Physiological and biochemical responses of seed vigor during common bean germination under water deficitABSTRACT:
Water deficit causes oxidative stress in plants through the overproduction of reactive oxygen species, whose mitigation depends on antioxidant activity. Considering that seed vigor influences tolerance to adverse conditions, this study evaluate the physiological response of seeds with contrasting vigor under water deficit to identify metabolic differences that influencing the performance of the seed lot. Seeds of the BAF07 genotype were subjected to five and ten days of artificial aging to obtain lots with contrasting vigor. Subsequently, germination and seedling performance tests were carried out, measuring root, hypocotyl, and total seedling length. The antioxidant components catalase, ascorbate peroxidase, guaiacol peroxidase, and proline, as well as the oxidative stress marker malondialdehyde, were quantified. The contrast between artificial aging and water deficit revealed physiological and metabolic differences among seed lots. Low-vigor seeds exhibited reduced germination performance and seedlings with greater lipid peroxidation and increased antioxidant enzyme activity. These results indicate that initial seed vigor is a key determinant tolerance to water deficit, highlighting its role in preserving metabolic balance essential for seedling establishment.
Index terms:
catalase; malondialdehyde; peroxidase; Phaseolus vulgaris L.; proline
RESUMO:
O déficit hídrico induz estresse nas plantas, promovendo a produção de espécies reativas de oxigênio, cuja mitigação depende da eficiência do sistema antioxidante. Considerando que o vigor das sementes influencia a tolerância a condições adversas, este estudo avaliou a resposta fisiológica de sementes com vigor contrastante sob déficit hídrico para identificar diferenças metabólicas que influenciam o desempenho dos lotes de sementes. Sementes do genótipo BAF07 foram submetidas a cinco e dez dias de envelhecimento artificial para obtenção de lotes com vigor contrastante. Posteriormente, realizaram-se testes de germinação e desempenho de plântulas, com mensuração do comprimento de raiz, hipocótilo e total da plântula. Foram quantificados os componentes do sistema antioxidante catalase, ascorbato peroxidase, guaiacol peroxidase e prolina, além do marcador de estresse oxidativo malondialdeído. O contraste entre envelhecimento artificial e déficit hídrico evidenciou diferenças fisiológicas e metabólicas entre os lotes. Sementes de baixo vigor apresentaram menor desempenho germinativo e plântulas com maior peroxidação lipídica e incremento na atividade de enzimas antioxidantes. Esses resultados indicam que o vigor inicial da semente é um fator determinante fundamental da tolerância ao déficit hídrico, destacando seu papel na preservação do equilíbrio metabólico essencial para o estabelecimento das plântulas.
Termos para indexação:
catalase, malondialdeído, peroxidase, Phaseolus vulgaris L.; prolina
INTRODUCTION
Common bean (Phaseolus vulgaris L.) is a vital legume, providing protein, carbohydrates, and fiber in many Latin American, African, and Asian countries (Los et al., 2018). Cultivated mainly in tropical and subtropical regions, it is frequently exposed to water deficit conditions (Assefa et al., 2019), which negatively affect germination, emergence, growth, photosynthesis, and nutrient uptake, reducing grain yield (Nadeem et al., 2019).
Seed germination and emergence are directly linked to physiological quality (Marcos-Filho, 2015). In general, high-quality seeds enhance germination and seedling performance even under stressful conditions (Padilha et al., 2022). Seed vigor can be defined as the sum of properties determining seed lot performance across environments (ISTA, 2014), and it’s crucial, since the physiological mechanisms under water deficit remain unclear.
Water deficit increases reactive oxygen species (ROS) production, including superoxide, hydroxyl radicals, hydrogen peroxide, and singlet oxygen, triggering oxidative stress (Nadeem et al., 2019). Hydroxyl radicals are particularly damaging to lipids, proteins, and DNA (Mittler, 2017). Plants mitigate ROS through a complex antioxidant system of enzymatic and nonenzymatic components (Morais et al., 2020).
Catalase (CAT), ascorbate peroxidase (APX), and guaiacol peroxidase (GPX) and others are enzymes that convert hydrogen peroxide into water, preventing oxidative damage (Van Doorn and Ketsa, 2014; Zulfiqar and Ashraf, 2021). In Vigna mungo L., stress tolerance correlates with high peroxidase activity (Ali et al., 2016). Similarly, stress-tolerant soybean cultivars show increased CAT, APX, and GPX activities alongside elevated proline levels (Begum et al., 2022). A similar trend was observed by Grahl et al. (2022) in soybean seeds, where water deficit stress during seed development reduced physiological quality and increased oxidative damage, reinforcing the role of antioxidant activity as a protective mechanism.
