The mixture was incubated at pH 5.0–7.0 for reaction times ranging from 1 to 6 h, while the temperature of the water bath was kept steady at a given temperature(reaction temperature ranged from 40 to 60◦C). The reaction was stopped by boiling the samples for 10 min, KOG thus obtained was concentrated with a rotary evaporator, and then mixed with 95%ethyl alcohol.
The KOG, which had been collected as a precipitate by centrifugation at 4000 rpm for 20 min, was resuspended in 95% ethyl alcohol three times. After being redissolved in distilled water, KOG was nanofiltrated using a polymer membrane (MWcut-off limit = 8000 Da) to remove undegradated KG and was then lyophilized in a freeze-dryer.