ekofertile and microfertile plant biostimulants enhanced wheat productivity and water use efficiency

David Tavi Agbor; Darina Štyriaková; Sena Pacci; Desmond Kwayela Sama; Salih Demirkaya; Abdurrahman Ay; Elis-Bright Iteke Molua; Endalamaw Dessie Alebachew; Orhan Dengiz

doi:10.1038/s41598-025-22728-2

. 2025 Nov 5;15:38812. doi: 10.1038/s41598-025-22728-2

ekofertile and microfertile plant biostimulants enhanced wheat productivity and water use efficiency

David Tavi Agbor ^1,^2,^3,^✉, Darina Štyriaková ², Sena Pacci ¹, Desmond Kwayela Sama ^3,⁴, Salih Demirkaya ¹, Abdurrahman Ay ¹, Elis-Bright Iteke Molua ¹, Endalamaw Dessie Alebachew ¹, Orhan Dengiz ¹

PMCID: PMC12589634 PMID: 41193682

Abstract

The extensive reliance on synthetic agrochemicals in modern agriculture has raised environmental concerns, prompting the need for sustainable alternatives. ekofertile and microfertile biostimulants, consisting of nutrients, organic acids, and beneficial microbes, offer eco-friendly alternatives to synthetic fertilizers. However, their potential remained unexploited. This study aimed to evaluate the effects of these biostimulants on wheat productivity and water use efficiency (WUE) under controlled greenhouse conditions. A split-plot experimental design was employed, incorporating two biostimulants (ekofertile and microfertile) and five treatment levels (control, inorganic fertilization, 2.5%, 5% and 10% biostimulant dosages), triplicated in the greenhouse. Results showed that ekofertile had a significantly higher tillering total chlorophyll content than microfertile. Treatment level 10% had the highest significant tillering and stem elongation total chlorophyll content (3.05 and 3.02 mg/g, respectively). microfertile x 10% stem elongation, total chlorophyll content was the highest significant (3.03 mg/g). ekofertile (9.4 t ha⁻¹) yielded more than microfertile (8.8 t ha⁻¹) in grain yield, while the 10% treatment level yielded the highest (10.4 t ha⁻¹). Water use efficiency was higher under ekofertile (3,279 g mL⁻¹) compared to microfertile (2,928 g mL⁻¹), with the 10% treatment level achieving higher WUE (3553 g mL⁻¹). These findings highlight the potential of ekofertile and microfertile biostimulants in enhancing wheat productivity at appropriate treatment levels, making them viable alternatives to synthetic fertilizers.

Supplementary Information

The online version contains supplementary material available at 10.1038/s41598-025-22728-2.

Keywords: Wheat productivity, Water use efficiency, Sustainable agriculture, Photosynthetic activity, ekofertile, microfertile

Subject terms: Biotechnology, Environmental sciences, Plant sciences

Introduction

The increasing demand for sustainable agricultural practices necessitates the development of innovative biostimulants that optimize crop performance while minimizing environmental impact. Biostimulants have gained significant attention in agricultural research due to their potential to enhance crop productivity and reduce the reliance on conventional fertilizers^1,2. Some biostimulants serve as stress modifiers, soil fertility enhancers through nutrient solubilization and fixation, adjuvants, and surfactants. These substances, typically derived from organic or mineral sources, stimulate plant growth by improving nutrient uptake and utilization, and enhancing plant stress tolerance³. Numerous studies have documented biostimulants’ importance in improving soil fertility and agricultural output⁴. Some biostimulants include moringa leaf extract, bacteria, and fungi. Among the various sources of biostimulants, ekofertile and microfertile plant biostimulants represent abundant and potentially valuable resources that have received limited attention in scientific investigations. Although ekofertile and microfertile have shown valuable results in increasing crop productivity at the farm level and in other trials, they have not been extensively investigated. Pacci et al.⁵, who showed significant soil quality improvement by these biostimulants. Similarly, these biostimulants are produced through the innovative innoBiotech ecological process which mimics the natural microbial weathering minerals in rhizosphere(https://ekolive.eu/).

The utilization of ekofertile and microfertile plant biostimulants for agricultural production offers a promising avenue to promote sustainable agriculture practices as they contain a wide range of minerals, including silica, which has been reported to have beneficial effects^6,7. Traditional biostimulants are classified into four categories: seaweed extracts, microbial extracts, humic substances, and amino acids and peptides^8,9. ekofertile and microfertile biostimulants due to their uniqueness are classified as waste-derived biostimulants¹⁰. They are a combination of the four categories, as they contain organic acid, nutrients, and plant growth-promoting microorganisms that directly promote plant growth and indirectly through metabolites¹¹. ekofertile and microfertile biostimulants are produced from an eco-bioleaching technology by ekolive company that leads to the efficient utilization of sand and waste from mining sites to enhance agricultural productivity while limiting environmental pollution from mining sites. This patented technology replicates the natural process of soil formation through microbial weathering of minerals. The process solubilises silicate, carbonate, oxides, and hydroxide minerals via microbial metabolism, releasing accessible macro and micronutrients along with bioactive metabolites, including phytohormones and organic acids. This technology has demonstrated the capability of mass production of these biostimulants as well as global applicability and accessibility, including trials in Slovakia, Turkey, Benin, and many other countries (https://ekolive.eu/), unlike other biostimulants like seaweed and humic acid, which are hardly massively produced and accessible. They act via stimulation of plant physiology through beneficial microorganisms, organic acids, and dissolved macro/micronutrients when either soil or foliar applied. Specifically promotion of root growth and nutrient uptake, regulating plant hormones and gene expression for development, increasing resistance to both biotic and abiotic stresses, and modulating plant metabolism for better photosynthesis and nutrient assimilation (https://ekolive.eu/).