The antioxidant system integrates enzymatic antioxidants (catalase, superoxide dismutase, ascorbate, and guaiacol peroxidase) and nonenzymatic molecules (proline, flavonoids, carotenoids, and anthocyanins) to mitigate oxidative stress (Mittler, 2017; Soares et al., 2019). Among these, proline contributes to oxidative balance by increasing antioxidant enzyme activities, likely due to its regulatory role in antioxidant gene expression (Carvalho et al., 2013; Rejeb et al., 2014).
Recent studies in commom bean under water deficit indicate that stress intensifies oxidative stress and alters antioxidant enzyme dynamics (Javornik et al., 2025). underscoring the importance of this system in maintaining seedling performance under limited water availability. Similarly, Padilha et al. (2025) demonstrated that seed vigor influences antioxidant enzyme activation in common bean under cold stress, suggesting that physiological quality modulates defense responses across different stress conditions.
Considering this evidence, we hypothesized that higher antioxidant activity or molecule concentrations protect against oxidative damage during germination under water deficit, promoting superior performance in high-vigor seeds. Therefore, this study evaluated the physiological response of seeds with contrasting vigor under water deficit and identified metabolic differences influencing seed lot performance.
MATERIAL AND METHODS
The genotype BAF07 used in this study belongs to the Bean Germplasm Active Bank (BAF) at the Agroveterinary Sciences Center, Universidade do Estado de Santa Catarina (UDESC). This cultivar was selected for its agronomic traits (Michels et al., 2014), genetic value for root distribution (Melo et al, 2016), and water deficit tolerance (Padilha et al., 2022).
The seed lot was produced in the 2020/2021 season in Lages, SC, Brazil, at CAV-UDESC. After harvest, seeds were cleaned, standardized, and classified into three vigor levels via artificial aging tests with saturated saline solution (Jianhua and McDonald, 1997). Seeds were aged at 41 °C for five or ten days and dried to 12-13% humidity. The resulting lots were high-vigor (HV, not aged), medium-vigor (MV, aged five days), and low-vigor (LV, aged ten days).
The germination test was conducted at 22 ± 2 °C with four replications of 50 seeds on Germitest® paper under a 12 h/12 h light/dark cycle. The paper was moistened with distilled water (0.0 MPa) or polyethylene glycol 6000 solution (-0.2 MPa) (Villela et al., 1991). Germination percentage was determined at four and eight days.
Seedling performance was evaluated at four and seven days by measuring root length (RL), hypocotyl length (HL), and total length (TSL) via a digital caliper. After measurements, seedlings were dried at 90 °C for 24 hours to determine dry mass (SLDM). The test used four replications of 10 seeds, considering only normal seedlings (Pereira et al., 2009). Cotyledon reserve reduction (CRR) was calculated as the difference between the initial seed dry mass and the remaining cotyledon mass.
At four and seven days, the proline, ascorbate peroxidase (APX), guaiacol peroxidase (GPX), catalase (CAT) and malondialdehyde (MDA) contents of normal control and water deficit-treated plants were analyzed. Initial evaluations included seedlings and cotyledons from HV, MV, and LV lots. HVs and LVs were later selected for separate analysis of root and hypocotyl samples at seven days.
The enzyme extract was obtained from 200 mg of fresh sample, ground in a mortar with 5 mL of 100 mM potassium phosphate buffer (pH 7.2) containing 1 mM EDTA, 3 mM DTT, and 1% PVP, then centrifuged (Azevedo et al., 1998).
Proline content was determined as described by Bates et al. (1973). Fresh material (0.2 g) was homogenized in 4 mL of 3% sulfosalicylic acid, centrifuged, and reacted with glacial acetic acid and acid ninhydrin in a water bath for 1 h. The chromophore was extracted with toluene and measured at 520 nm. Proline concentration was determined via a standard curve and expressed as µmol.g⁻¹ range of fresh weight leaf material.
APX activity was measured as described by Nakano and Asada (1981) via the addition of 50 mM potassium phosphate buffer (pH 7.2), 5 mM ascorbic acid, and 1 mM hydrogen peroxide, initiated by enzyme extract addition. Readings were taken at 290 nm for 120 s. One unit was defined as the amount required to oxidize 1 µmol ascorbate.min-1.
GPX quantification followed Nakano and Asada (1981) and Simões et al. (2015) with modifications. The reaction medium contained 200 mM sodium phosphate buffer (pH 6.0), 40 mM guaiacol, and 10 mM hydrogen peroxide and was initiated with enzyme extract. Readings were taken at 470 nm for 120 s. One unit was defined as the amount required to form 1 µmol tetraguaiacol per minute.
CAT activity was determined following Aebi (1984), with modifications. The reactions used 100 mM potassium phosphate buffer (pH 7.2), 75 mM hydrogen peroxide, and enzyme extract. Readings were taken at 240 nm for 120 s. One unit was defined as the amount required to degrade 1 µmol of hydrogen peroxide per minute. Enzyme activities were expressed as units per milligram of protein (U.mg⁻¹). Soluble protein content was determined following Bradford (1976).
