School of Life Science, Central South University, Changsha, 410013, China.
School of Chemistry and Chemical Engineering, Qilu University of Technology (Shandong Academy of Sciences), Jinan, 250353, China.
Appl Biochem Biotechnol. 2022 Nov;194(11):5522-5536. doi: 10.1007/s12010-022-04058-5. Epub 2022 Jul 6.
A simple and effective method for detecting the antioxidant activity by utilizing oxidative damage of pigment proteins was developed. In this method, phycocyanin and bovine hemoglobin pigment proteins were used as substrates attacked by free radicals; AAPH was used as a free radical initiator; and Trolox as a positive control; and the fermentation products of Lactobacillus plantarum 793, phycocyanin hydrolysates, salmon skin collagen hydrolysates, and synthetic peptides PMRGGYHY and FCVLRP are antioxidants inspected in this study. Because of being attacked by free radicals, the absorbance of the pigment proteins at their characteristic absorption peak changes with time. By recording the time-varying curve at the characteristic absorption peak of the pigment protein in the blank/negative control sample, the Trolox positive control sample, and the samples of inspected antioxidants, the antioxidant activity could be calculated by using the specific equation. The linear detection ranges of Trolox in the phycocyanin assay and the bovine hemoglobin assay were 1-4 μM and 4-24 μM, respectively. Compared with the ORAC assay, the antioxidant activities of the samples measured by this method were slightly lower. The method proposed in this study reflects the protective effects of inspected antioxidants on pigment proteins, which could potentially serve as new biomarkers of oxidative damage processes.
本研究利用色素蛋白的氧化损伤，开发了一种简单有效的抗氧化活性检测方法。在该方法中，藻蓝蛋白和牛血红蛋白色素蛋白被用作自由基攻击的底物；AAPH 被用作自由基引发剂；Trolox 作为阳性对照；并检测了植物乳杆菌 793 的发酵产物、藻蓝蛋白水解物、三文鱼皮胶原蛋白水解物、合成肽 PMRGGYHY 和 FCVLRP 等抗氧化剂。由于受到自由基的攻击，色素蛋白在其特征吸收峰处的吸光度随时间发生变化。通过记录空白/阴性对照样品、Trolox 阳性对照样品和待检测抗氧化剂样品中色素蛋白特征吸收峰处的时变曲线，可使用特定方程计算抗氧化活性。在藻蓝蛋白测定和牛血红蛋白测定中，Trolox 的线性检测范围分别为 1-4 μM 和 4-24 μM。与 ORAC 测定法相比，本方法测定的样品的抗氧化活性略低。本研究提出的方法反映了待检测抗氧化剂对色素蛋白的保护作用，这可能成为氧化损伤过程的新生物标志物。
