Acid In Amniotic Fluid Research Articles

Chemical analysis of succinylacetone and 4-hydroxyphenyllactate in amniotic fluid using selective ion monitoring.

A method for the measurement of the concentration of succinylacetone and 4-hydroxyphenyllactic acid in amniotic fluid was developed for the prenatal diagnosis of hereditary tyrosinemia. Succinylacetone was converted to 5-methyl-3-isoxazolepropionic acid and isolated with 4-hydroxyphenyllactic acid by liquid partition chromatography and the trimethylsilyl derivatives quantified by ammonia chemical ionization selected ion monitoring gas chromatography-mass spectrometry with 2-hydroxy-n-caproic acid as the internal standard. The concentration of 4-hydroxyphenyllactic acid in normal amniotic fluid was 1.97 +/- 0.75 (S.D.) mumol/l while succinylacetone was undetectable. A pregnancy at risk for tyrosinemia type II was monitored. The concentration of 4-hydroxyphenyllactic acid was within the normal range and a healthy child was born.

Prenatal diagnosis of glutaric aciduria type II by direct chemical analysis of dicarboxylic acids in amniotic fluid.

A method for the measurement of dicarboxylic acids in amniotic fluid was developed that utilizes isolation of the acids by liquid partition chromatography and quantification by ammonia chemical ionization selected ion monitoring, gas chromatography-mass spectrometry. The concentrations of dicarboxylic acids in ten normal samples of amniotic fluid (mumol/l +/- 1 S.D.) were glutaric acid 0.91 +/- 0.15, adipic acid 0.33 +/- 0.08, suberic acid 0.27 +/- 0.08, and sebacic acid 0.21 +/- 0.10. A highly elevated concentration of 14.48 mumol/l glutaric acid was found in the amniotic fluid of a pregnancy in which the fetus was affected with glutaric aciduria type II. Adipic, suberic and sebacic acids were also significantly elevated. The dicarboxylic acids were normal in the amniotic fluid of a pregnancy at risk for glutaric aciduria type II in which the fetus was unaffected. This method is suitable for the rapid prenatal diagnosis of glutaric aciduria types I and II and of potential value for the prenatal diagnosis of other inherited disorders in which dicarboxylic acids accumulate.

Stable isotope dilution analysis of 3-hydroxyisovaleric acid in amniotic fluid: contribution to the prenatal diagnosis of inherited disorders of leucine catabolism.

A quantitative assay for 3-hydroxyisovaleric acid in amniotic fluid was developed using D6-3-hydroxyisovaleric acid as an internal standard. 3-Hydroxyisovaleric acid was isolated by liquid partition chromatography and the amount determined by selected ion monitoring, ammonia chemical ionization gas chromatography-mass spectrometry of the trimethylsilyl derivatives. The concentration of 3-hydroxyisovaleric acid in ten normal amniotic fluid was 4.52 +/- 1.73 mumol/l. The level was elevated eight-fold in the amniotic fluid from a pregnancy resulting in the birth of a child with biotin-responsive multiple carboxylase deficiency. The stable isotope dilution assay of 3-hydroxyisovaleric acid in amniotic fluid is a rapid, sensitive and accurate method for the prenatal diagnosis of this disorder, and may be of value in the prenatal diagnosis of other inherited disorders of leucine catabolism.

