定量体积排除色谱测定高分子双水相系统的组成和分子量分布

doi:10.7503/cjcu20150567

摘要/Abstract

摘要：

利用定量体积排除色谱研究葡聚糖-聚乙二醇双水相系统相分离后上下两相中2种高分子组分的含量、分子量和分子量分布. 由定量体积排除色谱法得到的两相组成(即系线端点)与用浊点滴定法得到的浊点曲线几乎完全重合, 二者仅在靠近临界点的聚乙二醇富集相有一定偏差. 同时, 利用体积排除色谱测得两相中葡聚糖和聚乙二醇的分子量和分子量分布. 结果表明, 由系线端点得到的体系两相共存线与浊点曲线的偏差是由于相分离过程中, 不同分子量的高分子组分在两相的非均匀分配造成的. 聚乙二醇分子量分布较窄, 发生相分离后, 在两相的分子量和分子量分布相差不大. 而葡聚糖分子量分布较宽, 在相分离后两相中的分子量和分子量分布具有较大差异, 即葡聚糖组分在葡聚糖富集相中的分子量显著高于其在聚乙二醇富集相中的分子量. 随着葡聚糖-聚乙二醇体系初始浓度的增加, 两相中葡聚糖的分子量差异变大. 定量体积排除色谱可以准确得到高分子双水相系统的相平衡数据及两相中2组分的分子量和分子量分布信息, 其结果不仅为深入理解葡聚糖-聚乙二醇-水三元溶液的相平衡提供基础, 而且为双水相系统在萃取分离中的应用提供理论指导.

关键词: 体积排除色谱, 双水相系统, 相分离, 葡聚糖, 聚乙二醇

Abstract:

Quantitative size exclusion chromatography(SEC) was exploited to study the composition, molecular weight and molecular weight distribution of aqueous two-phase system of dextran and poly(ethylene glycol)(PEG) following phase separation. Tie lines constructed by SEC method were compared with the cloud point curve of the system obtained by titration. An excellent agreement was found between the tie line end points and the cloud point, except for the data points of the PEG-rich phases close to the critical point. The molecular weight and molecular weight distribution of the two polymer species in two co-existing phases obtained by SEC indicate that the mismatch is caused by the uneven distribution of macromolecular components between two phases upon phase separation. Having a broad molecular weight distribution, dextran in the two phases show prominent molecular mass difference. The molecular weight of dextran in the dextran-rich phase is significantly higher than that in the PEG-rich phase. As the initial polymer concentration of the system increases, the molecular weight difference of dextran in the two phases becomes more significant. However, we have not observed such a trend for PEG because of its narrow molecular weight distribution. Accurate data on the phase diagram and molecular weight of two polymer components in the co-existing phases can be obtained using quantitative SEC. The above results will not only help to understand the phase diagram of dextran-PEG-water ternary system, but also provide guidance to its application in extraction and separation of biological materials.

Key words: Size exclusion chromatography, Aqueous two-phase system, Phase separation, Dextran, Poly(ethylene glycol)

中图分类号:

O631.4

TrendMD:

赵梓良, 李琦, 薛彦虎, 姬相玲, 薄淑琴, 刘勇刚. 定量体积排除色谱测定高分子双水相系统的组成和分子量分布. 高等学校化学学报, 2016, 37(1): 167.

ZHAO Ziliang, LI Qi, XUE Yanhu, JI Xiangling, BO Shuqin, LIU Yonggang. Composition and Molecular Weight Determination of Aqueous Two-phase System by Quantitative Size Exclusion Chromatography^†. Chem. J. Chinese Universities, 2016, 37(1): 167.

图/表 7

Fig.1 Cloud point curve of the aqueous solution of dextran and PEG at (25±1) ℃ obtained by titration ^The homogeneous solution in the left vial has a composition in the one-phase region, the right one shows phase separation in the two-phase region. Au nanoparticles are dispersed in water and added to the mixed solution to enhance the color contrast.