MDA was quantified following Hodges et al. (1999). Samples (100 mg) were homogenized in 1.5 mL of 80% ethanol. A diluted extract was reacted with 20% trichloroacetic acid (TCA) containing 0.65% thiobarbituric acid (TBA) and 0.01% butylated hydroxytoluene (BHT). Samples were incubated at 95 °C for 25 min, and readings were taken at 440, 532, and 600 nm. MDA was calculated as described by Hodges et al. (1999).
The experiment followed a completely randomized design. The first stage used a 3×2 factorial arrangement with three seed lots (HV, MV, LV) and two germination conditions (0.0 MPa and -0.2 MPa), with four replications. The second stage used a 2×2 factorial arrangement with two seed lots (HV, LV) and two conditions (control, water deficit), with four replications. Data were subjected to normality tests when necessary, followed by ANOVA, and means were compared via Tukey’s test at 5% probability. The software Sisvar was used (Ferreira, 2011).
RESULTS AND DISCUSSION
Seed lots of the genotype BAF07 subjected to artificial aging, showed significant differences (p ≤ 0.05) in physiological quality, demonstranstrating the efficiency of the procedure in producing distinct vigor levels. The non-aged seed lot (HV) exhibited higher physiological quality compared to the lot aged for ten days (LV), while the lot aged for five days (MV) showed an intermediate response, particularly in percentage of germination under water deficit conditions (Table 1).
Such results are consistent with the concept that seed vigor is directly associated with the deterioration process (Marcos-Filho, 2015), and lower vigor seeds have a reduced capacity to withstand abiotic stress conditions. Similar patterns have been reported by Mondo et al. (2016) in common bean, where seed vigor significantly influenced both physiological and agronomic performance.
Seedling performance tests (root length - RL, hypocotyl length - HL, total length - TSL and dry mass - SLDM) confirm the differences in physiological quality among the lots. The high-vigor seed lot showed greater length (root and seedling) and mass accumulation under both evaluation conditions (i.e., absence and presence of water deficit) and at both evaluation times (Table 2). These results demonstrate that vigor differences remain evident even when germination occurs under suboptimal water potential, reinforcing the physiological basis of vigor expression.
Total length and seedling dry mass are variables positively associated with seed vigor, which makes them efficient for seed lot differentiation (Finch-Savage and Bassel, 2016). This relationship had also been observed for common bean seeds under water deficit (Padilha et al., 2022), indicating that the same vigor-dependent mechanisms operate across different genotypes and environmental conditions. Thus, a well-established contrast can be observed between the seed lots, especially between the high-vigor and low-vigor lots.
Total length and seedlings’ dry mass are also positively associated with mobilizing reserves stored in the reserve tissues such as endosperm (Andrade et al., 2019) and cotyledons (Ehrhardt-Brocardo and Coelho, 2016). This was confirmed by the reduction of cotyledon reserves (CRR), where high-vigor seeds showed greater reserve mobilization under both control and water deficit conditions, resulting in seedlings with greater vigor (Table 3).
Free proline content in the absence of water deficit showed no significant differences among seed lots in either evaluation time or seedling structures. However, under water deficit, there was an increase in free proline content in seedlings and cotyledons at both times. Low-vigor seed lots showed greater proline accumulation in all seedling structures and evaluation periods (Figure 1), indicating greater metabolic demand under stress.
Free proline content in seedlings and cotyledons from high-vigor (HV), medium-vigor (MV) and low-vigor (LV) seed lots during germination of common bean evaluated at four (A and B) and seven days (C and D) without stress (0.0 MPa) or deficit (-0.2 MPa). Different lowercase letters indicate significant differences by Tukey’s test (p ≤ 0.05).
The accumulation of free proline is a biochemical response to abiotic stress, where proline synthesized from glutamate (Rejeb et al., 2014), acting in osmotic adjustment, protecting cellular structures during dehydration, and functioning as a signaling molecule and antioxidant (Soares et al., 2019; Zulfiqar and Ashraf, 2021). Although this compound often correlates with stress tolerance (Chen et al., 2016), in this study, the relationship was inverse; the highest proline content occurred in low-vigor seeds, which also showed inferior performance under water deficit (Tables 2 and 3). As the lots belong to the same genotype, greater proline accumulation likely reflects a stress-compensatory mechanism rather than effective tolerance.
Similar evidence was reported by Padilha et al. (2022), where low-vigor seeds accumulated more proline, yet exhibited reduced seedling performance due to the metabolic cost of proline synthesis. Therefore, higher proline content should be interpreted as an indicator of stress intensity rather than of adaptive efficiency.
At four days of germination, malondialdehyde (MDA) content increased in seedlings and cotyledons under water deficit compared to the control (i.e., 0.0 MPa), indicating enhanced lipid peroxidation as a result of oxidative stress (Figures 2A and 2E). MDA accumulation is a known marker of membrane lipid degradation triggered by reactive oxygen species (ROS) (Mittler, 2017), and its intensity depends on both stress severity and genotype sensitivity (Begum et al., 2022). Accordingly, low-vigor seeds exhibited the highest MDA concentrations (Figures 2A and 2E), corroborating their higher susceptibility and explaining the lower physiological performance observed (Table 1).
