
Glucose
Lactate, pyruvate
Ammonia
Liver enzymes, prothrombin time, and partial
thromboplastin time
Quantitative amino acid analysis
Carnitine levels (total, free, esterified)
Acylcarnitine profile
Plasma for storage at 20C
Urine Tests
Routine urinalysis
Organic acids
Ferric chloride test, DNPH, Acetest,
reducing substances (see text)
Urine for storage at 20C
DNPH dinitrophenylhydrazine

being approximately 8 to 16 mEq/L (8 to 16 mmol/L).

However, the actual concentration of unmeasured anions (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

Normal Serum
Concentrations of
Unmeasured Cations and
Unmeasured Anions
Table 2.

Unmeasured Cations
(mEq/L [mmol/L])

Total

Decreased Unmeasured Cation

Increased Unmeasured Anion (MUDPILES)

Blood Tests

K
Ca2
Mg2

Causes of Increased
Anion Gap

Table 3.

4.5
5
1.5
11

Unmeasured Anions
(mEq/L [mmol/L])
Protein
PO42
SO42
Organic acids

15
2
1
5
23

Adapted with permission from Oh MS, Carroll HJ. The anion gap. N
Engl J Med. 1977;297:814 817. Copyright 1977 Massachusetts
Medical Society. All rights reserved.

and calculated unmeasured anions may be explained

when the equation used to calculate the anion gap is
rewritten to include the concept of electroneutrality.
The total cations, Na and the unmeasured cations
(UC), must equal the total anions of Cl, HCO32,
and the unmeasured anions (UA): NaUC(Cl
HCO32)UA. Therefore, Na(ClHCO32)
UAUC. When unmeasured cations are subtracted
from unmeasured anions, a value of 12 mEq/L
(12 mmol/L) is obtained (Table 2). Therefore, the
clinically used term of anion gap actually represents the
difference in concentration between unmeasured anions
and unmeasured cations, and is equal to Na(Cl
HCO32). It is this last subtraction that is used for
estimation in clinical circumstances.
An increased anion gap is caused by either an increase
in the concentration of unmeasured anions or a decrease
in the concentration of unmeasured cations (Table 3).
A decreased anion gap is rarer, but it may be seen if the
unmeasured anion concentration is decreased or if the
unmeasured cation concentration is increased. In practice, the most common cause of a decreased anion gap is
hypoalbuminemia, although an increased or decreased
anion gap also may result from laboratory errors.
Metabolic acidosis frequently is accompanied by an
increased anion gap, usually caused by the accumulation
of organic acids that titrate bicarbonate. Organic acids
typically responsible for causing an increase in the anion
gap include lactic acid, ketone bodies (acetoacetate and
beta-hydroxybutyrate), and unusual organic acids such as
methylmalonic acid, propionic acid, and their derivatives
(Table 4). On the other hand, when metabolic acidosis
occurs as a result of bicarbonate loss, the anion gap is
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inborn errors of metabolism laboratory findings

Inborn Errors of
Metabolism With Increased
Anion Gap

Table 4.

Organic Acidemias
Propionic acidemia
Isovaleric acidemia
Methylmalonic acidemia
Holocarboxylase synthetase deficiency
Multiple acyl-CoA dehydrogenase deficiency
3-Hydroxyisobutyric acidemia
3-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 deficiency
Very long-chain acyl-CoA dehydrogenase (VLCAD)
deficiency
Carnitine uptake deficiency
Carnitine-acylcarnitine translocase (CAT) deficiency
Carnitine palmitoyltransferase 2 (CPT-2) deficiency
Congenital Lactic Acidosis
Pyruvate dehydrogenase deficiency
Pyruvate carboxylase deficiency
Mitochondrial respiratory chain disorders
Tricarboxylic Acid Cycle Defects
Fumaric aciduria
Alpha-ketoglutarate dehydrogenase deficiency
Disorders of Gluconeogenesis
Phosphoenolpyruvate carboxykinase deficiency
Fructose-1,6-bisphosphatase deficiency

normal because of a concomitant increase in Cl. Hyperchloremic metabolic acidosis usually is caused by excessive gastrointestinal bicarbonate loss (eg, diarrhea) or
renal tubular acidosis. Metabolic diseases associated with
renal tubular acidosis are shown in Table 5. A basic
approach to metabolic acidosis is illustrated in Figure 1.