However, the physiological and agronomic effects of ekofertile and microfertile plant biostimulants remain largely unexplored. In particular, their role in enhancing chlorophyll biosynthesis, water use efficiency (WUE), and wheat productivity. This is because research has shown that micronutrients such as Fe and Si are essential for chlorophyll synthesis and plant resistance to stress^12,13. Si promotes the maintenance and synthesis of chlorophyll under water deficit conditions. At the same time, Fe is a crucial component for chlorophyll formation. Also, it enhances water use efficiency by influencing aquaporin expression, a process also supported by their beneficial bacteria and growth-promoting metabolites. Similar mechanistic influence is observed with Mn, Mg, beneficial bacteria, and metabolites, all of which are constituents of ekofertile and microfertile plant biostimulants¹⁴. As well, chlorophyll content, water use efficiency (WUE), and wheat yield have interconnected roles as reliable indicators of plant health, resilience, and ultimate agricultural productivity¹⁵. These directly measure the effectiveness of biostimulants in promoting sustainable agriculture by optimizing plant processes and minimizing environmental stress. Thus, this study aimed to evaluate the effects of ekofertile and microfertile biostimulants on wheat growth by assessing their influence on chlorophyll content, water use efficiency, and yield performance, offering sustainable alternatives to synthetic fertilizers. We hypothesize that ekofertile and microfertile plant biostimulants will significantly enhance wheat chlorophyll content, improve water use efficiency, and boost grain yield. This study contributes to developing eco-friendly alternatives to conventional fertilizers, quantifying their impact on key agronomic traits and establishing optimal application levels for enhanced wheat performance.

Result

Sole and interactive effect of biostimulant type and treatment level on wheat chlorophyll content

Table 1 showed the effect of biostimulant type, treatment level, and their interaction on wheat chlorophyll a, b, carotenoid, and total chlorophyll content at tillering and stem elongation stages. Biostimulant types are ekofertile and microfertile biostimulants, while treatment levels include control, inorganic fertilisation, and biostimulant concentrations of 2.5%, 5% and 10%. ekofertile exhibited a significantly increased level of chlorophyll a (2.12 mg/g, df = 1, F = 22.0, P = 0.043) and carotenoids (0.43 mg/g, df = 1, F = 130, P = 0.008) at the tillering stage, which differed from microfertile (Table 1). ekofertile biostimulant significantly (df = 1, F = 277, P = 0.004) increased total chlorophyll content by 2% (2.97 mg/g) more than microfertile biostimulant (2.81 mg/g) (Table 1). At stem elongation stage, the level of chlorophyll a (2.05 mg/g, df = 1, F = 26.4, P = 0.036), b (0.88 mg/g, df = 1, F = 217, P = 0.005), and carotenoids (0.52 mg/g, df = 1, F = 57.0, P = 0.017) exhibited significant differences with higher contents in ekofertile as compared to microfertile. However, no significant difference was observed in the total chlorophyll content between the two biostimulants; microfertile had the highest total chlorophyll (2.73 mg/g) compared with ekofertile (2.71 mg/g) (Table 1).

Table 1.

Sole and interactive effect of biostimulants on chlorophyll content.

Sampling period	Tillering stage				Stem elongation stage
Properties	Chl.-a	Chl.-b	Total Chl.	Carotenoid	Chl.-a	Chl.-b	Total Chl.	Carotenoid
Units	mg/g fresh matter
Biostimulant type (B)
ekofertile	2.12^a	0.86^a	2.97^a	0.43^a	2.05^a	0.88^a	2.71^a	0.52^a
microfertile	2.01^b	0.81^a	2.81^b	0.41^b	1.90^b	0.65^b	2.73^a	0.41^b
Treatment level (T)
Control	1.62^d	0.62^d	2.1^d	0.31^e	1.52^d	0.51^c	1.96^d	0.34^c
Inorganic fertilisation	2.13^a	0.74^c	2.87^bc	0.41^c	1.78^c	0.69^b	2.54^c	0.55^a
2.5%	1.96^c	0.76^bc	2.72^c	0.36^d	1.87^c	0.66^b	2.46^c	0.44^b
5.0%	2.05^b	0.85^ab	2.89^b	0.44^b	1.98^b	0.70^b	2.68^b	0.45^b
10.0%	2.17^a	0.90^a	3.05^a	0.46^a	2.08^a	0.93^a	3.02^a	0.49^b
B x T interactions
ekofertile x Control	1.62^a	0.62^a	2.10^a	0.31^a	1.52^a	0.51^e	1.96^e	0.34^d
ekofertile x Inorganic	2.13^a	0.74^a	2.87^a	0.41^a	1.78^a	0.69^cd	2.54^c	0.55^a
ekofertile x 2.5%	2.06^a	0.76^a	2.81^a	0.37^a	1.93^a	0.63^de	2.55^c	0.49^ab
ekofertile x 5.0%	2.07^a	0.90^a	2.97^a	0.45^a	2.04^a	0.52^e	2.56^c	0.50^ab
ekofertile x 10.0%	2.23^a	0.93^a	3.12^a	0.47^a	2.20^a	0.79^bc	3.02^a	0.56^a
microfertile x Control	1.62^a	0.62^a	2.10^a	0.31^a	1.52^a	0.51^e	1.96^e	0.34^d
microfertile x Inorganic	2.13^a	0.74^a	2.87^a	0.41^a	1.78^a	0.69^cd	2.54^c	0.55^a
microfertile x 2.5%	1.87^a	0.76^a	2.63^a	0.35^a	1.82^a	0.68^cd	2.37^d	0.39^cd
microfertile x 5.0%	2.02^a	0.80^a	2.81^a	0.43^a	1.92^a	0.89^b	2.8^b	0.39^cd
microfertile x 10.0%	2.12^a	0.87^a	2.98^a	0.45^a	1.97^a	1.06^a	3.03^a	0.43^bc
Significance level (ns = not significant, Chl = chlorophyll, columns with different letters are significantly different)
B	0.043	ns	0.01	0.008	0.036	0.005	ns	0.017
D	< 0.001	< 0.001	< 0.001	< 0.001	< 0.001	< 0.001	< 0.001	< 0.001
B x D	ns	ns	ns	ns	ns	0.001	0.014	0.044