Antioxidant enzymatic activity and accumulation of MDA and proline in the seedling (A-D) and cotyledons (E-H) of bean seedlings originating from high-vigor (HV), medium-vigor (MV), and low-vigor (LV) seeds after four days without stress (0.0 MPa) or deficit (-0.2 MPa). Different lowercase letters indicate significant differences by Tukey’s test (p ≤ 0.05). Malondialdehyde (A and E), guaiacol peroxidase (B and F), ascorbate peroxidase (C and G) and catalase (D and H).
At four days, antioxidant enzymes catalase (CAT), ascorbate peroxidase (APX) and guaiacol peroxidase (GPX) showed increased activity in seedlings under water deficit for all seed lots (Figures 2B, 2C and 2D). Oxidative stress results from an imbalance between the excessive generation of reactive oxygen species (ROS) and the capacity of the antioxidant system to detoxify them (Mittler, 2017). Under stress conditions, ROS such as superoxide anion (O₂•⁻), hydrogen peroxide (H₂O₂), and hydroxyl radical (•OH) can accumulate and damage lipids, proteins, and nucleic acids, unless efficiently scavenged by enzymatic defenses (Rao et al., 2025).
In this study, low-vigor seeds exhibited higher CAT and APX activities, consistent with the compensatory response observed under oxidative stress (Sarker and Oba, 2018). However, despite the increased activity of these enzymes, the antioxidant system in low-vigor seeds was insufficient to prevent lipid peroxidation or sustain physiological performance, indicating limited protective efficiency, since this seed lot displayed the lowest vigor and germination rates (Table 1 and 2).
In cotyledons, APX, GPX, and CAT were not responsive to water deficit, preventing the detection of enzymatic adjustment in this structure (Figures 2F, 2G and 2H). However, proline concentration was consistently higher in cotyledons of low-vigor seeds (Figure 1B), suggesting that osmotic regulation in this tissue relied more on compatible solute accumulation than on enzymatic detoxification. High-vigor seeds, in contrast, showed higher CAT and GPX activity, possibly contributing to the lower lipid peroxidation observed (Figure 2E) and to maintaining cotyledon integrity under stress.
This condition likely favored more efficient reserve mobilization toward the embryonic axis, ensuring a continuous supply of soluble metabolites for respiration and tissue synthesis (Table 3). Similar responses were reported by Catão et al. (2024), who observed that under water deficit, soybean seeds capable of maintaining efficient reserve translocation to the embryonic axis exhibited better seedling growth and stress tolerance. These findings reinforce that a greater translocation of reserves from the cotyledons contributes to superior embryo development, even under limited water availability.
The lower enzymatic activity in cotyledons compared to seedlings may be related to the participation of non-enzymatic antioxidants. According to Soares et al. (2019), the antioxidant system comprises both enzymatic and non-enzymatic components, including phenolic compounds, carotenoids, and sugars, all of which are present in common bean seeds (Los et al., 2018). These compounds may have acted synergistically to neutralize ROS, particularly in cotyledons where enzymatic activity was limited.
At seven days, MDA content remained higher in low-vigor seeds across both structures evaluated (Figures 3A and 3E). Enzyme activities (APX, GPX, and CAT) were generally higher under water deficit in seedlings and lower in cotyledons, corroborating previous results. The persistence of elevated APX activity in medium- and low-vigor seeds (Figures 3C and 3G) reinforces the hypothesis of higher oxidative stress in deteriorated seeds.
Antioxidant enzymatic activity and accumulation of MDA and proline in the seedling (A-D) and cotyledons (E-H) of bean seedlings originating from high-vigor (HV), medium-vigor (MV), and low-vigor (LV) seeds after seven days without stress (0.0 MPa) or deficit (-0.2 MPa). Different lowercase letters indicate significant differences by Tukey’s test (p ≤ 0.05). Malondialdehyde (A and E), guaiacol peroxidase (B and F), ascorbate peroxidase (C and G) and catalase (D and H).
Seed deterioration is directly associated with increased ROS production, leading to loss of protective capacity, reduced enzyme activity, lipid peroxidation, consumption of reserves and damage to genetic material (Ebone et al., 2019). Consequently, low-vigor seeds exhibited slower growth due to enhanced repair requirements of cellular structures (Marcos-Filho, 2015). Under water deficit, this sensitivity was intensified, leading to a higher requirement to repair and susceptibility to stress (Figures 3A and 3E) required a greater synthesis of proline (Figure 1), APX (Figures 2C and 3C), GPX (Figure 3F), and CAT (Figure 2D), trying to avoid greater damage to metabolism during germination.
