Urine Screening for Metabolic DisordersAng. Avena. Blas. Tagarao. 3FMT

Metabolic Substances in the

Urine Overflow Type

- disruption of normal metabolic pathway

- increase in plasma concentrations from the non-metabolized substances.

Renal Type

- abnormal accumulations of substances that are caused by malfunctions in the mechanism of tubular reabsorption

Amino Acid Disorders

• Phenylalanine-Tryptophan Disorders• Branched Chain Amino Acid Disorders• Cysteine and Homocysteine Disorders• Porphyrin Disorders• Mucopolysaccharide Disorders• Purine Disorders• Carbohydrate Disorders

Phenylalanine-Tryptophan Disorders

• Phenylketonuria • Tyroslyuria• Melanuria• Alkaptonuria

Phenylketonuria (PKU)

Inherited autosomal recessive trait

absence of activity of the enzyme phenylalanine hydroxylase (PAH)

catalyzes the conversion of phenylalanine to tyrosine

Catabolites: phenylpyruvic acid, phenyllactic acid, phenylacetic acid, phenylacetylglutamine

Types: Mild PKU (600-1200 μmol/L) Non-PKU mild hyperphenylalaninenemia (180-600 μmol/L; no

phenylketones)

Can cause:Mental retardationsConvulsionsBehavior problems Skin rashMusty body odor

Urine: Musty Odor

Test: Ferric Chloride Tube Test

Procedure: 5 drops 10% FeCl3 + 1mL urine sample

FeCl3 reacts w/ phenylpyruvic acid = Permanent blue-green color.

Tyrosyluria

accumulation of excess tyrosine in the plasma (tyrosinemia)

either inherited or metabolic defect

Urine: excess tyrosine or , p-hydroxyphenylpyruvic acid and p-hydroxy phenyllatic acid.

Three types of Tyrosinemia:

1) Type I – low levels of the enzyme fumarylacetoacetate hydrolase, the last of five enzymes needed to break down tyrosine

2) Type II –a deficiency of the enzyme tyrosine aminotransferase, the first in a series of five enzymes that converts tyrosine to smaller molecules

3) Type III – deficiency of the enzyme 4-hydroxyphenylpyruvate dioxygenase, ncluded in the five enzymes needed to break down tyrosine

Test: Nitroso-Napthol Test

Procudure:

1mL of 2.63N nitric acid + 1 drop of 21.5% sodium nitrite + 0.1mL 1-nitroso-2-napthol + 5 drops of urine

Mix and wait for 5 minutes

Result: Orange-red color which indicates tyrosine metabolites

Melanuria Darkening of the urine = increase of melanin in

the body

Melanin - is the pigment responsible for the dark color of hair, skin and eyes.

Serious finding: if urinary melanin is elevated - indicates proliferation of the normal melanin-producing cells (melanocytes)

Urine: Dark urine

5,6-dihydoxyindole - a colorless precursor of melanin

secreted by Malignant melanoma oxidizes to melanogen melanin

Alkaptonuria Inborn metabolic disease transmitted as an autosomal

recessive gene

HDG gene - the lack of the enzyme homogentisate oxidase (needed in the metabolism of tyrosine and phenylalanine)

Test: Homogentisic Acid Test

Procedure:

0.5 mL of urine + 4 mL of 3% silver nitrite and mix

Result: Black color after 24 hours

Other tests

Ferric Chloride Test: transient deep blue color

Clinitest: Yellow precipitate

Branched Chain Amino Acid Disorders

• Maple Syrup Urine Disorders• Organic Acidemia • Indicanuria• 5-Hydoxyindolacetic Acid

Maple Syrup Urine Disorders (MSUD)

rare autosomal recessive genetic inherited disorder

absence or reduced activity of the enzyme branched-chain-α-ketoacid decarboxylase

blocking the normal metabolism of the three essential branched-chain amino acids leucine, isoleucine and valine.

1:150,000 births

Urine: Maple syrup or burnt sugar odor

Test: 2,4-Dinitrophenylhydrazine (DNPH) Test

Procedure

10 drops of 0,2% 2,4-DNPH + 2N HCL +1 mL urine

Wait for 10 minutes

Result: Yellow turbidity and precipitate.

