Boiling-induced nanoparticles and their constitutive proteins from Isatis indigotica Fort. root decoction: Purification and identification

Colloidal particles are essential components of sun-dried Isatis indigotica Fort. roots (Ban-Lan-Gen in Chinese, BLG) decoction. Nanoparticles (NPs) were isolated from BLG decoction with size exclusion chromatography and characterized. Their average diameter is ∼120 nm, reversibly responding to pH and temperature changes. They promoted the growth of normal cells but suppressed that of cancerogenic cells and macrophages. Two constitutive glycated proteins were identified from the NPs, namely BLGP1 and BLGP2. Their N-terminal amino acid sequences were V-X-R-E-V-V-K-D-I and V-V-R-E-V-V-K-D-I-A-G-A-V-Q-T-N-E-Q-Y. Their full-length cDNA sequences were cloned to obtain the highly homological amino acid sequences of non-glycated proteins, whose theoretical molecular weights are 21831.64 Da and 21841.67 Da. Using pepsin hydrolysis and mass spectrometry, four possible glycation adducts were identified in BLGP1, whereas one in BLGP2. To conclude, bioactive nanoparticles isolated from the herbal decoction are intelligent nanoassemblies composed of a new boiling-stable protein. Glycation plays a critical role in heat-induced formation of these nanoassemblies. The novel, intelligent, safe and stable nano-carriers for drug delivery may be developed using BLG NPs as prototype.