Alanine is a major end product of metabolism by Giardia lamblia: a proton nuclear magnetic resonance study.

1H-NMR spectroscopy was used to monitor the major metabolic end products released by Giardia lamblia when maintained anaerobically in culture in Diamond's TYI-S-33 medium. Spectra were acquired for the cell-free medium and the resonances of metabolites utilised and produced during cell growth identified by the addition of pure compounds and by difference spectroscopy. The major metabolites produced by the parasite were alanine, ethanol and acetate, with increases in concentrations in the media after 4 days' growth (end of log phase) of 18, 15 and 4 mM, respectively. The production of both alanine and ethanol approximated to cell growth, with ethanol formation lagging behind alanine during log growth but predominating after the parasites entered stationary phase. Acetate was formed at a more constant rate during growth. Glucose utilisation was sufficient to account for only 50% of the total carbon appearing in alanine, ethanol and acetate. The aminotransferase inhibitors L-cycloserine and carboxymethoxylamine inhibited growth and selectively inhibited the production of alanine. Analysis of the amino acid composition of the medium by HPLC showed that the only amino acid produced, apart from alanine, was proline, which increased in concentration in the medium by 4 mM after 4 days. There was also a 7 mM increase in ammonia over the same period. The only amino acids that were utilised were arginine and the components of an unresolved peak comprising serine, asparagine and glutamine.(ABSTRACT TRUNCATED AT 250 WORDS)

Metabolic studies of the protozoan parasite, Crithidia luciliae, using proton nuclear magnetic resonance spectroscopy.

Proton nuclear magnetic resonance (NMR) spectroscopy was used to follow glucose metabolism in Crithidia luciliae. Parasites were grown aerobically and anaerobically in culture, with glucose as the major carbon source and 1H NMR spectra were acquired for the cell free medium. The 1H NMR resonances of metabolites utilised and produced during cell growth were identified by difference spectroscopy, and quantitated from standard curves using 3-trimethylsilyl propionate-2,2,3,3-d4 sodium salt as an internal standard. The major metabolites produced by C. luciliae grown aerobically on 8 mM glucose were succinate, pyruvate, acetate and ethanol, in final concentrations in the media when the cells entered stationary phase of 8.5 +/- 0.5, 5.0 +/- 0.3, 2.1 +/- 0.2 and 2.5 +/- 0.6 mM, respectively. The production of succinate and pyruvate, but not acetate and ethanol, followed closely the growth curve of the parasites. Succinate was also measured enzymically and glucose using an autoanalyser. In both cases the results correlated well with the NMR data. The amounts of end products formed were greater than could be accounted for by the utilisation of glucose or any other metabolite observable in the 1H NMR spectra. There was approximately one extra atom of carbon for each molecule of succinate formed, supporting the view that succinate is produced via phosphoenolpyruvate carboxykinase and carbon dioxide fixation. Anaerobically the same major metabolites were produced, but with a decreased ratio of succinate to acetate and ethanol. The formation of glycerol from glucose was not observed under these conditions.