Studies On The Identification Method And Biological Activity Of Mature Acacia Honey

Honey is the natural sweet substance produced by honeybees,which collect nectar from the flowers,transform and combine it with their own specific substances,store and leave it in the honeycomb to ripen and mature^[1].APIMONDIA’s statement on honey fraud clearly states that the maturation process of honey should be completed by bees alone,and beekeepers should not harvest immature honey,and then use vacuum concentration equipment to manually remove moisture,resulting in the loss of active components in honey.At present,the identification method of mature honey,immature honey and concentrated honey has not been established at home and abroad,and the difference of biological activity between mature honey and immature honey is not clear.In order to establish an effective method to identify mature honey and guide the production of mature honey,new methods based on physicochemical parameters,chromatographic electrochemical fingerprint and amino acid to identify mature honey were established in this paper.At the same time,the antioxidant activity and antibacterial activity of mature honey,immature honey and concentrated honey were researched.This thesis was divided into six chapters as follows:1. We established a method for the identification of natural mature acacia honey with eighteen physicochemical parameters combined with chemometric analysis.The analysis of variance showed significant differences between mature and immature acacia honey in physicochemical parameters.The principal component analysis（PCA）explained 82.64%of the variance among samples,and indicated that total phenolic content,total protein content,and total sugar（glucose,fructose,sucrose）were the major variables.Cluster analysis（CA）divided 50 honey samples into two categories,and the results were consistent with the results of the principal component score map.The model parameters R²X,R²Y and Q²of orthogonal partial least squares discriminant analysis（OPLS-DA）were 0.928,0.978 and 0.971,respectively.Four predicted principal components were extracted,and the recognition accuracy of 50 honey samples was 100%.2. A novel method for the identification of the mature acacia honey was established by HPLC-ECD.From the HPLC-ECD fingerprints,the areas of the common peaks were obtained and PCA and OPLS-DA were performed to classify honey samples.The results showed that PCA extracted a total of six principal components with a cumulative variance contribution rate of 96.60%;OPLS-DA effectively screened out five key common peaks（Peak5,Peak4,Peak 8,Peak13 and Peak7）,which are basically consistent with the PCA results;and the recognition accuracy of the 30 validation set sample regression OPLS-DA model was 100%.3. The waters AQC derivatization method was used to determine the 18 free amino acids in mature acacia honey,immature acacia honey,and concentrated acacia honey.The results showed that there were significant differences in the 18 free amino acids in the three types of honey.The levels of alanine,lysine,proline,glutamic acid and arginine were significantly reduced;glycine and isoleucine were not detected in concentrated acacia honey.In addition,with the 18 free amino acids content in the three kinds of honey as variables,combined with PCA and OPLS-DA,it is found that the three kinds of honey can be accurately classified based on amino acid differences,which provides new ideas for the identification of mature honey.4. The total phenol content and total flavonoid content of mature acacia honey,immature acacia honey and concentrated acacia honey were determined.The antioxidative activity of three kinds of honey in vitro were measured using DPPH radical scavenging activity,ferrous ion-chelating ability（FICA）and ferric reducing antioxidant power（FRAP）.The antibacterial activity of three types of honey against Gram-negative bacteria（Escherichia coli and Pseudomonas aeruginosa）and Gram-positive bacteria（Staphylococcus aureus and Bacillus subtilis）was measured by the Oxford Cup method.The results showed that the antioxidant activity and antibacterial activity of mature acacia honey were higher than that of immature and concentrated acacia honey,and the content of total phenol and total flavonoids decreased significantly after concentration.