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Phenylketonuria- An overview
Explain why phenylketonurics are warned against eating products containing the artificial sweetener aspartame (Nutrasweet; chemical name L-Aspartyl-L-Phenylalanine methyl ester)?
Discuss the biochemical defect, clinical manifestations, laboratory diagnosis and treatment of Phenylketonuria.
Aspartame contains Aspartic acid and phenyl alanine. The patients suffering from Phenylketonuria have high levels of phenyl alanine, any further increase in phenylalanine can prove harmful to the patient.
(See the details below )
Phenylketonuria (PKU) is an inherited error of metabolism caused by deficiency of the enzyme phenylalanine hydroxylase. Loss of this enzyme results in mental retardation, organ damage, and unusual posture and can, in cases of maternal PKU, result in severely compromised pregnancy.
Classic PKU and the other causes of hyperphenylalaninemia affect about one of every 10,000 to 20,000 Caucasian or Oriental births. The incidence in African Americans is far less. These disorders are equally frequent in males and females.
Deficiency of the enzyme phenylalanine hydroxylase or of its cofactor (Figure-1) causes accumulation of phenylalanine in body fluids and the central nervous system (CNS).
Figure-1- showing the conversion of phenyl alanine to Tyrosine. The reaction is catalyzed by phenyl alanine hydroxylase . The enzyme requires tetrahydrobiopterine as a cofactor.
Overview of Phenyl alanine metabolism –Phenyl alanine is metabolized through formation of Tyrosine. The first enzyme in the catabolic pathway for phenylalanine (Figure-1 and 2), phenylalanine hydroxylase, catalyzes the hydroxylation of phenylalanine to tyrosine. Phenylalanine hydroxylase inserts one of the two oxygen atoms of O2 into phenylalanine to form the hydroxyl group of tyrosine; the other oxygen atom is reduced to H2O by the NADH/NADPH also required in the reaction. This is one of a general class of reactions catalyzed by enzymes called mixed-function oxidases , all of which catalyze simultaneous hydroxylation of a substrate by O2 and reduction of the other oxygen atom of O2 to H2O. Phenylalanine hydroxylase requires a cofactor,tetrahydrobiopterin, which carries electrons from NADH/NADPH to O2 in the hydroxylation of phenylalanine. During the hydroxylation reaction the coenzyme is oxidized to dihydrobiopterin (Figure-2). It is subsequently reduced again by the enzyme dihydrobiopterin reductase in a reaction that requires NADH/NADPH.
Figure-2- Showing the metabolism of phenylalanine . The different enzyme deficiencies cause different disorders with different clinical manifestations. Under normal conditions phenylalanine is catabolized to produce fumarate and acetoacetate, thus it is both glucogenic as well as ketogenic.
The severity of hyperphenylalaninemia depends on the degree of enzyme deficiency and may vary from very high plasma concentrations (>20mg/dL, or >1200µM, “classic PKU”) to mildly elevated levels (2–6mg/dL or 120–360µM). In affected infants with plasma concentrations over 20mg/dL, excess phenylalanine is metabolized to phenylketones (phenylpyruvate and phenyl acetate through a secondary pathway of phenylalanine metabolism. In this minor pathway phenylalanine undergoes transamination with pyruvate to yield phenylpyruvate (Figure 3) Phenylalanine and phenylpyruvate accumulate in the blood and tissues and are excreted in the urine: hence the name of the condition, phenylketonuria. Much of the phenylpyruvate is either decarboxylated to produce phenylacetate or reduced to form phenyllactate. Phenylacetate is excreted in conjugation form with Glutamine that imparts a characteristic odor (Mousy odor) to the urine and that has been used to detect PKU in infants. The accumulation of phenylalanine or its metabolites in early life impairs the normal development of the brain, causing severe mental retardation. Excess phenylalanine may compete with other amino acids for transport across the blood-brain barrier, resulting in a depletion of some required metabolites.
Figure-3- Alternative pathways for catabolism of phenylalanine in phenylketonurics. Phenylpyruvate accumulates in the tissues, blood, and urine. Phenylacetate and phenyllactate can also be found in the urine.
- The affected infant is normal at birth.
- Mental retardation may develop gradually and may not be evident for the first few months. It is usually severe, and most patients require institutional care if the condition remains untreated. Mental retardation is due to direst toxic effect of phenyl alanine as well as due to impaired formation of catecholamines .
- Vomiting, sometimes severe enough to be misdiagnosed as pyloric stenosis, may be an early symptom.
- Older untreated children become hyperactive with purposeless movements, rhythmic rocking, and athetosis.
- On physical examination these infants are fairer in their complexion (Figure-5) than unaffected siblings. There is impaired formation of melanin, since tyrosine is a precursor of melanin (Figure-4).
- Some may have a seborrheic or eczematous rash, which is usually mild and disappears as the child grows older.
- These children have an unpleasant odor of phenyl acetic acid, which has been described as musty or mousy.
- There are no consistent findings on neurologic examination. However, most infants are hypertonic with hyperactive deep tendon reflexes.
- About 25% of children have seizures, and more than 50% have electroencephalographic abnormalities.
- Microcephaly, prominent maxilla with widely spaced teeth, enamel hypoplasia, and growth retardation are other common findings in untreated children.
