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Open AccessArticle

Colorimetric Determination of the Activity of Starch-Debranching Enzyme via Modified Tollens’ Reaction

Graduate School of Biotechnology & Department of Food Science and Biotechnology, College of Life Sciences, Kyung Hee University, Yongin 17104, Korea
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Nanomaterials 2019, 9(9), 1291; https://doi.org/10.3390/nano9091291
Received: 6 August 2019 / Revised: 3 September 2019 / Accepted: 6 September 2019 / Published: 10 September 2019
(This article belongs to the Special Issue Nanostructured Materials and Natural Extract)
Nelson–Somogyi and 3,5-dinitrosalicylic acid (DNS) assays are the classical analytical methods for the determination of activity of starch-debranching enzymes, however, they have a narrow detection range and do not adapt to the quantitative measurement of linear polysaccharides. Herein, we developed a simple and accurate colorimetric assay for determining the activity of starch-debranching pullulanase through the modified Tollens’ reaction in combination with UV irradiation. Silver nanoparticles (AgNPs) were formed by reducing aldehyde groups in short-chain glucans (SCGs) generated by debranching of waxy maize starch using pullulanase through the modified Tollens’ reaction. In addition to providing a reducing moiety to the Tollens’ reaction, the debranching product, SCGs, also enhanced the colloidal stability of synthesized AgNPs, of which the amplitude of its surface plasmon resonance (SPR) absorbance peak was proportional to the concentration of SCGs ranging from 0.01–10 mg/mL. The detection limit of this system was 0.01 mg/mL, which was found to be 100 times higher than that of the conventional DNS assay. The purification of SCGs by recrystallization and gelatinization improved the selectivity of this colorimetric assay for debranching products, which provides a simple and accurate means of monitoring the debranching process and characterizing the activity of starch-debranching enzymes. View Full-Text
Keywords: silver nanoparticles; pullulanase; starch-debranching; Tollens’ reaction; colorimetric assay; enzyme activity silver nanoparticles; pullulanase; starch-debranching; Tollens’ reaction; colorimetric assay; enzyme activity
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Luo, K.; Kim, N.-G.; You, S.-M.; Kim, Y.-R. Colorimetric Determination of the Activity of Starch-Debranching Enzyme via Modified Tollens’ Reaction. Nanomaterials 2019, 9, 1291.

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