Yi Zhao, Yi Zhong, Qi Zheng, Chun‐Yan Yin, Jia‐Li Cai, Dijun Wang, Li Zhu, Ji Cao, Xiao‐Jing Yan, Yuanpei Lian
ABSTRACT Indigo naturalis‐ processed Ophiopogon japonicus (IN‐OJ) is a common processing method in the Menghe medical school to enhance its heat‐clearing and detoxifying effects. However, its impact on the structure and bioactivity of the main active component—polysaccharides—remains unclear. In this study, polysaccharides were extracted from both raw and IN‐processed O. japonicus using hot reflux and ultrasound‐assisted methods. Their structural features were characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM), gel permeation chromatography (GPC), high‐performance liquid chromatography (HPLC), and Fourier‐transform infrared spectroscopy (FT‐IR). Monosaccharide composition, molecular weight, and microscopic morphology were systematically analyzed. Antioxidant activities were evaluated in vitro through DPPH, superoxide anion, and hydroxyl radical scavenging assays, using Vitamin C (Vc) as a positive control. The results indicated that IN processing significantly increased the polysaccharide and uronic acid contents. Molecular weight profiles revealed an additional high‐molecular‐weight fraction in the processed samples (U‐IP: 520,049 kDa; W‐IP: 356,167 kDa). Monosaccharide analysis showed notable increases in arabinose and fucose. Microscopic observations indicated a transition from a loose, honeycomb‐like structure to a denser, aggregated morphology, while FT‐IR spectroscopy confirmed the introduction of additional carboxyl groups. Furthermore, antioxidant assays demonstrated that IN processing significantly enhanced scavenging activities against DPPH and superoxide anion radicals, with W‐IP exhibiting the strongest DPPH radical scavenging capacity (IC 50 = 3.19 mg/mL). However, hydroxyl radical scavenging activity decreased post‐processing, likely due to steric hindrance within the aggregated structures. The positive control Vc exhibited markedly superior scavenging activity across all assays. These findings suggest that IN processing enhances specific antioxidant capacities of OJ polysaccharides through structural modification, with reflux extraction yielding the most active fractions, thus providing a scientific basis for optimizing this traditional method in functional food applications.