https://ijpb.ui.ac.ir/ Journal of Plant Biological Sciences en daily 1 Sun, 23 Aug 2026 00:00:00 +0330 Sun, 23 Aug 2026 00:00:00 +0330 https://ijpb.ui.ac.ir/article_30646.html To investigate the effect of foliar application of iron nanoparticles and putrescine (Put) on morphological, physiological, and biochemical traits of the medicinal plant Salvia officinalis L. under salinity stress, a factorial experiment was conducted based on a completely randomized design. The treatments included salinity at three levels (0, 75, and 150 mM sodium chloride), iron nanoparticles at four levels (0, 50, 100, and 200 ppm), and putrescine at three levels (0, 0.5, and 1 mM). The results showed that salinity stress significantly reduced morphological traits (plant height, leaf number, shoot fresh and dry weight) and chlorophyll content, while increasing the content of carotenoids, soluble sugars, proline, flavonoids, and the activity of catalase (CAT) and peroxidase (POD) enzymes. Application of iron nanoparticles (particularly at 200 ppm) and putrescine (1 mM) significantly mitigated the negative effects of salinity and enhanced all studied traits compared to the control treatment. The highest increase in shoot fresh and dry weight, total chlorophyll content, and antioxidant enzyme activity was observed in the combined treatment of iron nanoparticles (200 ppm) plus putrescine (1 mM). Based on the findings, the application of iron nanoparticles and putrescine can ameliorate the adverse effects of salinity stress in sage plants by improving growth, physiological, and biochemical traits. IntroductionSalvia officinalis L. is a valuable medicinal plant from the Lamiaceae family, widely used in pharmaceutical, food, and cosmetic industries due to its phenolic compounds, flavonoids, and essential oils. However, its cultivation faces salinity stress, which limits growth by inducing osmotic stress, causing ion imbalance, reducing photosynthesis, and increasing ROS production. The use of nanoparticles as nutritional elicitors and polyamines such as putrescine as growth regulators constitutes a novel strategy to enhance abiotic stress tolerance. Iron nanoparticles (Fe-NPs) reduce oxidative damage by enhancing the activity of antioxidant enzymes and chlorophyll synthesis. Putrescine contributes to osmotic adjustment, stabilizes membranes, and neutralizes free radicals. Despite previous studies, there is a lack of comprehensive data on their synergistic effects on Salvia officinalis under salinity. Thus, this study evaluated individual and combined effects of Fe-NPs and putrescine on improving salinity tolerance in Salvia officinalis. Materials and MethodsA completely randomized factorial design with three replications was conducted in a greenhouse in 2025. Disinfected Salvia officinalis seeds were germinated, and uniform seedlings were transplanted into pots with perlite to peat moss (1:1 v/v) and irrigated with Hoagland solution (pH 5.5–5.7). Treatments included salinity (NaCl: 0, 75, 150 mM) applied via irrigation; Fe-NPs (0, 50, 100, 200 ppm); and putrescine (0, 0.5, 1 mM), with foliar sprays applied at five‑day intervals (three total applications). Sampling was performed four weeks after the last treatment. Morphological traits—height, leaf number, shoot fresh weight, and dry weight (48 h at 75°C)—were recorded. Photosynthetic pigments were extracted from 0.5 g fresh leaf in 80% acetone, measured at 663, 645, and 470 nm, and calculated via Arnon's formulas. Soluble sugars were determined using the anthrone method at 620 nm with a glucose standard curve. Proline content was determined using the Bates method with ninhydrin reagent, and absorbance was read at 520 nm after toluene extraction. Total flavonoids were measured by aluminum chloride colorimetry at 506 nm using a catechin standard curve. Antioxidant enzymes were extracted from 0.5 g tissue in phosphate buffer containing EDTA. Catalase activity was assayed by monitoring the decrease in H₂O₂ absorbance at 240 nm according to Luck's method. Peroxidase activity was measured by monitoring tetraguaiacol formation at 470 nm according to Tang & Newton's method. DPPH scavenging activity was measured at 517 nm. All assays were performed with three replicates. Data were subjected to analysis of variance (ANOVA) using SAS 9.2, and means were compared by Duncan's multiple range test at the 5% significance level. Normality and homogeneity of variances were confirmed by Kolmogorov–Smirnov and Levene's tests, respectively. Results and DiscussionSalinity significantly reduced all morphological traits and photosynthetic pigment contents (p < 0.01). At 150 mM salinity, height decreased from 22.71 to 16.78 cm (26%), leaf number decreased from 9.67 to 6.25 (35%), fresh weight decreased from 1.01 to 0.70 g (31%), dry weight decreased from 0.21 to 0.13 g (38%), and total chlorophyll decreased from 2.72 to 2.20 mg/g FW (19%). These reductions resulted from impaired water and nutrient uptake, reduced turgor, chloroplast degradation, and decreased anabolic enzyme activity. Conversely, Fe-NPs at 200 ppm increased height by 36.26%, leaf number by 27.42%, fresh weight by 20.73%, and dry weight by 18.75%. Putrescine at 1 mM similarly increased height (20.92 cm), leaf number (8.30), fresh weight (0.94 g), and dry weight (0.19 g). The interaction between Fe-NPs and putrescine was significant for fresh and dry weight (p < 0.01), with the highest values in the NFe3Pu2 treatment (200 ppm Fe-NPs and 1 mM putrescine). For photosynthetic pigments, Fe-NPs at 100 and 200 ppm increased total chlorophyll by 20.45% and 20% over the control, respectively, while putrescine (1 mM) increased it by 12.28%. Salinity increased carotenoids by 20% (to 0.12 mg/g FW at 150 mM), indicating their protective role against oxidative stress. The Fe-NPs × salinity interaction showed that at 150 mM salinity, 200 ppm Fe-NPs raised total chlorophyll by 25% over the control, confirming the role of iron (supplied via Fe-NPs) in preserving chloroplast integrity. Biochemically, salinity elevated soluble sugars (11.67%), proline (12.68%), flavonoids (20.78%), catalase (28.33%), peroxidase (20.53%), and DPPH (10.04%), indicating activation of the plant's defense system. Fe-NPs and putrescine, individually or in combination, further enhanced these parameters. The NFe3Pu2 treatment exhibited the highest soluble sugars (172.91 mg/g FW), proline (38 μg/g FW), flavonoids (30.29 mg quercetin/g FW), catalase (18.09 ΔA/mg protein/min), and peroxidase (4.28 U/mg protein/min), corresponding to increases of 13.4%, 19.7%, 31.4%, 41%, and 24% over the control, respectively. The putrescine × salinity interaction was also significant for sugars, proline, and flavonoids; at 150 mM salinity, 1 mM putrescine increased soluble sugars by 13%. These increases can be attributed to iron‑mediated activation of catalase and peroxidase, which neutralizes ROS, and to putrescine‑induced membrane stabilization, which prevents lipid peroxidation and enhances osmotic regulation through proline and sugar accumulation. The synergistic effect is attributed to complementary mechanisms—enhanced antioxidant defense by iron and membrane protection by putrescine—leading to improved water uptake, enhanced photosynthesis, reduced oxidative damage, and increased secondary metabolite production. ConclusionSalvia officinalis showed relative tolerance to salinity up to 75 mM, whereas growth was severely restricted at 150 mM NaCl. The combined treatment of Fe-NPs (200 ppm) and putrescine (1 mM) was the most effective strategy to mitigate salinity stress, improving morphological traits, photosynthetic pigments, enzymatic and non‑enzymatic antioxidants, and compatible metabolites (proline and soluble sugars), which ultimately increased biomass. Considering the medicinal value of Salvia officinalis and the extensive saline lands in Iran, this combined treatment is recommended for integration into nutritional and stress management programs for cultivation in saline areas. Further field‑scale studies, along with investigations into essential oil quantity and quality, are recommended. https://ijpb.ui.ac.ir/article_30748.html Selenium (Se) can regulate plant growth and antioxidant metabolism, though its effects depend strongly on the form and concentration used. This study compared the effects of conventional selenium (sodium selenate) and selenium nanoparticles (SeNPs) on the physiological responses of saffron (Crocus sativus L.) callus cultured in vitro. Uniform calli were treated with 5 mg L-1 sodium selenate or SeNPs, while untreated calli served as the control. Both Se forms increased callus biomass, compatible solutes, phenolic compounds, total protein, and antioxidant enzyme activities. Sodium selenate reduced hydrogen peroxide and malondialdehyde contents, indicating improved redox regulation and reduced membrane lipid peroxidation. Conversely, SeNPs increased both oxidative-stress markers despite inducing stronger antioxidant responses, suggesting that the activated defense system was insufficient to prevent oxidative damage. Both treatments reduced photosynthetic pigment contents, with a more pronounced decline under SeNP exposure. Overall, sodium selenate produced a more favorable physiological response, whereas SeNPs at 5 mg L-1 induced both metabolic stimulation and oxidative toxicity. These findings emphasize the need to