Pore size of macroporous polystyrene microspheres affects lipase immobilization
Pore size of macroporous polystyrene microspheres affects lipase immobilization
复制标题
大孔聚苯乙烯微球孔径影响脂肪酶固定化
DOI：
10.1016/j.molcatb.2010.05.007
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发表时间：
2010-09-01
影响因子：
--
通讯作者：
Zhou, Wei-Qing
中科院分区：
文献类型：
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作者：
Li, Yan;Gao, Fei;Zhou, Wei-Qing
聚苯乙烯（PST）微球具有机械强度理想、粒径可调、化学稳定性好等优点，被广泛用于固定化载体。然而，利用大孔径微球（> ~ 100nm）进行固定化的报道很少。我们用一种新颖的方法成功地制备了具有大孔和千兆孔的PST微球。本研究采用千孔/宏孔/介孔PST微球（孔径分别为314 nm、104 nm和14.7 nm）通过强疏水相互作用固定化来自洋葱伯氏杆菌的脂肪酶，并详细研究了孔径对脂肪酶分布、相对活性、动力学行为、热稳定性、储存稳定性和可重复使用性的影响。激光扫描共聚焦显微镜（LSCM）观察，脂肪酶渗透到巨孔微球的中心。对于介孔微球，脂肪酶仅吸附在外壳上。千兆孔/大孔/介孔PST微球固定化脂肪酶的相对活性分别为146%、126%和50.9%。与游离脂肪酶的动力学常数（0.441 mM）相比，介孔pst -脂肪酶的K-m值（0.532 mM）较高，而千兆/大孔pst -脂肪酶的K-m值（0.402 mM和0.411 mM）相对较低，表明底物对酶活性位点的可及性是无限的。随着孔隙尺寸的增大，材料的热稳定性、贮存稳定性和可重复使用性均有显著提高。在刺激体系中，即使经过100次循环，固定化在巨孔和大孔PST微球上的脂肪酶的活性仍然接近100%和93%，而在14.7 nm孔的PST微球上固定化的脂肪酶的活性仅能保持64.1%。在实际体系中，巨孔pst -脂肪酶和大孔pst -脂肪酶在使用30次后分别保持73.5%和68.8%的活性，而中孔pst -脂肪酶仅保持49.4%的活性。因此，与其他微球相比，巨孔PST微球作为一种潜在的酶载体在工业上具有明显的优势。(C) 2010 Elsevier B.V.版权所有
Polystyrene (PST) microspheres are commonly chosen as immobilization carriers due to their unique advantages such as ideal mechanical strength, adjustable particle size, and favorable chemical stability. However, there were few reports on immobilization by using microspheres with large pore sizes (>100 nm). We have successfully prepared the PST microspheres with macropores and gigapores by a novel method. In this study, giga-/macro-/meso-porous PST microspheres (314 nm, 104 nm, and 14.7 nm in pore sizes) were employed to immobilize lipase (from Burkholderia cepacia) by strong hydrophobic interactions, and the effects of pore sizes on lipase distribution, relative activity, kinetic behavior, thermal stability, storage stability, and reusability were also investigated in detail. According to laser scanning confocal microscope (LSCM) observation, lipase penetrated into the center of those giga-/macro-porous microspheres. With regard to the mesoporous microspheres, lipase only adsorbed to the external shell. The relative activities of immobilized lipase were 146%, 126%, and 50.9% for giga-/macro-/meso-porous PST microspheres, respectively. Comparing with the kinetic constants of free lipase (0.441 mM), the K-m value for mesoporous PST-lipase (0.532 mM) was higher, whereas for giga-/macro-porous PST-lipase (0.402 mM and 0.411 mM), the K-m values were comparatively lower suggesting the accessibility of substrate to the enzyme active sites was unlimited. The thermal stability, storage stability, and reusability were all improved significantly with the increase of pore sizes. In stimulant system, even after 100 times of recycling, the activity of lipase immobilized on gigaporous and macroporous PST microspheres remained nearly 100% and 93%, respectively, while that of lipase-PST with 14.7 nm pores could only keep 64.1%. In real system, 73.5% and 68.8% activity of gigaporous PST-lipase and macroporous PST-lipase, respectively, were retained after being used 30 times, whereas only 49.4% activity was remained for mesoporous PST-lipase. Therefore, compared with other microspheres, the gigaporous PST microsphere exhibited obvious advantages as a potential enzyme support in industry. (C) 2010 Elsevier B.V. All rights reserved.