Inborn Errors of
Metabolism Without Increased
Anion Gap (Renal Tubular
Acidosis)

Table 5.

Galactosemia
Hereditary fructose intolerance
Glycogen storage disease, types I and III
Phosphoenolpyruvate carboxykinase deficiency
Tyrosinemia, type I
Cystinosis
Carnitine palmitoyltransferase 1 deficiency
Mitochondrial respiratory chain disorders
Lowe syndrome*
Carbonic anhydrase II deficiency
*Phosphatidylinositol-4,5-bisphosphate 5-phosphatase deficiency.

Osteopetrosis with renal tubular acidosis.

disease, cardiomyopathy), pulmonary hypertension, and

severe anemia may cause lactic acidosis. However, once
the underlying cause is treated, the lactic acidosis resolves
relatively quickly. Inborn errors of metabolism that are
associated with lactic acidosis include disorders of pyruvate metabolism, mitochondrial respiratory chain function, gluconeogenesis, and fatty acid oxidation and some
organic acidemias (Table 6).
When evaluating a neonate who has an anion gap
metabolic acidosis, it is helpful to determine if the gap is
explained by lactate. In other words, if the anion gap
becomes normal when lactate is taken into account
Na(ClHCO32lactate)8 to 16 mEq/L (8 to
16 mmol/L)the acidosis is accounted for by the increase in blood lactate concentration. Conversely, if the
gap remains substantially elevated after the lactate concentration has been considered, other unmeasured anions must be contributing to the increased anion gap. In
the latter case, an organic acidemia or cause of significant
ketosis (diabetic ketoacidosis) is a diagnostic possibility.
The lactate-to-pyruvate (L/P) ratio may help in diagno-

Lactic Acidosis
Abnormal accumulation of lactic acid is a common cause
of metabolic acidosis with increased anion gap in the
neonate. Most often, lactic acidosis is caused by tissue
hypoxia due to inadequate circulation or diminished
oxygen supply. Sepsis, heart failure (congenital heart
e194 NeoReviews Vol.2 No.8 August 2001

Figure 1. Approach to the investigation of metabolic acidosis.

Adapted from Clarke JTR, 1996, p. 78 with permission of
Cambridge University Press.

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inborn errors of metabolism laboratory findings

Inborn Errors of
Metabolism Associated With
Lactic Acidosis

Table 6.

Primary Lactic Acidosis

Defects of pyruvate metabolism
Pyruvate dehydrogenase deficiency
Pyruvate carboxylase deficiency
Mitochondrial respiratory chain defects
Secondary Lactic Acidosis
Gluconeogenesis disorders
Phosphoenolpyruvate carboxykinase (PEPCK)
deficiency
Fructose-1,6-bisphosphatase deficiency
Carbohydrate disorders
Glycogen storage disease, type I
Hereditary fructose intolerance
Fatty acid oxidation defects
Organic acidemias
Holocarboxylase synthetase deficiency
Biotinidase deficiency*
Propionic, methylmalonic, isovaleric acidemias
3-Hydroxy-3-methylglutaryl-CoA (HMG-CoA)
lyase deficiency
Multiple acyl-CoA dehydrogenase deficiency
*Tends to present later in infancy.

sis. An elevated L/P ratio (25) may be present in

mitochondrial disorders or pyruvate carboxylase deficiency type A, and the ratio is normal in pyruvate dehydrogenase deficiency or gluconeogenesis disorders.
(Note that the L/P ratio also may be elevated in hypoxemic states.) It may be necessary to repeat the lactate
measurement several times because spurious elevations
may result from a difficult blood draw and local tissue
hypoxia. An approach to the evaluation of a child who
has persistently elevated lactate is shown in Figure 2.

Figure 2. Approach to the investigation of lactic acidosis.

L/Plactate/pyruvate ratio, TCAtricarboxylic acid cycle,
PCpyruvate carboxylase, GSDglycogen storage disease,
PDHpyruvate dehydrogenase. Adapted with permission from
Scriver et al, eds, 2001.

Figure 3. Approach to the investigation of hypoglycemia.

