Urine Screening for Metabolic

Disorders
Urine Screening for Metabolic Disorders

 Many abnormal results obtained in routine

urinalysis are related to metabolic disorders
rather than renal diseases
 Urine as an end product of body metabolism
contain additional abnormal substances not
tested by routine urinalysis
 Positive urine tests proceed with other
diagnostic procedures
Abnormal Metabolic Constituents
Detected in Routine Urinalysis

 Color: homogentisic acid, melanin, porphyrins

 Odor: phenylketonuria, maple syrup urine disease,

homocystinuria
 Crystals: cystine, leucine, tyrosine, Lesch-Nyhan disease
Categories of Disorders with Urine Metabolic Abnormalities
Overflow vs. Renal Disorders

Overflow: Renal:
-from disruption of normal -abnormal accumulations
metabolic pathway caused by malfunctions
-causes increase plasma in tubular reabsorption
concentrations of
nonmetabolized
substances
Overflow vs. Renal Disorders

 Most frequently encountered metabolic

abnormalities are associated with protein &
carbohydrate metabolism
 Inborn error of metabolism – disruption of
enzyme function due to failure to inherit gene
to produce the enzyme
Major Disorders of Protein &
Carbohydrate Metabolism

 Inherited phenylketonuria, tyrosinemia, Maple syrup

urine disease, porphyria
 Metabolic tyrosinemia, melanuria, indicanuria,
porphyria
 Renal cystinuria, Hartnup disease
Amino Acid Disorders
Phenylketonuria

 Most well-known of aminoacidurias

 May cause severe mental retardation
 PKU is caused by failure to inherit the gene (autosomal
recessive) to produce enzyme, phenylalanine
hydroxylase
 Initial screening: Increased blood levels of
phenylalanine
 Urine test as follow-up procedure
 Ferric Chloride Tube test – nonspecific reaction
 Addition of ferric chloride with urine containing phenylpyruvic
acid produces permanent blue-green color
Tyrosyluria

 Accumulation of excessive tyrosine in plasma

produces urinary overflow
 Seen in premature infants with
underdeveloped liver function
 Ferric Chloride test: green color fades when
tyrosine is present
 Nitroso-naphthol test: positive test is red-
orange color
Alkaptonuria

 Darkened urine after becoming alkaline from

standing at room temperature
 From failure to inherit the gene to produce
enzyme homogentisic acid oxidase
 High % of persons with this disease develop
liver & cardiac problems
 Homogenistic acid reacts in screening tests
Alkaptonuria

 Ferric Chloride test – transient deep blue

color is produced
 Benedict’s test/Clinitest – produce yellow
precipitate
 Adding alkali to urine produce darkening of
the color of urine
 Adding Silver nitrate & ammonium hydroxide
produce black urine
Melanuria

 2nd metabolic pathway for tyrosine produces

melanin
 Melanin – for dark color of hair, eyes, skin
 Deficient melanin  albinism
 Increased melanin  darkening of urine after
being exposed to air
 Increased urine melanin indicates
overproliferation of melanocytes
Melanuria

 Melanin reacts with ferric chloride &

produces gray/black precipitate
 Sodium nitroprusside test provides additional
screening test for melanin & produces red
color
Branched Amino Acid Disorders:
Maple Syrup Urine Disease

 Rare
 Due to inborn error of metabolism, no
enzyme to produce oxidative decarboxylation
of ketoacids
 Involves leucine, isoleucine & valine
 DNPH: 2,4dinitrophenylhydrazine –
screening test for ketoacids
 Produce yellow turbidity/precipitate
Organic Acidemia

 Generalized symptoms: vomiting with

metabolic acidosis, hypoglycemia, ketonuria,
increased serum ammonia
 3 most common acidemia encountered are
– Isovaleric: characteristic of odor of sweaty feet
– Propionic
– Methylmalonic: screening test is p-nitroaniline &
produce emerald green color
Tryptophan Disorders:
Indicanuria

 Normally, tryptophan enters intestine &

reabsorbed by the body or converted to
indole by intestinal bacteria & excreted in
feces
 If with intestinal disorders or Hartnup
disease, increased amounts of tryptophan
are converted to indole
 Excess indole reabsorbed, converted to
indican & excreted in urine
Tryptophan Disorders:
Indicanuria

 Indican excreted in urine is colorless until

oxidized to the dye indigo blue by exposure
to air
 Early diagnosis of Hartnup is reported with
presence of blue stain in diapers (blue diaper
syndrome)
 Urine indican reacts with acidic ferric chloride
 deep blue/violet color that can be
extracted into chloroform
Tryptophan Disorders:
Indicanuria

 Inherited Hartnup disease affects both

intestinal & renal reabsorption of amino
acids, resulting in a generalized
aminoaciduria (Fanconi’s syndrome)
 Defective renal reabsorption does not affect
tubular function
 Dietary corrections by giving niacin in
patients with Hartnup provides good outcome
5-Hydroxyindoleacetic Acid

 2nd metabolic pathway for trytophan is

production of serotonin used in stimulation of
smooth muscles
 Serotonin produced from tryptophan by
argentaffin cells in intestine & carried in the
body by platelets
 Normally, body uses most of serotonin &
small amount of 5-HIAA is excreted in urine
5-Hydroxyindoleacetic Acid