optimize the concentration of each Se form independently before its application in saffron tissue culture.IntroductionSaffron (Crocus sativus L.) is a sterile, vegetatively propagated geophyte of considerable economic and medicinal importance. Propagation through corms is slow and may transmit pathogens; therefore, tissue culture is valuable for rapid clonal propagation, production of pathogen-free material, and controlled production of bioactive metabolites. Its success depends on plant growth regulators and the mineral composition of the culture medium. Selenium (Se), although not essential for higher plants, can exert beneficial effects at low concentrations by promoting growth and stimulating antioxidant systems. At excessive concentrations, however, Se may become pro-oxidant and impair cellular functions. Selenium nanoparticles (SeNPs) have attracted attention because their distinctive physicochemical properties may alter cellular uptake and biological activity. Nevertheless, plant responses depend on the form, concentration, species, and culture conditions of Se, and comparative information for saffron callus remains limited. This study compared the effects of sodium selenate and SeNPs, applied at the same nominal concentration, on growth, metabolites, oxidative-stress markers, and antioxidant enzymes in saffron callus cultured in vitro. Materials and MethodsHealthy saffron corms were washed under running water and surface-disinfected sequentially with 70% ethanol for 5 min and 0.1% mercuric chloride for 10 min, followed by three rinses with sterile distilled water. Corm-derived explants were cultured on Murashige and Skoog (MS) medium containing 3% sucrose and 1 mg L⁻¹ each of 1-naphthaleneacetic acid (NAA) and 6-benzylaminopurine (BAP) to induce callus formation. After five subcultures, morphologically uniform calli were transferred to MS media with the same plant-growth-regulator composition and assigned to three treatments: an untreated control, 5 mg L-1 conventional Se supplied as sodium selenate, and 5 mg L-1 SeNPs. Cultures were maintained in darkness under controlled conditions in a culture room. At the end of the treatment period, calli were harvested, and samples intended for biochemical analyses were stored at −80 °C.Fresh and dry weights were recorded as indices of biomass accumulation. Photosynthetic pigments, including chlorophyll a, chlorophyll b, total chlorophyll, and carotenoids, were determined spectrophotometrically. Soluble sugars and free proline were measured as indicators of carbon status and osmotic adjustment. Total phenolics, flavonoids, and anthocyanins were assayed to evaluate non-enzymatic antioxidant and secondary-metabolic responses. Malondialdehyde (MDA) and hydrogen peroxide (H₂O₂) were quantified as indicators of lipid peroxidation and oxidative status, respectively. Total soluble protein was determined using the Bradford assay. The activities of peroxidase (POD), catalase (CAT), and superoxide dismutase (SOD) were also measured spectrophotometrically. Data were analyzed using SPSS, and treatment means were compared using Duncan’s multiple range test at P ≤ 0.05. Results and DiscussionBoth Se treatments significantly affected most of the evaluated physiological and biochemical traits, but the magnitude and direction of the responses differed between sodium selenate and SeNPs. Application of either Se form increased callus fresh and dry weights relative to the control, indicating that Se at the tested nominal concentration could promote biomass accumulation under the present culture conditions. This response may reflect improved cellular redox regulation and metabolic activity. Nevertheless, the growth-promoting effect should be interpreted alongside the oxidative-stress responses, because increased biomass does not necessarily imply the absence of cellular stress.The highest concentrations of chlorophyll a, chlorophyll b, total chlorophyll, and carotenoids were observed in untreated calli. Both sodium selenate and SeNPs reduced pigment contents, with SeNPs generally exerting the stronger negative effect, particularly on carotenoids. Because the cultures were maintained in darkness and callus is a largely undifferentiated tissue, pigment values should be regarded as biochemical characteristics of the callus rather than direct measures of photosynthetic performance. The decline may be associated with inhibition of pigment biosynthesis or reduced pigment stability under Se-induced changes in cellular redox status. The stronger response to SeNPs is consistent with their potentially greater interaction with cellular surfaces; however, nanoparticle uptake and intracellular localization were not