基于双流体喷嘴的磁性琼脂糖微球的制备及其蛋白吸附性能探究

doi:10.11949/0438-1157.20230211

摘要/Abstract

摘要：

针对现有磁性琼脂糖微球（MAM）制备技术工艺流程长、重复性差、难以实现规模化及连续化生产等问题，提出了一种基于双流体喷嘴雾化技术的新型磁性琼脂糖微球制备方法；探究了不同雾化条件下，液滴尺寸变化的规律；在优化后的喷雾条件下，成功制备出球形度高、粒径小、磁响应迅速的磁性琼脂糖微球；将筛分后的微球制备成DEAE阴离子交换剂（DEAE-MAM），并以牛血清白蛋白（BSA）为模型蛋白，探究了不同粒径下的DEAE-MAM的蛋白吸附性能。结果表明，在保证水相性质一致的前提下，喷雾条件通过改变气液比影响液滴尺寸，进而影响微球粒径；粒径最小的DEAE-MAM（d₃₂ = 36 μm）离子交换容量最大（192.5 μmol/ml），饱和吸附量最高（150.0 mg/ml）。

关键词: 气液两相流, 凝胶, 造粒, 雾化, 离子交换, 蛋白吸附

Abstract:

Aiming at the problems of the existing magnetic agarose microspheres (MAM) preparation technology, such as long process flow, poor repeatability, and difficulty in achieving large-scale and continuous production, a new preparation method of magnetic agarose microspheres based on two-fluid nozzle atomization technology was proposed. The variation of droplet size under different atomization conditions was investigated. Moreover, under the optimized spray conditions, MAM with high sphericity, small particle size and fast magnetic response were successfully prepared. The microspheres after sieving were prepared as DEAE anion exchanger (DEAE-MAM), and bovine serum albumin (BSA) was used as a model protein to explore the protein adsorption performance of DEAE-MAM with different particle sizes. The results show that the spray conditions affected the droplet size and thus the particle size by changing the gas-liquid ratio under the premise that the water phase properties were consistent. The DEAE-MAM with the smallest particle size (d₃₂ = 36 μm) had the largest ion exchange capacity (192.5 μmol/ml) and the highest saturated adsorption capacity (150.0 mg/ml).

Key words: gas-liquid flow, gel, granulation, atomization, ion exchange, protein adsorption

中图分类号:

O 636.1

高燕, 伍鹏, 尚超, 胡泽君, 陈晓东. 基于双流体喷嘴的磁性琼脂糖微球的制备及其蛋白吸附性能探究[J]. 化工学报, 2023, 74(8): 3457-3471.

Yan GAO, Peng WU, Chao SHANG, Zejun HU, Xiaodong CHEN. Preparation of magnetic agarose microspheres based on a two-fluid nozzle and their protein adsorption properties[J]. CIESC Journal, 2023, 74(8): 3457-3471.

图/表 18

图1 用于制备MAM的喷雾装置1—钢瓶减压阀;2—N2钢瓶;3~5—气路调压阀,对应的气压分别为开枪气压(PS)、雾化气压(PZ)和水相气压(PM);6~8—气路开关;9—料液罐;10—机械搅拌装置;11—双流体喷嘴;12—承接容器(直径18 cm);13—水浴加热装置

Fig.1 Spraying device for preparing MAM1—pressure reducing valve of steel cylinder; 2—N2 cylinder; 3-5—pressure regulating valve for gas pipelines, the corresponding nitrogen pressure is opening pressure (PS), atomization pressure (PZ), and water phase pressure (PM) respectively; 6-8—switches of gas pipelines; 9—liquid tank; 10—mechanical stirring device; 11—two-fluid nozzle; 12—receiving container (diameter 18 cm); 13—water bath heating device

图2 “外混式”双流体喷嘴的结构示意图

Fig.2 Schematic diagram of the “external mixing” two-fluid nozzle

图3 CL-MAM的制备

Fig.3 Preparation of CL-MAM

图4 DEAE-MAM的制备

Fig.4 Preparation of DEAE-MAM

图5 雾化区域对微球粒径的影响(a) 实验示意图;(b),(c) 粒径分布曲线;(d) 不同雾化气压下d32随径向距离的变化及不同区域中MAM收集量的体积分数

Fig.5 Effect of atomization area on particle size of MAM(a) experimental schematic; (b),(c) particle size distribution curves; (d) variation of d32 with radial distanceat differentatomization pressure and volume fraction of MAM collected in different areas