Similar patterns have been reported in other species, where antioxidant activity varies by plant structure (Chen et al., 2016; Van Doorn and Ketsa, 2014). Considering this, the high and low vigor seed lots that showed the greatest contrast in physiological quality (Table 1 and Table 2) were evaluated under the same conditions in the different structures (i.e., roots and hypocotyl) at seven days of germination.
MDA content in roots and hypocotyls mirrored that of whole seedlings, with higher accumulation in low-vigor seeds under water deficit (Figures 4A and 4F). The enzymes evaluated in the aerial part showed no difference between the seed vigor levels; only the proline content can be associated with seed vigor, as low-vigor seeds showed higher proline concentration (Figure 4E).
Antioxidant enzymatic activity and accumulation of MDA and proline in the hypocotyl (A-E) and roots (F-J) of bean seedlings originating from high-vigor (HV) and low-vigor (LV) seeds, after seven days without stress (0.0 MPa) or deficit (-0.2 MPa). Malondialdehyde (A and F), guaiacol peroxidase (B and G), ascorbate peroxidase (C and H), catalase (D and I), and proline (E and J) contents.
In roots, the activies of GPX, APX, and CAT increased was observed for the high and low-vigor seed, seeds with low-vigor presented higher activity of these enzymes (Figures 4G, 4H, and 4I), and greater proline accumulation (Figure 4J). According to Ebone et al. (2019), seeds with a higher level of deterioration present a greater accumulation of reactive oxygen species, making it necessary for the antioxidant system to act to control them.
In the present study, the higher oxidative challenge faced by low-vigor seeds during germination under water deficit likely induced both enhanced antioxidant enzyme activity and the synthesis of compatible solutes such as proline (Figure 4J), which act synergistically to mitigate oxidative stress (Figure 4F). Proline accumulation is a well-documented osmoprotective and cytoprotective response to water deficit, contributing to osmotic adjustment, stabilization of proteins and membranes, and indirect ROS scavenging, in addition to acting as a stress signal (Hayat et al., 2012).
In roots, increased proline may assist in maintaining water uptake and root elongation under stress, whereas its accumulation in aerial tissues supposrs turgor maintenance and cell expansion. In common bean, Padilha et al. (2022) also reported increased proline levels under water deficit, which were associated with improved osmotic adjustment and maintenance of seedling growth, reinforcing the protective role of this metabolite under limited water conditions.
The obtained results showed that greater antioxidant activity is not necessarily associated with higher seed vigor. In general, low-vigor seeds exhibited higher antioxidant enzyme activity, indicating the need for intensified synthesis of these enzymes to overcome water deficit. This compensatory response, howerever, was insufficient to prevent the detrimental effects of previous deterioration, resulting in lower germination (Table 1) and seedling performance (Table 2). Thus, the performance of the antioxidant system during germination under water deficit is strongly dependent on the initial seed vigor level. Seedlings originating from high-vigor seeds maintained superior performance due to their greater capacity for reserve hydrolysis and mobilization (Table 2 and Table 3).
CONCLUSIONS
High-vigor seeds exhibited greater reserve mobilization efficiency and lower lipid peroxidation during germination indicating a better aclimatation to water limitation. In contrast, higher proline accumulation and antioxidant activity mainly in roots of low-vigor seedlings reflects a stronger stress response to cytotoxic effect. Therefore, seed vigor defines the metabolic capacity required to sustain physiological performance under water deficit.
ACKNOWLEDGMENTS
The authors would like to thank the financial support from Programa de Bolsas Universitárias de Santa Catarina (UNIEDU) and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) for granting doctoral scholarships, and to FAPESC (FAPESC/2023TR332/UDESC) for the financial support. The corresponding author (Coelho, C.M.M.) thanks the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) for the productivity grant. This article is part of the doctorate obtained by Padilha, M. S.
