Aqueous biphasic systems (ABSs) are an interesting alternative for separating industrial enzymes due to easy scale-up and low operational cost. The proteases of Pseudomonas sp. M211 were purified through ABS platforms formed by polyethylene glycol (PEG) and citrate buffer salt. Two experimental designs 2³ + 4 were performed to evaluate the following parameters: molar mass of PEG (M_PEG), concentration of PEG (C_PEG), concentration of citrate buffer (C_Cit), and pH. The partition coefficient (K), activity yield (Y), and purification factor (PF) were the responses analyzed. The best purification performance was obtained with the system composed of M_PEG = 10,000 g/mol, C_PEG = 22 wt%, C_Cit = 12 wt%, pH = 8.0; the responses obtained were K = 4.9, Y = 84.5%, PF = 15.1, and tie-line length = 52.74%. The purified proteases of Pseudomonas sp. (PPP) were used to obtain hydrolysates of Lupinus mutabilis (Peruvian lupin cultivar) seed protein in comparison with the commercial protease Alcalase® 2.4L. A strong correlation between hydrolysis degree and radical scavenging activity was observed, and the highest antioxidant activity was obtained with Alcalase® (1.40 and 3.47 μmol Trolox equivalent/mg protein, for 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) and oxygen radical absorbance capacity, respectively) compared with PPP (0.55 and 1.03 μmol Trolox/mg protein). Nevertheless, the IC₅₀ values were lower than those often observed for antioxidant hydrolysates from plant proteins. PEG/citrate buffer system is valuable to purify Pseudomonas proteases from the fermented broth, and the purified protease could be promising to produce antioxidant protein hydrolysates.

中文翻译：

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假单胞菌属的纯化。蛋白酶通过水性双相系统作为获得生物活性蛋白水解物的替代来源

由于易于放大和低运营成本，水相双相系统 (ABS) 是分离工业酶的有趣替代方案。假单胞菌蛋白酶。M211 通过由聚乙二醇 (PEG) 和柠檬酸盐缓冲盐形成的 ABS 平台进行纯化。进行了两个实验设计 2 ³  + 4 以评估以下参数：PEG 的摩尔质量 ( M _PEG )、PEG 的浓度 ( C _PEG )、柠檬酸盐缓冲液的浓度 ( C _Cit ) 和 pH。分配系数（K）、活性产率（Y) 和净化因子 (PF) 是分析的响应。由M _PEG  = 10,000 g/mol、C _PEG  = 22 wt%、C _Cit  = 12 wt%、pH = 8.0组成的系统获得了最佳纯化性能；获得的响应为K  = 4.9、Y  = 84.5%、PF = 15.1 和联络线长度 = 52.74%。纯化的假单胞菌蛋白酶。(PPP) 用于获得羽扇豆的水解产物（秘鲁羽扇​​豆品种）种子蛋白与商业蛋白酶 Alcalase® 2.4L 的比较。观察到水解程度与自由基清除活性之间有很强的相关性，对于 2,2'-azino-bis(3-ethylbenzothiazolin-6-)，使用 Alcalase®（1.40 和 3.47 μmol Trolox 当量/mg 蛋白质）获得了最高的抗氧化活性磺酸）和氧自由基吸收能力）与 PPP（0.55 和 1.03 μmol Trolox/mg 蛋白质）相比。然而，IC ₅₀值低于通常观察到的来自植物蛋白的抗氧化水解物的值。PEG/柠檬酸盐缓冲系统对于从发酵液中纯化假单胞菌蛋白酶很有价值，纯化的蛋白酶有望产生抗氧化蛋白水解物。