directly measured in this study and require further investigation.Both Se forms enhanced the accumulation of phenolics, flavonoids, and anthocyanins, and the highest phenolic levels were generally recorded in SeNP-treated calli. These compounds contribute to non-enzymatic antioxidant capacity and may accumulate when secondary-metabolic pathways are stimulated by moderate redox perturbation. Selenium may influence phenylpropanoid metabolism, including processes associated with phenylalanine ammonia-lyase activity, whereas the pronounced response to SeNPs may reflect a stronger elicitation signal. Proline and soluble sugars also increased following both treatments, with the greatest accumulation under SeNP exposure. In cultured tissues, these compatible solutes may support osmotic balance, stabilize macromolecules, store carbon, and buffer redox. Their accumulation, however, may represent either improved protective capacity or a response to treatment-induced stress; thus, it should not be interpreted solely as evidence of enhanced tolerance.The clearest distinction between the two Se forms was observed in oxidative-stress markers. Sodium selenate reduced both H₂O₂ and MDA contents relative to the control, suggesting improved control of reactive oxygen species and lower membrane lipid peroxidation. In contrast, SeNPs at the same nominal concentration markedly increased H₂O₂ and MDA levels, indicating increased oxidative stress and membrane damage. These findings show that equal mass concentrations of conventional and nanoparticulate Se are not necessarily physiologically equivalent. Differences in dissolution, surface reactivity, cellular contact, and effective availability could account for the contrasting responses, although these mechanisms were not directly assessed. Consequently, the increase in protective metabolites and antioxidant enzymes under SeNP treatment should be viewed as a compensatory defense response rather than unequivocal evidence of a beneficial effect.Total soluble protein increased in response to both Se forms. Moreover, POD, CAT, and SOD activities were enhanced, with the highest activities generally occurring in SeNP-treated calli. SOD provides an initial line of enzymatic defense by converting superoxide radicals to H₂O₂, whereas CAT and POD participate in H₂O₂ detoxification. The coordinated induction of these enzymes indicates activation of the antioxidant defense network. Under sodium selenate treatment, this activation coincided with reduced H₂O₂ and MDA, suggesting that antioxidant capacity was sufficient to constrain oxidative damage. Under SeNP treatment, however, elevated enzyme activities did not prevent H₂O₂ accumulation or lipid peroxidation, suggesting that reactive oxygen species production exceeded the tissue's detoxification capacity. Thus, the stronger biochemical response elicited by SeNPs included both defense stimulation and evidence of toxicity at 5 mg L-1. ConclusionConventional Se and SeNPs both modified growth and metabolism in saffron callus, but they produced distinct redox outcomes. Sodium selenate promoted biomass, compatible-solute accumulation, phenolic metabolism, protein content, and antioxidant activity while reducing H₂O₂ and MDA. SeNPs also stimulated biomass and defense-related traits, often more strongly, but simultaneously decreased pigment contents and increased oxidative-stress markers. Therefore, the SeNP response at 5 mg L-1 cannot be regarded as uniformly beneficial. The results emphasize that Se form and concentration must be optimized independently before using Se-based treatments in saffron tissue culture. Dose-response experiments, physicochemical characterization of SeNPs in the culture medium, and direct assessment of Se uptake and localization are recommended to distinguish beneficial elicitation from nanoparticle-induced toxicity and to define a safe and effective application range. https://ijpb.ui.ac.ir/article_30735.html This study was conducted to evaluate the effects of Cytokinin (CK) and Salicylic Acid (SA) application on photosynthetic characteristics, antioxidant activity, and yield performance of sunflower under drought stress conditions. The experiment was arranged as a split-plot based on a randomized complete block design with three replications at Shahid Chamran University of Ahvaz during the 2022–2023 growing season. The main factor included two irrigation levels: 80% and 40% of soil field capacity, representing optimal and deficit water conditions, respectively. The sub-factor consisted of five hormonal treatments: control (no application), CK 25 µM + SA 200 µM, CK 25 µM + SA 400 µM, CK 50 µM + SA 200 µM, and CK 50 µM + SA 400 µM. Results