Microsoft Word - 2--赵丽_news_.doc

Size: px

Start display at page:

Download "Microsoft Word - 2--赵丽_news_.doc"

演筌禄
5 years ago
Views:

1 酒石酸钾钠双水相体系分离纯化 C- 藻蓝蛋白 1, 赵丽 2, 彭一良 2, 康天 2 1, 蔡伟民 (1., ; 2., ) 摘要 : 采用廉价的聚乙二醇 (PEG)/ 酒石酸钾钠建立双水相体系, 从干螺旋藻中分离纯化藻蓝蛋白 C-PC 以纯度( 目的蛋白在 620 nm 处的特征吸光值和总蛋白在 280 nm 处吸光值的比值 ) 为指标, 优化体系参数研究结果表明, 体系 ph 8.06, 系线长为 21.65% 体积比为 0.22 的 PEG1000 和酒石酸钾钠双水相体系分离纯化效果最好, 纯度由 0.42 提高到 1.27, 高于食品级纯度 0.7, 纯化因子 3.04, 回收率 85.11% 硫酸铵盐析提高粗提液纯度后, 二次双水相纯度达到 3.28, 高于药品级 3.0 得到的 C-PC 具有较高的生物活性关键词 : C- 藻蓝蛋白 ; 纯度 ; 酒石酸钾钠 ; PEG1000; 双水相中图分类号 : Q81, Q51 文献标识码 : A 文章编号 : (2015) doi: /hykx (Aqueous two-phase systems, ATPS) [1], [2], 75% 90%,,, [3-4], (C-phycocyanin, C-PC ), C-PC α β, α β 1 (Phycobilins) [5], C-PC, C-PC [6], C-PC,, [7], 50 /mg [8], t 70%,, 20% [9],, C-PC, C-PC [10-11] [12-13],, PEG4000/ [14], PEG4000,, C-PC PEG4000, PEG PEG2000/ [15], 1.2 PEG1000/, C-PC,,,,, 1 材料与方法 1.1 材料试剂与仪器 ( : ; : : (C201219) : (1976-),,,, :, lili_zhao_cc@126.com;,,,, wwwbbll@sina.com 10 / 2015 / 39 / 9

) UV-2102 PC -, ; (TGL-16MI) 1.2 方法 1.2.1 6 g, 0.01mol/L, : 20,, 6 10 000 r /min 10 min,, 4 1.2.2 PEG /,,,,,, 4,,, 1.2.3 [12-14, 16-17] (P): 620 nm 280 nm (F): (K): C-PC PEG 0.1 g/l, 0.

2 ) UV-2102 PC -, ; (TGL-16MI) 1.2 方法 g, 0.01mol/L, : 20,, r /min 10 min,, PEG /,,,,,, 4,,, [12-14, 16-17] (P): 620 nm 280 nm (F): (K): C-PC PEG 0.1 g/l, g/l C-PC (TLL): ATPS (Vr): ATPS PEG,, 4,,, ATPS , 260 nm, 700 nm C-PC 560 nm 580 nm 770 nm ORIGN 8.5, MATLAB R2008a 2 结果与分析 2.1 螺旋藻藻粉的细胞学形态 (10 ) 1 C-PC Fig. 1 The morphology of the dry spirulina platensis powder 2.2 不同盐相对 C-PC 纯度的影响 TLL 27%±1% Vr 1, PEG, ( ) C-PC, 2 C-PC 4.0 ph (ph ) [16], C-PC,, 2 C-PC Fig. 2 The influence of different salts phase on the purity of C-PC Marine Sciences / Vol. 39, No. 9 /

