The objectives of this study were to analyze the influence of chemical water quality on fish guilds, pollution tolerance and the multi-metric ecological health, based on the Fish Assessment Index (FAI) in the main stream of Mangyeong River between 2009−2016. The quality of water with specific conductivity, TP, and NH4-N got worse dramatically in the down region. During the study, a total of 50 species were collected and the most dominant species was Zacco platypus. Also known as tolerant species, accounted 22.9% of the total abundances, thus indicating a trophic degradation. The downstream region (S5) had the highest number of fish external abnormalities, indicating a degradation of ecological health, based on the fish assemblages. Pearson correlation analysis indicated that relative abundance of tolerant fish species and omnivore fish species had a significant positive correlation (r>0.30, p<0.05) with values of BOD, conductivity and NH4-N. Whereas, the relative abundance of the sensitive species and insectivore species had a significant negative relations (r<−0.30, p<0.001) with the parameters. The mean obtained from the multi-metric fish model, based on the FAI of all sites was 47 (n=40). This indicated a “fair condition” in the ecological health, and the downstream regions (S3−S5) were judged as “bad condition”, indicating an influence of the chemical degradation on the ecological health.