| 研究実績の概要 |
In this research, an enzyme will be immobilized inside the chemically modified mesoporous silica, MCM-41, to enhance both its stability and reactivity. The enzyme of interest in this work is protocatechuate 3,4-dioxygenase, which can selectively cleave a strong C-C bond in the catechol aromatic ring.
In year 2021, preparation of mesoporous silica and modification of its surface using silane coupling agent were achieved. As a choice of mesoporous silica, SBA-15 was synthesized, instead of MCM-41. This was due to the fact that (1) the protocol of its synthesis was well-established in the Ogura lab, the group where this research was carried out and (2) SBA-15 has a similar pore structure as MCM-41 with controlled pore size. The synthesized SBA-15 with average pore size of 8 nm was then subjected to surface modification by silane coupling to obtain the modified SBA-15, m-SBA-15.
To prove the stabilizing effect of the unstable enzyme via immobilization into mesoporous silica, a more commonly and widely used enzyme compared to protocatechuate 3,4-dioxygenase, lipase, was immobilized first into m-SBA-15. As a result, the increase in stability of lipase was successfully observed through immobilization into m-SBA-15. Moreover, methods to measure and quantify the amount of active lipase were explored and established.
In addition, SBA-15 with pore size of 10 nm and 12 nm that are commercially available were also purchased, and subjected to surface modification to compare the amount of active, immobilized lipase.
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| 現在までの達成度 (区分) |
現在までの達成度 (区分)
3: やや遅れている
理由
The progress of the research is slightly delayed. This is due to the fact that the synthesis of MCM-41 with large pore size required special techniques, which turned out to be not as straightforward as originally expected. To solve this issue, more suitable choice of mesoporous silica, SBA-15, was selected as a substitute material for its (1) well-established synthetic protocols at Ogura lab, where this research was carried out and (2) similar structure as MCM-41.
In addition, immobilization of enzyme into mesoporous silica turned out to require careful handling techniques due to its unstable nature. To establish the proper techniques to treat unstable enzymes, the research project of well-studied lipase was carried out before the immobilization of protocatechuate 3,4-dioxygenase, as indicated in the summary.
Through this approach, acquired protocols for treating lipase can be applied to the immobilization of protocatechuate 3,4-dioxygenase from year 2022, which will result in smooth transition of the research.
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| 今後の研究の推進方策 |
In year 2022 and 2023, completion of lipase immobilization project as well as exploration of protocatechuate 3,4-dioxygenase immobilization project will be carried out.
First, immobilized lipase will be utilized to explore its stability and reaction efficiency in details. At the same time, immobilization of protocatechuate 3,4-dioxygenase will be explored by applying the acquired protocols for handling unstable enzymes from the lipase project. During this transition, adjustment of the established protocols will be examined by optimizing reaction conditions for immobilization of protocatechuate 3,4-dioxygenase, which includes the screening of mesoporous silica as well as silane coupling agents for surface modifications.
Once the protocatechuate 3,4-dioxygenase is successfully immobilized into the right choice of mesoporous silica, the obtained material will be used to explore the degradation reactions of catechol moieties via C-C bond activation of aromatic rings.
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| 次年度使用額が生じた理由 |
The amount of grant to be used in the next fiscal year in year 2021 was 3,438 yen. This was mainly due to the fact that all of the conferences were held online instead of on-site, which reduced the travel expenses to 0 yen.
In the next fiscal year, more expense is expected to be used as travel expenses.
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