Study On Formation Mechanism And Rapid Determination Technology Of 5-hydroxymethyl-furfural In Apple Juice Concentrate

China is a big producer of apple juice concentrate（AJC）.5-hydroxymethyl-furfural（5-HMF）is an important quality index,which has been widely studied not only as the indicator of thermal treatment,but also as a potential food pollutant.During juice production,5-HMF acts as the indicator of heating degree to identify the technic level.High concentrations of 5-HMF indicate the apple juice has been overheated,reducing the color value of juice and affecting the taste and aroma.Furthermore,high concentrations of 5-HMF can affect human health.Therefore,the formation mechanism and the rapid determination methods of 5-HMF becomes the research emphasis and hotspot.In this study,the formation of 5-HMF and the changes of other physicochemical indexes were monitored throughout the AJC production process.The correlativity between 5-HMF and other physicochemical indexes were observed.The formation characteristic of 5-HMF was monitored in seven simulated solutions,and the formation dynamics of 5-HMF was studied in the sugar-amino acid simulated solutions.In addition,the rapid detection methods of 5-HMF were constructed based on the electronic nose,electronic tongue and Fourier transform near infrared spectroscopy.This study provides support for the explanation of 5-HMF formation.The main research contents and conclusions were as follows:（1）The changing profile of 5-HMF and physicochemical indexes were monitored,and the correlativity between 5-HMF and other physicochemical indexes were observed throughout the AJC processing.In the three production batches of September,October and November in 2016,the samples of different process stages were collected.The soluble solids content showed an increasing trend,and the content in the final product was 70°Brix.The other indexes of samples were determined under the same soluble solids content,which was11.5°Brix.The p H values stabilized at around 3.6-4.0,and were not affected by different treatment.The color value of apple juice significantly increased,while the browning index markedly decreased.The total sugar and acid content showed little change.The content of5-HMF showed the rising trend,and the final contents in the production batches of September,October and November were 0.741 mg/L,0.228 mg/L,0.212 mg/L,respectively.The main sugars in the apple juice were glucose,fructose and sucrose.Under the same soluble solids content,the different sugars showed little change after the processing.The change patterns of different kinds of amino acids were diverse.Their contents obtained increase after preconcentration.They showed little change after enzymatic hydrolysis clarification,ultra-filtration and decoloration.In the final products,the contents of aspartic acid and asparagine were the highest.The contents of the polyphenols changed greatly during the AJC processing.The content of phlorizin was the highest in the crushed juice,and most of the polyphenols experienced increase of different degrees after preconcentration.Their contents decreased markedly after resin adsorption,and in the final products,the most important polyphenol was chlorogenic acid.The formation of 5-HMF showed a positive correlation with soluble solids content,color value,total sugar,fructose,glucose,glutamic acid,asparagine,threonine,alanine,methionine,proline,gallic acid,and rutin content,and showed a negative correlation with browing index,histidine,cinnamic acid,p-coumaric acid and ferulic acid.（2）Seven kinds of simulated systems were established to study the effect of different sugars,amino acids and polyphenols on the formation of 5-HMF under different temperatures.The results showed that the formation of 5-HMF was related to temperature,sugars and amino acids.Fructose was much easier to induce caramelization reaction and maillard reaction to generate 5-HMF than glucose and sucrose.Furthermore,with higher temperature,the 5-HMF content would increase.Under the same conditions,there was only caramelization reaction under the temperature of 60℃or 70℃.However,there were both caramelization reaction and maillard reaction while the temperature was 80℃.In acidic conditions,the difference between the effect of aspartic acid on the formation of 5-HMF and that of glutamic acid,was not significant.In addition,the polyphenols inhibited the formation of 5-HMF.