Open in a new tab

The 10% treatment level resulted in the highest chlorophyll a (2.17 mg/g, df = 4, F = 84.5, P < 0.001), b (0.90 mg/g, df = 4, F = 9.45, P < 0.001), and carotenoid (0.46 mg/g, df = 4, F = 324, P < 0.001) in contrast to other treatment levels at the tillering stage. Also, 10% treatment level significantly (df = 4, F = 50.9, P < 0.001) increased wheat’s total chlorophyll content at the tillering stage the highest (Table 2) with 22% (3.05 mg/g) increase compared to 21% (2.89 mg/g) for 5% treatment level and 20% (2.72 mg/g) for 2.5% treatment level (Table 2). The trend continued for chlorophyll a (2.08 mg/g, df = 4, F = 39.2, P < 0.001) and b (0.93 mg/g, df = 4, F = 24.2, P < 0.001) contents at stem elongation except for carotenoid content, which was highest at inorganic fertilization (0.55 mg/g, df = 4, F = 18.9, P < 0.001). The treatment level of 10% biostimulant significantly affected wheat total chlorophyll content (3.02 mg/g). It varied significantly (df = 4, F = 111, P < 0.001) from the control (1.96 mg/g) at the stem elongation stage (Table 1).

Table 2.

Sole and interactive effect of biostimulant type and treatment level on wheat yield and yield components.

Properties	1000 grains weight (g)	Grain yield (t ha⁻¹)	Straw yield (t ha⁻¹)	Biological yield (t ha⁻¹)	Harvest index	Water use (ml)	Water use efficiency (g/ml)
Biostimulant type (B)
ekofertile	70.4^a	9.4^a	13.3^a	22.70^a	41.5^a	7332^a	3279^a
microfertile	69.8^a	8.8^b	13.2^a	21.94^a	39.9^b	7402^a	2928^b
Application dose (D)
Control	63.7^d	7.3^d	12.3^a	19.66^c	37.2^a	8381^a	2510^c
Inorganic fertilisation	71.2^ab	9.1^b	13.3^a	22.38^b	40.7^a	7421^a	2928^b
2.5%	66.5^cd	8.0^c	12.7^a	20.66^c	38.8^a	7364^a	2809^bc
5.0%	69.7^bc	9.0^b	12.8^a	21.85^b	41.5^a	7362^a	3008^b
10.0%	74.1^a	10.4^a	14.2^a	24.46^a	41.9^a	7376^a	3553^a
B x D interactions
ekofertile x Control	63.7^a	7.3^a	12.3^a	19.66^a	37.2^a	8381^a	2510^a
ekofertile x Inorganic	71.2^a	9.1^a	13.3^a	22.38^a	40.7^a	7421^a	2928^a
ekofertile x 2.5%	66.9^a	8.3^a	12.7^a	20.98^a	39.5^a	7345^a	2863^a
ekofertile x 5.0%	70.0^a	9.2^a	12.9^a	22.13^a	41.6^a	7330^a	3100^a
ekofertile x 10.0%	74.4^a	10.8^a	14.3^a	25.01^a	43.4^a	7320^a	3873^a
microfertile x Control	63.7^a	7.3^a	12.3^a	19.66^a	37.2^a	8381^a	2510^a
microfertile x Inorganic	71.2^a	9.1^a	13.3^a	22.38^a	40.7^a	7421^a	2928^a
microfertile x 2.5%	66.2^a	7.7^a	12.6^a	20.33^a	38.1^a	7407^a	2754^a
microfertile x 5.0%	69.3^a	8.9^a	12.7^a	21.56^a	41.3^a	7395^a	2916^a
microfertile x 10.0%	73.7^a	9.6^a	14.1^a	23.92^a	40.3^a	7408^a	3234^a
Significance level (ns = not significant; columns with different letters are significantly different)
B	ns	0.005	ns	ns	0.02686	ns	0.004
D	< 0.001	< 0.001	ns	< 0.001	ns	ns	< 0.001
B x D	ns	ns	ns	ns	ns	ns	ns

Open in a new tab

Table 1 showed no significant effect of biostimulant type and treatment level interaction on chlorophyll content at the tillering stage. Chlorophyll a was insignificant at the stem elongation stage (Table 1). Significant differences were observed for chlorophyll b (1.06 mg/g, df = 4, F = 7.87, P = 0.001), with the highest at microfertile x 10% and carotenoid (0.56 mg/g, df = 4, F = 3.13, P = 0.044) with the highest at ekofertile x 10% across the interactions at the stem elongation stage (Table 1). Biostimulant type and treatment level interactions yielded significant differences (df = 4, F = 4.40, P = 0.014) in wheat total chlorophyll content at the stem elongation stage (Table 1). As the biostimulant’s treatment level increased, so did the total chlorophyll content. The highest values were observed at the 10% treatment level for both ekofertile^® (3.03 mg/g) and microfertile (3.02 mg/g), in which ekofertile × 10% was highest.