Fig.2 Volume fraction(φd) of the dextran-rich phase as a function of the normalized distance from the cloud point(w/wcl-1) for polymer solutions of different mass ratios(wd/wp) between dextran and PEG(A) and dependence of the volume fraction φd on the mass ratio wd/wp at w/wcl=1.01(B)

Fig.3 Dependence of the RI peak area(ARI) as a function of polymer concentration(cinj) of the solutions injected into the SEC columns for dextran(a) and PEG(b)

Fig.4 SEC chromatograms of the coexisting dextran-rich(A) and PEG-rich phases(B) at

Fig.5 Phase diagram of the ternary dextran-PEG-water system^The cloud point curve(same data as in Fig.1) is shown as a solid curve. The compositions of the initial solutions(with mass ratio wd/wp=1.23) for which SEC measurements after phase separation are performed are indicated by black crosses. The end points of the respective tie lines(dashed lines) consist of red crosses indicating the compositions of the dextran-rich phases and green crosses indicating the compositions of the PEG-rich phases. The midpoints(blue circles) of the tie lines were extrapolated to determine the critical point.

Table 1 Molecular weight of dextran in dextran-rich and PEG-rich phases

No.	ε	Dextran-rich phase			PEG-rich phase
No.	ε	10^-3 M_w	10^-3 M_n	M_w/M_n	10^-3 M_w	10^-3 M_n	M_w/M_n
Native	______	380.0	173.7	2.19	380.0	173.7	2.19
1	0.0098	453.9	192.9	2.35	249.3	152.6	1.63
2	0.030	426.9	182.0	2.35	208.9	138.3	1.51
3	0.092	394.1	172.1	2.29	167.6	120.8	1.39
4	0.134	397.9	177.8	2.24	152.3	112.5	1.35
5	0.200	380.6	170.3	2.23	136.0	103.1	1.32
6	0.302	369.8	164.2	2.25	116.9	90.0	1.30
7	0.389	373.3	167.1	2.23	109.4	84.8	1.29
8	0.508	371.3	164.6	2.26	97.1	76.9	1.26
9	0.599	370.6	163.1	2.27	95.9	69.4	1.38
10	0.729	375.2	166.0	2.26	82.2	66.9	1.23
11	0.982	372.4	163.7	2.28
12	1.552	368.7	161.8	2.28
13	2.087	368.6	155.8	2.37
14	2.825	374.7	160.5	2.34
15	3.605	368.7	161.8	2.28

Table 2 Molecular weight of PEG in dextran-rich and PEG-rich phases

No.	ε	Dextran-rich phase			PEG-rich phase
No.	ε	10^-3 M_w	10^-3 M_n	M_w/M_n	10^-3 M_w	10^-3 M_n	M_w/M_n
Native	_____	8.45	7.64	1.11	8.45	7.64	1.11
1	0.0098	8.38	7.57	1.11	8.35	7.58	1.10
2	0.030	8.12	7.33	1.11	8.35	7.61	1.10
3	0.092	8.08	7.25	1.11	8.30	7.52	1.10
4	0.134	8.18	7.31	1.12	8.41	7.65	1.10
5	0.200	8.02	7.06	1.14	8.39	7.63	1.10
6	0.302	8.02	7.00	1.15	8.28	7.53	1.10
7	0.389	7.98	6.89	1.16	8.40	7.63	1.10
8	0.508	7.93	6.69	1.19	8.38	7.60	1.10
9	0.599	7.88	6.55	1.20	8.50	7.70	1.10
10	0.729	7.78	6.22	1.25	8.29	7.49	1.11
11	0.982				8.44	7.65	1.10
12	1.552				8.63	7.82	1.10
13	2.087				8.43	7.64	1.10
14	2.825				8.40	7.57	1.11
15	3.605				8.56	7.76	1.10

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