Thermoresponsive horseradish peroxidase nanocapsules (nHRP) were prepared via in situ precipitation polymerization using N-isopropyl acrylamide (NIPAM) as the monomer.As an example

nHRP was prepared by a NIPAM/HRP mass ratio of 6

under the reaction temperature of 37 ℃

it showed a monodispersed size distribution centered at 13.7 nm and a negative zeta potential centered at (-3.70.3) mV.The modification degrees of HRP were characterized by MALDI-TOF mass spectrometer (MALDI-TOF MS)

while the enzymatic activity of the free native HRP

modified HRP and encapsulated nHRP were determined by conducting a standard enzyme activity assay and followed by monitoring absorbance change through UV-Vis spectrometer.The particle size and morphology of nHRP were characterized using dynamic light scattering (DLS) and transmission electron microscopy (TEM).The results showed a monodispersed nHRP was prepared successfully.Compared with the native HRP

nHRP exhibits significantly improved thermal stability

according to the HRP activity assay.Thermoresponsive behavior of nHRP was examined from 25 ℃ to 45 ℃ by DLS studies

where nHRP showed a responsive temperature at 33 ℃ and a reversible response property.Accordingly

high-speed centrifugation was applied to separate and recycle nHRP at 50 ℃ with a well-retained activity.This work provides a new delicate way to immobilize enzyme based on thermoresponsive enzyme nanocapsules

which is of great interest for effective enzyme separation and recycling and other applications.