[Objective] Grain size significantly influences the yield, marketability and processing attributes of quinoa. studying the differences of seed phenotypes, filling characteristics and starch synthase activity of large- and small-grained quinoa during the filling period to provide theoretical guidance for the selection and breeding of large-grained quinoa varieties. [Methods] Two quinoa samples, one with a thousand grain weight exceeding 5.0 g and the other is below 3.0 g, were selected for a field experiment at the Innovative Experimental Base of Germplasm Resources of Qinghai Academy of Agricultural and Forestry Sciences. The study comparing the differences in grain phenotypes, filling characteristics, and starch synthase activity between large- and small-grained quinoa at 7, 14, 21, and 28 days of the during the filling stage. [Results] (1) The phenotypic traits of large- and small-grained quinoa characteristics, including grain area, circumference, diameter, kernel length, and kernel width, significantly increased with reproductive period, and there were significant differences between grain types, the largest differences were in seed area and circumference, with large-grained quinoa being significantly higher than small-grained quinoa by 9.12% to 11.54% and 21.49 to 23.92%, respectively. (2) Throughout the filling period, the dry weight of 100 grains of large-grained quinoa consistently exceeded that of small-grained quinoa in the same period, with an average increase of 21.23% to 31.04%; the filling rate of large and small-grained quinoa first rose and subsequently declined during the reproductive period, which was in line with the pattern of change of "slow-fast-slow", however the time to reach the peak and the peak height were obviously different, with large-grained quinoa appearing early and high, while small-grained quinoa was low and late. The peak of large grain appeared early and high, while small grain was low and late. (3) The starch branching enzyme (SBE), sucrose synthase (SS), soluble starch synthase (SSS) and ADPG pyrophosphorylase (AGP) varied during for both grain sizes, with SBE and SS activities being stronger in small-grained quinoa than in large-grained quinoa, while SSS and AGP activities were stronger in large-grained than in small-grained quinoa. [Conclusion] Variations in the activities of four starch synthases during quinoa grain filling contribute to differences in the accumulation of starch synthesis and peak filling rate, which in turn leads to differences in grain phenotypic traits, whereas SSS and AGPase are the key enzymes affecting quinoa grain size formation.