Organic Acidemia

Three most frequently encountered disorders:

Isovaleric

Propionic

Methylmalonic

The presence of these three can be detected by newborn screening programs using MS/MS.

Isovaleric academia autosomal recessive metabolic disorder from a

deficiency of the enzyme isovaleryl-CoA dehydrogenase

preventing normal metabolism of the amino acid leucine.

1:250,000 births

Urine: Sweaty feet odor urine

Propionic and Methylmalonic Acidemia

due from the errors in the metabolic pathway converting isoleucine, valine and threonine and methionine to succinyl CoA.

Indicanuria Due to certain intestinal disorders, presence of

abnormal bacteria, malabsorption syndrome

Hartnup disease - increase amounts of tryptophan are converted to indole

Excess indole is then reabsorbed from the intestine into the bloodstream and circulated to the liver, converted to indican and then excreted in the urine.

Urine: Colorless

Urine exposed to air: oxidized to Indigo Blue.

5-Hydoxyindolacetic Acid (5-HIAA)

The degradation product of serotonin that is excreted in the urine normally in small amounts.

Serotonin - produced from the second metabolic pathway of tryptophan by the argentaffin cells in the intestine.

If there are carcinoid tumors involving the argentaffin cells develop = increase in the production of serotonin results in the elevation of urinary 5-HIAA levels.

Cysteine and Homocysteine

Disorders

• Cystinuria• Cystinosis• Homocystinuria

Cystinuria

inherited renal tubular disorder.

Two forms:

1. patients cannot reabsorb lysine, ornithine, arginine, or cysteine

2. only cysteine and lysine cannot be reabsorbed.

more likely to have pathological urinary cysteine crystals and stones composed of cysteine.

Cystine forms urinary calculi (insoluble, unless alkalinize)

Test: Cyanide nitroprusside test

Result: Red Purple Color

Treatment: More water intake (4L/day)PenicillinasePercutaneous nephrolithotripsy

Cystinosis

Inborn Errors of Metabolism (IEM) – can range from from severe and fatal in infancy to a milder adult form.

defect in the lysosomal membrane

prevents release of cystine into the cellular cytoplasm for metabolism

Leads to: Deposition of cysteine crystals in many cells of the body.

 Patients w/Fanconi syndrome deposits of cysteine in the cells of the proximal convoluted tubule interfere with reabsorption and of these crystal deposits in cells can lead to early renal failure.

Mild Form: No kidney involvement

polyuria and positive urine tests for reducing substances

Urinalysis tests = lack of ability to vary specific gravity.

Test: cyanide nitroprusside test

Result: Positive  

  

Homocystinuria Lack of the enzyme cystathionine β-synthase

necessary for the metabolism of the amino acid methionine

increased plasma and urine levels of methionine and of the precursor homocysteine.

Result to: failure to thrive mental retardation Cataractsincreased thrombosis riskDeath

Prevention: Diet modification with reduction in methionine

sourceshigh doses of Vitamin B6

Test: Cyanide nitroprusside test

Result: Positive

Other test:Silver Nitroprusside test – needed to differentiate

from the cystine disorders.

Homocystinuria = postive

Cystinuria = negative

Cystinosis = negative

Porphyrin Disorders

• Porphyrins – complex iron-fee cyclic substances; intermediates in the biosynthetic pathway of heme.

• Porphyrins differ in the side chains present at the eight available positions on the pyrrole rings.

• Major sites for Porphyrin production are:1. bone marrow - intermediates in the synthesis of

hemoglobin2. liver and other tissues - produce intermediates for

other heme proteins like myoglobin.

Porphyrins can be seen elevated in: urine, feces, and/or blood

Porphobilinogen (PBG) and the porphyrinogens -colorless, non-fluorescent substances

Oxidized forms or the Porphyrins have red pigments under the microscope

Urine: port wine / burgundy color

Common method for separating individual porphyrin: high performance liquid chromatography

Delta-aminolevulenic Acid Dehydrogenase Deficiency Porphyria Delta-aminolevulinic acid dehydratase (ALAD),

also known as porphobilinogen synthase, catalyzes the second step of heme synthesis. Deficiency of this enzyme produces ALAD deficiency porphyria (ADP), an extremely rare cause of acute porphyria.

a rare form of acute porphyria

aminolevulenic acid is increased

PBG is not increased

Acute porphyriasrapid testing is important.