3 ,, C-PC PEG , C-PC,, C-PC PEG, 2.3 不同 PEG 分子质量对 C-PC 纯度的影响 Vr 1, TLL 21.65% PEG /, PEG ( ), C-PC, C-PC, PEG, 3 PEG , TLL 27%±1%, : PEG 1000 PEG PEG, PEG,,,,, PEG C-PC, PEG C-PC PEG, PEG,,,,, C-PC, PEG, C-PC PEG 1 000, PEG , TLL 40%( ) [18] PEG1 000, TLL( ): 14.66% 21.65% 26.51% 30.26% 33.47% Vr, TLL C-PC, C-PC, Vr, 4 C-PC TLL Vr, C-PC TLL,, TLL, C-PC TLL 14.66%, C-PC,, TLL,, TLL 21.65%, C-PC TLL 14.66%,,, TLL 21.65% 4 TLL C-PC Fig. 4 The influence of TLL on the purity of C-PC 2.5 不同 Vr 对 C-PC 纯度的影响 3 PEG C-PC Fig. 3 The influence of PEG molecular weight on the purity of C-PC 2.4 TLL 对 C-PC 纯度的影响 TLL, Vr C-PC TLL, Vr C-PC C-PC Z, Vr Y, TLL X, 5 C-PC Vr TLL, C-PC Vr, Vr, C-PC TLL 21.65%, Vr 0.22, C-PC 1.27 Vr,,,,,, C-PC [8] PEG4000 [14] PEG, 12 / 2015 / 39 / 9

4 C-PC Z, PEG1 000 Y, X, 7 PEG % 21%, ph 8.06, 21.65% 0.22, , 3.04, 85.11% Fig. 5 5 Vr C-PC The influence of Vr on the purity of C-PC 2.6 不同 TLL 和 Vr 对 C-PC 回收率的影响 TLL Vr C-PC C-PC Z, Vr Y, TLL X, 6 Vr C-PC TLL, Vr, Vr, TLL 33.47%, Vr 2.11 C-PC 95.64% 1.27, 85.11% C-PC TLL 33.47%, Vr, TLL 85% 7 Fig. 7 The influence of components in the ATPS on the purity of C-PC 2.8 多次 ATPS 对纯度的影响, 1.46, PEG/ ATPS, ATPS Z, (ATPE) X, 8 2 ATPE 3.28, ATPE 2.9 C-PC 的吸收光谱 TLL Vr C-PC Fig. 6 The influence of TLL and Vr on the yield of C-PC 2.7 不同相组成质量分数对 C-PC 纯度的影响 TLL PEG1 000, Fig. 8 8 ATPS C-PC The influence of multiple ATPE stages on the purity of C-PC Marine Sciences / Vol. 39, No. 9 /

5 ATPS C-PC, 260 nm, 700 nm, 9, 2 ATPS, C-PC 3.28, 3.0 C-PC, : Fig. 9 9 C-PC The absorption spectrum of the recovery C-PC [1] Show P L, Tan C P, Shamsul A M, et al. Extractive fermentation for improved production and recovery of lipase derived from Burkholderia cepacia using a thermoseparating polymer in aqueous two-phase systems [J].Bioresour Technol, 2012, 116: [2] Naganagouda K, Mulimani V H. Aqueous two-phase extraction (ATPE): An attractive and economically viable technology for downstream processing of Aspergillus oryzae α-galactosidase [J]. Process Biochem, 2008, 43: C-PC 的荧光测定 2 ATPS C-PC, 10 C-PC, 643 nm ATPS C-PC,, 643 nm, 155.5, [3] Garza-Madrid M, Rito-Palomares M, Serna-Saldívar S O, et al. Potential of Aqueous Two-Phase Systems constructed on flexible devices: Human serum albumin as proof of concept [J]. Process Biochem, 2012, 45: [4] Benavides J, Rito-Palomares M. Review practical experiences from the development of aqueous two-phase processes for the recovery of high value biological products [J]. J Chem Technol Biotechnol, 2008, 83: [5] Choi J W, Nam Y S, Kim J M, et al. Biomolecular photonic device consisting of Chl a/chl b/phycoerythrin /phycocyanin hetero structure [J]. Nanoscience and Nanotechnology, 2006, 6(11): [6] Yoshida A, Takagaki Y, Nishimune T. Enzyme immunoassay for phycocyanin as the main component of spirulina color in foods [J]. Biosci Biotechnol 10 C-PC Fig. 10 The fluorescence emission spectrum of the recovery C-PC 3 结论 PEG1000/, C-PC, 7.76% (w/w) PEG %(w/w) TLL 21.65% Vr 0.22, ph 8.06, 1.27, 85.11% Biochem, 1996, 60: [7] Bhat V B, Madyastha K M. Scavenging of peroxynitrite by phycocyanin and phycocyanobilin from Spirulina platensis: protection against oxidative damage to DNA [J]. Biochem Biophys Res Commun, 2001, 285: [8] Benavides J, Rito-Palomares M. Potential Aqueous Two Phase Processes for the Primary Recovery of Colored Protein from Microbial Origin [J]. Life. Sci, 2005, 5 (3): [9] Ogbonda K H, Aminigo R E, Abu G O. Influence of 14 / 2015 / 39 / 9

