The Applicability Study Of Techniques In Evaluating The Quality Changes Of Repeated Freeze-thaw Beef

Frozen meat is one of the important forms of international meat trade,national strategic reserve and enterprise production stock^[1].With the increase of beef consumption in China,the quality of frozen beef has gradually attracted much attention.To date,it is inevitable that the temperature fluctuation always happen with long-term storage,long-distance transportation and multiple turnover of beef before it moves to an end consumer,which leads to the repeated freeze-thaw.Many studies have shown that repeated freeze-thaw accelerates the deterioration of beef quality,such as the deterioration of smell,water holding capacity?WHC?,color,texture and oxidation?lipid and protein?^[2].Therefore,the quality control of frozen meat food has become an important part of the production in the food industry.To date,the traditional quality eavaluation of meat mainly include physical,chemical,sensory and biological methods.Although these methods are important standards and recommended methods,they are difficult to adapt to the development of modern meat industry due to their destructive,tedious and time-consuming characteristics.The development of non-destructive,rapid,accurate and online evaluation techniques in meat quality has become an urgent need for the development of meat industry.Therefore,this thesis studies the applicability of spectroscopy,nuclear magnetic resonance and electronic nose techniques in evaluating the quality changes of repeated freeze-thaw beef.Five parts were included:1.The application of Raman spectroscopy and electron spin resonance?ESR?technology in characterizing the lipid oxidation process of repeated freeze-thaw beef.ESR was used to monitor the changes of free radicals,the results showed that the generation time of free radical signals was advanced with the number of freeze-thaw cycles increased.Besides,the intensity of free radical signals increased with the increase of freeze-thaw cycles.Those results indicated that the degree of lipid oxidation increased with the increase of freeze-thaw cycles.The Raman spectra results showed that the peak intensity of 1655 cm^–1 decreased obviously,which involves the stretching vibration of the C=C double bond.In addition,the Raman intensity ratios of I₁₆₅₅/I₁₄₄₂???C=C?/??CH₂??and I₁₆₅₅/I₁₇₄₅???C=C?/??C=O??related to lipid unsaturation decreased with the increase of freeze-thaw cycles.Finally,the correlation analysis results of conventional parameters of lipid oxidation,the ESR radical intensity data and Raman spectra data showed that the correlation coefficients?r?of Raman data with conventional parameters were between-0.791 to-0.928?P<0.01?,and the r of ESR data and conventional parameters were between 0.826 to 0.914.It suggested that Raman spectroscopy or ESR has a good potential in characterizing lipid oxidation process during repeated freeze-thaw.2.Fourier transform attenuated total reflection infrared?ATR-FTIR?and Raman spectroscopy were used to in-situ monitor the oxidation/denaturation process of repeated freeze-thaw beef protein.The results showed that the content of protein secondary structure components characterized by ATR-FTIR and Raman spectra changed significantly with the decrease of?-helix and the increase of?-sheet after freeze-thaw cycles.In addition,the peak intensity ratios at 850 cm^–1 and 830 cm^–1 of Raman spectra indicated that the tyrosine residues were exposed to polar environment during freeze-thaw.The results of correlation analysis showed that carbonyl content,Ca²⁺-ATPase activity and the enthalpy of myosin denaturation??H₁?had a good correlation with protein secondary structure.In particular,the?-sheet content of ATR-FTIR spectroscopy and the?-sheet/?-helix ratio of Raman spectroscopy were highly negative correlation with?H₁?r of-0.841 and-0.906,respectively?.It indicated that the content changes of protein secondary structure calculated by ATR-FTIR and Raman spectroscopy can be used as a good index for in situ tracking thermal properties of myosin during repeated freeze-thaw.3.The texture prediction of repeated freeze-thaw beef using Raman and Fourier near infrared?FT-NIR?spectroscopy.The results showed that the prediction accuracy of tenderness?R_P²=0.81,RMSEP=2.57 N?,chewiness?R_P²=0.80,RMSEP=942 g.s?,firmness?R_P²=0.81,RMSEP=11.5 g?and hardness?R_P²=0.82,RMSEP=12.8 g?was good by Raman spectroscopy based on the partial least squares?PLS?algorithm model.The good prediction accuracy of chewiness?R_P²=0.73,RMSEP=1110 g.s?,firmness?R_P²=0.81,RMSEP=12.6 g?and hardness?R_P²=0.74,RMSEP=13.2 g?was also obtained using FT-NIR