Antenatal Diagnosis of Defects of Ureagenesis

The approach usually taken to the intrauterine diagnosis of inborn errors of metabolism is to assay a particular enzyme in cultured amniotic cells, the rationale being that if the enzyme is normally present, its absence can be used to indicate fetal disease. Of the five inborn errors of ureagenesis, however, only two, ie citrullinemia and argininosuccinic aciduria, can be diagnosed in this manner because argininosuccinic acid synthetase and argininosuccinic acid lyase are the only urea cycle enzymes normally found in these cells. Both of these disorders have now been detected in utero,87,88 (S. D. Cederbaum and C. R. Scott, unpublished observations, 1980), and several other fetuses at risk have been correctly diagnosed as unaffected88-90 (S. I. Goodman, unpublished observations, 1980). Enzyme deficiency in cultured amniotic cells can be demonstrated directly21,22,91 or by examining the incorporation of 14C into arginine or cell protein.21,73,88,89,92 Further, fetuses with argininosuccinic aciduria have shown increased concentrations of argininosuccinic acid in amniotic fluid at amniocentesis87,88 (S. D. Cederbaum, unpublished observations, 1980) and delivery,93 with its amount remaining undetected when the fetus was not affected88 (S. I. Goodman, unpublished observations, 1980). Argininosuccinic acid is accumulated by the affected fetus, eg it is the most prominent free amino acid in its liver87,94 and, as it is poorly reabsorbed from the glomerular filtrate by cells of the proximal tubule,71 it probably enters the amniotic fluid in fetal urine.95 Thus, provided that a fetus has an intact urinary tract, whether or not it has argininosuccinic aciduria can probably be established within hours of amniocentesis merely by examining the fluid for the acid and/or its anhydrides by ion-exchange chromatography.

Isotope dilution analysis of methylcitric acid in amniotic fluid for the prenatal diagnosis of propionic and methylmalonic acidemia

A stable isotope dilution assay for methylcitric acid in amniotic fluid was developed to provide rapid prenatal diagnosis of the inherited disorders propionic acidemia and methylmalonic acidemia. The method utilizes two 2H 3-labeled diastereoisomers of methylcitric acid as internal standards, isolation by liquid partition chromatography and quantitation of the trimethyl esters by chemical ionization selected ion monitoring gas chromatography-mass spectrometry. Methylcitric acid at a concentration of 0.38 ± 0.10 μmol/1 was detected in normal amniotic fluid. Highly elevated levels of 7.87 and 9.16 μmol were found in the fluids surrounding fetuses affected with propionic acidemia and levels of 1.79, 2.72 and 12.27 μmol were found for fetuses with methylmalonic acidemia. Methylcitric acid was not elevated in the amniotic fluid of a fetus heterozygous for propionic acidemia. In the five pregnancies at risk for propionic acidemia, and three pregnancies at risk for methylmalonic acidemia, the levels of methylcitric acid in amniotic fluid gave the diagnosis in all cases. Measurement of methylcitric acid in amniotic fluid therefore provides a rapid and reliable method for the prenatal diagnosis of these genetic disorders.

Gas chromatographic determination with flash methylation of palmitic acid in amniotic fluid in prediction of fetal lung maturity

A new method for determination of palmitic acid in amniotic fluid was developed. The dipalmitoyl lecithin was hydrolysed, and after extraction the palmitic acid was flash methylated in the injector of the gas Chromatograph. The between batch coefficient of variation was 6.0% at 40 and 70 μmol/1 of palmitic acid concentrations in 500 μ1 amniotic fluid. The accuracy of the method was demonstrated by linearity, recovery and correlation studies. The correlation coefficient was 0.98 between the new and bortrifluoride methylating method. The concentrations of lecithin and palmitic acid in amniotic fluid showed a correlation coefficient of 0.97. If the concentration of palmitic acid falls below 80 μmol/1 respiration distress syndrome is likely. The new method was simple and easy to carry out and could be adjusted for haemoglobin contamination of the amniotic fluid. It was concluded that the determination of palmitic acid in amniotic fluid was a reliable method for the assessment of fetal lung maturity that could replace the determination of lecithin concentration.

Pattern of branched-chain α-keto acids in amniotic fluid

Pattern of branched-chain α-keto acids in amniotic fluid

Quantitation of organic acids in amniotic fluid by gas chromatography

The concentrations of 13 major organic acids in normal human amniotic fluid have been determined by gas chromatography. Each acid showed a large range and an irregular frequency distribution.

Prenatal Diagnosis of Propionic Acidemia

Prenatal diagnosis of a fetus with propionic acidemia has been accomplished by the detection of methylcitrate, a unique metabolite, in the amniotic fluid by liquid partition chromatography and gas chromatography-mass spectrometry. The diagnosis was confirmed by demonstration of deficient activity of propionyl-CoA carboxylase in cells cultured from the amniotic fluid and in fetal tissues. In two subsequent pregnancies, methylcitrate was not present in amniotic fluid. Enzyme assay indicated that one fetus was heterozygous and the other was normal, and healthy infants were born at term. The analysis of organic acids in amniotic fluid permits very rapid prenatal diagnosis, usually within 48 hours after obtaining the fluid. It may provide a general method in conditions in which an unusual metabolite can be identified.