REFERENCES
-
AEBI, H. Catalase in vitro. Methods in Enzymology, v.105, p.121-126, 1984. https://doi.org/10.1016/S0076-6879(84)05016-3
» https://doi.org/https://doi.org/10.1016/S0076-6879(84)05016-3 -
ALI, Q.; HAIDER, M.Z.; IFTIKHAR, W.; JAMIL, S.; JAVED, M.T.; NOMAN, A.; IQBAL, M.; PERVEEN, R. Drought tolerance potential of Vigna mungo L. lines as deciphered by modulated growth, antioxidant defense, and nutrient acquisition patterns. Brazilian Journal of Botany, v.39, n.3, p.801-812, 2016. https://doi.org/10.1007/s40415-016-0282-y
» https://doi.org/https://doi.org/10.1007/s40415-016-0282-y -
ANDRADE, G.C.; COELHO, C.M.M.; PADILHA, M.S. Seed reserves reduction rate and reserves mobilization to the seedling explain the vigour of maize seeds.Journal of Seed Science, v.41, p.488-497, 2019. http://dx.doi.org/10.1590/2317-1545v41n4227354
» https://doi.org/http://dx.doi.org/10.1590/2317-1545v41n4227354 -
ASSEFA, T.; MAHAMA, A.A.; BROWN, A.V.; CANNON, E.K.S.; RUBYOGO, J.C.; RAO, I.M.; BLAIR, M. W.; CANNON, S.B. A review of breeding objectives, genomic resources, and marker-assisted methods in common bean (Phaseolus vulgaris L.). Molecular Breeding, v.39, n.2, p.1-23, 2019. https://doi.org/10.1007/s11032-018-0920-0
» https://doi.org/https://doi.org/10.1007/s11032-018-0920-0 -
AZEVEDO, R.A.; ALAS, R.M.; SMITH, R.J.; LEA, P.J. Response of antioxidant enzymes to transfer from elevated carbon dioxide to air and ozone fumigation, in the leaves and roots of wild-type and catalase-deficient mutant of barley. Physiologia Plantarum, v.104, n.2, p.280-292, 1998. https://doi.org/10.1034/j.1399-3054.1998.1040217.x
» https://doi.org/https://doi.org/10.1034/j.1399-3054.1998.1040217.x -
BATES, L.S.; WALDREN, R.P.; TEARE, I.D. Rapid determination of free proline for water-stress studies. Plant and Soil, v.39, n.1, p.205-207, 1973. https://doi.org/10.1007/BF00018060
» https://doi.org/https://doi.org/10.1007/BF00018060 -
BEGUM, N.; HASANUZZAMAN, M.; LI, Y.; AKHTAR, K.; ZHANG, C.; ZHAO, T. Seed germination behavior, growth, physiology and antioxidant metabolism of four contrasting cultivars under combined drought and salinity in soybean. Antioxidants, v.11, n.3, p.498, 2022. https://doi.org/10.3390/antiox11030498
» https://doi.org/https://doi.org/10.3390/antiox11030498 -
BRADFORD, M.M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein dye binding. Analytical Biochemistry, v.72, p.248-254, 1976. https://doi.org/10.1006/abio.1976.9999
» https://doi.org/https://doi.org/10.1006/abio.1976.9999 -
CATÃO, H.C.R.M.; PONTES, B.S.; PINHEIRO, D.T.; OLIVEIRA-FILHO, M.A.; SANTOS, A.L.C.; ZOLLA, M. C. Chemical treatment and mobilization of reserves of soybean seeds under water deficit. Journal of Seed Science , v.46, e202446005, 2024. https://doi.org/10.1590/2317-1545v46278828
» https://doi.org/https://doi.org/10.1590/2317-1545v46278828 -
CARVALHO, K.; CAMPOS, M.K.F.; DOMINGUES, D.S.; PEREIRA, L.F.P.; VIEIRA, L.G.E. The accumulation of endogenous proline induces changes in gene expression of several antioxidant enzymes in leaves of transgenic Swingle citrumelo. Molecular Biology Reports, v.40, n.4, p.3269-3279, 2013. https://doi.org/10.1007/s11033-012-2402-5
» https://doi.org/https://doi.org/10.1007/s11033-012-2402-5 -
CHEN, J.; WU, J.; LU, Y.; CAO, Y.; ZENG, H.; ZHANG, Z.; WANG, L.; WANG, S. Molecular cloning and characterization of a gene encoding the proline transporter protein in common bean (Phaseolus vulgaris L.). The Crop Journal, v.4, n.5, p.384-390, 2016. https://doi.org/10.1016/j.cj.2016.05.009
» https://doi.org/https://doi.org/10.1016/j.cj.2016.05.009 -
EBONE, L.A.; CAVERZAN, A.; CHAVARRIA, G. Physiologic alterations in orthodox seeds due to deterioration processes. Plant Physiology and Biochemistry, v.145, p.34-42, 2019. https://doi.org/10.1016/j.plaphy.2019.10.028
» https://doi.org/https://doi.org/10.1016/j.plaphy.2019.10.028 -
EHRHARDT-BROCARDO, N.C.M.; COELHO, C.M.M. Padrão de hidratação e qualidade fisiológica de sementes de feijão. Semina: Ciências Agrárias, v.37, n.4, p.1791-1800, 2016. https://doi.org/10.5433/1679-0359.2016v37n4p1791
» https://doi.org/https://doi.org/10.5433/1679-0359.2016v37n4p1791 -
FERREIRA, D.F. Sisvar: a computer statistical analysis system. Ciência e Agrotecnologia, v.35, n.6, p.1039-1042, 2011. https://doi.org/10.1590/S1413-70542011000600001
» https://doi.org/https://doi.org/10.1590/S1413-70542011000600001 -
FINCH-SAVAGE, W.E.; BASSEL, G. W. Seed vigor and crop establishment: extending performance beyond adaptation. Journal of Experimental Botany, v.67, n.3, p.567-591, 2016. https://doi.org/10.1093/jxb/erv490
» https://doi.org/https://doi.org/10.1093/jxb/erv490 -
GRAHL, T. D.; ROCKENBACH, A. P.; CASASSOLA, A. Outcomes of water deficit stress on the physiological quality of soybean seeds. Journal of Seed Science , v.44, e202244030, 2022. https://doi.org/10.1590/2317-1545v44260716
» https://doi.org/https://doi.org/10.1590/2317-1545v44260716 -
HAYAT, S.; HAYAT, Q.; ALYEMENI, M.N.; WANI, A.S.; PICHTEL, J.; AHMAD, A. Role of proline under changing environments: A review. Plant Signaling & Behavior, v.11, n. 11, p. 1456-1466, 2012. https://doi.org/10.4161/psb.21949
» https://doi.org/https://doi.org/10.4161/psb.21949 -
HODGES, D.M.; DELONG, J.M.; FORNEY, C.F.; PRANGE, R.K. Improving the thiobarbituric acid-reactive-substances assay for estimating lipid peroxidation in plant tissues containing anthocyanin and other interfering compounds. Planta, v.207, n.4, p.604-611, 1999. https://doi.org/10.1007/s004250050524
» https://doi.org/https://doi.org/10.1007/s004250050524 - ISTA. Seed vigor testing. International Rules for Seed Testing Zurich: International Seed Testing Association, 2014.
