Effect on relative water content of seedlings
In this experiment, the relative water content (RWC) in of 8 treatment groups decreased during freeze-thaw cycle. As shown from Fig. 1, RWC in H. vulgare L. seedlings had a maximum decrease under combined stresses of basic salt and drought. The RWC in seedlings of groups FB0, F0D and F00 had no significant differences compared to that in groups 0B0, 00D and 000, respectively. However, RWC in seedlings of group FBD showed significant differences compared with that of group 0BD after T3. Notably, the RWC in H. vulgare L. seedlings of 4 freeze-thaw groups (F**) showed a sequence as F00 > FB0 > F0D > FBD. Consistently, a similar order of RWC in seedlings can be observed among 4 non-freeze-thaw groups (0**) as well, that is, 000 > 0B0 > 00D > 0BD.
Effect on soluble protein content of seedlings
It can be observed that the soluble protein (SP) content in H. vulgare L. seedlings of 4 treatment groups under freeze-thaw stress (F**) was higher than that of 4 treatment groups without freeze-thaw stress (0**), respectively (Fig. 2). The SP content in seedlings of groups 0B* was significantly higher than that of groups 00* (P < 0.05), which indicated that under non-freeze-thaw conditions, the SP content in seedlings increased due to the occurrence of basic salt stress. Nevertheless, the SP content in seedlings of groups under either single freeze-thaw stress or single drought stress had no significant difference compared with that of control group (P > 0.05). In the case of freeze-thaw stress, the SP content in seedlings of groups F** increased during the period of freeze-thaw (T1-T7). Among them, group FAD reached the maximum at T5, which exhibited a further 41.2% increase than the minimum (T3). Somewhat differently, the other 3 groups FB0, F0D and F00 all reached the maximum value at T6, and were 46.4%, 86.6% and 72.7% higher than the minimum value at T5, T1 and T5, respectively.
Effect on MDA content of seedlings
Figure 3 shows that the malondialdehyde (MDA) content in H. vulgare L. seedlings of all experimental groups was higher than that of the control group. The non-freeze-thaw groups (0**) fluctuated little during a 14-hour freeze-thaw period, however the MDA content in H. vulgare L. seedlings of groups 0B* was significantly higher than that of other groups 00* (P < 0.05). Besides, we have noticed that MDA content in H. vulgare L. seedlings in response to group 00D was significantly higher than that in response to blank treatment (P < 0.05). Under the freeze-thaw stress, the MDA content in H. vulgare L. seedlings of single freeze-thaw group (F00) was significantly lower than that of groups F0D and FBD (P < 0.05).
Effect on SOD activity
The SOD activity in H. vulgare L. seedlings of groups *BD had no significant difference compared with control group (000) (Fig. 4). The SOD activity significantly enhanced owing to the occurrence of drought stress (00D) in H. vulgare L. seedlings (P < 0.05), while significantly weakened due to the basic stress (0B0) in seedlings (P < 0.05). Under freeze-thaw stress, the SOD activity in H. vulgare L. seedlings of groups F** decreased at first and then increased. During the freeze-thaw cycle, except for T4, the SOD activity in H. vulgare L. seedlings of group F0D was significantly higher than that of groups FBD, FB0 and F00 (P < 0.05).
Effect on CAT activity
It can be seen from Fig. 5 that, under non-freeze-thaw conditions, the CAT activity in H. vulgare L. seedlings of groups 0B0, 00D and 0BD was significantly lower than that of the control group (000) (P < 0.05). CAT activity in seedlings of either group F00 or group FB0 showed a trend of initially increasing and then decreasing, while that of group F0D showing a general downward trend, and that of group FBD showing an upward trend. Take if further, the CAT activity in seedlings of group FBD was significantly lower than that of group F00 (P < 0.05), and that of the groups FB0 and F0D was significantly lower than that of group F00 only in the latter thawing case (P < 0.05).
Effect on POD activity
In this experiment, during freeze-thaw cycle, the POD activity in H. vulgare L. of groups F00 and F0D showed a trend of initially increasing and then decreasing, while that of groups FB0 and FBD showing an increasing trend (Fig. 6). When the temperature dropped to 10 °C (T1), the POD activity in H. vulgare L. seedlings of groups F00, 0B0 and 00D were significantly lower than that of control group (P < 0.05). Under basic salt stress, the POD activity in seedlings significantly decreased within a freeze-thaw cycle (P < 0.05). Nevertheless, no significant difference was observed in POD activity between groups F0D and 00D. Accordingly, under freeze-thaw stress, there was a significant reducing of POD activity of groups subjected to basic salt treatment, but no effect on that of groups subjected to drought treatment.
Correlation Analysis
It can be observed from Table 1 that the content of SP and MDA in seedlings were significantly positively correlated under freeze-thaw conditions (P < 0.01). There was a significant negative correlation between SP content and antioxidant enzyme activity (P < 0.01). MDA was significantly negatively correlated with antioxidant enzyme activity and RWC, while CAT and POD were positively correlated (P < 0.01).
Table 1
Pearson correlation analysis between relative water content (RWC), soluble protein (SP) content, malondialdehyde (MDA) content, SOD, CAT and SOD activity in H. vulgare L. seedlings of freeze-thaw treatment groups (F**)
|
RWC
|
SP
|
MDA
|
SOD
|
CAT
|
POD
|
RWC
|
1
|
-0.524**
|
-0.427*
|
-0.341
|
0.726**
|
0.640**
|
SP
|
|
1
|
0.750**
|
-0.351
|
-0.799**
|
-0.868**
|
MDA
|
|
|
1
|
-0.407*
|
-0.663**
|
-0.798**
|
SOD
|
|
|
|
1
|
0.052
|
0.213
|
CAT
|
|
|
|
|
1
|
0.833**
|
POD
|
|
|
|
|
|
1
|
* Significant correlation at 0.05 level (both sides). ** Significant correlation at the 0.01 level (both sides). |