Lysozyme in amniotic fluid during Rivanol-induced second trimester abortion

The concentrations of lysozyme and free arachidonic acid in amniotic fluid were determined in eight patients during induction of second trimester abortion. Abortion was induced by extraamniotic instillation of 150 ml 0.1% Rivanol. Lysozyme increased in all patients during the induction reaching significancy after 60% of the induction-abortion time. This suggests a leakage of lysosomal enzymes from the fetal membranes. Free arachidonic acid relative to total free fatty acids increased parallell to lysozyme.

Arachidonic acid in amniotic fluid after extra-amniotic instillation of rivanol for midtrimester abortion

Free arachidonic acid was measured by gas liquid chromatography in amniotic fluid in 13 women during induction of midtrimester abortion. The abortions were induced by extraamniotic instillation of 0.1% Rivanol. Serial sampling of amniotic fluid were performed through a trans-abdominal catheter up to 22 hrs. Free, total arachidonic acid showed a significant increase from 26±8 ng/ml to 293±102 ng/ml at 22 hrs. The percentage of free arachidonic acid in total free fatty acids increased significantly from 2.2±0.5 to 6.1±1.6% during the same time. The C results suggest a selective release of arachidonic acid during Rivanol-induced abortion.

Organic acids in amniotic fluid

The organic acids of human amniotic fluid were studied by gas chromatography and mass spectrometry. About 30 acids were identified. A “normal” metabolic profile of acids was established which can provide the basis for the identification of abnormal patterns.

Rapid estimation of foetal lung maturity

A 40 min assay of total palmitic acid in amniotic fluid has been developed for routine use. The assay is simpler and more rapid than similar, published assays because of: (1) no initial extraction of lipids, (2) rapid evaporation of fatty acid extracts, and (3) no derivatization of fatty acids. Main stages include alkaline hydrolysis of whole amniotic fluid, and GLC using 10% SP-216-PS on Supelcoport, n-Pentadecanoic acid is used as the internal standard. Recoveries of added free or esterified palmitic acid are highly reproducible and almost quantitative. Palmitic acid levels increase rapidly after 37 wk gestation. 0.073 mmol/l is used as the threshold, higher values indicating foetal lung maturity. This assay has been compared with L/S estimations using 40 amniotic fluids. In 6 cases in which this and the L/S assay gave rise to different conclusions, the palmitic acid result coincided with the clinical state. The only false positive arose in one liquor containing meconium.

Fetal lung maturity, as assessed by gas-liquid chromatographic determination of phospholipid palmitic acid in amniotic fluid.

We describe a new and specific method for measurement of lecithin palmitic acid in amniotic fluid. Dipalmitoyl lecithin, the major alveolar surfactant, has previously been estimated by measuring the lecithin-sphingomyelin ratio, total lecithin, total phospholipid phosphorus, and (or) total palmitic acid. Our method is more specific for estimation of dipalmitoyl lecithin, because nonphospholipid sources of palmitic acid are removed by solvent extraction. Using a hexane/2-propanol/sulfuric acid system, we obviated the major interferences from triglycerides and free fatty acids. The palmitic acid derived from the phospholipid fraction is measured by gas-liquid chromatography of its methyl ester. No contribution appears to be made by sphingomyelin palmitic acid--probably owing to the mild hydrolysis conditions. The measured palmitic acid therefore appears to be derived from lecithins, principally dipalmitoyl lecithin. The value for palmitic acid determined by this method correlates well with the lecithin-sphingomyelin ratio and total phospholipid phosphorus. Infants are unlikely to develop respiratory distress syndrome when the measured palmitic acid in amniotic fluid exceeds 8.0 mg/liter, which corresponds to an lecithin-sphingomyelin ratio of 2.0.

Identification of 2-hydroxybutyric acid in human amniotic fluid

2-Hydroxybutyric acid has been identified in human amniotic fluid by combined gas chromatography and mass spectrometry. It was present in all 22 specimens examined of gestation 15 to 20 weeks and 30 to 40 weeks. The concentration of 2-hydroxybutyric acid in amniotic fluid was about 100 times less than that of the predominant acid, lactic acid.

