Phenotypic analysis
During the storage period of pomegranates, the colour of the skin changes slowly, and the skin loses water, resulting in a wrinkled shape. Then, from the surface to the inside, separation occurs between the placenta and the inner skin. Due to water loss, the fruit quickly becomes soft and even rots. Fig. 1 shows the macroperformance of pomegranate(P. granatum L.) during cold storage. Within a storage period of 28 days, soft seed pomegranates stored at both 0 °C and 8 °C showed no decay. After 112 days of storage, the soft seed pomegranate stored at 8 °C did not rot, while the soft seed pomegranate stores at 0 °C severely rotted.
As shown in Fig. 2A, after 28 days at 0 °C, the pomegranates showed cold damage affecting 1/4 of the fruit surface, yielding a cold damage index of 0.1666, while those stored at 8 °C showed no cold damage spots and therefore had a cold damage index of 0. Pomegranates stored at 8 °C had fewer cold damage spots and a lower cold damage index than those stored at 0 °C. After 56 days at 0 °C, two pomegranates had cold damage spots affecting 2/3 of the fruit surface, and one pomegranate had cold damage spanning 1/2 of the fruit surface, resulting in a cold damage index of 0.833. At 8 °C, the pomegranates had no cold damage spots and a cold damage index of 0. After 84 days at 0 °C, all three pomegranates showed cold damage affecting 2/3 of the fruit surface, yielding a cold damage index of 1. At 8 °C, the pomegranates showed no cold damage and had a cold damage index of 0. The cold damage index of the three pomegranates stored at 0 °C was 4 at 112 d, whereas those stored at 8 °C had no cold damage spots, and their cold damage index was 1. With increasing storage time, the cold damage index of the pomegranates stored at 0 °C significantly increased compared to their initial value. Pomegranates stored at 8 °C had fewer cold damage spots and a lower cold damage index than those stored at 0 °C.
As shown in Fig. 2B, at 28 days, the pomegranates stored at 0 °C had browning spanning 1/4 of the fruit surface, resulting in a browning index of 1/6, while those stored at 8 °C, showed no browning, and their colour was normal; the cold damage index was 0. At 56 days, one pomegranate stored at 0 °C showed browning affecting 1/4 of the fruit surface, one pomegranate showed browning affecting 1/2 of the fruit surface, and one pomegranate showed browning affecting 3/4 of the fruit surface, yielding a browning index of 2/3. The pomegranates stored at 8 °C exhibited no browning, and their colour was normal; the browning index was 0. At 84 days, all three pomegranates stored at 0 °C showed browning affecting 1/2 of the fruit surface, with a browning index of 1. At 8 °C, one pomegranate had browning covering approximately 1/4 of the fruit surface, while the other two showed a normal colour, resulting in a browning index of 0.33. At 112 days, all three pomegranates stored at 0 °C had browning spanning 1/2 of the total area or more, with a browning index of 1. Among the pomegranates stored at 8 °C, one had browning affecting approximately 1/4 of the fruit surface, and one had a normal colour; the browning index was 0.22. With increasing storage time, the browning index of the pomegranates stored at 0 °C significantly increased compared to their initial index, while the browning index of the pomegranates stored at 8 °C did not significantly change compared to their initial index.
As shown in Fig. 3, with increasing storage time, the hardness of the fruits stored at 0 °C and 8 °C decreased. The decrease in fruit hardness with storage at 0 °C was more significant after 56 days, possibly due to decreases in phenolic substances in the fruit, an increase in the fruit browning index, and reduced cell membrane permeability, ultimately leading to decreased fruit hardness. The increase in fruit hardness with storage at 0 °C for 84 days may be due to cold damage to the flesh, which leads to the formation of ice crystals in the intercellular spaces of the tissues. Overall, the hardness of the pomegranates stored at 0 °C was lower than that of the pomegranates stored at 8 °C, and the hardness of the pomegranates decreased with decreasing temperature.
According to the changes in chlorophyll content shown in Fig. 4, the chlorophyll content in the fruit peel at both temperatures initially increased but gradually decreased with increasing storage time until the end of the storage period. In the early stage of storage, the chlorophyll content in the fruit peel sharply decreased, followed by a transitional stage where the reductions in chlorophyll content in the fruit peel slowed. The rate of chlorophyll reduction in the fruit peel following storage at 0 °C was greater than that following storage at 8 °C.