Sole and interactive effect of biostimulant type and treatment level on wheat yield and yield components

Table 2 showed the effect of biostimulant type, treatment level, and their interaction on wheat yield, yield component, water use, and efficiency. Biostimulant types are ekofertile and microfertile biostimulants, while treatment levels include control, inorganic fertilisation, and biostimulant concentrations of 2.5%, 5% and 10%. There were insignificant differences between ekofertile and microfertile for 1000-grain weight, straw yield, and biological yield. Grain yield, harvest index and water use efficiency (Table 2) showed significant differences across ekofertile and microfertile with a significant higher influence from ekofertile having grain yield (9.4 t ha⁻¹, df = 1, F = 210, P = 0.005), harvest index (41.5, df = 1, F = 35.7, P = 0.027), and water use efficiency (3279 g/ml df = 1, F = 276, P = 0.004), compared to microfertile.

Treatment level yielded significant differences for water use efficiency (3553 g/ml, df = 4, F = 8.56, P < 0.001), 1000 grains weight (df = 4, F = 12.4, P < 0.001), grain yield (df = 4, F = 39.5, P < 0.001) and biological yield (24.46 t ha⁻¹, df = 4, F = 22.5, P < 0.001 with the highest at 10% treatment level (Table 2). Meanwhile, treatment level did not significantly modulate straw yield, water use, and harvest index (Table 2).

Table 2 showed that there was no significant effect of biostimulant type and treatment level interaction on water use efficiency, harvest index, straw yield, 1000 grains weight, and grain yield, with ekofertile x 10% exerting a greater effect compared to the other biostimulant and treatment level interactions.

Correlation

Table 3 shows a mild, positive and significant (P = 0.0313) correlation between 1000-grain weight and grain yield (0.3938). A similar trend existed between 1000-grain weight and harvest index (0.4852), and tillering total chlorophyll (0.3748). Significant (P = 0.0427) positive mild correlation existed between grain yield and straw yield (0.3725). While a significant positive strong correlation existed between grain yield with harvest index (0.7797), water use efficiency (0.6548), tillering (0.8015), and stem elongation (0.8079) total chlorophyll. Straw yield and water use efficiency had a significant (P = 0.0002) moderate correlation (0.6303), while a strong positive and significant (P = 0.0001) correlation was found with tilering and stem elongation total chlorophyll (0.8633). Whereas harvest index showed a significant positive moderate correlation with tilering (0.5439) and stem elongation (0.4809) total chlorophyll, a similar trend was observed with water use efficiency and tilering and stem elongation total chlorophyll.

Table 3.

Correlation matrix.

	1000 grains of weight	grain yield	Straw yield	Harvest index	water use efficiency	Tillering total chlorophyll	Stem elongation total chlorophyll
1000 grains weight	1.0000
grain yield	0.3938 (0.0313)	1.0000
Straw yield	−0.0621 (0.7443)	0.3725 (0.0427)	1.0000
Harvest index	0.4852 (0.0066)	0.7797 (0.0001)	−0.2855 (0.1261)	1.0000
water use efficiency	0.0247 (0.8970)	0.6548 (0.0001)	0.6303 (0.0002)	0.2477 (0.1868)	1.0000
Tillering total chlorophyll	0.3748 (0.0413)	0.8015 (0.0001)	0.4530 (0.0119)	0.5439 (0.0019)	0.5910 (0.0006)	1.0000
Stem elongation total chlorophyll	0.3042 (0.1022)	0.8079 (0.0001)	0.5545 (0.0015)	0.4809 (0.0071)	0.6336 (0.0002)	0.8633 (0.0001)	1.0000

Open in a new tab

Discussion

This study revealed that the biostimulant type significantly enhanced chlorophyll activity. A study reported increased pepper chlorophyll content treated with biologically digested humic acid biostimulant, which supports this result¹⁶. Similarly, treatment of gravevine with several exogenous biostimulants elevated chlorophyll content significantly, which aligns with our study¹⁷. This result revealed that at the tillering stage, ekofertile plant biostimulant had a significantly higher effect on chlorophyll a, carotenoid, and total chlorophyll content than microfertile. However, neither biostimulant showed any difference in chlorophyll b content. Similar results were observed for chlorophyll a and carotenoids at the stem elongation stage. ekofertile modulated chlorophyll b content higher than microfertile, with no different effect from the biostimulant on total chlorophyll content. This variation in performance on chlorophyll content by different biostimulants observed in our study has been shown by Irani et al.¹⁷. They used different biostimulants like fulvic, amino, and humic acids, as well as seaweed extract from grapevine, resulting in differences in chlorophyll content. This is supported by the variation in biostimulant organic acid, nutrient, and microbial content⁵. Interestingly, the higher chlorophyll content in ekofertile compared to microfertile is consistent with its higher magnesium (Mg) content, as Mg forms an integral part of the chlorophyll molecule^4,18. Mg plays a central role in chlorophyll synthesis via being the Mg²⁺ that is chelated at the center of the porphyrin ring¹⁹. This structural incorporation stabilises the chlorophyll molecule, facilitating light energy absorption, enabling the electron transport chain vital for photosynthesis²⁰. During the synthesis process, Mg chelatase, an enzyme complex, requires Mg²⁺ in an ATP-dependent process to insert the Mg²⁺ into the protoporphyrin IX molecule, which is the immediate precursor to chlorophyll²¹. This increase in chlorophyll content could be attributed to the biostimulant’s ability to reduce chloroplast biogenesis and chlorophyll degradation, as suggested by Nair et al.²² and Jannin et al.²³. This is because the biostimulants likely influence chloroplast development by increasing chlorophyll content. They promote green mass formation and enhance overall plant vigor and photosynthesis essential for chloroplast function and biogenesis (ekolive)²⁴. They act via mechanisms like hormonal regulation, enhanced nutrient and water use efficiency, and improved stress tolerance, supporting the development and function of chloroplasts as the sites of photosynthesis¹⁴. Treatment level had a significant influence on chlorophyll content. Control had the least effect on chlorophyll content compared to the other treatment levels. This is due to no input, possibly facilitating the increase of chloroplast biogenesis and chlorophyll degradation as reported by Nair et al.²² and Jannin et al.²³. The 10% was the best result, significantly increasing chlorophyll a, b, carotenoids, and total chlorophyll content at the tillering and stem elongation stages. This result aligns with Chrysargyris et al.²⁵, who experienced increased chlorophyll content after treating tomatoes with 0–3% biostomulant containing essential oils of rosemary and eucalyptus. Hidangmayum and Sharma²⁶ showed that different concentrations of seaweed extract biostimulant enhanced chlorophyll content differently in onion, with the best results as the concentration increased until diminishing returns. From this result, 10% with either ekofertile or microfertile biostimulant generally enhanced chlorophyll content better than the other treatment combinations. The best chlorophyll content result at the tillering and stem elongation stage came at the ekofertile^® x 10% interaction²⁶.

