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Enzyme specificity and particularity is needed not only in enzymatic separation methods, but also in enzymatic determination methods for plant compound extraction. Stevioside, rubusoside, and rebaudioside A are natural sweet compounds from plants. These compounds have the same skeleton and only contain different side-chain glucosyl groups, making them difficult to separate. However, enzymes that target diterpenoid compounds and show specific activity for side-chain glucosyl groups are rare. Herein, we report the identification and characterization of an enzyme that can target both diterpenoid compounds and sophorose, namely, β-glucosidase SPBGL1 from Sphingomonas elodea ATCC 31461. SPBGL1 displayed high specificity toward sophorose, and activity toward stevioside, but not rebaudioside A. The stevioside conversion rate was 98%. SPBGL1 also operated at high substrate concentrations, such as in 50% crude steviol glycoside extract. Glucose liberated from stevioside was easy to quantify using the glucose oxidase method, allowing the stevioside content to be determined.
Copyright © 2019 Elsevier Ltd. All rights reserved.
Enzymatic determination method; Plant content measurement; Stevioside; β-Glucosidase
Highly specific sophorose β-glucosidase from Sphingomonas elodea ATCC 31461 for the efficient conversion of stevioside to rubusoside.
Lan Q1, Tang T1, Yin Y1, Qu X1, Wang Z1, Pang H2, Huang R1, Du L3.
2019 Oct 15
Diabetes is one of the most severe chronic diseases worldwide. It is widely accepted that apoptosis of the pancreatic beta cell is an important cause for the induction of hyperglycemia and high levels of free fatty acids (FFAs), also called lipotoxicity associated with pancreatic beta cell dysfunction. Lipotoxicity has been proven to be an important pathogenic factor of diabetes. However, until now, the mechanism of FFA-induced lipotoxicity in INS-1 cells has not been fully understood. Current anti-diabetic drugs that protect islet cells are often toxic to healthy cells, resulting in negative side effects. Thus, there is an urgent need to identify more effective and safer anti-diabetic agents to protect pancreatic islet cells. Rubusoside (RUB) is a major ingredient in the leaves of Rubus suavissimus S. Lee, which decreases blood glucose levels by protecting pancreatic islet cells. However, the exact mechanism of this effect is unknown. In this study, metabolomics experiments based on UPLC-Q/TOF MS characterized a total of 15 metabolites as potential biomarkers associated with lipotoxicity induced by palmitic acid in INS-1 cells. According to the metabolic pathway analysis, pentose and glucuronate interconversions, and glycerophospholipid metabolism were recognized as the most influenced metabolic pathways associated with lipotoxicity. Unexpectedly, deviations of 14 metabolites in lipotoxic INS-1 cells were regulated by RUB, suggesting synergistic mediation of the abnormal metabolic pathways. The metabolomics strategy based on UPLC/Q-TOF MS analysis provides a new insight into the mechanisms of lipotoxicity induced by palmitic acid and the anti-lipotoxic activity of RUB in INS-1 cells.
Metabolomics analysis of the protective effect of rubusoside on palmitic acid-induced lipotoxicity in INS-1 cells using UPLC-Q/TOF MS.
Zheng H1, Wu J2, Huang H3, Meng C1, Li W1, Wei T2, Su Z2.
2019 Jun 1;
Diterpenoid UDP-Glycosyltransferases from Chinese Sweet Tea and Ashitaba Complete the Biosynthesis of Rubusoside.
Sun Y1, Chen Z2, Li J3, Li J1, Lv H1, Yang J4, Li W4, Xie D4, Xiong Z1, Zhang P3, Wang Y5.
2018 Oct 8