Study on browning mechanism of fresh-cut eggplant (Solanum melongena L.) based on metabolomics, enzymatic assays and gene expression

Abstract

Enzymatic browning is one of the crucial problems compromising the flavor and texture of fresh-cut fruit and vegetables. In this study, an untargeted metabolomics approach based on liquid chromatography-mass spectrometry (LC-MS) was used to explore the browning mechanism in fresh-cut eggplant. Metabolomics studies showed that with the increase of fresh-cut time, the contents of 946 metabolites changed dynamically. The metabolites having the same trend share common metabolic pathways. As an important browning substrate, the content of chlorogenic acid increased significantly, suggesting that may be more important to fresh-cut eggplant browning; all 119 common differential metabolites in 5 min/CK and 3 min/CK contrastive groups were mapped onto 31 KEGG pathways including phenylpropanol metabolism, glutathione metabolism pathway, et al. In physiological experiments, results showed that the Phenylpropanoid-Metabolism-Related enzymes (PAL, C4H, 4CL) were changed after fresh-cut treatment, the activities of three enzymes increased first and then decreased, and reached the maximum value at 5 min, indicating the accumulation of phenolic substances. At the same time, ROS were accumulated when plant tissue damaged by cutting, the activities of related antioxidant enzymes (SOD, APX and CAT) changed dynamically after oxidative damage. SOD and APX content increased significantly and reached the maximum value at 10 min after cutting, and then showed a downward trend. However, CAT activity increased sharply and reached the maximum value within 3 min after cutting, then maintained the same activity, and showed a downward trend after 30 min. These data fully demonstrated that the activities of browning related enzymes and gene expression increased with the prolonging of fresh cutting time. We explained the browning mechanism of fresh-cut eggplant by combining metabolomics and physiology, which may lay the foundation for better understanding the mechanism of browning during the fruits and vegetables during processing.

Figure 1

Score scatter plots from PCA, PLS-DA, OPLS-DA and validation plots of OPLS-DA for three comparative groups. (a) PLS-DA for 3 min/CK comparative group. (b) OPLS-DA for 3 min/CK comparative group. (c) Validation plots of OPLS-DA for 3 min/CK comparative group. (d) PLS-DA for 5 min/CK comparative group. (e) OPLS-DA for 5 min/CK comparative group. (f) Validation plots of OPLS-DA for 5 min/CK comparative group. (g) PLS-DA for 3 min/5 min comparative group. (h) OPLS-DA for 3 min/5 min comparative group. (i) Validation plots of OPLS-DA for 3 min/5 min comparative group. (j) Score scatter plots of PCA for 3 min/CK comparative group, 5 min/CK comparative group and 3 min/5 min comparative group.

Figure 2

Venn diagram of the differential metabolites among three comparative groups.

Figure 3

Heatmap showing the differences in expression levels of metabolites.

Figure 4

Contents of phenolic substances related to browning.

Figure 5

KEGG enrichment of the differential metabolic pathways.

Figure 6

H₂O₂ content of Fresh-Cut eggplants.

Figure 7

Rate of O₂·− production of Fresh-Cut eggplants.

Figure 8

MDA content of Fresh-Cut eggplants.

Figure 9

Activities of polyphenol oxidase (PPO) (A), peroxidase (POD) (B), and lipoxygenase (LOX) (C) and gene expression levels of SmePPO (D), SmePOD (E), and SmeLOX (F) of fresh-cut eggplant. Data represent the means of three replicates and their standard errors.

Figure 10

Activities of phenylalanine ammonium lyase (PAL) (A), cinnamate-4-hydroxylase (C4H) (B), and 4-coumarate coenzyme A ligase (4CL) (C) and the gene expression levels of SmePAL (D), SmeC4H (E), and Sme4CL (F) of fresh-cut eggplant. Data represent the means of three replicates and their standard errors.

Figure 11

Activities of superoxide dismutase (SOD) (A), ascorbate peroxidase (APX) (B), and catalase (CAT) (C) and gene expression levels of SmeCu/ZnSOD (D), SmeAPX (E), and SmeCAT (F) of fresh-cut eggplant. Data represent the means of three replicates and their standard errors.