| Immature vegetable soybean is prone to deterioration during storage, which has limited the scope of its markets and tenure. Therefore, seeking new ways to effectively extend the shelf life is currently pressing of immature vegetable soybean industry bottlenecks. This paper aims to employ metabolite analysis of the different varieties samples under low temperature storage mode, corresponding to physico-chemical, biochemical detection of the metabolism system in seeds, so as to screen metabolites target that is relevant with quality deterioration of immature vegetable soybean. Vegetable soybean treated with exogenous spermine for tracing and filtering was studied to illuminate quality change and control mechanism. The results could provide theory foundation for the further development of the new method for preservation. The results are as follows:1. Morphological, sensory evalation and component parameters of immature vegetable soybean varieties were measured, Xindali No.l was selected.1H NMR metabolomics approach was applied to study the metabolite profiles of eight varieties of immature vegetable soybean, combined with pattern recognition methods such as PCA, the results showed different metabolic characteristics, especially including free sugar content (15.13 to 33.98 mg/g DW), free amino acids (4.58 to 10.18 mg/g DW) and organic acid content (3.75 to 6.75 mg/g DW). Through cluster analysis, the samples could be broadly classified into four categories. Among them, Tongdou No.6 and Ningdou No.4 genotypes were rich in malic acid and asparagine, Huaidou No.8 contained higher sugar contents, while Sucai No. 254 and Xudou 17 showed higher alanine, glutamic acid, glycine and serine contents.2. Cold storage delayed immature vegetable soybean seeds from green to yellow, weakened transpiration, maintained grain hardness. Under storage temperatures, sucrose, glutamic acid, asparagine, alanine and malic acid made important contributions for storage quality distinction of immature vegetable soybean through metabolomics. Low temperature reduced the metabolic rate of immature vegetable soybeans, effectively maintained the quality.Sucrose, glucose and fructose in postharvest immature vegetable soybean seeds showed an overall downward trend, different storage temperatures had different effects, sucrose metabolism was significantly inhibited at 5℃, which was coregulated by AI, NI, SS and SPS, and SPS and NI were key enzymes. In the glycolytic pathway, glucose kinase activity was significantly increased, no other significantly changed. As the storage temperature increases, especially in storage at 20℃, PFK, PFP and PK activity had increased, but with the storage period, the trend did not change significantly. Fermentation metabolism, acetaldehyde levels were significantly increased five-fold in the first four days. Associated with the fermentation pathway, PDC activity increased four-fold, ADH activity increased 2-fold, LDH activity in the first 7 days of storage also had a weak increase.Throughout the storage period, content of citric acid in immature vegetable soybean had a significant increase; malic acid had an inconsistent trend. Related enzymes in organic acid metabolism, PEPC activity increased significantly in the first seven days of storage, the activity increased by 1 times. PCK activity is decreased in the first 4 days of storage. By examining changes in malic enzyme activity, we found NADP-ME and NAD-ME activity did not change, and activity trends under different storage temperatures were consistent.Immature vegetable soybean seed aging was closely related to carbon metabolism, particularly carbohydrates, organic acids and fermentation products, carbon metabolism seriously impacted on the quality of fresh grains and storage time. Low temperature regulated the choice of sucrose utilization, shunt of glycolysis and fermentation pathways.3. Immature vegetable soybean under low temperature during long-term storage conditions was susceptible to chilling injury, during postharvest storage with a 1℃, ethylene production dynamics,1-Aminocyclopropanecarboxylic acid Metabolism (ACC and MACC), oxidation behavior (H2O2 content) and potential antioxidant enzymes (SOD, CAT and POX) was related. Low temperature inhibited the synthesis of MACC, which mainly used for regulating ACC and ethylene production capacity. Although spermine inhibited ethylene production and accumulation of ACC refrigerated, but spermine significantly inhibited POD activity increased during cold storage. These results revealed ACC metabolism should play a key role for cold vegetable soybean during senescence, the MACC and malonyl transferase activity was key regulatory factors for controlling vegetable soybean aging.At low temperature (1℃) storage conditions, spermine can delay the sucrose to fructose and glucose conversion process, while suppressing decreased malic acid and citric acid content; it showed that spermine might limit grain internal signal transduction, carbohydrate metabolism and TCA rate. Effects of spermine on amino acids of immature vegetable soybean were not significant.4. Using proteomics technology based on two-dimensional gel electrophoresis and matrix-assisted laser desorption-ionization time of flight mass spectrometry to study the changes in protein expression of immature vegetable soybean in different storage periods under the condition of low-temperature storage. Results showed that the total of 64 protein points accumulated differentiately in whole storage duration,63 of which matched soybean protein. Based on the existed classification information, the identificated proteins consisted of energy and metabolism, response to environmental stress, and participation in the defense. In addition, we classified protein changes during storage, and found that fructose diphosphate kinase, citrate synthase and malate dehydrogenase acted as an important role in the regulation of storage quality of vegetable soybean, the expressions of stress-related proteins were upregulated, such as LEA protein, formate tetrahydrofolate ligase and isoflavone reductase. During low temperature storage of vegetable soybean, trypsin inhibitor expression was found in more effectively so as to prevent catabolism in soybean seed itself. Exogenous spermine showed a positive protective effect at the protein level, which significantly increased the expression of part of the low temperature down proteins (such as ATP synthase a-subunit, Cu/Zn-SOD and APX). |
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