HFIhereditary fructose intolerance, FAODfatty acid oxidation disorders, HMG3-hydroxy-3-methylglutaryl-CoA lyase
deficiency, CDGcongenital disorders of glycosylation (formerly carbohydrate-deficient glycoprotein syndrome),
FBPfructose-1,6-bisphosphatase deficiency, PEPCKphosphoenolpyruvate carboxykinasae deficiency, SCOTsuccinylCoA:3-oxoacid-CoA transferase deficiency, SCHADshortchain 3-hydroxyacyl-CoA dehydrogenase deficiency [SCHAD
deficiency also may be associated with hypoketotic hypoglycemia, and abnormal urine organic acids suggestive of a FAOD
may be present].

Hypoglycemia
The sick neonate commonly exhibits hypoglycemia regardless of the etiology of the illness. Hypoglycemia
associated with severe systemic illness caused by sepsis or
asphyxia is usually relatively easy to control by the administration of glucose at, or slightly above, the basal neonatal glucose oxidation rate (4 to 6 mg/kg per minute). In
contrast, hypoglycemia due to inborn errors of metabolism may be somewhat recalcitrant to therapy; intravenous glucose supplementation may be required at high
levels or it may be difficult to wean the infusion.
Hypoglycemia normally stimulates ketone body formation by increased mitochondrial beta-oxidation of
fatty acids. Disorders of fatty acid oxidation and hyperinsulinism are associated with low plasma and urine
ketone body concentrations (hypoketotic hypoglycemia). Some disorders of gluconeogenesis, including glycogen storage disease type I, phosphoenolpyruvate carboxykinase deficiency, and fructose-1,6-bisphosphatase
deficiency, may cause secondary inhibition of ketone
formation and, therefore, also may be associated with
hypoketotic hypoglycemia. On the other hand, the presence of urinary ketones in the neonate is unusual. If a
prominent ketosis is observed in association with hypoglycemia and an anion gap metabolic acidosis, an organic
acidemia should be suspected. Ketosis and hypoglycemia
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inborn errors of metabolism laboratory findings

also may be present in maple syrup urine disease

(MSUD), but metabolic acidosis is not a typical finding
early in the course of disease. An approach to the evaluation of hypoglycemia is shown in Figure 3.

Hyperammonemia
Hyperammonemia is a life-threatening condition that
may occur in the absence of other obvious metabolic
derangements. Respiratory alkalosis due to central stimulation of ventilation by ammonium is often present. The
ammonia level should be evaluated immediately in all
infants who have altered consciousness or a child who
has encephalopathy may be misdiagnosed as having
sepsis or another more common disorder. Significant
neonatal hyperammonemia (200 mcmol/L and often
2,000 mcmol/L) most frequently is associated with a
defect of the urea cycle. However, infants who have
transient hyperammonemia of the newborn (THAN),
organic acidemias (eg, propionic, isovaleric, and methylmalonic acidemias), and some fatty acid oxidation defects
(eg, carnitine uptake defect, carnitine palmitoyl transferase 1 deficiency, and carnitine-acylcarnitine translocase deficiency) may have similar ammonia elevations
caused by a secondary inhibition of the urea cycle by
toxic metabolites.
Measurement of blood gases, electrolytes, and urine
ketones may help to distinguish hyperammonemia due
to a urea cycle defect from that due to an organic
acidemia. Unlike organic acidemias, urea cycle defects
usually are not associated with significant metabolic acidosis or ketosis. However, tissue hypoxia may supervene
in the critically ill neonate, making a distinction difficult
in practice. Precise diagnosis of urea cycle disorders requires serum amino acid quantitation, urine organic
acid analyses (especially for orotic acid), and fibroblast
or hepatocyte measurement of enzyme activity. Other
causes of neonatal hyperammonemia include the
hyperammonemia-hyperornithinemia-homocitrullinuria
syndrome, the hyperinsulinism-hyperammonemia syndrome, and lysinuric protein intolerance. A guide to the
differential diagnosis of neonatal hyperammonemia is
shown in Figure 4.

Simple Urine Tests

Neonates have a mild physiologic hyperketonemia that
does not result in ketonuria significant enough to be
detected by dipstick methods. This normal ketogenesis
may be attenuated in small-for-gestational-age and preterm infants. Routine urinalysis or dipstick test for ketones may offer critical information. The presence of
significant ketonuria in a neonate who has an anion gap
e196 NeoReviews Vol.2 No.8 August 2001

Figure 4. Approach to the investigation of hyperammonemia.

