This study aims to establish a quality grading standard that combines the conventional quality evaluation based on morphological characteristics of traditional Chinese medicine with the modern quality evaluation. Based on the existing standards and market circulation of Isatidis Radix, the diameter and color of Isatidis Radix decoction pieces were selected as the appearance traits for preliminary grading. The effects of internal quality indexes such as moisture, total ash, acid-insoluble ash, ethanol-soluble extractives, and 9 water-soluble components on different grades of decoction pieces were comprehensively compared, and the key grading indexes were determined by t-test. The relevance between appearance traits and the internal quality indexes was investigated by multi-dimensional statistical analyses such as correlation analysis, principal component analysis(PCA), orthogonal partial least squares-discriminant analysis(OPLS-DA), and hierarchical cluster analysis(HCA). Furthermore, the fluorescent substrate was employed to investigate the inhibitory effects of different grades of Isatidis Radix decoction pieces on the neuraminidase of influenza virus. The rationality of the selected grading index components was verified by network pharmacology. The results showed that the diameter of decoction pieces had significantly positive correlations with other index components except moisture. Considering the existing commodity specifications, grading standards, and actual market circulation of Isatidis Radix, the quality grading standard of Isatidis Radix decoction pieces was developed with diameter as the appearance trait and(R,S)-goitrin as the limit index component of being qualified. On this basis, the content of ethanol-soluble extractives, total sinigrin, and total nucleoside were selected for quality grading. The in vitro enzyme activity evaluation showed that the first grade of Isatidis Radix decoction pieces had stronger inhibitory effect on the neuraminidase of influenza virus, and the network pharmacology results showed that the selected index components for quality grading had significant correlations with influenza. This study established a quality grading standard combining appearance traits and internal quality indexes, which provides a scientific basis for improving the existing grading standards and the price and quality of Isatidis Radix decoction pieces.