The structure of the native pteridine in Tetrahymena pyriformis was determined as (6R)-5,6,7,8-tetrahydrod-monapterin ((6R)-2-amino-5,6,7,8-tetrahydro-6-[(1R,2R)-1,2,3-trihydroxypropyl]pteridin-4(3H)-one; 4). First, the configuration of the 1,2,3-trihydroxypropyl side chain was confirmed as d-threo by the fluorescence-detected circular dichroism (FDCD) spectrum of its aromatic pterin derivative 2 obtained by I2 oxidation (Fig. 1). The configuration at the 6-position of 4 was determined as (R) by comparison of its hexaacetyl derivative 6 with authentic (6R)- and (6S)-hexaacetyl-5,6,7,8-tetrahydro-d-monapterins 6 and 7, respectively, in the HPLC, LC/MS, and LC-MS/MS (Figs. 3 ± 6). (6R)-5,6,7,8-Tetrahydro-d-monapterin (4) is a newly discovered natural tetrahydropterin.