（3）The kinetics of 5-HMF formation in four kinds of monosaccharide-amino acid simulated solutions（glucose-aspartic acid,glucose-glutamic acid,fructose-aspartic acid,fructose-glutamic acid）and two kinds of monosaccharide solution（glucose and fructose）,were explored.The results indicated that the formation of 5-HMF was related to the type of sugar and amino acids,as well as the temperature and time.Fructose was much easier to induce caramelization reaction and maillard reaction to generate 5-HMF than glucose and sucrose.Furthermore,with higher temperature and longer heating time,the 5-HMF content would increase.At 60℃,there was only caramelization reaction in the three simulated solution of glucose during 0-47 h,while both caramelization reaction and maillard reaction existed in the simulated solutions during 47-95 h,and the content of 5-HMF in the reactions of both the two amino acids with glucose showed no significant difference.At 70℃,there was only caramelization reaction during 0-15 h,while both caramelization reaction and maillard reaction existed in the simulated solutions during 15-95 h,and the content of5-HMF in the reactions of both the two amino acids with glucose showed no significant difference.At 80℃,there were both caramelization reaction and maillard reaction existed in the simulated solutions during 7-95 h,and the reaction between aspartic acid and glucose formed more 5-HMF than the reaction between glutamic acid and glucose.In the three simulated solutions of fructose,there were both caramelization reaction and maillard reaction existed during 7-95 h at every tested temperature,and the content of 5-HMF in the reactions of both the two amino acids with fructose showed no significant difference.At temperature of 60℃,70℃or 80℃,the formation of 5-HMF in the six simulated systems increased linearly with the passage of heating time,which conformed to the zero-level reaction kinetics model.（4）The electronic nose and electronic tongue were used to establish qualitative and quantitative models for analysis of 5-HMF in apple juices.The characteristic signals during the stabilization phase of sensors were extracted for statistical analysis.The results indicated that the principle component analysis based on e-nose exhibited better distinguishing ability than Fisher discriminant analysis.The sensors of 2（W5S）、10（W3S）、4（W6S）、6（W1S）and 3（W3C）,which were sensitive to organic aromatic substances,played important role in the recognition of apple juices with different concentration of 5-HMF.In the partial least square（PLS）models,the R²values for validation and prediction were 0.942 and 0.926,respectively,while the RMSEV and RESEP values were 0.3023 and 0.4168,respectively.These results demonstrated that the PLS models based on e-nose signals could predict the concentration of 5-HMF in the apple juices very well.On the other hand,the principle component analysis and Fisher discriminant analysis based on e-tongue both showed great abilities to distinguish apple juices with different concentration of 5-HMF.The sensors of HA,JE and CA contributed most to the models,and played important role in the recognition of apple juices with different content of 5-HMF.In the PLS models,the R²values for validation and prediction were 0.956 and 0.914,respectively,while the RMSEV and RESEP values were 0.3703 and 0.5836,respectively,indicating that the PLS models based on e-tongue signals could predict the concentration of 5-HMF in the apple juices very well.（5）The Fourier transform near infrared spectroscopy（FT-NIR）was used to establish partial least square（PLS）and support vector machine（SVM）models for quantitative analysis of 5-HMF in apple juices.In the PLS model,the raw spectral were pretreated by methods of first derivative（FD）and multiplicative scatter correction（MSC）,and the optimized wavelength ranges were 8800～7195.5 cm^-1 and 6400.9～5596.7 cm^-1.Under these optimal modelling conditions,the best models were obtained.The Rcv²value of validation model was 0.9769,and the RMSECV value was 1.36.The Rp²values for prediction was0.9697,and the RMSEP value was 1.62.In the SVM model,the raw spectral were pretreated by methods of Savitzky-Golay convolution smoothing coupled with direct orthogonal signal correction（DOSC）,and the optimized wavelength ranges were 8800～7195.5 cm^-1.The penalty parameter C was 7.286 and the kernel parameterγwas 0.383.Under these optimal modelling conditions,the best models were obtained.The Rp²values for prediction was0.9577,and the RMSEP value was 1.91.This work demonstrated that the models based on NIR spectroscopy explained great ability to predict the content of 5-HMF in the apple juice,and NIR spectroscopy was a feasible technique for real-time analyzing and predicting5-HMF content during AJC processing.