The trophodynamics of toxic trace metals is significant for assessing the threat of toxic trace metals to the aquatic ecosystem and human safety. However, due to the difficulty of accurately calculating the trophic positions of freshwater aquatic organisms in the food web, the comprehensive process of trophodynamics of toxic trace metals in freshwater ecosystems was still rarely known. By integrating the compound-specific nitrogen stable isotopic analysis of amino acids (CSIA-AAs) and the Bayesian stable isotope mixing model (SIMM) as a novel approach, the present study investigated the trophodynamics of five harmful toxic trace metals (Zn, As, Cr, Cu, and Hg) in the food web of the YangZong Lake, a plateau freshwater lake that was once heavily polluted by arsenic in Yunnan Province, China. The results revealed that Hg tended to be efficiently biomagnified in the food web with a trophic magnification factor of 1.75; As, Cr, and Cu were biodiluted significantly, while Zn showed no biomagnification or biodilution trends. The dietary health risk assessment indicated the potential health risk of toxic trace metals for the local residents of long-term fish consumption. The present work highlights the accuracy and reliability of the novel CSIA-AAS+SIMM approach in the calculation of the trophic positions of freshwater organisms.