ASAargininosuccinic acid, THANtransient hyperammonemia of the newborn, LPIlysinuric protein intolerance, HHH
hyperammonemia-hyperornithinemia-homocitrullinuria syndrome, OTCornithine transcarbamylase, CPScarbamyl phosphate synthetase, NAGSN-acetylglutamate synthetase. Urine
orotic acid also tends to be elevated in LPI and HHH, but
citrulline concentration is normal.

metabolic acidosis may occur in organic acidemias or

later stages of MSUD. Alternatively, hypoketosis in the
face of severe distress may point to an underlying fatty
acid oxidation disorder.
Urine color and odor also may yield diagnostic clues
to an underlying genetic defect. Other simple urine
screening tests include the Clinitest (reducing sub-

Causes of Urine Reducing

Substances
Table 7.

Compound

Disorder/Source

Glucose

Diabetes mellitus
Renal Fanconi syndrome
Galactosemia
Severe liver disease
Hereditary fructose
intolerance
Essential fructosuria
Pentosuria
Tyrosinemia

Galactose
Fructose
Xylose
4-Hydroxyphenylpyruvic
acid
Homogentisic acid
Oxalic acid
Uric acid
Ascorbic acid
Salicylates

Alkaptonuria
Hyperoxaluria
Hyperuricosuria
Exogenous administration
Exogenous administration

Adapted with permission from Blau et al, 1996.

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inborn errors of metabolism laboratory findings

The Urine Ferric Chloride Test

crease in conjunction with results

from urine organic acid analysis
lead to a diagnosis of metabolic disColor
Compound
Disorder
ease in many cases. For example,
Blue-green
Phenylpyruvic acid
Phenylketonuria
the branched-chain amino acids
Imidazolepyruvic acid
Histidinemia
leucine, isoleucine, and valine are
Homogentisic acid
Alkaptonuria
Greenish-gray
Branched-chain oxoacids
MSUD
increased in MSUD; glycine is eleGreen
Hydroxyphenylpyruvate
Tyrosinemia
vated in many organic acidemias;
Cherry red
Acetoacetic acid
3-Oxothiolase deficiency
and methionine is often high in tyDiabetic ketoacidosis
rosinemia. Characteristic organic
Purple
Ketones
3-Oxothiolase deficiency
acid findings confirm the diagnosis
The ferric chloride test detects oxo-acids, and differently colored complexes may form depending on the
in each case. The citrulline concencompound. MSUDmaple syrup urine disease. Adapted with permission from Blau et al, 1996.
tration is helpful in differentiating
urea cycle defects. Very high levels
are present in argininosuccinic acid
synthetase deficiency (citrullinemia); high levels in
stances), ferric chloride test (oxoacids), dinitrophenylhyargininosuccinic acid lyase deficiency; and low or undedrazine (DNPH) test (2-oxoacids), Acetest (ketones),
tectable levels in N-acetylglutamate synthetase, carbamyl
and nitroprusside test (sulfur-containing acids). Comphosphate synthetase, and ornithine transcarbamylase
pounds screened for and the corresponding disorders
deficiencies. An elevated glutamine concentration from
detected are listed in Tables 7 through 10.
transamination of glutamic acid is also a clue to underlySpecialized Investigations
ing hyperammonemia. Similarly, elevated alanine levels
These investigations are not available in many centers
may reflect lactic acidosis; an increased lactate concentraand require specialized equipment and expert interpretation leads to the formation of pyruvate via lactate dehytion. Nevertheless, all children suspected of having indrogenase, and pyruvate can be converted to alanine by
born errors of metabolism should have plasma and urine
alanine aminotransferase. A CSF-to-plasma glycine ratio
obtained for these analyses and sent to a reference labogreater than 0.08 is diagnostic of nonketotic hyperglyratory that has expertise in biochemical genetic diagnocinemia, a disorder in which other metabolic laboratory
sis.
findings are typically normal. Urine amino acids are
analyzed less frequently, but such analysis may detect
Plasma Amino Acid Analysis
patterns of dibasic or neutral amino acid elevations charPlasma amino acid analysis is indicated for any infant
acteristic of specific amino acid transport defects (eg,
suspected of having an inborn error of metabolism.
Hartnup disease, cystinuria). Generalized aminoaciduria
is seen in the renal Fanconi syndrome, but it also may be
Characteristic patterns of amino acid elevation or depresent in normal neonates due to
immaturity of tubular transport.
Table 9.
Table 8.