Amino acid composition of amniotic fluid in intrahepatic cholestasis of pregnancy, pre-eclampsia and rhesus incompatibility.

The levels of free amino acids in amniotic fluid were determined in twenty patients with normal pregnancy, five with intrahepatic cholestasis of pregnancy, twelve with pre-eclampsia, and four with rhesus incompatibility. In pre-eclampsia there was no significant change in the amino acid composition of the amniotic fluid. In recurrent intrahepatic cholestasis the concentration of 15 out of the 22 amino acids studied was significantly higher than normal and the mean concentration of 4 others was more than 20 per cent above normal without this being statistically significant. In rhesus incompatibility there was a significant decrease in the concentration of 13 amino acids, and a decrease of more than 20 per cent in the mean concentration of 5 others.

Developmental changes of free amino acids in bovine fetal fluids with gestational age and the interrelationships between the amino acid concentrations in the fluid compartments.

Thirty ninhydrin-positive compounds were determined in the sera, amniotic fluid, stomach content and allantoic fluid from 29 bovine fetuses of various gestational ages. Fetal serum was found to contain about 3-fold higher concentrations of free amino acids (FAA) than maternal serum, and allantoic fluid contained about 3-fold higher concentration of FAA than fetal serum. Decreasing concentrations of FAA were found in serum as a function of the crown-rump length for the amino acids taurine, aspartic acid, threonine, serine, ornithine and lysine. Decreasing concentrations of FAA in allantoic fluid were found for threonine, alanine and ornithine, whereas increasing concentrations were found for phosphoserine and methionine as a function of the crown-rump length. Correlations were found between the concentrations of most amino acids in amniotic fluid and stomach content, but fewer correlations were found between the other fluid compartments. The transport of amino acids between compartments is discussed.

Creatinine, urea and uric acid in amniotic fluid, maternal and umbilical cord blood at delivery.

Amniotic fluid, maternal vein serum and umbilical cord serum were obtained almost simultaneously from 31 women after an uneventful course of pregnancy. Specimens of amniotic fluid contaminated by blood or meconium were excluded. Creatinine, urea and uric acid were measured by autoanalyser techniques. The concentrations of creatinine, urea and uric acid in amniotic fluid were significantly higher when compared with the levels in maternal vein serum or umbilical cord serum. Between maternal and umbilical cord serum there were only small or no differences. The linear correlations of creatinine, urea and uric acid between maternal and umbilical cord serum were greater than those between amniotic fluid and maternal serum of amniotic fluid and cord serum. It is suggested that the high levels of creatinine, urea and uric acid in amniotic fluid are primarily determined by fetal urine.

Amniotic fluid composition in malformations of the fetal central nervous system.

SummaryThe results are presented of measurements of optical density at 450nm (ΔOD450), 5‐hydroxyindole acetic acid (5‐HIAA), α‐fetoprotein (AFP) and amino acids in amniotic fluid at various stages of gestation in over 60 pregnancies in which the fetus had a neural tube defect namely anencephaly and/or spina bifida. Before 20 weeks' gestation neither ΔOD450 nor 5‐HIAA levels were abnormal. AFP levels were more reliable than amino acid concentrations for the antenatal diagnosis of neural tube defects, particularly in early pregnancy. Various interpretations for these findings are discussed including the possibility that ΔOD450 and AFP levels in amniotic fluid might reflect altered maternal metabolism.

The effect of frusemide on uric acid levels in maternal blood, fetal blood and amniotic fluid.

SummaryConcurrent measurements of uric acid levels in maternal serum, fetal serum and amniotic fluid were made in 52 patients. The patients with pre‐eclampsia who were not on frusemide had higher uric acid levels in all fluids studied than did the untreated controls and frusemide increased the uric acid levels further in both types of patient. The potentially harmful effect on the fetus of high uric acid levels induced by frusemide should be considered. Raised levels of uric acid in amniotic fluid may invalidate the use of this measurement as an index of fetal maturity in patients who have pre‐eclampsia or who are on diuretics.