Department of Applied Chemistry, Northeast Agricultural University, Harbin, 150030, China.
Sci Rep. 2017 Oct 17;7(1):13320. doi: 10.1038/s41598-017-13507-9.
An aqueous two-phase system (ATPS) consisting of poly(ethylene glycol-ran-propylene glycol) monobutyl ether (UCON)/phosphate was developed for the separation of the antioxidant peptides from pepsin hydrolysate of Whey Protein Isolate (WPI). The efficiency of the separation was evaluated based on the DPPH radical scavenging activity, ABTS radical scavenging activity and ferric reducing antioxidant power (FRAP) of the separated peptides. The effects of some parameters on the partition of antioxidant peptides were investigated. An efficient separation of antioxidant peptides was achieved using ATPS with pH of 4.0, 4 mL of UCON solution (40%, w/w), 4 mL of KHPO solution (15.5%, w/w), 2 mL of WPI hydrolysate and 0.40 g/10 mL of NaCl. Reversed-phase high-performance liquid chromatography (RP-HPLC), amino acid analyzer and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) were used to characterize the purified peptides separated by the ATPS. The peptides in top phase were less polar than those in bottom phase. More antioxidative and hydrophobic amino acids were extracted to the top phase of ATPS, and the peptides with the amino acid sequences with antioxidant activities moved to the top phase as well. In conclusion, antioxidant peptides were successfully separated from the WPI hydrolysate by UCON/phosphate ATPS.
建立了一种由聚（乙二醇-丙二醇）单丁醚（UCON）/磷酸盐组成的双水相体系（ATPS），用于从乳清蛋白分离物（WPI）胃蛋白酶水解物中分离抗氧化肽。根据分离肽的 DPPH 自由基清除活性、ABTS 自由基清除活性和铁还原抗氧化能力（FRAP）评估分离效率。研究了一些参数对抗氧化肽分配的影响。在 pH 值为 4.0、4 mL UCON 溶液（40%，w/w）、4 mL KHPO 溶液（15.5%，w/w）、2 mL WPI 水解物和 0.40 g/10 mL NaCl 的条件下，使用 ATPS 可有效分离抗氧化肽。采用反相高效液相色谱（RP-HPLC）、氨基酸分析仪和基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）对 ATPS 分离得到的纯化肽进行了表征。上层的肽比下层的肽极性小。更多的抗氧化和疏水性氨基酸被提取到 ATPS 的上层，具有抗氧化活性的肽序列也移动到了上层。总之，成功地从 WPI 水解物中通过 UCON/磷酸盐 ATPS 分离出了抗氧化肽。
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