ekofertile and microfertile biostimulants showed insignificant differences across harvest index, straw yield, biological yield, and 1000-grain weight. The results showed that ekofertile biostimulant performed better than microfertile biostimulant. Wheat grain yield significantly differed across the two biostimulants, with ekofertile biostimulant showing a higher yield than microfertile biostimulant. This is mainly attributed to the higher nutrient, microbial, and organic acids contents of ekofertile as shown by PACCI et al.⁵. Generally, most biostimulants contain one or two of the constituents above. However, ekofertile and microfertile biostimulants containing all the constituents above could have contributed to the insignificant differences between the two biostimulants. ekofertile biostimulant has better results than microfertile biostimulant, likely due to the higher nutrient and microbial contents observed by AGBOR et al.¹¹. Mironenko et al.²⁷ observed a 1000-grain weight increase of wheat after liquid protein hydrolysate biostimulant application, aligning with this study. Similarly, Popko et al.²⁸ found an increase in wheat yield by applying a new plant-growth biostimulant based on amino acids, which supports our results. 1000-grain weight, biological yield, and grain yield were significantly influenced by treatment level, illustrating the treatment level dependency of biostimulant, with the best result at the 10%. This is likely attributed to a higher concentration of nutrients, organic acids, and microbial content as the concentration increases²⁶. The higher concentrations resulted in higher yields than inorganic fertilization, again throwing more light on the biostimulant constituents. Yaldiz and Camlica²⁹ documented the highest fresh and dry weight of cotton at the highest concentration of organic manure, supporting this study’s findings. Popko et al.²⁸ also found the best wheat yield at the highest amino acid concentration application, further supporting this result. Ali et al.³⁰ discovered that a low dosage of Cuscuta reflexa biostimulant extract enhances wheat yield better than higher dosages, which is contrary to our study, affirming that beyond a threshold, higher biostimulant concentration generates an adverse effect on yield. The direct reason for the variation in higher or lower doses is not apparent, but literature has attributed it to variation in manufacturing material, formulation concentration, and frequency of application³¹. This suggests that since microfertile and ekofertile biostimulants are made from silicified rock residues and sand, and their composition is higher, higher doses are preferable with no plausible adverse effect. Thus, the concentrations applied in this study are still below the threshold for the biostimulants used. The treatment level showed an insignificant effect on straw yield and harvest index, with better results for the concentrations than the control, 10% better than inorganic fertilization. Biostimulant type and treatment level interaction yielded insignificant differences across wheat yield and yield components. We observed better performance of ekofertile and treatment level interactions compared to microfertile biostimulant-dosage interactions. This depicts the higher nutrients and microbial content in ekofertile compared to microfertile^5,11. While the greenhouse results are promising, a more mechanistic study, including field applications, will be important.

The result showed that ekofertile had a higher water use efficiency than microfertile. The highest treatment level, 10% had the highest water use efficiency, confirming that biostimulants are concentration dependent as suggested by Lopes et al.³². Although biostimulants and treatment interactions result in insignificant differences in enhancing water use efficiency, it could be observed that biostimulants at higher concentrations lead to better water use efficiency. This further shows the biostimulants constituent of organic acids, nutrients, and microorganisms’ potential to stimulate metabolic processes that increase the plant’s ability to use water efficiently. Seciu et al.³³ enhanced the water use efficiency of cabbage plants using biostimulants, which supports our study result. Similarly, Chen et al.³⁴ showed how applying biostimulants enhances agronomic performance and water use efficiency in maize, supporting our study premise. This has shown that ekofertile and microfertile biostimulants offer great potential in sustainable agriculture by minimizing water use while enhancing crop productivity, as shown by our study in conformity with the literature³⁵.

Materials and methods

Description of the site

The experiment was conducted at the greenhouse of Ondokuz Mayis University’s Department of Soil Science and Plant Nutrition, Faculty of Agriculture, Samsun, Turkey. The site location is coordinated at 264,201°E and 4,582,754°N (WGS-84, Zone 36, UTMm). The average yearly highest and lowest temperatures are 5 ℃ to 27.7 ℃, and the relative humidity is 73%. The average annual precipitation is 937.26 mm per year.

Biostimulants

This study used two biostimulants, ekofertile plant and microfertile plant, derived from the ekolive company in Slovakia. PACCI et al.⁵ showed the organic acid, chemical, and biological constituents of ekofertile plant and microfertile plant, also attached here as supplementary material A. The biostimulants are classified as waste-derived biostimulants from sand and silicified rock residues¹⁰.

