Year
1976
Degree Name
Doctor of Philosophy
Recommended Citation
Gan, Ignatius Eng Tho, The application of gas chromatography and mass spectrometry for the diagnosis and study of genetic disorders - the use of stable isotope labelled substrates for the study of two patients with non-hepatic tyrosinaemia, Doctor of Philosophy thesis, , University of Wollongong, 1976. https://ro.uow.edu.au/theses/4475
Abstract
Deuterated tyrosine has been used to study tyrosine metabolism in one normal and two patients with rare forms of non-hepatic tyrosinaemia. The labelled tyrosine was ingested orally, after which urine and serum samples were examined by GC-MS at regular intervals for residual labelled tyrosine and tyrosine metabolites.
The derivatives of choice for GC-MS were the O-methyl ether of the N-neopentylidene tyrosine methyl ester, the trimethyl silyl derivatives of the phenolic acids and the N-trifluoroacetyl O-trimethyl silyl derivatives of the catecholamines.
It was shown that loading rates of 220 μmoles of deuterated tyrosine/kg body weight were adequate for precise measurement of the stable isotope content of urinary and serum tyrosine and tyrosine metabolites. This load approximates to that provided by a protein containing meal, so these stable isotope load studies can be claimed to mimic physiological conditions.
The amount of deuterated substrate used in this study was not considered a health risk, as even complete isotope retention in vivo would only increase the natural abundance in the body by less than 20%.
Deuteration experiments were made with a combined GC-MS instrument operating under computer control using time averaging and mass fragmentometry. The studies on the two patients and a normal control were carried out under conditions of high and low tyrosine loads. The results of these studies defined the biochemical lesion in one patient at the site of p-hydroxyphenylpyruvic acid oxidase. Although the site of the primary defect in the other patient has not been established. It is clear that it is not at the site of p-hydroxyphenylpyruvic acid oxidase. This patient suffered from substrate inhibition of p-hydroxyphenylpyruvic acid oxidase when on a normal diet. Resolution of the clinical and biochemical symptoms with disappearance of the inhibition of p-hydroxyphenylpyruvic acid oxidase occurred with the implementation of a diet low in phenylalanine and tyrosine. Although the fundamental defect in this patient is not resolved, the findings appear compatible with a defect in hepatic soluble tyrosine aminotransferase. The rates of deuteration of tyrosine metabolites have allowed us to distinguish between individuals having different clinical manifestations but similar biochemical abnormalities.
Unless otherwise indicated, the views expressed in this thesis are those of the author and do not necessarily represent the views of the University of Wollongong.