The results revealed that the germination percentage exhibited significant difference among treatments, period of storage and its interaction. Between the treatments, the highest germination percentage of 74% after 8 months of storage was recorded in the seeds stored in vacuum packaging and kept at -5°C(T
4) which was on par with modified atmospheric storage (0% CO
2, 100% N
2, 0% O
2) at -5°C (T
13) and (40% CO
2, 60% N
2, 0% O
2) at -5°C(T
10) while the lowest value (61%) was observed in seeds stored in ambient condition (T
1). Irrespective of the treatments, the germination percentage declined with increased period of storage (Table 1). The results were in accordance with
Specht and Borner (1998) who observed a maintenance of germination in rye seeds when stored at high N
2 concentration than vacuum and high CO
2 condition. Groundnut seed kernels when exposed to modified atmospheric storage in gaseous combination of (60%) N
2 + (40%) CO
2 + (0%) O
2 in 700 gauge polyethylene bag had a higher germination and vigour upto ten months followed by vacuum storage
(Vasudevan et al., 2014).
The seeds stored under MAS (0% CO
2, 100% N
2, 0% O
2) and stored under -5°C maintain 74% germination after 8 months of storage. Similarly, MAS (0% CO
2, 100% N
2, 0% O
2) and kept under ambient condition, it could also maintain the germination above Indian Minimum Seed Certification Standards (IMSCS). Irrespective of the packaging, the seeds stored under -5°C recorded superiority over the other temperature conditions.
Similarly, the seed vigour exhibited higher vigour (308) in seeds stored in modified atmospheric storage (0% CO
2, 100% N
2, 0% O
2) at -5°C (T
13) while the lowest (231) was estimated in seeds stored in ambient condition (T
1) and it decreased with increasing storage period (Fig 1).
Similar results were observed for dehydrogenase activity also
i.e., seeds stored in modified atmospheric storage (0% CO
2, 100% N
2, 0% O
2) at 25°C (T
11) (2.819) and at (0% CO
2, 100% N
2, 0% O
2) at -5°C (T
13) (2.824) recorded higher activity depicting maintenance of seed viability even after 8 months of storage while the lowest (2.687) was observed in seeds stored in ambient condition (T
1) (Table 2). The viability loss in the seeds stored under low temperature condition (-5°C) in seeds stored under both vacuum packaging and MAS was recorded. The results were in accordance with
Borem (2001) who concluded that the factors such as moisture, emergence, viability and insect after were kept at a check when stored at 5°C in bean seeds.
The electrical conductivity of seed leachate was high (0.336 dS/m) in seeds stored in ambient condition (T
1) indicating the increased seed deterioration while the low electrical conductivity was observed in (0% CO
2, 100% N
2, 0% O
2) at 5°C (T
12) (0.208 dS/m) and at (0% CO
2, 100% N
2, 0% O
2) at -5°C (T13) (0.204 dS/m) (Table 3). The results were in accordance with the results estimated in onion, where the seeds stored under different combination of CO
2, N
2 and O
2 recorded less electrolyte leakage
(Demirkaya et al., 2010).
The viability and vigour loss is associated with declining cell membrane permeability. Even though the loss of viability and vigour is less in values in case of modified atmospheric storage, loss will depict the accumulation of harmful metabolic changes in the seeds (
Delouche and Baskin, 1973). These changes are due to the production of ROS which attack on polyunsaturated fatty acids in the cell membrane resulting in the rupture of seed coat leading to the loss of cell membrane integrity. It can be determined by increasing electrical conductivity
i.e., more number of efflux of solute in the medium along with decreasing dehydrogenase activity in the living cells
(Kumar et al., 2015).