showed that drought stress significantly reduced photosynthetic rate, stomatal conductance, chlorophyll pigments, carotenoid content, seed yield, and seed oil percentage. In the control treatment, photosynthetic rate decreased from 4.4 to 1.0 mmol m⁻² s⁻¹ and stomatal conductance declined from 113 to 44 mmol m⁻² s⁻¹ under drought stress. In contrast, antioxidant enzyme activities including catalase, peroxidase, and superoxide dismutase markedly increased under water deficit conditions. The combined application of cytokinin and salicylic acid significantly improved physiological and biochemical traits under both irrigation conditions. The highest photosynthetic rate (8.8 mmol m⁻² s⁻¹), total chlorophyll content (2.34 mg g⁻¹ FW), catalase activity (1000 mg protein min⁻¹), and seed yield (3213 kg ha⁻¹) were obtained from the application of 50 µM cytokinin + 400 µM salicylic acid. Overall, combined application of cytokinin and salicylic acid enhanced drought tolerance in sunflower through improving photosynthetic capacity and antioxidant defense mechanisms. https://ijpb.ui.ac.ir/article_30782.html Introduction “Zinc (Zn) deficiency is a widespread nutritional constraint in agricultural soils worldwide, including Iran.”. It particularly disrupts plant growth, photosynthesis, and yield, especially in calcareous and saline-alkaline soils with high pH. “Sunflower provides a useful system for investigating genotypic variation in responses to Zn deficiency.”due to its high sensitivity to zinc deficiency. This study investigated the physiological and biochemical responses of two groups of sunflower genotypes, Zn-efficient and Zn-inefficient to zinc deficiency conditions. Materials and Methods The experiment was conducted as a factorial design in a completely randomized design (CRD) with three replications in spring and summer 2023 in the greenhouse of the Research Center for Natural Resources and Agriculture of West Azarbaijan Province. The first factor was two zinc levels — no Zn application, and application of Zn at 5 mg kg-1 soil (elemental Zn), supplied as 21.94 mg ZnSO4.7H2O per kg soil based on the nutrient solution composition — and the other factor was Zn-efficient (Diamantis and Suzuka) and Zn-inefficient (Shams and Master) sunflower cultivars. The planting substrate was prepared from the “Khaneh Arkh” River sand, sieved through a 2 mm sieve after preparation, and then washed five times with tap water. Subsequently, it was rinsed with deionized water to remove soluble salts, air-dried, and supplemented with the necessary nutrients for plant growth. It was then air-dried again. In addition to the aforementioned nutrients, half of the treatments received Zn from (ZnSO₄.7H₂O). Irrigation was performed daily to field capacity. Thirty days after planting, root and leaf samples were collected, and biochemical indices including hydrogen peroxide (H2O2), malondialdehyde (MDA), total phenol, flavonoid, total chlorophyll, and the activity of the enzymes catalase (CAT), superoxide dismutase (SOD), guaiacol peroxidase (GPX), and ascorbate peroxidase (APX) were measured. Results and Discussion The results showed that zinc deficiency conditions increased the amount of hydrogen peroxide. This increase was significantly higher in the Zn-inefficient cultivar Shams than in the Zn-efficient cultivar Suzuka. No significant difference was observed between cultivars with different zinc efficiencies in the amount of phenol, either under sufficient zinc conditions or under zinc deficiency conditions. Zinc deficiency significantly increased flavonoid levels; however, cultivars were not differentially affected by zinc conditions. Total chlorophyll levels were severely reduced under zinc deficiency conditions. However, this reduction was much lower in Zn-efficient cultivars than in Zn-inefficient cultivars. Variance analysis of antioxidant enzyme activity results showed that, except for guaiacol peroxidase, there was no interaction effect between cultivars and zinc conditions for any of the enzymes. Comparison of means revealed that zinc deficiency significantly increased the activities of catalase, guaiacol peroxidase, and ascorbate peroxidase, whereas superoxide dismutase activity significantly decreased under zinc-deficient conditions. Among the antioxidant enzymes, guaiacol peroxidase showed the greatest increase in activity (approximately 388%), followed by catalase (150%) and ascorbate peroxidase (31%). In contrast, superoxide dismutase activity decreased by 11.11% under zinc deficiency. Conclusion These findings suggest that Zn-efficient cultivars can respond more effectively to oxidative stress caused by zinc deficiency by utilizing a more effective