关键词: 色素蛋白复合体, 双水相萃取系统, 分离因素, 萃取率, 生物活性

Abstract: To develop a novel purification method for the pigment protein complexes(PPCs),PPCs were separated and purified using aqueous two-phase system(ATPS) of polyethylene glycol(PEG)1000/potassium citrate controlled by pH.The extraction condition of PPCs was optimized.The distribution behavior of PPCs in ATPS was detected by UV-Vis spectroscopy,and the purity and biological activity of product were investigated.The results indicated that the optimal extraction condition were 19.0%(W/W) PEG1000,20.0%(W/W) potassium citrate,the proteins adding amount was 3.42 mg/g,and with pH 9.0.Under these conditions,the distribution coefficient(K) and the extraction rate of PPCs reached to the maximum at 8.8 and 86.0%,respectively.Response surface methodology analysis revealed that the phase compositions of PEG1000 and potassium citrate,pH condition had the significant effects on K and extraction rate.In reverse extraction process,the K of the reverse extraction came to the maximum of 0.15,the extraction rate of 86.6% at the pH 7.0.And the total protein yield was to 74.2%.The results of UV-Vis analysis discovered that the distribution behavior of PPCs can be regulated by the pH of system.PPCs tended to concentrate in the top phase solution under the condition of pH>8.5,otherwise in bottom phase solution.The factors of PEG1000,potassium citrate and proteins added had not affected on the distribution behavior in ATPS.The electrophoresis analysis result found that there were two protein components in top phase(pH 9.0) with molecular weight(MW) about 7.0 kD and 14.0 kD.After reverse extraction(pH 7.0),only a component with MW 7.0 kD was back to potassium citrate solution,which was the β- subunit of LH2.The biological activity of the purified PPCs was stable after the routines of the extraction and reextraction.PPCs can be separated and purified by the ATPS of PEG1000 / potassium citrate through controlling the pH of the system.

Key words: pigment-protein complexes, aqueous two-phase system, distribution coefficient, extraction rate, biological activity