Effects Of Modified Technique On The Structure Of Goat Milk ?-Lactoglobulin And Its Peptidomic Profile After Enzymatic Hydrolysis

Due to its unique structural and physiochemical properties,?-lactoglobulin is a relatively small protein with high nutritional value and has certain biological properties.Compact structure of natural?-lactoglobulin makes it difficult to be digested by pepsin.Bovine milk?-lactoglobulin has been extensively studied,much less is known about physiochemical and structural properties of?-lactoglobulin of other milks.Due to the differences in amino acids sequence,the physical and chemical properties of goat and cow milk?-lactoglobulin were different.The objective of this study was to study the structural and digestive properties of goat milk?-lactoglobulin after heat and ultrasound treatments.Effects of modification treatment on the digestive characteristics of?-lactoglobulin were analyzed by using peptidomics techniques.Combined with bioinformatics technology and UWM database,bioactive peptides released from the digestion of?-lactoglobulin were identified.Additionally,effects of food matrix on the digestive properties of goat milk?-lactoglobulin were also investigated in this study.The main conclusions were drawn as follows:(1)This study aimed to compare the effects of high intensity ultrasound(HIU)applied at various amplitudes(20?40%)and for different durations(1?10 min)on the physiochemical and structural properties of goat milk?-lactoglobulin.No significant change was observed in the protein electrophoretic patterns by sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE).Deconvolution and second derivative of the Fourier Transform Infrared spectra(FTIR)showed that the percentage of?-sheet of goat milk?-lactoglobulin was significantly decreased while those of?-helix and random coils was increased after HIU treatment The surface hydrophobicity index and intrinsic fluorescence intensity of samples was enhanced and increased with increasing HIU amplitude or time.Differential scanning calorimetry(DSC)results exhibited that HIU treatments improved the thermal stability of goat milk?-lactoglobulin.Transmission electron microscopy(TEM)of samples showed that the goat milk?-lactoglobulin molecules interactions had occurred and it contained larger aggregates when compared with the untreated goat milk?-lactoglobulin sample.Data suggested that HIU treatments resulted in secondary and tertiary structural changes of goat milk?-lactoglobulin and improved its thermal stability.(2)Sequence of amino acids,secondary and tertiary structure of goat milk?-lactoglobulin were highly close to those of bovine milk?-lactoglobulin.Effects of mild heat treatment combined with high-intensity ultrasonic treatment(20 k Hz,40%amplitude,30 min,60±3 ?)on the digestion ability of goat and bovine milk?-lactoglobulin were investigated.Results shown that the primary structure of both goat and bovine milk?-lactoglobulins were remained stable after treatment.For the analysis of secondary structure of protein,content of?-sheet was significantly decreased from48.80±0.56%(before treatment)to 37.37±0.21%(after treatment).Base on the results described above,the tertiary structure of all?-lactoglobulin samples were also changed after treatment.Most of hydrophobic groups in protein molecules were exposed and the values of surface hydrophobicity index for goat milk?-lactoglobulin were increased from 353.77±1.25 to 828.96±8.43.Values of free sulfhydryl groups of both goat and cow?-lactoglobulin were decreased(especially in goat milk?-lactoglobulin,50.18±1.32?mol/g to 32.71±0.84?mol/g)after mild heat combined with high-intensity ultrasonic treatments.Treated goat and bovine milk?-lactoglobulin were more sensitive,and easy to be digested by pepsin,and the degradation ratio of two proteins were accelerated in the simulated intestinal.Peptidomics was used to identify the composition of the peptides in the digestive products.Results shown that the digestibility of goat milk?-lactoglobulin was significantly higher than that of cow milk?-lactoglobulin.Compared with bovine milk?-lactoglobulin,the peptides chain of goat milk?-lactoglobulin were more loosened which made it more sensitive to be digested by chymotrypsin.The relative abundance of the DPP IV inhibitory peptide TPEVDDEALEK in the final digestion product of goat milk?-lactoglobulin were increased when treated by heating coupled with high intensity ultrasound.(3)Effects of high-heat(85 ?,30 min)combined with high-intensity ultrasonic treatment(20k Hz,40%amplitude,30 min,12±3 ?)on the proteolysis of goat milk?-lactoglobulin were investigated.Results shown that high-heat combined with high-intensity ultrasonic treatment(regardless of the order)has a greater influence on the structure of goat milk?-lactoglobulin than just ultrasonic treatment or heat treatment.High-heat interact with high-intensity ultrasonic treatment could make?-lactoglobulin to combine with each other to form polymers.Peptides obtained from goat milk?-lactoglobulin after in-vitro digestion were analyzed by peptidomics platform.Compared with high-intensity ultrasonic treatment,high-heat treatment could significantly influence the digestive characteristics of goat milk?-lactoglobulin.Cluster analysis indicated that a highly peptides similarity has been found for the samples treated by high-heat(before or after treatment)combined with high-intensity ultrasound.Relative abundance of bioactive peptides which was obtained from in-vitro digestion was decreased for the goat milk?-lactoglobulin sample when was treated by high-heat temperature.Additionally,for the high-heat treated goat milk?-lactoglobulin samples,low abundance of peptides which has toxicity were found after simulate gastric digestion,indicating that high-heat treatment of goat milk?-lactoglobulin may produce more toxic proteolytic products.(4)Food processing may lead to the interactions between proteins.Goat milk whey protein products are widely used in infant formulas.The infant gastrointestinal digestive system should be considered when analyzing the digestive characteristics of goat milk?-lactoglobulin.Effects of heating combined with high-intensity ultrasonic treatment(20 k Hz,40%amplitude,30 min,60±3?)on the adult or infant digestive ability of goat milk?-lactoglobulin were investigated.Results showed that the effects of heating combined with high-intensity ultrasonic treatment on the digestive characteristics of?-lactoglobulin in goat milk whey protein system were more significant than in pure system.The peptides(belonging to goat milk?-lactoglobulin)obtained from simulate infant stomach digestion was only one ninth of that of from the one of adult,and the proportion of hydrophilic peptides in the digestive products of infant stomach was significantly higher than that of hydrophobic peptides which was opposite of that from adult.Poor digestive ability of simulated infant gastric juice maybe caused by low content of pepsin and high value of p H,which lead to the most of the hydrophobic amino acids located in the?-lactoglobulin molecules were not degraded into short peptides.For the goat milk?-lactoglobulin which was in food matrix system(goat milk whey protein system),after in-vitro digestion,abundance of bioactive peptides which has ACE inhibitory activity(such as LDAQSAPLR and VLDTDYK)were found.However,the relative abundance of these peptides may be reduced after treated by heat or high-intensity ultrasound.It is worth noting that a unique peptide ALPMHIR which has ACE inhibitory activity were only found from goat milk?-lactoglobulin samples which was digested by simulated infant gastric juice.