Experimental design

The greenhouse experimental design is a split-plot design consisting of two factors. Factor 1, treatment had 5 levels (control, inorganic fertilization, 2.5%, 5%, and 10% biostimulant), the range of concentrations were chosen to determine the optimal dose-response for the specific biostimulant to aid the identification of the most effective rate for promoting wheat growth and yield⁵. This allows for optimization and will help establish a practical and effective application strategy. Biostimulant type factor 2 served as the main plots with 2 levels (ekofertile and microfertile plant biostimulants). This gave 10 treatment combinations, each being replicated individually 3 times in the greenhouse (Table 4), giving 30 experimental units. A 150 kg of soil was collected from the field, Samsun, Turkiye, Bafra plain. The soil was placed in the shade to air dry for two weeks. The hefty clumped soils were crushed and sieved through a 4 mm sieve to obtain fine particle soil for growing crops in the greenhouse. Three kilograms of soil were placed in a 5 L bucket of 0.031 m² surface area without perforations to avoid leaching. The soil moisture content was calculated to estimate the soil’s field capacity (FC). Here, a 1 m² area was irrigated to saturation and covered with polythene for 3 days. After 3 days, the soil sample was collected at the center, taken to the laboratory, and oven-dried for 24 h³⁶.

Table 4.

Factor interactions.

Biostimulant type treatment level	Biostimulant type
Biostimulant type treatment level	Ekofertile plant	Microfertile plant
Control	ekofertile x control	microfertile x control
Inorganic fertilization	ekofertile x inorganic	microfertile x inorganic
2.5% biostimulant	ekofertile x 2.5%	microfertile x 2.5%
5% biostimulant	ekofertile x 5%	microfertile x 5%
10% biostimulant	ekofertile x 10%	microfertile x 10%

Open in a new tab

FC (%) = ((soil fresh weight – soil oven dry weight)/soil oven dry weight) × 100.

Since 500 wheat seeds are sown per m², 15 were sown per pot according to the treatment combination and watered after seeding. The wheat seeds were obtained from the Department of Field Crops, Faculty of Agriculture, Ondokuz Mayis University, Samsun, Turkiye. The liquid ekofertile plant and microfertile plant dosages were foliar applied as percentages at 3 intervals: planting, tillering, and stem elongation stage. This is to ensure the microbial activity begins before plant growth to enable the timely availability of fixed nutrients and reduce stress. Meanwhile, inorganic fertilization was supplied during tillering and stem elongation stages, as nutrients are readily available and most needed. The dosage was soil applied according to the pre-soil results, which was 0.121gN₂ (0.06 gKNO₃ and 0.06 gNH₂SO₄).

Greenhouse management

The wheat plants were irrigated to field capacity in the evening periods of the day at 2-day intervals to prevent drought stress, by subtracting the 2-day weight from the initial weight (threshold) and adding water till the initial weight limit. At the experiment’s establishment, the weight of the pot, including the soil and water added to the field capacity, was recorded, and this was set as the upper limit for continuous moisture monitoring at 2-day intervals. Weeding was done manually.

Collection of data

Collection of plant data

Randomly tagged five plants from each unit were used for vegetative and yield data collection and plant sample collection for photosynthesis analysis.

Yield data were collected on 1000-grain weight (g), grain yield in tons per hectare (t ha⁻¹), straw yield (t ha⁻¹), biological yield (t ha⁻¹), which is the total dry weight of the entire plant, and harvest index (grain yield/biological yield) ×100.

Plant samples were collected at the wheat tillering and stem elongation stage to analyze chlorophyll a, b, and carotenoids

The analysis was done following the method described by^37,38. The analysis was done early in the morning in a dark room to avoid light degradation of pigment and the calculation done as below.

Chlorophyll a (mg/g) = [(12.70 x A₆₆₃) - (2.69 x A₆₄₅)] x V ÷ (1000 x W).

Chlorophyll b (mg/g) = [(22.90 x A₆₄₅) - (4.68 x A₆₆₃)] x V ÷ (1000 x W).

Total chlorophyll (mg/g) = [(20.2 x A₆₄₅) + (8.02 x A₆₆₃)] x V ÷ (1000 x W).

Carotenoid (mg/g) = (A₄₈₀ x V) ÷ (250 x W).

Where A₆₆₃ = absorbance reading at 663 nm, A₆₄₅ = absorbance reading at 645 nm, A₄₈₀ = absorbance reading at 480 nm, V_(ml) = final volume 25 ml, W_(g) = weight of sample.

Water use efficiency (WUE)

Water used efficiency was calculated as

Where biological yield is the whole biomass produced.

Water used was obtained by adding the difference between the initial weight of the pot including the soil and water added to the field capacity, and the 2-day interval weight recorded for the entire experiment.

Analysis of data

All data collected on yield, chlorophyll a, b, total, carotenoid, and WUE were keyed into Excel version 16 and later analyzed using the R programming package. A split-plot model at P ≤ 0.05 was used to test the effect of biostimulant type and treatment level as categorical predictors. Significantly different data means were separated using post hoc LSD (P ≤ 0.05). A split-plot model was used because the biostimulant was the main plot while the treatment level was the minor³⁹. Assumption of ANOVA for normal distribution of data was verified using the Q-Q plot, in which the expected and observed z-score values fall within the trendline.

