In humans and pigs, hydrolysis of dietary polyglutamyl folates is carried out by intestinal brush border folate hydrolase (GCPII), while the transport of the monoglutamyl folate derivatives occurs via the intestinal brush border reduced folate carrier (RFC). The study objective was to measure the expression of intestinal GCPII and RFC during postnatal development of pigs and their effects on plasma and liver folate concentrations. Duodenum, jejunum, ileum, liver, and plasma samples were collected from female Yorkshire pigs at birth, 24 hours, 1 week, 3 weeks, and 6 months (n=6 at each time point). GCPII mRNA transcripts and protein (normalized using -actin), and enzyme activity (normalized per mg mucosal protein) were highest in all segments of small intestine at birth and were undetectable in ileum after 1 wk, while jejunal protein and activity predominated at 6 mo. RFC mRNA transcripts were present in all segments of small intestine at birth and declined significantly throughout development to 6 mo. Conversely, RFC protein increased 2 fold during the first 24 hours and remained constant throughout development in all segments of small intestine. Liver RFC mRNA transcripts were detected at birth but were reduced by 6mo. Liver folate concentration increased throughout postnatal development, whereas plasma folate levels increased during the first 24 hours, but decreased over time, reflecting the pattern of RFC expression in small intestine. These findings show that intestinal GCPII and intestinal and hepatic RFC all exhibit ontogenic changes in the pig which is reflected in post-natal folate status.