 If with carcinoid tumor affecting argentaffin cells

present, excess serotonin produced resulting in
elevation of urinary 5-HIAA
 Adding nitrous acid & 1-nitroso-2-naphthol to urine
with 5-HIAA  appearance of purple to black color
 Normal excretion of 5-HIAA per day is 2 to 8 mg, if
with tumor, 160 to 628 mg per day
 Test done random or early in morning to have little
chance of false negative
Cystine Disorders:
Cystinuria

 Condition with elevated amounts of amino

acid cystine in urine
 Does not affect cystine metabolism, but the
inability of renal tubules to reabsorb cystine
filtered by glomerulus
 2 modes of inheritance:
– Defective Reabsorption of 4 amino acids (cystine,
lysine, arginine, ornithine)
– Defective reabsorption of only 2 (cystine & lysine)
Cystine Disorders:
Cystinuria

 Cystine is less soluble than the 3 amino

acids, so observation of cystine crystals is
determined in specimen
 Screening test: Cyanide-nitroprusside
– Reduce cystine by sodium cyanide then addition
of nitroprusside will produce red-purple color
– False reactions if ketones & homocystine are
present
Cystine Disorders:
Cystinosis

 Inborn error
 Has 3 variations raning from severe fatal
disorder in infancy to benign form appearing
in adulthood
 Incomplete metabolism of cystine results in
increased crystalline deposits of cystine in
cornea, bone marrow, lymph nodes &
internal organs.
 Fanconi’s syndrome: major defect in renal
tubular reabsorption mechanism
Cystine Disorders:
Homocystinuria

 Defects in metabolism result in failure to

thrive, cataracts, mental retardation,
thromboembolic problems & death
 Increased urine homocystine produce
positive cyanide-nitroprusside test
 Additional test is silver-nitroprusside test in
which only homocystine will react
Cystine Disorders:
Homocystinuria

 Silver nitrate will reduce homocystine to

nitroprusside reactive form but will not
reduce cystine
Porphyrin Disorders

 Porphyrins are intermediate compounds in

production of heme
 Blockage in the pathway will produce
accumulation of of product formed
 Solubility of porphyrin compound varies
 ALA, porphobilinogen & uropophyrin are
most soluble & readily appear in urine
 Protoporphyrin cannot be seen in urine
Porphyrin Disorders

 Observation of red/port wine color to urine is

an indication for possible presence of
porphyrinuria
 2 screening tests:
– Ehrlich reaction detects ALA & porphobilinogen
– Fluorescent technique used for other porphyrins
– Ehrlich & Watson-Schwartz test to differentiate
between porphobilinogen & urobilinogen
Porphyrin Disorders

 Testing porphobilinogen is most useful when

patients have acute attacks
 Fluorescent screening for other porphyrins
– Uses extraction into a mixture of glacial acetic
acid & ethyl acetate
– Solvent layer is examined
– Positive reaction will fluoresce as violet, pink, red
(depending on the concentration)
Mucopolysaccharide Disorders

 Mucopolysaccharides/glycosaminoglycan are
group of large compounds in the connective
tissues
 They consist of protein core with numerous
polysaccharide branches
 Inherited disorders of metabolism prevent
complete breakdown of polysaccharides
resulting in accumulation in lysosomes of
connective tissues
Mucopolysaccharide Disorders

 Products most frequently seen in urine:

 Dermatan sulfate
 Keratan sulfate
 Heparan sulfate
 Types of Mucopolysaccharidoses:
 Hurler’s syndrome (abnormal skeletal structure & severe mental retardation)
 Hunter’s syndrome (abnormal skeletal structure & severe mental retardation)
 Sanfilippo’s syndrome (severe mental retardation)
Mucopolysaccharide Disorders

 Most frequently used screening tests:

 CTAB/ cetyltrimethylammonium bromide turbidity test
 Acid-albumin turbidity test
 In both, end result is thick, white turbidity
 Turbidity is graded 0 to 4 after 30 minutes in acid-
albumin and after 5 minutes in CTAB
 Metachromatic staining test uses basic dyes to react
with acidic mucopolysaccharides
– Positive test will produce blue spot that cannot be washed
away by methanol solution
Purine Disorders

 Lesch-Nyhan Syndrome
– Disorder of purine metabolism
– Inherited sex-linked recessive
– Results in massive excretion of urinary uric acid
crystals
– Due to failure to inherit gene to produce enzyme
hypoxanthine guanine phosphoribosyltransferase
that is responsible for accumulation of uric acid
Purine Disorders

 Clinical manifestations:
 Severe motor defects
 Mental retardation
 Tendency toward self-destruction
 Gout
 Renal calculi/stones
 1st symptom – observation of uric acid
crystals resembling orange sand in diapers
Carbohydrate Disorders

 Majority of meliturias cause no disturbance to

body metabolism
 Melituria (presence of increased urinary sugar due to
inherited disorder)
 Galactosuria (inability to metabolize galactose to glucose)
 Lactosuria (seen during pregnancy & lactation)
 Fructosuria (associated with parenteral feeding)
 Pentosuria ( associated with ingestion of large amount of fruits)
Carbohydrate Disorders

 Pediatric urine should be routinely screened

using reducing substances:
 Benedict’s or Clinitest copper reduction test
 Suggestive of carbohydrate metabolism
disorder
 (+) copper reduction test combined with
 (-) reagent strip glucose oxidase test
end