antioxidant system. Therefore, selecting and cultivating Zn-efficient genotypes in soils with zinc deficiency, especially in arid and semi-arid regions of Iran, or utilizing these genotypes genomes in modern and advanced breeding programs such as genetic engineering, can play a key role. https://ijpb.ui.ac.ir/article_30826.html Abstract To evaluate the effects of chemical and biofertilizers on photosynthetic pigments, total antioxidant capacity, and the activity of some antioxidant enzymes of black cumin (Nigella sativa L.) under different irrigation regimes, an experiment was conducted using a split-factorial scheme in a randomized complete block design at two locations in May 2025. The main plots included three irrigation regimes (100%, 75%, and 50% of crop water requirement) and the subplots included the factorial of seven fertilizer treatments, including: control, basal chemical fertilizer (based on soil analysis), brown seaweeds (Ascophyllum nodosum) + basal fertilizer, brown seaweeds (Sargassum glaucescens) + basal fertilizer, red seaweed (Gracilaria corticata) + basal fertilizer, soil application of mycorrhiza + basal fertilizer, and seed inoculation with Biofarm (including Pseudomons, Azospirillum and Azotobacter bacteria) + basal fertilizer, along with three black cumin ecotypes (Golestan, Semirom, and Lordegan). The results showed that the mycorrhiza + basal fertilizer treatment under full irrigation produced the highest content of chlorophyll a, b, and total chlorophyll (0.68, 0.27, and 0.95 mg/g fresh weight, respectively), and under 50% stress, the highest content of carotenoids (0.17 mg/g fresh weight), total antioxidants (88%), and catalase, ascorbate peroxidase, and guaiacol peroxidase activities (1.35, 2.5, and 0.6 Unit/mg protein, respectively). Semirom ecotype showed the highest enzyme activity at 50% stress. With reduced irrigation, the chlorophyll a/b ratio, carotenoid content, antioxidant capacity, and the activity of the studied enzymes increased, while the content of other pigments decreased. Overall, the Semirom ecotype and the mycorrhiza combined with basal fertilizer exhibited more favorable physiological responses and antioxidant defense mechanisms regarding the studied traits, and demonstrated greater adaptability to drought stress under water-limited conditions. Keywords: Antioxidant enzymes, Black cumin, Drought stress, Mycorrhiza and Seaweed. Introduction Black cumin is an annual, medicinal and oilseed plant belonging to the Ranunculaceae family. Its seeds contain valuable compounds, including oil, protein, and fatty acids. Water limitation is one of the most important factors reducing its growth and physiological stability. Reduced water availability can disrupt the plant’s metabolic balance by limiting water and nutrient uptake and altering the content of photosynthetic pigments. In addition, drought stress increases the production of reactive oxygen species, leading to oxidative stress and damage to cell membranes and the photosynthetic apparatus. To cope with these conditions, plants employ enzymatic and non-enzymatic defense mechanisms, among which increased activities of enzymes such as catalase, ascorbate peroxidase, and guaiacol peroxidase are considered important defensive responses to oxidative stress. In recent years, the use of biological inputs in combination with chemical fertilizers has received increasing attention as an approach to sustainable plant nutrient management. Seaweeds, owing to their growth-promoting compounds, nutrients, and bioactive substances, can enhance physiological processes and the antioxidant defense system of plants. Arbuscular mycorrhizal fungi can also increase the effective root absorption area and improve plant access to water and nutrients, thereby enhancing plant tolerance to water deficit. Moreover, plant growth-promoting bacteria can contribute to improving plant responses to environmental stresses through nitrogen fixation, increasing nutrient availability, and producing growth-promoting compounds. Despite several studies on the individual application of these inputs, information on the combined effects of biological and chemical fertilizers on the physiological responses and antioxidant defense of different black cumin ecotypes under different irrigation regimes remains limited. Therefore, this study was conducted to investigate the effects of integrated water and nutrient management on photosynthetic pigments, total antioxidant capacity, and the activities of some antioxidant enzymes in three black cumin ecotypes under different irrigation regimes in two locations. Materials and Methods The experiment was conducted in May 2025 at two