Objective:To analyse the effect of HIV-1 infection on the glucose metabolism of human intestinal epithelial cells.Methods:HT-29 cells were infected with HIV-1NDKand studied 3 weeks (acutely infected cells) or 9 months (chronically infected cells) post-infection. Perchloric acid extracts were analysed by high-resolution1H,31P and13C nuclear magnetic resonance spectroscopy. Metabolite concentrations and specific13C enrichments were quantified for chronically infected, acutely infected and control cells grown in Dulbecco's modified Eagle's medium containing natural-abundance or 1-13C-enriched glucose to determine significant differences between infected and non-infected cells.Results:Chronically HIV-infected cells showed alterations in glycerol-3-phosphate (+40%), fructose-1,6-diphosphate (−66%), uridine diphosphate glucuronic acid (−33%), lactate (+75%) and [1-13C]glucose (+181%) levels, and in specific lactate 3-13C enrichment (+19%) when compared with controls. Acutely infected cells exhibited decreased fructose-1,6-diphosphate (−58%) and increased nicotinamide adenine dinucleotide (+33%) levels relative to controls.Conclusion:HIV-1 infection results in a disturbance of glycolytic and oxidative activities in human intestinal epithelial cells. This finding supports the concept that HIV-1 may directly impair some metabolic functions of the intestinal epithelium, and that it can be considered a potential aetiological agent for HIV-associated enteropathy.