تأثیر مدیریت تلفیقی کودهای شیمیایی و زیستی تحت تیمارهای مختلف آبیاری بر رنگدانه‌های فتوسنتزی و فعالیت برخی آنزیم‌های آنتی‌اکسیدانی سه اکوتیپ سیاهدانه (Nigella sativa L.)

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.