The Urine Dinitrophenylhydrazine (DNPH)

and Acetest
DNPH

Acetest

Positive Compound

Disorder/Source

Phenylpyruvic acid
2-Oxoisovaleric acid
2-Oxoisocaproic acid
2-Oxo-3-methylvaleric acid
Imidazolepyruvic acid
Acetone
2-Methylacetoacetate
Butanone
4-Hydroxyphenylpyruvate
Pyruvate

Phenylketonuria
MSUD
Histidinemia
Organic acidemias*
Tyrosinemia, liver disease
Lactic acidosis

*Propionic acidemia, methylmalonic acidemia, 3-oxothiolase deficiency. MSUDmaple syrup urine

disease. Adapted with permission from Blau et al, 1996.

Urine Organic Acid Analysis

The analysis of urine organic acids
by gas chromatography-mass spectrometry is often key to diagnosing
or excluding the presence of an inborn error of metabolism. If an organic acidemia is severe enough to
present in a neonate, characteristic
compounds are likely to be detected as long as the urine sample
was collected while the infant was in
a catabolic state. Diagnostic metabolites may be absent or difficult to
detect once therapy has been initiNeoReviews Vol.2 No.8 August 2001 e197

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inborn errors of metabolism laboratory findings

Table 10.

The Urine Nitroprusside

Test
Compound

Disorder/Source

Cystine

Cystinuria
Generalized aminoaciduria
Hyperargininemia
Homocystinuria
Cobalamin C and D disease
MTHFR deficiency
Vitamin B12 deficiency
Glutathionuria
Dehydration

Homocystine

Glutathione
Ketones high creatinine

Nitroprusside reacts with sulfur acids to form a pink to purple colored

complex. MTHFRmethylene tetrahydrofolate reductase. Adapted
with permission from Blau et al, 1996.

ated and metabolic balance has been restored. Diagnosis

is based on pattern recognition of specific organic acid
elevations. For example, elevations of methylmalonate,
3-hydroxypropionate, and methylcitrate are characteristic of methylmalonic acidemia. Dicarboxylic acids are
present in fatty acid oxidation defects, but also may be
seen after valproate administration or in patients receiving supplemental feedings containing medium-chain
triglycerides (MCT). However, the pattern of dicarboxylic acid elevation tends to be different in MCT supplementation (sebacicsubericadipic) than in fatty acid
oxidation disorders (adipicsubericsebacic). Disorders
of pyruvate metabolism may show only high lactate and
pyruvate peaks, whereas mitochondrial respiratory chain
disorders, in addition to increased lactate and pyruvate,
often show multiple nonspecific elevations, including
ethylmalonic acid and tricarboxylic acid cycle intermediates (eg, 2-oxoglutaric, fumaric, and succinic acids).

Carnitine Levels
Carnitine (hydroxytrimethylaminobutyric acid) transports long-chain fatty acids across the inner mitochondrial membrane and, therefore, is essential for the proper
function of the fatty acid oxidation cycle. Carnitine is
synthesized by the liver and kidney and present in the
diet, but secondary deficiency is relatively common. Low
carnitine levels are common in preterm infants and neonates receiving total parenteral nutrition without adequate carnitine supplementation for long periods. Some
metabolic disorders also may result in secondary carnitine deficiency. Because carnitine has an additional role as
a metabolic scavenger, acylcarnitine esters accumulate
in times of decompensation in fatty acid oxidation defects or inborn errors of organic acid metabolism and
e198 NeoReviews Vol.2 No.8 August 2001

then are excreted in the urine. Under normal circumstances, the concentration of acylcarnitine esters is low,
with most of the plasma carnitine being in a free, unesterified form. An elevation of carnitine esters (an
esterified-to-free carnitine ratio 0.30) may be seen in
fatty acid oxidation defects, organic acidemias, and ketosis. Such an elevation in the esterified fraction may be a
clue to diagnosing an underlying inborn error.

Acylcarnitine Profile
The plasma acylcarnitine profile is determined by fastatom bombardment or electrospray tandem mass spectrometry (MS/MS). Whereas free and esterified carnitine
levels may offer a clue to the presence of a fatty acid
oxidation defect or organic acidemia, the acylcarnitine
profile allows determination of the biochemical composition of the elevated esterified fraction. For example,
elevated isovalerylcarnitine is seen in isovaleric acidemia,
and long-chain fatty acylcarnitines (C14, C14:1, C16, and
C18 acylcarnitines) are characteristic of very long-chain
acyl-CoA dehydrogenase (VLCAD) or carnitine palmitoyl transferase 2 (CPT-2) deficiency. MS/MS is being
used increasingly in newborn screening programs to
detect an additional 15 to 30 metabolic disorders. Such
MS/MS newborn screening could provide critical diagnostic information.