The seed moisture increased throughout the storage period irrespective of the packaging material and storage temperature. But, the percent increase in moisture content was less when the seeds were stored in modified atmospheric storage (0% CO
2, 100% N
2, 0% O
2) at -5°C (T
13) while the high percent of moisture was estimated in seeds stored in ambient condition (T
1) (Fig 2). Similar trend in moisture content was observed in onion seeds stored under modified atmospheric storage condition
(Demirkaya et al., 2010). The increasing trend of seed moisture irrespective of packaging might be due to moisture absorption by seeds by attaining equilibrium with environment during ageing process.
The percentage of oil extracted from the seeds was high in seeds stored in modified atmospheric storage (0% CO
2, 100% N
2, 0% O
2) at -5°C (T
13) with 48% even after 8 months of storage, while the lowest oil content was observed in seeds stored in ambient condition (T
1) which recorded 45.1% oil content. The total oil content decreased with increased storage period (Table 4). Reduction in oil content in seeds are due to the seed deterioration caused by the ROS attack in the lipid membranes.
Quantification of MDA and H
2O
2 content in the embryo exhibited an increasing trend in accumulation of MDA and H
2O
2 content with increasing storage period. But the percent increase was less in T
13 (64% for MDA and 10% for H
2O
2 content) while high percent increase was observed in T
1 (82% for MDA and 26% for H
2O
2 content) (Fig 3 and 4). Accumulation of MDA and H
2O
2 content during ageing are reported in wheat
(Lehner et al., 2008), cotton
(Goel et al., 2003) and soybean
(Sharma et al., 2013). McDonald (1999) suggested that decrease in the initiation of free radicals, extending the longevity of the seeds with a reduction in the levels of lipid peroxidation and production of damaging compounds could be caused by O
2 elimination for storage atmosphere of seed.
Lipid peroxidation and auto-oxidation were highly dependent on seed moisture content. If the seeds have moisture less than 6%, lipid auto-oxidation will occur and produce ROS, whereas oxidative enzymes will be responsible for ROS production (lipid peroxidation) when the moisture is more than 14% (
McDonald, 1999). As the seed moisture reached 7% under MAS and 8% under ambient condition which are above 6%, the reduction in oil content and accumulation of MDA and H
2O
2 content (final products of lipid peroxidation) in the embryo is due to lipid peroxidation leading to seed deterioration with reduced viability and vigour (
El-Maarouf-Bouteau et al., 2011).
The seeds stored under modified atmospheric storage registered a less increase in MDA and H
2O
2 content as it had lesser increase in seed moisture content and had a less metabolic changes compared to seeds stored under ambient condition. And also the storage temperature had a greater influence on storage behaviour of seeds. The seeds packed by modified atmospheric storage with 100% N
2 and eliminating O
2, CO
2 stored under -5°C showed a lesser increase in moisture content with less accumulation of MDA and H
2O
2 content in the embryo even after 8 months of storage.
Similarly, declining trend was observed in antioxidant enzymes like catalase and peroxidase activity. But the percent decrease was less in T
13 (15% for peroxidase and 4% for catalase activity) while the high per cent increase was observed in T
1 (59% for peroxidase and 21% for catalase activity) (Fig 5 and 6).
Begum et al., (2014) stated that reduction in catalase and peroxidase activity was highly evident in case of groundnut where the reduction in peroxidase enzyme activity was from 0.236 to 0.444 OD 10 min
-1 during higher storage periods. The reduced antioxidant defense system was associated with inability of the seeds to repair the damage caused by ageing and seed deterioration (
Mittler, 2002;
Mullan and McDowell, 2011).
The metabolic activity
i.e., lipid peroxidation and antioxidant activity in seeds stored under MAS (100% N
2 and eliminating O
2, CO
2) was less than seeds stored under MAS (60% CO
2, 40% N
2, 0% O
2) and (40% CO
2, 60% N
2, 0% O
2) which might be due to the less influence of carbon-dioxide and oxygen in seed respiration and the inert activity of nitrogen over the seeds which has no influence on seed metabolic activity (
Specht and Borner, 1998;
Manolopoulou and Varzakas, 2016).