Nutritional quality
The TSS contents of the soft seed pomegranates stored at two different temperatures tended to first decrease and then increase with increasing storage time. However, the TSS contents of the soft seed pomegranates stored at 8 °C eventually reached their lowest value. As shown in Fig. 5A, during the experiment, the fruit juice of the pomegranates stored at 8 °C had a bright colour, while the fruit juice of those stored at 0 °C had a dark colour. Ascorbic acid is an important indicator of a fruit’s nutritional quality and plays an important role in the antioxidant, anti-ageing, and anti-browning properties of fruits. The ascorbic acid contents of the soft seed pomegranates stored at different temperatures decreased continuously with increasing storage time. The rate of reduction in the ascorbic acid contents of the soft seed pomegranates stored at 8 °C was lower than that of the soft seed pomegranates stored at 0 °C, indicating that 8 °C is more suitable for storing soft seed pomegranates (Fig. 5B). According to Fig. 5C, the TA contents of the soft seed pomegranates stored at different temperatures showed a similar trend with increasing storage time—first decreasing, then increasing, and finally decreasing. During the same storage period, little difference in TA contents was noted between the soft seed pomegranates stored at the two temperatures, indicating that different temperatures have little effect on the TA contents of soft seed pomegranates and that temperature is therefore not an important factor in TA content changes. However, as the storage time increased, the flavour of the soft seed pomegranates changed significantly, and the red seeds also showed dullness and a lack of lustre during ageing. Changes in soft seed pomegranates were more significant when stored at 0 °C, indicating that storage at 8 °C was more effective.
Changes in the Membrane Lipid Peroxide Product MDA and Cell Membrane Permeability
As shown in Fig. 6A, the accumulation of the membrane lipid peroxidation product MDA reflects the degree of damage to the cell membrane system. MDA accumulation in pomegranate peels stored at 0 °C was greater than that in pomegranate peels stored at 8 °C. At 56 days, the MDA content with storage at 0 °C was 52% greater than the prestorage value. At this time, the membrane permeability of the fruit peel cells increased sharply, indicating that the membrane system may have been severely damaged. MDA accumulation in the early stage of storage at 8 °C was relatively slow. In the later stage, with the ageing and browning of the fruit, the degree of membrane lipid peroxidation gradually increased, and the MDA content also increased. During fruit ageing or cold injury, the fluidity and integrity of cell membranes are disrupted, leading to increased membrane permeability. The degree of membrane permeability is generally expressed by relative conductivity. During storage, the relative conductivity of the pomegranate peels gradually increased at the different storage temperatures. As shown in Figure 5, after 28 days of storage at 0 °C, the relative conductivity of the fruits increased rapidly, and membrane permeability increased. At this time, the low temperature of 0 °C may have caused damage to the cell membrane system of the fruits. At 8 °C, the relative conductivity of the fruits also showed an overall upward trend, with a significant increase in the later stages of storage.
Changes in phenolic substances and polyphenol oxidase (PPO)
As shown in Fig. 7A, the trend of changes in phenolic substances in the flesh of postharvest pomegranates stored under different temperatures was the same, but the amplitude of changes in phenolic substances in the flesh of the soft seed pomegranates stored at 8 °C was greater than that in the soft seed pomegranates stored at 0 °C. Phenolic substances play a major role in the enzymatic browning of fruit peels, and the browning index of fruit peels is negatively correlated with phenolic substance levels. The content of phenolic substances varied at different temperatures. Storage at 8 °C had a relatively small impact on the browning index of the fruit peel and appears to be better for long-term pomegranate preservation. As shown in Fig. 7B, the PPO activity of the pomegranates stored at 0 °C and 8 °C showed a similar trend. PPO activity increased before storage for 84 days, peaked at 84 days, and decreased after 84 days until the end of the storage period. However, the reasons for the changes in enzyme activity at the two temperatures are different. The change at 8 °C was caused by ageing, while the change at 0 °C was caused by low-temperature cold damage. Compared with 0 °C, 8 °C is more conducive to maintaining the freshness of pomegranates and reducing the probability of browning. Phenolic substances are substrates for the enzymatic browning of fruits and vegetables.