Conclusion

This study has shown that ekofertile and microfertile biostimulants, treatment level, and interactions significantly enhanced wheat photosynthetic activity, yield, and yield components and water use efficiency. ekofertile had a significantly higher effect on chlorophyll content at the tillering and stem elongation stages, except for total chlorophyll content, where microfertile was higher though not significant. ekofertile at tillering; total chlorophyll 2.97 mg/g significantly higher than microfertile (2.81 mg/g), similar for carotenoid 0.43 mg/g to 0.41 mg/g. Total chlorophyll was not significantly different at stem elongation, but carotenoid was significantly higher at ekofertile (0.52 mg/g) compared with microfertile (0.41 mg/g). Treatment level 10% biostimulant dosage had the highest significant effect on total chlrophyll content at tillering (3.05 mg/g) and stem elongation (3.02 mg/g) stages compared with lower values for the other treatment levels. The ekofertile biostimulant interaction with 10% had the highest effect. A similar trend was observed for wheat yield and components; ekofertile (9.4 t ha⁻¹) had a significantly higher grain yield than microfertile (8.8 t ha⁻¹). The result suggest that application of ekofertile biostimulant at 10% will significantly increase chlorophyll content and wheat yield. ekofertile and microfertile biostimulants likely enhance wheat performance by stimulating plant hormonal regulation, improving nutrient uptake and utilization through microbial activity and enhanced root development, increasing photosynthetic efficiency via improved chlorophyll content and antioxidant activity, and improving water use efficiency through mechanisms like osmotic adjustment and stomatal regulation. These mechanisms reduce assimilate diversion to stress responses and enhance biomass allocation, leading to higher wheat yields and improved components like grain number and size. Thus, these biostimulants’ uniqueness, consisting of beneficial microbes, mineral nutrients, and organic acids, are valuable inputs for sustainable agriculture. Also, higher ekofertile organic acids, plant growth-promoting microorganisms, and mineral nutrients content translated to the better result in chlorophyll content and wheat yield. Further mechanistic study of the biostimulaants, other doses, field experiment and crops will be valuable.

Supplementary Information

Below is the link to the electronic supplementary material.

Supplementary Material 1^{(21.8KB, docx)}

Acknowledgements

Acknowledgment: We thank the ekolive Company in Slovakia for contributing to the biostimulant and sharing its expertise. We would also like to acknowledge the research support received from the Erasmus Mundus Joint Master’s Degree in Soil Science (emiSS) program of the European Union. The present study also received support from the Scientific Research Projects Coordination Unit of Ondokuz Mayıs University, grant number PYO.ZRT.1904.23.008. Special appreciation to Bambe Bertrand AS for his support in experiment establishment, management, and laboratory analysis.

Author contributions

All authors contributed to the study’s conception and design. David Tavi Agbor: Conceptualization, Methodology, Software, and Writing- Original draft preparation. Sena Pacci: Conceptualization, Methodology, Data Curation. Salih DEMIRKAYA: Conceptualization, Methodology, Software, Visualization. Abdurrahman AY: Conceptualization, Methodology, Software. Desmond Kwayela Sama: Data curation, Writing- Original draft preparation. Elis-Bright Iteke Molua: Data Curation, Reviewing, and Editing. Endalamaw Dessie ALEBACHEW: Data Curation, Reviewing, and Editing. Darina ŠTYRIAKOVÁ: Investigation, Reviewing, Editing, and Validation. Orhan Dengiz: Supervision, Writing- Reviewing and Editing, and Validation. All authors have read and approved the final version of the manuscript.

Funding

The funding for this research was provided by the Erasmus Mundus Joint Master’s Program in Soil Science of the European Union Scholarship through the Erasmus Mundus Master Program (emiSS; https://emissmaster.omu.edu.tr), ekolive Company in Slovakia, and the Scientific Research Projects Coordination Unit of Ondokuz Mayıs University under project number PYO.ZRT.1904.23.008.

Data availability

The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.

Declarations

Competing interests

The authors declare no competing interests.

