It has recently been proposed that basal muscle protein synthesis can be effectively assessed by measuring the background enrichment in total plasma protein, thereby omitting the initial biopsy, and determining the difference in enrichment from a single muscle biopsy obtained during a primed continuous infusion of isotope-labeled amino acids. We determined the reliability of calculating basal mixed muscle protein fractional synthetic rates (FSRs) from mixed plasma proteins and a single muscle biopsy compared against the sequential muscle biopsy approach. Ten men (age, 23 +/- 1 years; body mass index, 22 +/- 1 kgm(-2)) received muscle biopsies of the vastus lateralis after 2 and 4 hours of a primed continuous infusion of l-[ring-(13)C(6)]phenylalanine. Mixed muscle protein FSR was calculated from baseline plasma enrichments and muscle protein enrichments determined from the biopsy at 2 hours (1BX SHORT) or 4 hours (1BX LONG), or between muscle protein enrichments at 2 and 4 hours (2BX) of the infusion. No differences (P = .50) were observed in mixed muscle protein FSR, using plasma [ring-(13)C(6)]phenylalanine enrichments as the precursor, between the 1BX SHORT (0.031% +/- 0.010%h(-1)), 1BX LONG (0.032% +/- 0.007%h(-1)), or 2BX (0.035% +/- 0.011%h(-1)) approach. A significant correlation was observed between the calculated muscle protein FSR assessed using the 1BX LONG and 2BX approach (r = 0.7, P = .02). Our data demonstrate that the single-biopsy approach, irrespective of whether the biopsy is obtained at 2 or 4 hours, can be used as a surrogate for the sequential-biopsy approach to determine basal muscle protein synthesis in a group.