spectroscopy.It was found that the variable contributions of Raman spectroscopy to beef texture prediction model were mainly located in the regions of 1000 cm^–1,1420 cm^–1 and 1550–1680 cm^–1.These regions are related to the changes of protein secondary structure and polarity microenvironment of side chain amino acid residues during repeated freeze-thaw.The main variable contributions of FT-NIR spectroscopy located in the regions of 6600–7700 cm^–1 and 8500–9000 cm^–1,the results showed that water absorption and the changes of protein and lipid components played an important role in beef texture properties during repeated freeze-thaw.4.The application of Raman spectroscopy and nuclear magnetic resonance technology in evaluating color and moisture related traits of repeated freeze-thaw beef.The results showed that Raman spectroscopy has good performance in predicting the color L^*value?R_P²=0.72,RMSEP=12.72?and thawing loss?R²_P=0.70,RMSEP=4.25%?of repeated freeze-thaw beef based on PLS model.The prediction results of color a^*value,color b^*value and moisture content were general.It was found that the change of protein amide I band(1600–1700 cm^–1)has great contribution to the prediction of color a^*value and thawing loss,which indicated that the changes of protein secondary structure caused by repeated freeze-thaw affected the redness and thawing loss of beef meat.In addition,low field nuclear magnetic resonance technology was used to monitor the change of water migration and distribution of repeated freeze-thaw beef.The results showed that the transverse relaxation time T₂₁ decreased significantly after seven freeze-thaw cycles,while regular changes of the transverse relaxation time T_2b and T₂₂ were not found.The correlation analysis results showed that there was a strong correlation between T₂₁ and thawing loss?r=-0.775,P<0.0001?.It suggested that repeated freeze-thaw promoted the transformation of T₂₁ water to free water and eventually lost in the form of juice.Combining with the contribution of protein vibration information in predicting thawing loss,this result may be related to the destruction of myofibrillar protein network and the protein denaturation during repeated freeze-thaw,which leads to the decrease of the binding capacity of protein to water.5.The application of electronic nose technology in identifing the fresh and repeated freeze-thaw beef.The results of principal component analysis and discriminant factor analysis using electronic nose showed that there was a significant difference between fresh beef and repeated freeze-thaw beef,but it was difficult to distinguish samples between different freeze-thaw cycles.The results of conventional gas chromatography-mass spectrometry showed that there were differences in the types and contents of volatile organic compounds between fresh and repeated freeze-thaw beef.Finally,the volatile basic nitrogen?TVB-N?value and aerobic plate count of beef after freeze-thaw cycles changed slightly and were within the safety limits,hexanal could be regarded as an important volatile marker of beef odor deterioration due to lipid oxidation during repeated freeze-thaw based on the high correlation?r=0.821?with thiobarbituric acid value.In conclusion,the results of this study showed that Raman spectroscopy showed good sensitivity in monitoring the vibration information changes of lipid,protein secondary structure and the side chain amino acid of beef during repeated freeze-thaw.The good prediction accuracy for texture properties was also obtained using Raman.However,the prediction effects of color and moisture content of repeated freeze-thaw beef were general using Raman.Nuclear magnetic resonance technology can supplement the deficiency of Raman spectroscopy in water related studies,showing a good sensitivity to the change of water migration and distribution caused by repeated freeze-thaw.In addition,infrared spectroscopy was also sensitive to the changes of protein secondary structure during repeated freeze-thaw.Meanwhile,infrared spectroscopy can provide the contribution of muscle composition such as water and lipid in the prediction of texture properties.In addition,the whole freeze-thaw cycle was performed under the condition of 4?refrigeration for thawing.The TVB-N value and aerobic plate count of beef were within the safety limits.The electronic nose still showed a certain ability for distinguishing fresh and repeated freeze-thaw beef.Finally,the study results of Raman spectroscopy,infrared spectroscopy,nuclear magnetic resonance and electronic nose techequies will provide theoretical and applied basic data in evaluating frozen meat quality in the future.