Stable-isotopic methods were employed to evaluate the utilization of dietary pyridoxine-5'-beta-D-glucoside (PN-glucoside), a major form of vitamin B-6 in plant-derived foods, as a source of available vitamin B-6 for adult men (20-35 y old, n = 5). Deuterium-labeled forms of free pyridoxine (PN) and PN-glucoside were compared using the urinary excretion of labeled forms of the vitamin B-6 metabolite 4-pyridoxic acid as the main index of absorption and metabolism. When comparing orally administered, isotopically labeled PN and PN-glucoside in separate groups of subjects, similar bioavailability was observed although within-group variability was high. A dual-label study designed to examine the bioavailability of these compounds when administered simultaneously indicated that the utilization of deuterated PN-glucoside was 58 +/- 13% (mean +/- SEM) relative to that of deuterated PN. PN-glucoside was detected in all urine samples, which provided additional evidence of incomplete metabolic utilization. In contrast, intravenously administered PN-glucoside underwent approximately half the metabolic utilization of oral PN-glucoside. These studies indicate that the bioavailability of dietary PN-glucoside, although incomplete, is substantially greater in humans than previously found in rats. In addition, the difference between oral and intravenous routes suggests a role of beta-glucosidase(s) of the intestinal mucosa, microflora, or both in the release of free PN from dietary PN-glucoside.