Figure-4- showing the metabolic role of tyrosine. There is impaired formation of melanin and catecholamines , manifested by blond hair, lighter skin and mental retardation.
MILDER FORMS OF PKU
In any screening program for PKU, a group of infants are identified in whom initial plasma concentrations of phenylalanine are above normal (2mg/dL, 120µM) but less than 20mg/dL (1200µM). These infants do not excrete phenylketones. Clinically, these infants may remain asymptomatic but progressive brain damage may occur gradually with age. These patients have milder deficiencies of phenylalanine hydroxylase or its cofactor tetrahydrobiopterin (BH4) than those with classic PKU.
2.Hyperphenylalaninemia from Deficiency of the Cofactor Tetrahydrobiopterin (BH4)-
In 1–2% of infants with hyperphenylalaninemia, the defect resides in one of the enzymes necessary for production or recycling of the cofactor BH4 .These infants are diagnosed as having PKU, but they deteriorate neurologically despite adequate control of plasma phenylalanine. BH4 is the cofactor for phenylalanine, tyrosine, and Tryptophan hydroxylase. The latter two hydroxylase are essential for biosynthesis of the neurotransmitters dopamine and serotonin.
Plasma phenylalanine levels may be as high as those in classic PKU or in the range of milder forms of hyperphenylalaninemia. Neurologic manifestations, such as loss of head control, truncal hypotonia (floppy baby), drooling, swallowing difficulties, and myoclonic seizures, develop after 3 months of age despite adequate dietary therapy.
3. Maternal Phenylketonuria
A number of women with Phenylketonuria who have been treated since infancy will reach adulthood and become pregnant. If maternal phenylalanine levels are not strictly controlled before and during pregnancy, their offspring are at increased risk for congenital defects and Microcephaly. After birth, these children have severe mental and growth retardation. Pregnancy risks can be minimized by continuing lifelong phenylalanine-restricted diets and assuring strict phenylalanine restriction 2 months prior to conception and throughout gestation.
Figure-5- showing blond hair and eczematous rashes in a child suffering from PKU
Because of gradual development of clinical manifestations of hyperphenylalaninemia, early diagnosis can only be achieved by mass screening of all newborn infants.
NEONATAL SCREENING FOR HYPERPHENYLALANINEMIA
- The bacterial inhibition assay of Guthrie,(Figure-6) which was the first and still the most widely used method for the purpose, is being replaced by more precise and quantitative methods (fluorometric and tandem mass spectrometry).
- Blood phenylalanine in affected infants with PKU may rise to diagnostic levels as early as 4hr after birth even in the absence of protein feeding. It is recommended, however, that the blood for screening be obtained in the first 24–48hr of life after feeding protein to reduce the possibility of false-negative results, especially in the milder forms of the condition
- Plasma Phenyl Alanine levels In infants with positive results from the screen for hyperphenylalaninemia, diagnosis should be confirmed by quantitative measurement of plasma phenylalanine . A normal blood phenylalanine level is about 1 mg/dl. In classic PKU, levels may range from 6 to 80mg/dl, but are usually greater than 30mg/dl. Levels are somewhat less in the other disorders of hyperphenylalaninemia. .
- Identification and measurement of phenylketones in the urine has no place in any screening program. However, in countries and places where such programs are not in effect, identification of phenylketones in the urine by ferric chloride may offer a simple test for diagnosis of infants with developmental and neurologic abnormalities.
- Once the diagnosis of hyperphenylalaninemia is established, deficiency of cofactor (BH4) should be ruled out in all affected infants.
- BH4 loading test. An oral dose of BH4 (20mg/kg) normalizes plasma phenylalanine in patients with BH4 deficiency within 4–8hr.
- Enzyme assay-The activity of Dihydropteridine Reductase can be measured in the dry blood spots on the filter paper used for screening purposes. The other enzymes required for the synthesis of BH4 can also be similarly estimated.
Figure-6- showing Guthrie card
The goal of PKU treatment is to maintain the blood level of phenylalanine between 2 and 10 mg/dl. Some phenylalanine is needed for normal growth. This requires a diet that has some phenylalanine but in much lower amounts than normal.
High protein foods, such as: meat, fish, poultry, eggs, cheese, milk, dried beans, and peas are avoided. Instead, measured amounts of cereals, starches, fruits, and vegetables, along with a milk substitute are usually recommended.
In some clinics, a phenylalanine ‘challenge’ may be suggested to evaluate whether or not the child continues to require a low phenylalanine diet. This test identifies those few persons with a transient or ‘variant’ form of the disorder.
No dietary restriction is currently recommended for infants whose phenylalanine levels are between 2–6mg/dL. Plasma concentrations of phenylalanine in treated patients should be maintained as close to normal as possible.
Because phenylalanine is not synthesized by the body, “over treatment” may lead to phenylalanine deficiency manifested by lethargy, failure to thrive, anorexia, anemia, rashes, diarrhea, and even death; moreover, tyrosine becomes an essential amino acid in this disorder and its adequate intake must be ensured.
The current recommendation is that all patients be kept on a phenylalanine-restricted diet for life, in order to promote maximal development and cognitive abilities.
Oral administration of the cofactor (BH4 ) to patients with milder forms of hyperphenylalaninemia from phenylalanine hydroxylase deficiency may produce significant reductions.
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