locations, including the research farm of Shahrekord University and a farm in Faradonbeh, Borujen County, using a split-factorial experiment based on a randomized complete block design with three replications. The main factor consisted of three irrigation regimes supplying 100, 75, and 50% of the crop water requirement, while the sub-factor comprised the factorial combination of seven fertilizer treatments and three black cumin ecotypes (Golestan, Semirom, and Lordegan). The fertilizer treatments included: control, basal chemical fertilizer (based on soil analysis), three types of seaweed (Ascophyllum nodosum, Sargassum glaucescens, and Gracilaria corticata), arbuscular mycorrhizal fungi (Glomus mosseae), and Biofarm biofertilizer containing Pseudomonas, Azospirillum, and Azotobacter bacteria. Except for the control, all fertilizer treatments were applied in combination with the basal chemical fertilizer. Chlorophyll a, chlorophyll b, the chlorophyll a/b ratio, total chlorophyll, carotenoid content, total antioxidant capacity, and the activities of catalase, ascorbate peroxidase, and guaiacol peroxidase enzymes were measured. Statistical analysis of the data was performed using SAS software, and means were compared using the LSD test at the 5% probability level. Results and Discussion Reduced irrigation altered the photosynthetic pigment profile and antioxidant defense responses of black cumin. Under full irrigation, mycorrhiza + basal fertilizer produced the highest chlorophyll a, chlorophyll b, and total chlorophyll contents, with values of 0.68, 0.27, and 0.95 mg g-1 FW, respectively. Reducing irrigation to 50% of crop water requirement increased carotenoid content, total antioxidant capacity, and antioxidant enzyme activities. Under this condition, the highest carotenoid content, total antioxidant capacity, and activities of catalase, ascorbate peroxidase, and guaiacol peroxidase were recorded with mycorrhiza + basal fertilizer, reaching0.17 mg g-1 FW, 88%, and 1.35, 2.5, and 0.6 Unit mg-1 protein, respectively. The Semirom ecotype exhibited higher activities of the evaluated enzymes under 50% irrigation, potentially indicating greater antioxidant defense capacity and physiological adaptation to water limitation. Overall, reduced irrigation increased the chlorophyll a/b ratio, carotenoid content, total antioxidant capacity, and antioxidant enzyme activities, while decreasing the contents of the other photosynthetic pigments. Conclusion The results showed that black cumin responses to water limitation were jointly influenced by ecotype genetic background and nutritional management. Drought stress altered photosynthetic pigments and activated the antioxidant defense system, whereas mycorrhizal application combined with basal fertilizer alleviated some unfavorable effects of stress. The Semirom ecotype exhibited superior antioxidant enzyme activity under water-limited conditions, highlighting the importance of genetic variation in drought adaptation. Therefore, integrated nutritional management together with selection of adapted ecotypes could enhance the physiological resilience of black cumin under water-deficit conditions. https://ijpb.ui.ac.ir/article_30858.html The CAB11E rootstock is one of the cherry and sour cherry rootstocks introduced into the country recently. This study aimed to optimize the micropropagation of this rootstock. During the shoot multiplication stage, virus-free CAB 11E plantlets were cultured in a factorial experiment based on a completely randomized design using three basal culture media (MS, WPM, and DKW) and four hormone combinations, including 0.5, 1, and 2 mg L-1 BA, and a combination of 1 mg L-1 BA and 0.1 mg L-1 IBA. The results showed that MS medium supplemented with 1 mg L-1 BA and 0.1 mg L-1 IBA was the most effective, resulting in a multiplication rate of 4.4 and a shoot length of 3.5 cm. To evaluate the effect of the bioreactor system on shoot proliferation, plantlets were cultured in the optimal medium from the previous stage as liquid culture in a bioreactor. The results demonstrated that the multiplication rate in the bioreactor system increased to 14.5. For rooting, plantlets were cultured on MS basal medium containing different iron sources, full and half-strength macronutrients, and varying concentrations of IBA. In the optimal rooting treatment, the use of MS medium supplemented with Fe-EDDHA, half-strength macronutrients, and 0.1 mg L-1 IBA resulted in 97% rooting of the plantlets. These findings suggest that utilizing a bioreactor system, combined with the optimization of culture medium composition and iron source, can offer an efficient approach for micropropagating the CAB11E rootstock.