Ultrasound-assisted Extraction Of Walnut Meal Protein And Its In-situ Real-time Monitoring Technology

Walnut meal is a by-product from walnut oil-extraction industry.It is rich in walnut meal protein and is a high-quality raw protein source for food.However,this high-quality protein resource has not been fully developed and utilized due to insufficient researches on the protein extraction and process-monitoring techniques.In view of these reasons,this study introduces the modern ultrasound technology combined with the micro-optical near-infrared spectroscopy technique to extract walnut meal proteins.The extraction efficiency and the process-monitoring accuracy of the near infrared analysis model are also taken into account.The main research results and conclusions are as follows:?1?Ultrasound-assisted extraction of walnut meal proteins:The purpose of this part is to improve the efficiency of protein extraction using different ultrasound modes.A six-frequency slit-type and five-frequency S-type ultrasound devices were used to compare the extraction effects of walnut meal proteins.The ultrasound working modes and extraction parameters were used to evaluate the impacts on the protein yield,purity,and weighted comprehensive score?Y?.The results show that the S-type ultrasound mode was superior to the slit-type one.The optimal frequency combination dual frequency mode was 20 and 28 kHz.Under the optimal working mode,the single-factor and orthogonal optimization experiments were carried out.The optimal ultrasound-assisted extraction parameters were ultrasound power density of 120 W/L,extraction time of 30 min,feed liquid temperature of 55°C,material-to-liquid ratio of 1:25 g/mL and pH value of 9.0.Under these conditions,the protein yield was 71.00%,the purity was 72.69%,and the Y value reached 71.68%.Compared with the non-ultrasound group?traditional alkali extraction group?,the extraction time reduced by 50%,the protein yield and purity increased by 7.30%and4.79%,respectively,and the comprehensive score Y increased by 6.29%.It can conclude that ultrasound-assisted extraction show obvious advantages in shortening the extraction time of walnut meal protein compared to triditional extration.?2?Study on the structure changes of walnut meal proteins during ultrasound assisted extraction:The purpose is to explore the mechanisms of ultrasound-assisted protein extraction.For this reason,the effects of ultrasound working time and power density on the molecular structures??-helix,?-turn,?-sheet,random coil,sulfhydryl,disulfide bond and surface hydrophobicity?and surface microscopic morphologies of walnut meal proteins were studied using Fourier transform infrared spectroscopy,fluorescence spectrophotometer and scanning electron microscopy.The results showed that ultrasound-assisted extraction could loosen the structures of walnut meal proteins,weaken the hydrogen bonding forces,and transformed the proteins from ordered structures to disordered structures,resulting in an increase in the relative contents of?-turn and random coil and a decrease in the relative contents of e?-helix and?-sheet.With the increase of extraction time and ultrasound intensity,the spatial structures of walnut meal protein were destroyed,and the disulfide bonds were broken,resulting in an increase in the content of sulfhydryl groups.Meanwhile,proteins were denatured,and hydrophobic groups were constantly exposed to the surfaces of the molecules,resulting in the increase of hydrophobicity?maximum at 30 min?.However,excessive extraction time would make protein molecules re-aggregate,causing the re-formation of disulfide bonds and the decrease of hydrophobicity.The results of scanning electron microscopy also showed that moderate ultrasound treatment could make the protein bonds break more thoroughly,the specific surface area smaller,resulting in more favorable for extraction,compared with the traditional alkali extraction.?3?Establishment of a quantitative association model between protein structure change and extraction yield:The purpose is to establish the spectral monitoring model.This work used Pearson correlation coefficient analysis,principal component analysis and stepwise regression analysis to study the correlation between walnut meal protein structure and extraction index in the ultrasound-assisted extraction process.The results of Pearson correlation analysis showed that there was a significant positive correlation between the sulfhydryl content of walnut meal proteins and protein yield?p<0.01?,indicating that the change of sulfhydryl content during ultrasound extraction was the main reason for the changes of protein yield.Results of the principal component analysis showed that the cumulative contribution rates of the three principal components reached 84.684%,reflecting all the changes of the original variables.The first main components were?-sheet,?-turn,random curl and hydrophobicity,the second main components were?-helix and sulfhydryl,and the third main component was disulfide bond.Through stepwise regression analysis,the established protein yield and thiol content equation was y?protein yield?=24.715+4.711 X₆?sulfhydryl content?,R²=0.966,indicating"sulfhydryl"was the key factor in the ultrasound-assisted extraction of walnut meal protein.The"characteristic structure"provides theoretical basis for the further establishment of an intelligent monitoring model for the ultrasound-assisted extraction process of walnut meal proteins.?4?In-situ real-time monitoring technology of sulfhydryl content in the process of ultrasound assisted extraction of walnut meal protein:In this paper,the in-situ real-time monitoring system and spectral acquisition method of protein structure in the process of ultrasound-assisted extraction of walnut peony protein were established using micro-fiber near-infrared spectroscopy.Several pretreatments selected for the spectra obtained from the ultrasound assisted extraction were Standard Normal Variate transformation?SNV?,Savizky-Golay?SG?,First Derivative(1^stDer)and Second Derivative(2^ndDer).The best pretreatment method for predicting the sulfhydryl content was evaluated by the correlation coefficient?Rc and Rp?and root mean square error?RMSECV and RMSEP?values.Under the optimal preprocessing method,the methods of Partial Least Squares?PLS?,interval Partial Least Squares?iPLS?and Synergy interval Partial Least Squares?Si-PLS?were used to build models.The results showed that the best pretreatment method was SG,and the best modeling method was Si-PLS.The optimal bands of the–SH based structure were946.946¹018.601 nm,1103.101¹174.448 nm,1496.646¹561.283 nm and1950.876²020.772 nm.The Rc,Rp,RMSECV and RMSEP values of the model were 0.9736,0.9342 and 0.446,0.807,respectively.The establishment of this model provides technical support for the design of intelligent system for extraction process in industrial production.