It Is Known That Fetal Urine Is Produced at 14Th Week of Gestation and Increases in Amount with Gestational Age (Jeffecoate and Scott 1959)

Tohoku J. exp. Med., 1974, 113, 169-171

Free Amino Acid Levels in Amniotic Fluid of Fetuses Affected with Lowe's Syndrome or Phenylketonuria

KEIYA TADA,* SHINOBU HIGAMI,* AKIE FUJIMOTO* and YUKIO OGITA •õ

Department of Pediatrics* and Department of Obstetrics and Gyneeology,•õ Osaka City University Medical School, Osaka

TADA, K., HIGAMI, S., FUJIMOTO, A. and OGITA, Y. Free Amino Acid Levels in Amniotic Fluid of Fetuses Affected with Lowe's Syndrome or Phenylketonuria. Tohoku J. exp. Med., 1974, 113 (2), 169-171 -Amniotic fluid obtained at 34th week of gestation of the fetus, which was proved to be affected with Lowe's syndrome after delivery, revealed elevated levels of most of free amino acids. This suggests a possibility of antenatal diagnosis of Lowe's syndrome by amino acid analysis of amniotic fluid, although accumulation of data is necessary especially in the early stage of gestation. Amniotic fluid at 19th week of gestation of a fetus at risk which was subsequently confirmed to be a homozygote of phenylketonuria showed normal levels of phenylalanine and tyrosine. This indicates the difficulty of prenatal diagnosis of phenylketonuria. Amniotic fluid was obtained at 20th week of gestation from a pregnant woman who had previously borne a child with hyperalaninemia with pyruvicemia due to a deficiency of pyruvate carboxylase in the liver. The concentrations of alanine, pyruvate and lactate in amniotic fluid were found to be within normal range; consequently she was delivered of a healthy girl. prenatal diagnosis; Lowe's syndrome; phenylketonuria

It is known that fetal urine is produced at 14th week of gestation and increases in amount with gestational age (Jeffecoate and Scott 1959). Therefore, the free amino acids in amniotic fluid may be of diagnostic value for fetuses at risk for inborn errors of amino acid metabolism. But this possibility remains unsolved becaues of the paucity of information. We had the oppotunities to investigate free amino acid levels in amniotic fluid of the fetuses at risk for Lowe's syndrome, phenylketonuria or hyperalaninemia.

MATERIALSAND METHODS 1. Case E.O., a boy, who showed glaucoma, cataract, muscular hypotonia, failure to thrive and mental retardation, was diagnosed as Lowe's syndrome (oculocerebrorenal syndrome) at the age of two months. His brother also had similar symptomsand died in infancy. Laboratory findings of Case E.O. revealed metabolicacidosis (bloodpH, 7.33; base excess,-9.5 mEq/liter; buffer base, 36.5 mEq/liter; standard bicarbonate, 16.8 mEq/ liter) and generalizedaminoaciduria of renal type. An amniocentesishad been made by obstetrical indication at 34th week of gestation of the patient. Using the amniotic fluid kept in freezing, free amino acids were retrospectivelyanalyzed. 2. An amniocentesiswas performedat 19th week of gestation in a pregnant woman, who had a previous child with phenylketonuria,for the purpose of chromosomeanalysis

Received for publication, April 15, 1974. 169 170 K. Tada et al.

TABLE 1. Free amino acids in amniotic fluid (ƒÊmoles/ml)

* Normal range was obtained from 6 samples .

according to parent's desire. Chromosome analysis of the cultured amniotic fluid cells showed no abnormality (46, XX). Free amino acids in amniotic fluid was also investi gated. 3. An amniocentesis was made at 20 week of gestation in a pregnant woman who had previously borne a child with hyperalaninemia with pyruvicemia due to a deficiency of pyruvate carboxylase in the liver. The details of the previous child were reported else where (Tada et al. 1973). The amniotic fluid was analyzed for free amino acids, pyruvate and lactate. Amniotic fluid obtained for chromosome analysis or obstetrical indications served as controls. Amino acids were quantitated with use of an automatic amino acid analyzer (Nihon densi JIC-6AH). One ml of amniotic fluid was treated with 3 ml of 4% sulfosalicylic acid and was centrifuged; the protein-free supernatant was applied to amino acid analyzer. The concentrations of pyruvate and lactate were determined by enzymatic method with use of Boehringer's reagent Kits.

RESULTS AND COMMENTS

Table 1 showed free amino acid levels in amniotic fluid of the fetus which was proved to be affected with Lowe's syndrome after delivery, in comparison with those of control specimens obtained from 32nd to 36th week of gestation. Most of amino acids in amniotic fluid of the fetus with Lowe's syndrome were found to be increased beyond normal range. This suggests a possibility of prenatal diag nosis of Lowe's syndrome by amino acid analysis of amniotic fluid, although more accumulation of data is necessary, especially in early stages of gestation. Table 2 showed free amino acid levels in amniotic fluid from a mother at risk for phenylketonuria and from a mother at risk for hyperalaninemia. The former Free Amino Acids in Amniotic Fluid 171

TABLE 2. Free amino acids in amniotic fluid (ƒÊmoles/ml)

* Normal range was obtained from 10 samples. showed normal levels of phenylalanine and tyrosine. After delivery, the child was confirmed to be a homozygote of phenylketonuria by elevation of blood phenylalan ine (22 mg/100 ml) at the fifth day of life when the dietary treatment low in phenylalanine was started. At the age of six months, the girl is developing normally under the dietary regimen. This finding indicates that phenylketonuria is difficult to detect antenatally. The fact that the activity of phenylalanine hydroxylase is physiologically very weak in the fetal liver and increases rapidly at the birth is in accord with the above observation. The latter case showed normal levels of alanine, pyruvate and lactate. Her pregnancy was continued and resulted in the delivery to a healthy girl who had normal levels of alanine, pyruvate and lactate in blood.

References 1) Jeffecoate,T.N. & Scott, J.S. (1959) Polyhydraminosand oligohydramnios. Canad. med. Ass. J., 80, 77-86. 2) Tada, K., Sugita, K., Fujitani, K., Uesakai, T., Takada, G. & Omura, K. (1973) Hyperalaninemiawith pyruvicemiain a patient suggestiveof Leigh's encephalomyelo pathy. TohokuJ. exp. Med., 109, 13-18.