-
JAVORNIK, T.; CAROVIC-STANKO, K.; GUNJACA, J.; ŠATOVIC, Z.; VIDAK, M.; SAFNER, T.; LAZAREVIC, B. Phenotyping common bean under drought stress: high-throughput approaches for enhanced drought tolerance. Agronomy, v.15, 1344, 2025. https://doi.org/10.3390/agronomy15061344
» https://doi.org/https://doi.org/10.3390/agronomy15061344 - JIANHUA, Z.; MCDONALD, M. B. The saturated salt accelerated aging test for small-seeded crops. Seed Science and Technology, v.25, n.1, p.123-131, 1997.
-
LOS, F.G.B.; ZIELINSKI, A.A.F.; WOJEICCHOWSKI, J.P.; NOGUEIRA, A.; DEMIATE, I.V. Beans (Phaseolus vulgaris L.): whole seeds with complex chemical composition. Current Opinion in Food Science, v.19, p.63-71, 2018. https://doi.org/10.1016/j.cofs.2018.01.010
» https://doi.org/https://doi.org/10.1016/j.cofs.2018.01.010 -
MARCOS-FILHO, J. Seed vigor testing: an overview of the past, present and future perspective. Scientia Agricola, v.72, n.4, p.363-374, 2015. https://doi.org/10.1590/0103-9016-2015-0007
» https://doi.org/https://doi.org/10.1590/0103-9016-2015-0007 -
MELO, R.C.; SCHMIT, R.; CERUTTI, P.H.; GUIDOLIN, A.F.; COIMBRA, J.L.M. Genetic variation in the trait root distribution over segregating generations of common bean. Euphytica, v.207, n.3, p.665-674, 2016. https://doi.org/10.1007/s10681-015-1568-z
» https://doi.org/https://doi.org/10.1007/s10681-015-1568-z -
MICHELS, A.F.; SOUZA, C.A.; COELHO, C.M.M.; ZILIO, M. Qualidade fisiológica de sementes de feijão crioulo produzidas no Oeste e Planalto Catarinense. Revista Ciência Agronômica, v.45, n.3, p.620-632, 2014. https://doi.org/10.1590/S1806-66902014000300025
» https://doi.org/https://doi.org/10.1590/S1806-66902014000300025 -
MITTLER, R. ROS are good. Trends in Plant Science, v.22, n.1, p.11-19, 2017. https://pubmed.ncbi.nlm.nih.gov/27666517/
» https://pubmed.ncbi.nlm.nih.gov/27666517/ -
MONDO, V.H.V.M.; NASCENTE, A.S.; NETO, M.O.C. Common bean seed vigor affecting crop grain yield. Journal of Seed Science , v.38, n.4, p.365-370, 2016. http://dx.doi.org/10.1590/2317-1545v38n4166814
» https://doi.org/http://dx.doi.org/10.1590/2317-1545v38n4166814 -
MORAIS, T.C.; PINHEIRO, D.T.; MARTINEZ, P.A.H.; FINGER, F.L.; DIAS, D.C.F.S. Physiological and antioxidant changes in sunflower seeds under water restriction. Journal of Seed Science , v. 42, e202042008, 2020. https://doi.org/10.1590/2317-1545v42225777
» https://doi.org/https://doi.org/10.1590/2317-1545v42225777 -
NADEEM, M.; LI, J.; YAHYA, M.; SHER, A.; MA, C.; WANG, X.; QIU, L. Research progress and perspective on drought stress in legumes: a review. International Journal of Molecular Sciences, v.20, n.10, p.2541, 2019. https://doi.org/10.3390/ijms20102541
» https://doi.org/https://doi.org/10.3390/ijms20102541 -
NAKANO, Y.; ASADA, K. Hydrogen peroxide is scavenged by ascorbate-specific peroxidase in spinach chloroplasts. Plant and Cell Physiology, v.22, n.5, p.867-880, 1981. https://doi.org/10.1093/oxfordjournals.pcp.a076232
» https://doi.org/https://doi.org/10.1093/oxfordjournals.pcp.a076232 -
PADILHA, M.S.; COELHO, C.M.M.; SOMMER, Â.S. Seed vigor, genotype and proline in common bean seedling formation under drought and saline stress. Revista Ciência Agronômica , v.53, e20228350, 2022. https://doi.org/10.5935/1806-6690.20220056
