Efficient production of isoquercitin, icariin and icariside II by a novel thermostable α-l-rhamnosidase PodoRha from Paenibacillus odorifer with high α-1, 6-/α-1, 2- glycoside specificity.

Minor flavonoids, such as isoquercitin, icariin and icariside II, have desired bioactivity and bioavailability. To enhance the production efficiency on converting rutin and epimedin C into minor flavonoids, a novel GH family 78 α-l-rhamnosidase PodoRha from the Paenibacillus odorifer was cloned and heterologous over-expressed in E. coli BL21(DE3). PodoRha exhibited a high catalytic ability in cleaving outer α-1, 2-rhamnopyranosyl moieties of epimedin C and α-1, 6- rhamnopyranosyl of rutin. It displayed the highest activity at 45 °C and pH 6.5, and has great thermostability at 40 °C. Under the optimal transformation condition (40 °C, pH 6.5, 1 h incubation), 10 g/L of Rutin was 99.86% transformed into 6.95 g/L of isoquercitin by PodoRha with a crude productivity of 6.95 g·L·h; 1.964 g/L icariin was detected after the epimedin C of the total Epimedium flavonoids (10 g/L) were 99.98% converted by PodoRha. Moreover, by using a two-stage conversion system with a crude icariside II productivity of 2.081 g·L·h, 5 g/L epimedin C was 99.85% converted into 3.122 g/L of icariside II with 1 h incubation under pH 6.5 at 40 °C after addition of 5.5 U/mL PodoRha; then with 30 min incubation under pH 6.5 at 90 °C after addition of 0.8 U/mL IagBgl1.