6 temperature and ph on biomass production and protein biosynthesis in a putative Spirulina sp [J]. Bioresour Technol, 2007, 98: [10] Kula M R, Kroner K H, Hustedt H. Adv. Purification of enzymes by liquid liquid extraction [J]. Biochem. Eng, 1982, 24: [11] Ranjitha K, Kaushik, B D. Putification of phycobiliproteins from Nostoc nuscorum [J]. Sci Ind Res, 2005, 64: [12] Anamika P, Sandhya M, Richa P, et al. Purification and characterization of C-Phycocyanin from cyanobacterial species of marine and freshwater habitat [J]. Protein Exp. Purif, 2005, 40: [13] Patil G, Chethana S, Sridevi A S, et al. Method to obtain C-phycocyanin of high purity [J]. Chromatogr A, 2006, 1127: [14] Patil G, Raghavarao K S M S. Aqueous two phase extraction for purification of C-phycocyanin [J]. Biochem Eng J, 2007, 34: [15],,,. - [J]..2012, 11: [16] Zhao L, Peng Y, Gao J, et al. Bioprocess intensification: an aqueous two-phase process for the purification of C-phycocyanin from dry Spirulina platensis [J]. Eur. Food. Res. Technol, 2014, 238 (3): [17]. [M]. :, 1997: [18] Cabezas, H. Theory of phase formation in aqueous two phase systems [J]. Chromatogr B, 1996, 680: Separation and purification of C-phycocyanin using aqueous two-phase extraction technique of potassium sodium tartrate ZHAO Li 1, 2, PENG Yi-liang 2, KANG Tian 2, CAI Wei-min 1 (1. School of Municipal and Environmental Engineering, Harbin Institute of Technology, Harbin , China; 2.College of Life Sciences and Biotechnology, Harbin Normal University, Harbin , China) Received: Sep., 12, 2014 Key words: C-phycocyanin; purity; potassium sodium tartrate; PEG1000; aqueous two- phase system Abstract: This paper used commercial aqueous two-phase system (ATPS) to efficiently extract and purify C-PC from the dry spirulina platensis. The purity of C-PC was defined as the relation between of 620 and 280 nm absorbance, which was as the target to optimize the system parameters. The optimal conditions were proved in polyethylene glycol (PEG) 1000 and potassium sodium tartrate, system ph of 8.06, the tie-line length (TLL) of 21.65% (w/w) and the volume ratio (Vr) of 0.22 to increase the purity from the initial purity of 0.42 to 1.27 after the first extraction. The purity was over the food grade 0.7 and the purification factor was The recovery yield was 85.11%. The purity was achieved up 3.28 after the saturation of ammonium sulfate in the second ATPS extraction. It was more than the pharmaceutical grade 3.0. ( 本文编辑 : 康亦兼 ) Marine Sciences / Vol. 39, No. 9 /

标题

第卷第期年月哈尔滨工业大学学报 JOURNAL OF HARBIN INSTITUTE OF TECHNOLOGY Vol No Jun doi:9 jissn 双水相分离纯化 R- 藻红蛋白, 赵丽, 彭一良, 付秋爽, 蔡伟民 ( 哈尔滨工业大学市政环境工程学院,9 哈尔滨 ; 哈尔滨师范大学生命科学与技术学院, 哈尔滨 ) 摘要 : 为了获得优于食品级的 R- 藻红蛋白 (R-PE),

Microsoft Word - 2--赵 丽_news_.doc

Microsoft Word - 2--赵丽_news_.doc