Abstract
Objective
To improve the efficiency of reactions of β-glucuronidase (GUS)-assisted glucuronic acid (GluA) removal within a microfluidic system.
Results
β-glucuronidase from Helix pomatia was immobilised and characterised in silica-based sol–gel monoliths. Efficiency of the GUS-doped silica monoliths was tested for hydrolysis of p-Nitrophenyl-β-d-glucuronide (pNP–GluA) in both ml-scaled medium via batch reactions and microfluidic environment via continuous-flow reactions. In the microfluidic platform, within a duration of 150 min of continuous operation (flow rate: 1 µL/min), the obtained highest pNP yield was almost 50% higher than that of the corresponding batchwise reaction. However, increased flow rates (3, 5, and 10 µL/min) resulted in lower conversion yields compared to 1 µL/min. The microfluidic platform demonstrated continuous hydrolytic activity for 7 days with considerable reaction yields while using a small amount of the enzyme.
Conclusion
These results revealed that usage of the microreactors has considerable potential to efficiently obtain bioactive GluA-free aglycons from various plant-derived β-glucuronides for pharmaceutical applications.
Graphical Abstract
Similar content being viewed by others
References
Bolivar JM, Wiesbauer J, Nidetzky B (2011) Biotransformations in microstructured reactors: more than flowing with the stream? Trends Biotechnol 29:333–342
Chen Q, Schönherr H, Vancso GJ (2009) Block-copolymer vesicles as nanoreactors for enzymatic reactions. Small 5:1436–1445
Chen L, Wei B, Zhang X, Li C (2013) Bifunctional graphene/γ-Fe2O3 hybrid aerogels with double nanocrystalline networks for enzyme immobilization. Small 9:2331–2340
Choi R, Ha I, Choi J, Park Y, Kim Y (2010) Biotransformation of flavonoid-7-O-glucuronides by β-glucuronidases. Nat Prod Sci 16:1–5
Cichna M (2003) Applicability of enzyme columns prepared by co-immobilising β-glucuronidase and arylsulfatase by the sol-gel method for deconjugation of glucuronides and sulfates in urine. J Sol-Gel Sci Tech 26:1159–1164
Cumana S, Ardao I, Zeng A-P, Smirnova I (2014) Glucose-6-phosphate dehydrogenase encapsulated in silica-based hydrogels for operation in a microreactor. Eng Life Sci 14:170–179
Fernandes P (2010) Miniaturization in biocatalysis. Int J Mol Sci 11:858
Huang HZ, Feng B, Song XB, Ma BP (2011) Purification and characterization of glycyrrhizin-β-d-glucuronidase and baicalin-β-d-glucuronidase from a commercial enzyme preparation. Biocatal Biotransform 29:179–185
Jensen KF (2001) Microreaction engineering—is small better? Chem Eng Sci 56:293–303
Kaneno J et al (2003) A simple method for surface modification of microchannels. New J Chem 27:1765–1768
Kanno KI, Maeda H, Izumo S, Ikuno M, Takeshita K, Tashiro A, Fujii M (2002) Rapid enzymatic transglycosylation and oligosaccharide synthesis in a microchip reactor. Lab Chip 2:15–18
Kazan A et al (2017) Formulation of organic and inorganic hydrogel matrices for immobilization of β-glucosidase in microfluidic platform. Eng Life Sci 17:714–722
Kim HS, Kim JY, Park MS, Zheng H, Ji GE (2009) Cloning and expression of beta-glucuronidase from Lactobacillus brevis in E. coli and application in the bioconversion of baicalin and wogonoside. J Microbiol Biotechnol 19:1650–1655
Lai J-K, Chuang T-H, Jan J-S, Wang SS-S (2010) Efficient and stable enzyme immobilization in a block copolypeptide vesicle-templated biomimetic silica support. Colloid Surf B 80:51–58
Miller SA, Hong ED, Wright D (2006) Rapid and efficient enzyme encapsulation in a dendrimer silica nanocomposite. Macromol Biosci 6:839–845
Morana A, Di Lazzaro A, Di Lernia I, Ponzone C, De Rosa M (2002) Enzymatic production of 18-β-glycyrrhetinic acid from Glycyrrhiza glabra L. Biotechnol Lett 24:1907–1911
Nakagawa K, Tamura A, Chaiya C (2014) Preparation of proteolytic microreactors by freeze-drying immobilization. Chem Eng Sci 119:22–29
Sheldon RA, van Pelt S (2013) Enzyme immobilisation in biocatalysis: why, what and how. Chem Soc Rev 42:6223–6235
Siddharth T (2014) Evaluation of wild type and mutants of β-Glucuronidase (GUS) against natural and synthetic substrates. Dissertation, University of Saskatchewan
Woo HH, Jeong BR, Hawes MC (2005) Flavonoids: from cell cycle regulation to biotechnology. Biotechnol Lett 27:365–374
Yesil-Celiktas O (2014) Patenting trends in enzyme related microfluidic applications. Biochem Eng J 92:53–62
Yesil-Celiktas O, Cumana S, Smirnova I (2013) Silica-based monoliths for enzyme catalyzed reactions in microfluidic systems with an emphasis on glucose 6-phosphate dehydrogenase and cellulase. Chem Eng J 234:166–172
Yildiz-Ozturk E, Yesil-Celiktas O (2015) Diffusion phenomena of cells and biomolecules in microfluidic devices. Biomicrofluidics 9:052606
Zhang Y, Wu H, Li L, Li J, Jiang Z, Jiang Y, Chen Y (2009) Enzymatic conversion of Baicalin into Baicalein by β-glucuronidase encapsulated in biomimetic core-shell structured hybrid capsules. J Mol Catal B Enzym 57:130–135
Acknowledgements
The financial support provided by the Scientific and Technological Research Council of Turkey (TUBITAK, 113M050) is highly appreciated. Special thanks are offered to Dr Barbaros Cetin from the Bilkent University Microfluidics & Lab-on-a-chip Research Group for microfabrication studies, and Dr Bogdan Parakhonskiy from the Ghent University for his valuable comments on the manuscript.
Supporting information
Supplementary Material Section A—methods of activity measurements for enzyme kinetics and stability.
Supplementary Material Section B—procedures regarding fabrication of the microfluidic apparatus.
Supplementary Fig. 1—Stability of immobilised GUS at 4 and 37 °C.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of interest
The authors have no conflicts of interest in relation to the manuscript.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Muderrisoglu, C., Sargin, S. & Yesil-Celiktas, O. Application of β-glucuronidase-immobilised silica gel formulation to microfluidic platform for biotransformation of β-glucuronides. Biotechnol Lett 40, 773–780 (2018). https://doi.org/10.1007/s10529-018-2530-7
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10529-018-2530-7