Porphobilinogen is increased patients with symptoms of acute porphyrias

PBG testing is both sensitive and specific

Measurement of PBG is often combined with measurments for delta aminolevulenic acid and total urine porphyrins.

Cutaneous Porphyria

measuring total plasma Porphyrins is effective for screening patients with skin photosensitivity

Plasma Porphyrins are rarely increased in other medical conditions.

Mucopolysaccharide Disorders

• Mucopolysaccharidoses Hurler Syndrome Hunter Syndrome Sanfilippo Syndrome

Mucopolysaccharidesaka Glycosaminoglycans

group of large compounds located primarily in connective tissue (CT)

consists of a protein core with numerous polysaccharide branches

products most frequently found in urine are dermatan sulfate, keratan sulfate, and heparan sulfate

Mucopolysaccharidoses

inherited disorders in the metabolism of mucopolysaccharides

preventing complete breakdown of polysaccharide portion

accumulation of incompletely metabolized polysaccharide portion in lysosomes of CT cells

increased excretion in urine

Types

1. Hurler Syndrome

abnormal skeletal structure

severe mental retardation

mucopolysaccharides accumulate in the cornea of the eye 

2. Hunter Syndrome

abnormal skeletal structure

severe mental retardation

sex-linked recessive (rarely seen in females)

3. Sanfilippo Syndrome

mental retardation

 

Diagnosis: Urinary screening test – newborn

1. acid-albumin and CTAB turbidity test

2. metachromatic staining spot test

Result: white turbidity (+)

Purine Disorders• Lesch-Nyhan Disease

Carbohydrate Disorders

• Melituria

Lesch-Nyhan Syndrome inherited, sex-linked recessive

massive excretion of urinary uric acid crystals

severe motor defects

mental retardation

tendency towards self-destruction, gout, and renal calculi

Diagnosis

normal development for the first 6-8 months

First Sign: uric acid crystals in diaper resembling orange sand

Increased uric acid crystals in pediatric urine specimen

Melituria

increased urinary sugar

usually caused by an inherited disorder

Most cause no disturbance in body metabolism

Type:

1. Pentosuria

2. Galactosuria

3. Lactosuria

4. Fructosuria

Pentosuria presence of pentose sugars in urine

one of Garrod’s original six IEMs

ingestion of large amounts of fruit

Galactosuria inability to properly metabolize galactose to

glucose

deficiency in any of the enzymes:

1. galactose-1-phosphate uridyl transferase (GALT) - causes severe, possible fatal symptoms

2. galactokinase - result in cataracts in adulthood

3. UDP-galactose-4-epimerase

asymptomatic or produce mild symptoms

results to galactosemia with toxic intermediate metabolic products

infant failure to thrive

liver disorder, cataracts, severe mental retardation

Lactosuria

may be seen during pregnancy and lactation

Fructosuria

associated with parenteral feeding

Diagnosis

Tests:

1. Copper reduction test – postive

2. Reagent strip glucose oxidase test – negative

3. Newborn screening tests

• Graff’s Textbook of Routine Urinalysis and Body Fluids by Mundt 2nd ed.• Concise Book of Medical Laboratory Technology: Methods and

Interpretations• William Clarke’s Contemporary Practice in Clinical Chemistry, 2nd ed.• Henry’s Clinical Diagnosis and Management by Laboratory Methods, 22nd

ed. by Mcpherson• Uniralysis and Other Body Fluids by Strasinger 6th ed.• Clinical Laboratory Medicine by Richard Ravel• Fundamentals of Urine and Body Fluids Analysis by Brunzel, 3rd ed.• Clinical Laboratory Science: the Basics and Routine Techniques, 6th ed. by

Turgeon• Inborn Metabolic Diseases: Diagnosis and Treatment by Saudubray 5th ed• Medical Biochemistry: Human Metabolism in Health and Disease by

Rosenthal• Clinical Chemistry: Principles, Techniques and Correlations by Bishop 7th

ed. 

References