Catalysis Division, National Chemical Laboratory, Pune 411008, India.
Biochemical Sciences Division, National Chemical Laboratory, Pune 411008, India.
Int J Biol Macromol. 2014 Feb;63:218-24. doi: 10.1016/j.ijbiomac.2013.11.008. Epub 2013 Nov 25.
We describe herein a simple and effective strategy for immobilization of bile salt hydrolase enzyme by grafting glutaraldehyde groups inside channels of APTES functionalized SBA-15. The increase in glutaraldehyde concentration prevents leakage of enzyme but showed a steep decrease in enzyme activity in the immobilized matrix. So the degree of cross-linking should be the minimum possible to ensure sufficient stability without loss of activity. Cross-linking carried out with 0.1% glutaraldehyde concentration showed the highest activity, so this was used in all further experiments. Physico-chemical characterizations of the immobilized enzyme were carried out by XRD, N2 adsorption, TEM, FTIR and (29)Si CP-MAS NMR techniques. Immobilized BSH exhibits enhanced stability over a wide pH (3-11) and temperature range (40-80 °C) and retains an activity even after recycling experiments and six months of storage. From our in vivo research experiment toward co-precipitation of cholesterol, we have shown that immobilized BSH enzyme may be the promising catalyst for the reduction of serum cholesterol levels in our preliminary investigation. Enhancement in pH stability at the extreme side of pH may favor the use of immobilized BSH enzyme for drug delivery purpose to with stand extreme pH conditions in the gastrointestinal conditions.
我们在此描述了一种简单有效的策略，通过在 APTES 功能化 SBA-15 的通道内接枝戊二醛基团，将胆汁盐水解酶固定化。戊二醛浓度的增加可以防止酶泄漏，但在固定化基质中酶活性急剧下降。因此，交联程度应尽可能低，以确保在不损失活性的情况下具有足够的稳定性。用 0.1%戊二醛浓度进行交联显示出最高的活性，因此在所有进一步的实验中都使用了该浓度。通过 XRD、N2 吸附、TEM、FTIR 和 (29)Si CP-MAS NMR 技术对固定化酶进行了物理化学表征。固定化 BSH 在较宽的 pH（3-11）和温度范围（40-80°C）下表现出增强的稳定性，即使在重复使用实验和六个月的储存后仍保持活性。从我们针对胆固醇共沉淀的体内研究实验中，我们已经表明，固定化 BSH 酶可能是降低血清胆固醇水平的有前途的催化剂，这是我们初步研究的结果。在 pH 值极端一侧的 pH 值稳定性的提高可能有利于将固定化 BSH 酶用于药物输送目的，以在胃肠道条件下耐受极端 pH 值条件。