» https://doi.org/https://doi.org/10.5935/1806-6690.20220056 -
PADILHA, M.S.; COELHO, C.M.M.; SIEGA, Y.P. Antioxidant response of bean seeds with contrasting vigor to cold stress. Pesquisa Agropecuária Tropical, v. 55, e80995, 2025. https://doi.org/10.1590/1983-40632025v5580995
» https://doi.org/https://doi.org/10.1590/1983-40632025v5580995 -
PEREIRA, W.A.; SÁVIO, F.L.; BORÉM, A.; DIAS, D.C.F.S. Influência da disposição, número e tamanho das sementes no teste de comprimento de plântulas de soja. Revista Brasileira de Sementes, v.31, n.1, p.113-121, 2009. https://doi.org/10.1590/S0101-31222009000100013
» https://doi.org/https://doi.org/10.1590/S0101-31222009000100013 -
RAO, M.J.; DUAN, M.; ZHOU, C.; JIAO, J.; CHENG, P.; YANG, L.; WEI, W.; SHEN, Q.; JI, P.; YANG, Y. CONTEH, O.; YAN, D.; YUAN, H.; RAUF, A.; AI, J.; ZHENG, B. Antioxidant Defense System in Plants: Reactive Oxygen Species Production, Signaling, and Scavenging During Abiotic Stress-Induced Oxidative Damage. Horticulturae, v.11, n.5, 477, 2025. https://doi.org/10.3390/horticulturae11050477
» https://doi.org/https://doi.org/10.3390/horticulturae11050477 -
REJEB, K.B.; ABDELLY, C.; SAVOURÉ, A. How reactive oxygen species and proline face stress together. Plant Physiology and Biochemistry , v.80, p.278-284, 2014. https://doi.org/10.1016/j.plaphy.2014.04.007
» https://doi.org/https://doi.org/10.1016/j.plaphy.2014.04.007 -
SARKER, U.; OBA, S. Catalase, superoxide dismutase and ascorbate-glutathione cycle enzymes confer drought tolerance of Amaranthus tricolor Scientific Reports, v.8, n.1, p.1-12, 2018. https://doi.org/10.1038/s41598-018-34944-0
» https://doi.org/https://doi.org/10.1038/s41598-018-34944-0 -
SIMÕES, N.A.; MOREIRA, S.I.; MOSQUIM, P.R.; SOARES, N.F.F.; PUSCHMANN, R. The effects of storage temperature on the quality and phenolic metabolism of whole and minimally processed kale leaves. Acta Scientiarum Agronomy, v.37, n.1, p.101-107, 2015. https://doi.org/10.4025/actasciagron.v37i1.18123
» https://doi.org/https://doi.org/10.4025/actasciagron.v37i1.18123 -
SOARES, C.; CARVALHO, M.E.A.; AZEVEDO, R.A.; FIDALGO, F. Plants facing oxidative challenges - a little help from the antioxidant networks. Environmental and Experimental Botany, v.161, p.4-25, 2019. https://doi.org/10.1016/j.envexpbot.2018.12.009
» https://doi.org/https://doi.org/10.1016/j.envexpbot.2018.12.009 -
VAN DOORN, W.G.; KETSA, S. Cross reactivity between ascorbate peroxidase and phenol (guaiacol) peroxidase. Postharvest Biology and Technology, v.95, p.64-69, 2014. https://doi.org/10.1016/j.postharvbio.2014.04.002
» https://doi.org/https://doi.org/10.1016/j.postharvbio.2014.04.002 - VILLELA, F.A.; DONI-FILHO, L.; SEQUEIRA, E.L. Tabela de potencial osmótico em função da concentração de polietileno glicol 6000 e da temperatura. Pesquisa Agropecuária Brasileira, v.26, n.12, p.1957-1968, 1991.
-
ZULFIQAR, F.; ASHRAF, M. Bioregulators: unlocking their potential role in regulation of the plant oxidative defense system. Plant Molecular Biology, v.105, n.1, p.11-41, 2021. https://doi.org/10.1007/s11103-020-01077-w
» https://doi.org/https://doi.org/10.1007/s11103-020-01077-w
Additional data will be made available by the authors upon reasonable request.