Additional Analyses
Other studies may be required based on the clinical
presentation. Examples include plasma very long-chain
fatty acids, phytanic acid, and erythrocyte plasmalogen
analyses for the diagnosis of peroxisomal disorders; glycosylated transferrin analysis for congenital disorders of
glycosylation; and urine bile acids for disorders of bile
acid synthesis. Final confirmation of the presence of an
inborn error of metabolism may require enzymology on
fibroblast, liver, or muscle or molecular genetic (DNA)
analysis.

Postmortem Diagnosis
Appropriate specimens should be collected from all infants who die unexpectedly from unknown causes. Preand perimortem blood and urine samples should be sent
for metabolic investigations, as outlined in Table 1. The
neonatal blood spot also may be retrieved for MS/MS
analysis. If consent is given for an autopsy, portions of
muscle, heart, and liver should be snap-frozen immediately for possible future analysis. A skin, diaphragm,
and/or lung biopsy should be obtained for establishing a
fibroblast cell line. Only by making a diagnosis can the
family be provided with accurate genetic counseling for

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inborn errors of metabolism laboratory findings

es), hypoglycemia, metabolic acidosis, and lactate and

pyruvate elevations suggest a defect of carbohydrate metabolism (Fig. 5). Confirmation of the presence of an
inborn error of metabolism, however, requires specific
biochemical analysis by specialized centers and often
additional enzymology or DNA studies. The physician
caring for an acutely ill neonate must consider metabolic
disorders upon initial presentation. In many instances,
only rapid diagnosis and management can prevent death
or significant morbidity.

Suggested Reading

Figure 5. Diagnostic approach to inherited metabolic disorders during infancy. Reprinted with permission from Packman
S, 1996.

future pregnancy and for evaluations of additional family

members at risk.

Summary
Simple laboratory tests can yield valuable information
and lead the clinician to a preliminary diagnosis of an
underlying inborn error of metabolism in the acutely ill
neonate. For example, an infant who has isolated hyperammonemia can be presumptively considered to have a
urea cycle disorder or THAN and treated accordingly.
A baby who has some combination of hyperammonemia,
hypoglycemia, and metabolic ketoacidosis, with or without lactic acidosis, may be suspected of having an organic
acidemia. If hypoketotic hypoglycemia is present, especially if associated with liver failure (a Reye syndrome-like
presentation) or cardiomyopathy, a disorder of fatty acid
oxidation should be strongly considered. Finally, various
combinations of mellituria (positive reducing substanc-

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Clarke JTR. Acute metabolic illness in the newborn. In: A Clinical
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Greene CL, Blitzer MG, Shapira E. Inborn errors of metabolism
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814 817
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inborn errors of metabolism laboratory findings

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 other
unmeasured anions), is useful in evaluating a newborn who is suspected of having a metabolic disorder. Of
the following, a decreased anion gap is most consistent with:
A.
B.
C.
D.
E.

Hyperchloremic metabolic acidosis.

Hypoalbuminemia.
Lactic acidosis.
Methylmalonic acidemia.
Renal tubular acidosis.

6. Hypoglycemia normally stimulates ketone body formation by increased mitochondrial beta-oxidation of

fatty acids. Failure of this compensatory response results in hypoketotic hypoglycemia. Of the following,
the most likely cause of hypoketotic hypoglycemia is:
A.
B.
C.
D.
E.

Hyperinsulinism.
Lactic acidosis.
Maple syrup urine disease.
Methylmalonic acidemia.
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 the
following, the deficient enzyme most likely to yield a very high plasma concentration of citrulline is:
A.
B.
C.
D.
E.

Argininosuccinic acid synthetase.

Carbamyl phosphate synthetase.
N-acetylglutamate synthetase.
Ornithine transcarbamylase.
Pyruvate dehydrogenase.

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Diagnosing Inborn Errors of Metabolism in the Newborn: Laboratory

Investigations
Gregory M. Enns and Seymour Packman
NeoReviews 2001;2;192
DOI: 10.1542/neo.2-8-e192

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