Study On The Mechanism Of Thermal Protection Of Protein To Betanin And Its Application In Red Pitaya Juice

Betanin(BN)is an edible natural pigment with a variety of biological activities,appearing rose color to purple red.However,instability limits the application of betanin in the food industry.Temperature,p H,oxygen and other factors would impact the stability of betanin.High temperature is the most threatening factor to the stability of betanin,and it is easy to discolor betanin to the light yellow and inactivate it.Food protein is a safe and edible carrier,which can interact with small natural pigment molecules and improve their stability.Three animal proteins(whey protein isolate,WPI;lactoferrin,BLF;β-lactoglobulin,BLG),three plant proteins(rice protein,RP;pea protein,PP;soybean protein isolate,SPI)and two complex plant proteins(ricesoybean complex protein,RP-SPI;rice-pea complex protein,RP-PP)were selected.The influence of those proteins on the thermal stability of betanin was explored,and the mechanism of improving the thermal stability of betanin was explained from the perspective of interaction.Finally,the optimal stabilization technology was applied to red pitaya juice.This study provides a new idea for the study of the stability of betanin,broadens the field of protein-betanin interaction,and provides a theoretical basis for the color protection of food systems rich in betanin,such as red pitaya juice.The main research results are as follows:(1)The addition of protein increased the retention rate of betanin at 80 ℃ for 60 min from 6.26% to 12.8%(RP),11.79%(PP),16.04%(SPI),27.26%(WPI),27.01%(BLF),13.86%(BLG).The effect of improving thermal stability was: WPI > BLF >SPI > BLG > RP > PP.The improvement of the thermal stability of protein to betanin was related to the non-covalent interaction,which was characterized by turbidimetry,fluorescence spectroscopy,molecular docking simulation and other means.The main interaction force of WPI-BN and BLG-BN were hydrogen bond.The main interaction force of BLF-BN and SPI-BN were hydrogen bond and electrostatic interaction.The main interaction force of RP-BN and PP-BN were hydrophobic interaction.(2)Complex plant proteins RP-SPI and RP-PP could significantly improve the thermal stability of betanin,and increase the retention rate of betanin at 80 ℃ for 60 min from 6.26% to 48.08% and 43.52%.The results of fluorescence spectrum,ITC,FTIR and molecular docking showed that RP-SPI interacted with betanin through hydrogen bond,and RP-PP bound with betanin through hydrophobic interaction,with binding-site number about 1.The binding constant of RP-SPI-BN was larger than RPPP-BN at room temperature.Binding with betanin altered the protein’s secondary structure content and reduced its surface hydrophobicity.The results of scanning electron microscopy showed that RP-SPI and RP-PP were porous network structural proteins,which may be more conducive to the transport and encapsulation of bioactive small molecules.It was observed by transmission electron microscope that the protein was aggregated and the particle size increased after interacting with betanin.In protein-betanin complex system,betanin acted as a bridge between proteins through non-covalent interaction and was enclosed by surrounding proteins,which weakened the impact of external factors on betanin and improved its stability.(3)The whole syrup of red pitaya prepared by industry-scale microfluidization system was an even fine juice system rich in betanin.After centrifugal filtration,the pigment stability of the clear juice decreased,and the order of pigment stability was:whole syrup of red pitaya(YJ)> clear juice of red pitaya(QZ)> betanin aqueous solution,which may be because of the positive effect from the interaction between betanin and protein,polysaccharide in the juice system.The addition of different content of RP-SPI and RP-PP to QZ decreased the degradation rate constant from0.0628 to 0.0383,and increased the half-life of betanin from 11.04 min to 21.87 min(5%RP-SPI)and 18.10 min(5% RP-PP)at 80 ℃,respectively.Complex plant protein could effectively reduce the loss of betanin in the heat sterilization process,and the degradation and discoloration of red pitaya juice in the storage process.The sequence of betanin protection of QZ by complex plant protein was : 5% RP-SPI > 5% RP-PP >2% RP-PP ≈ 2% RP-SPI.