Amino acid disorders screening is performed to detect inherited and acquired disoders in amino acid metabolism. Tests are most commonly done in the neonatal period.
Twenty of the 100 known amino acids are the main building blocks for human proteins. Of these 20 amino acids, ten (essential amino acids) are not made by the body and must be acquired through diet. Proteins regulate every aspect of cellular function. The goal of amino acid disorder screening is early detection and intervention. Errors of amino acid metabolism range from asymptomatic or benign to life-threatening. In some cases, management by dietary restriction of the amino acid(s) involved can prevent damage or symptoms.
All neonates should be screened for the most prevalent aminoacidurias within a few days of birth. Usually this includes tests for phenylketonuria, tyrosinemia, lysinemia, maple syrup urine disease, and homocystinuria using a dried spot of blood collected on a paper card. When an infant displays symptoms such as failure to thrive, repeated vomiting, or nervous system impairment and an amino acid disorder is suspected, testing may be done on either plasma or urine. The patient should be fasting for the blood test. Blood samples are usually collected by heel puncture. The nurse or phlebotomist should observe universal precautions for the prevention of transmission of bloodborne pathogens when collecting the sample. Blood samples show considerable variance depending upon the time of sampling, and urine testing avoids this problem. Urine testing should be performed on a first morning voided sample or a 24-hour urine sample. Some drugs may affect the results of amino acid tests by certain methods of analysis. Any medications being given to the patient should be reported to the laboratory performing the test. Antibiotics will adversely affect the growth of the bacterium used in the Guthrie bioassay test for phenylalanine. Mothers of breastfeeding infants should report any medications they are taking, since these can pass from mother to child in breast milk.
An enzyme deficiency that blocks the metabolism of an amino acid is called a primary aminoaciduria. An error in absorption or transport of an amino acid is called a secondary aminoaciduria. Both primary and secondary aminoacidurias may be inherited (congenital) or acquired. Aminoacidurias may be grouped into two types, overflow and renal, based on the mechanism by which the amino acids accumulate. In the overflow type, the plasma level of one or more amino acids will be increased (aminoacidemia). The most common overflow type is phenylketonuria (PKU). The incidence of PKU varies with race and ethnic origin and there are 5 different forms of the disease. The most prevalent is type I which occurs in approximately 1 in 10,000 births in the United States, and accounts for about 50% of PKU cases. Because of its high prevalence all states screen newborns for PKU. This condition is caused by an inherited deficiency of the enzyme phenylalanine hydroxylase. The enzyme is needed to convert phenylalanine to tyrosine. Phenylalanine accumulates in the plasma and is oxidized by phenylalanine transaminase to phenylpyruvic acid which is excreted in the urine. Phyenylpyruvic acid is a ketoacid, hence the name phenylketonuria. Mental retardation occurs as a consequence of PKU, but can be prevented by withholding phenylalanine from the diet. There are many other primary aminoacidurias. Several are caused by enzyme deficiencies of the urea cycle. These give rise to increased blood ammonia levels and an increase of one or more amino acids.
In renal type aminoacidurias, one or more amino acids are excreted in excess in the urine but plasma levels are not increased. Some renal type aminoacidurias are inherited, but many are acquired or secondary to other diseases. A common inherited renal type aminoaciduria is cystinuria which results from a defect in the reabsorption of dibasic aminoacids by the kidney.
Newborn screening for PKU, tyrosinemia, lysinemia, maple syrup urine disease, and homocystinuria are often performed on a spot of whole blood collected from a heelstick. The test most often used is a bioassay called the Guthrie test. The test is based upon the ability of blood with an excess of the respective amino acid to overcome the effect of a specific inhibitor of the bacterium B. subtilis. For example, B. subtilis requires phenylalanine for growth. The growth of a culture of the organism can be prevented by adding thienylalanine to the culture medium. However, if excessive phenylalanine is present in the blood, it will overcome the effect of the inhibitor permitting the organism to grow. An alternative screening procedure for excessive phenylalanine that can be performed on a blood spot is the measurement of fluorescence after extraction and reaction with a reagent containing ninhydrin.
Young children with acidosis (accumulation of acid in the body), severe vomiting and diarrhea, or urine with an abnormal color or odor, are also tested for abnormal levels of amino acids. The most widely used method is high-performance liquid chromatography because it separates and quantifies all amino acids in the sample. Urine is usually the preferred specimen since it will detect both overflow and renal type aminoacidurias. The most common approach is to separate the amino acids on an ion exchange column using stepwise elution by changing the pH of the mobile phase. As amino acids emerge from the column they are reacted with ninhydrin and the absorbance of derivative formed is measured using a photometric detector.
Before the blood test, the patient must not eat or drink for at least four hours. Failure to fast will alter the results of the test. The patient should eat and drink normally before the urine test.
The patient may feel discomfort when blood is drawn from a heel capillary or vein. Bruising may occur at the puncture site or the person may feel dizzy or faint. Pressure should be applied to the puncture site until the bleeding stops to reduce bruising. Warm packs can also be placed over the puncture site to relieve discomfort.
There are no particular risks associated with either blood or urine tests for amino acids. Occasionally minor bruising may occur at the site where the blood was taken.
Guthrie test results are reported as either positive or negative. Results of quantitative assay for each amino acid should be compared to age-related reference ranges. Representative normal ranges for phenylalanine are shown below.
- Guthrie test: Negative or less than 2 mg/dL.
- Plasma: neonate 1.2-3.4 mg/dL (40-110 micromol/L) (premature infant 2.0-7.5 mg/dL).
- Urine: neonate 1.2-1.7 mg/day (0-200 micromol/gram creatinine).
Heath care team roles
Amino acid tests are ordered and interpreted by a physician. The heel stick or venipuncture is performed by a nurse or phelbotomist. Amino acid analysis is performed by a clinical laboratory scientist/medical technologist. If an aminoacidopathy is suspected, the family is typically referred to a genetic counselor to discuss prognosis and recurrence risks. DNA tests are available for prenatal testing of some aminoacidurias.
- Amino acid
- An organic compound containing both an amino group and an acidic carboxyl group; amino acids are the basic building blocks of proteins.
- A condition involving abnormal amino acid levels in the blood or urine.
- A family of laboratory techniques that separate mixtures of chemicals into their individual components.
- A biological catalyst that increases the rate of a chemical reaction without being used up in the reaction.
- The sum of all the chemical and energy reactions that take place in the human body.