Effect And Mechanism Of Ultrasonication Assisted With Glycation On The Functional Properties Of Proteins

Protein can provide an essential nutrient for human,and provide the different colores,smell,status,taste of food.The role of protein depends upon its functional characteristics.Food technology and processing affects the functional properties of protein is always the focus of research at home and abroad.Ultrasonication can promote protein glycation,but the relationship between structural and functional properties of glycoprotein before and after ultrasonic treatment,and the effect and mechanism of ultrasonication on the functional properties of glycoprotein are still unclear.Therefore,mechanism of ultrasonication assisted with glycation on the functional properties of proteins were researched,it can provide theoretical support and technical guidance for the modified proteins,and is of great significance to improve the nutrition of the processed food.In this thesis,bovine serum albumin?BSA?and?-Lactalbumin??-La?were used as the research object.The ultrasonic treatment,ultrasonic pretreatment assisted with glycation were used to processing BSA and?-La.The spectroscopy,chromatography,mass spectrometry and proteomics research methods were used to measure the changes in the structural and functional properties of BSA and?-La treated by ultrasonic pretreatment assisted with glycation and the reasons for the changes was analyzed.The intrinsic molecular mechanism of the improved antioxidant capacity and decreased allergenicity of?-La treated by ultrasonic pretreatment assisted with glycation,and glycation properties would be clarified from the molecular level.The main conclusions of this thesis ware as follows:?1?The influences of the different ultrasonic time,power and temperature assisted with glycation on the functional properties of BSA were researched.The results showed that the different ultrasonic time,power and temperature assisted with glycation modification can increase molecular weight of BSA and reduce its intrinsic fluorescence intensity.The free amino acids content of the BSA-galactose conjugates was gradually reduced and its DG value,surface hydrophobicity?H₀?,emulsifying ability and thermal stability increased with the increase of ultrasonic power and temperature.When the prolonged ultrasonic time,the free amino acids content,H₀,emulsifying ability,thermal stability of BSA-galactose conjugates initially increased and then decreased.With the increase of ultrasonic power and time,foaming ability of BSA-galactose conjugates gradually increased,and its foaming capacity initially increased and then decreased with the increase of temperature.Ultrasonic pretreatment assisted with glycation can improve the functional properties of BSA by changing its structure.Moreover,the different ultrasonic parameters can significantly affect the functional properties.?2?The influences of the ultrasonic power assisted with glycation?isomer of glucose and galactose?on the functional properties of BSA were researched.The results showed that ultrasonic pretreatment can significantly increase the free amino acids,emulsifying and foaming ability of BSA.The molecular weight,degree of browning of BSA-galactose conjugates were greater than those of the BSA-glucose conjugates,and ultrasonic pretreatment can improve them.Free amino acids content,intrinsic fluorescence intensity,H₀,emulsifying and foaming ability of BSA-galactose conjugates were lower than those of the BSA-glucose conjugates,and ultrasonic pretreatment can significantly reduce them.The glycation reaction between BSA and glucose and galactose,the reaction rate was:galactose>glucose.Ultrasonic pretreatment was significantly improved the reaction rate.Different conformational structure of sugar can significantly affect protein glycation reaction,change the structure of protein,leading to improve its functional properties.?3?The immunogenic properties were estimated by the IgG/IgE-binding abilities to evaluate the effect of different ultrasonic power on allergenicity of?-La.Research the correlation among changed structure and allergenicity of?-La treated by ultrasonic treatment.The results showed that the ultrasonic treatment can gradually increase the allergenicity of?-La.With the increase of the ultrasonic power,the allergenicity gradually was enhanced.Ultrasonic treatment does not change the molecular weight of?-La and make it polymerization reaction.Ultrasonic treatment can unfold?-La,increase the surface hydrophobicity and?-spiral content.ESI-MS results showed that?-La contains four components,including calcium,succinyllysine,N-acetylglucosamine and N-acetylneuraminic acid.Interestingly,the ultrasonic treatment can affect their relative abundance.Therefore,the structural changes of?-La treated by ultrasonic treatment can improve the allergenicity of?-La to some extent.?4?The effects of ultrasonic pretreatment assisted with dry heating-induced glycation between?-La and galactose on the IgE/IgG-binding ability and glycation extent of?-La were investigated,which determined by inhibition ELISA assays and high-resolution mass spectrometry respectively.The IgE/IgG-binding ability of glycated?-La was significantly decreased as a result of ultrasonic pretreatment,while average molecular weight,incorporation ratio?IR?value,the location and number of glycation sites,and degree of substitution per peptide?DSP?value were elevated.When the mixtures of?-La and galactose pretreated by ultrasonic power at 150W/cm²,the glycated?-La possess seven glycation sites,the highest IR and DSP value,and the lowest IgE/IgG-binding ability.Therefore,the decrease in IgE/IgG-binding ability of?-La depend not only on the glycation of Lys13,Lys16,Lys58,Lys93 and Lys98 sites leading to a shielding effect of the epitopic areas but also on the intensified glycation extent,which reflected in the increase of IR value,the number of glycation sites and DSP value.Moreover,allergenic proteins and monosaccharides pretreated by ultrasonication then undergo dry-state glycation was revealed as a promising way of achieving lower allergenicity of proteins in food processing.?5?The relationship between bovine?-La subjected to ultrasonication and glycation treatment with respect to antioxidant activity and glycation extent were investigated?determined by ABTS⁺Radical Scavenging Activity?.After?-La was pretreated by ultrasonication assisted with glycation,the treated?-La showed low intrinsic fluorescence emission and high antioxidant activity at increased ultrasonic power levels.Prior to ultrasonic pretreatment,three glycated sites were identified,whereas the number of glycation sites was increased to four,four,five and six after ultrasonic power at 60,90,120,and 150 W/cm²,respectively.All the samples pretreated by ultrasonication exhibited a higher DSP compared with the untreated samples.Therefore,the intensified glycation extent and the conformational changes of protein were responsible for the increase of antioxidant activity of?-La.?6?The glycation properties?molecular weight,glycation sites and number,and DSP values,etc.?of?-La treated by the different ultrasonic time assisted with glycation modification were investigated.The results showed that the different ultrasonic time assisted with glycation modification can significantly change the primary and tertiary structure of?-La.Prior to ultrasonic pretreatment,three glycated sites were identified,whereas the number of glycation sites was increased to four,four,five and five after ultrasonic time at 5,10,15,and 20 min,respectively.All the samples pretreated by ultrasonication exhibited a higher DSP compared with the untreated samples.Therefore,different ultrasonic time assisted with glycation can affect the glycation properties of?-La.Moreover,the prolonged ultrasonic time can dramatically affect its glycation properties to extent.