图6 水相黏度对MAM粒径的影响(a) CMC溶液的流量-CMC浓度的关系曲线;(b) 琼脂糖溶液的流变曲线(90℃);(c) 粒径分布曲线;(d) d32随琼脂糖浓度cA的变化

Fig.6 Effect of water phase viscosity on particle size of MAM(a) relationship curve between the flow rate of CMC solution and the concentration of CMC; (b) rheological curves of the agarose solution (90℃); (c) particle size distribution curves; (d) variation of d32 with agarose concentration cA

图7 水相气压对MAM粒径的影响(a) 水相流量-水相气压的关系曲线;(b) 粒径分布曲线;(c) d32随水相气压PM的变化

Fig.7 Effect of water phase pressure on particle size of MAM(a) relationship curve between the flow rate of water and the pressure of water phase; (b) particle size distribution curves; (c) variation of d32 with water phase pressure PM

图8 雾化气压对MAM粒径的影响(a) 气相流量-雾化气压的关系曲线;(b)~(d) 粒径分布曲线;(e) 不同喷嘴孔径下 d32随雾化气压的变化

Fig.8 Effect of atomization pressure on particle size of MAM(a) relationship curve between the flow rate of gas and the atomization pressure; (b)-(d) particle size distribution curves; (e) variation of d32 with atomization pressure at different nozzle aperture

图9 喷嘴孔径对MAM粒径的影响(a) 水相流量-喷嘴孔径的关系曲线;(b)~(e) 粒径分布曲线;(f) 不同雾化气压下 d32随喷嘴孔径的变化

Fig.9 Effect of the bore diameter of the nozzles on particle size of MAM(a) relationship curve between the flow rate of water and the bore diameter of the nozzles; (b)-(e) particle size distribution curves; (f) variation of d32 with the bore diameter of the nozzles at different atomization pressure

图10 MAM的磁响应性能

Fig.10 Magnetic response performance of the MAM

图11 不同粒径大小MAM的光学显微镜照片

Fig.11 Optical microscope images of the MAM with different sizes

图12 不同冻干方式制备的微球的扫描电子显微镜照片(a) AB,水冷冻干燥;(b) MAM,水冷冻干燥;(c) MAM,叔丁醇冷冻干燥;(d) CL-MAM,叔丁醇冷冻干燥

Fig.12 SEM images of MAM prepared using different freeze-drying methods(a) AB, freeze-drying with water; (b) MAM, freeze-drying with water; (c) MAM, freeze-drying with tert-butyl alcohol; (d) CL-MAM, freeze-drying with tert-butyl alcohol

图13 琼脂糖（agarose）、MNPs及MAM的傅里叶红外光谱图

Fig.13 FT-IR spectra of agarose, MNPs and MAM

图14 MAM 和MNPs的XRD谱图

Fig.14 X-ray diffraction patterns of MAM and MNPs

图15 经沸水浴加热后的微球的显微镜照片

Fig.15 Optical microscope images of the microspheres heated by boiling water

图16 不同粒径的DEAE-MAM的BSA吸附等温线

Fig.16 Adsorption isotherms of BSA on DEAE-MAM with different particle sizes

表1 DEAE-MAM对BSA吸附的平衡参数

Table 1 Equilibrium parameters of DEAE-MAM for BSA adsorbtion

DEAE-MAM	平均粒径d₃₂/μm	E/(μmol/ml)	Q_m/(mg/ml)	K_d	R²
喷雾法（自制）	36	192.5	150.0	0.0345	0.9927
	86	148.2	143.4	0.0348	0.9729
	142	136.9	121.9	0.1029	0.9406
乳化-冷却法^[36]（自制）	50	100.6	129.4	0.0245	0.9941
文献^[37]	—	168.0	109.3	—	—

图17 BSA在DEAE-MAM上的分布(a) 激光共聚焦显微镜照片;(b) 荧光强度分布曲线

Fig.17 Distribution of the BSA on DEAE-MAM(a) CLSM image; (b) fluorescence intensity distribution curve

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