Footnotes

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

References

1.Jardin, P. Plant biostimulants: Definition, concept, main categories and regulation. Sci. Hortic. (Amsterdam). 196, 3–14. 10.1016/j.scienta.2015.09.021 (Nov. 2015).
2.Bulgari, R., Franzoni, G. & Ferrante, A. Biostimulants application in horticultural crops under abiotic stress conditions. Agronomy9 (6), 306. 10.3390/agronomy9060306 (Jun. 2019).
3.Colla, G., Rouphael, Y., Canaguier, R., Svecova, E. & Cardarelli, M. Biostimulant action of a plant-derived protein hydrolysate produced through enzymatic hydrolysis, Front. Plant Sci., vol. 5, no. SEP, p. 110917, Sep. (2014). 10.3389/fpls.2014.00448 [DOI] [PMC free article] [PubMed]
4.Savarese, C. et al. Combination of humic biostimulants with a microbial inoculum improves lettuce productivity, nutrient uptake, and primary and secondary metabolism. Plant. Soil.481, 1–2. 10.1007/s11104-022-05634-8 (Aug. 2022).
5.PACCI, S. & AGBOR, D. O. DENGIZ, and Impact of biostimulants on soil quality, in International soil science symposium on soil science & plant nutrition (8th International Scientific Meeting), Samsun, Turkey: Federation of eurasian soil science societies, pp. 153–161. Accessed: May 05, 2025. [Online]. (2023). Available: https://www.researchgate.net/profile/SenaPacci/publication/377627483_Impact_of_biostimulants_on_soil_quality/links/65affba37fe0d83cb562a344/Impact-of-biostimulants-on-soil-quality.pdf
6.Vashi, J., Saravaiya, S., Patel, A. & Chaudhari, B. Silicon – The most under-appreciated element for vegetables. Int. J. Chem. Stud.8 (4), 2122–2127. 10.22271/chemi.2020.v8.i4w.9941 (2020). [Google Scholar]
7.Swoboda, P., Döring, T. F. & Hamer, M. Remineralizing soils? The agricultural usage of silicate rock powders: A review. Sci. Total Environ.807, 150976. 10.1016/j.scitotenv.2021.150976 (Feb. 2022). [DOI] [PubMed]
8.Elnahal, A. S. M. et al. Correction: The use of microbial inoculants for biological control, plant growth promotion, and sustainable agriculture: A review (European Journal of Plant Pathology, 162, 4, (759–792), (2022). 10.1007/s10658-021-02393-7), Eur. J. Plant Pathol., vol. 162, no. 4, p. 1007, Apr. 2022, doi: 10.1007/s10658-022-02472-3.
9.Battacharyya, D., Babgohari, M. Z., Rathor, P. & Prithiviraj, B. Seaweed extracts as biostimulants in horticulture, Sci. Hortic. (Amsterdam)., vol. 196, pp. 39–48, Nov. (2015). 10.1016/j.scienta.2015.09.012
10.Johnson, R., Joel, J. M. & Puthur, J. T. Biostimulants: The futuristic sustainable approach for alleviating crop productivity and abiotic stress tolerance, J. Plant Growth Regul., vol. 43, no. 3, pp. 659–674, Mar. (2024). 10.1007/s00344-023-11144-3
11.Agbor, D., Štyriaková, D. & Eedings, O. D. U. Agroecological significance of ekofertile™ plant biostimulant on tropical soils and crop improvement, in International soil science symposium on soil science & plant nutrition (7th International Scientific Meeting), Samsun, Turkey: Federation of Eurasian Soil Science Societies, 2022, pp. 63–68. Accessed: May 05, 2025. [Online]. (2022). Available: https://www.fesss.org/uploadfile/1671012304.pdf#page=80
12.Mazhar, M. W. et al. Optimizing water relations, gas exchange parameters, biochemical attributes and yield of water-stressed maize plants through seed priming with iron oxide nanoparticles. BMC Plant. Biol.24 (1), 1–17. 10.1186/S12870-024-05324-W/TABLES/4 (Dec. 2024). [DOI] [PMC free article] [PubMed]
13.Ahmed, N. et al. Micronutrients and their effects on horticultural crop quality, productivity and sustainability. Sci. Hortic. (Amsterdam). 323, 112512. 10.1016/J.SCIENTA.2023.112512 (Jan. 2024).
14.Haghaninia, M., Javanmard, A., Rasouli, F., Radicetti, E. & Memarzadeh Mashhouri, S. Optimizing camelina performance under drought conditions through the combined action of nanochitosan-encapsulated rosemary oil and arbuscular mycorrhizal fungi, Biocatal. Agric. Biotechnol., vol. 67, p. 103682, Jul. (2025). 10.1016/J.BCAB.2025.103682
15.Sellami, M. H., Di Mola, I. & Mori, M. Evaluating wheat response to biostimulants: a 25-year review of field-based research (2000–2024). Front. Sustain. Food Syst.9, 1543981. 10.3389/FSUFS.2025.1543981/BIBTEX (Mar. 2025).
16.Antón-Herrero, R. et al. Biostimulant Effects of Micro Carbon Technology (MCT^®)-Based Fertilizers on Soil and Capsicum annuum Culture in Growth Chamber and Field, Agronomy, vol. 12, no. 1, p. 70, Dec. (2022). 10.3390/agronomy12010070
17.Irani, H., ValizadehKaji, B. & Naeini, M. R. Biostimulant-induced drought tolerance in grapevine is associated with physiological and biochemical changes. Chem. Biol. Technol. Agric.8 (1), 1–13. 10.1186/s40538-020-00200-9 (Dec. 2021).
18.Giordano, M. et al. Jul., Stand-alone and combinatorial effects of plant-based biostimulants on the production and leaf quality of perennial wall rocket, Plants, vol. 9, no. 7, pp. 1–15, (2020). 10.3390/plants9070922 [DOI] [PMC free article] [PubMed]
19.Ahmed, N. et al. The power of magnesium: unlocking the potential for increased yield, quality, and stress tolerance of horticultural crops. Front. Plant. Sci.14, 1285512. 10.3389/FPLS.2023.1285512/FULL (Oct. 2023). [DOI] [PMC free article] [PubMed]
20.Adriaje, L. Role of magnesium in the process of photosynthesis. J. Plant. Biochem. Physiol.13 (1), 1–2. 10.35248/2329-9029.25.13.342 (Feb. 2025).
21.Rissler, H. M., Collakova, E., DellaPenna, D., Whelan, J. & Pogson, B. J. Chlorophyll biosynthesis. Expression of a second Chl i gene of magnesium chelatase in Arabidopsis supports only limited Chlorophyll synthesis. Plant. Physiol.128 (2), 770. 10.1104/PP.010625 (2002). [DOI] [PMC free article] [PubMed] [Google Scholar]
22.Nair, P. et al. Transcriptional and metabolomic analysis of ascophyllum nodosum mediated freezing tolerance in Arabidopsis Thaliana. BMC Genom.13 (1), 1–23. 10.1186/1471-2164-13-643 (Nov. 2012). [DOI] [PMC free article] [PubMed]
23.Jannin, L. et al. Mar., Brassica napus growth is promoted by ascophyllum nodosum (L.) Le Jol. seaweed extract: microarray analysis and physiological characterization of N, C, and S metabolisms, J. Plant Growth Regul., vol. 32, no. 1, pp. 31–52, (2013). 10.1007/s00344-012-9273-9
24.microfertile^® plant EN – ekolive. Accessed: Sep. 12, 2025. [Online]. Available: https://ekolive.eu/agriculture-en/microfertile-plant/
25.Chrysargyris, A. et al. Assessing the biostimulant effects of a novel plant-based formulation on tomato crop, Sustain., vol. 12, no. 20, pp. 1–15, Oct. (2020). 10.3390/su12208432
26.Hidangmayum, A. & Sharma, R. Effect of different concentrations of commercial seaweed liquid extract of Ascophyllum nodosum as a plant bio stimulant on growth, yield and biochemical constituents of onion (Allium cepa L.), J. Pharmacogn. Phytochem., vol. 6, no. 4, pp. 658–663, [Online]. (2017). Available: https://www.phytojournal.com/archives/2017.v6.i4.1409/effect-of-different-concentrations-of-commercial-seaweed-liquid-extract-of-ascophyllum-nodosum-as-a-plant-bio-stimulant-on-growth-yield-and-biochemical-constituents-of-onion-allium-cepa-l
27.Mironenko, G. A., Zagorskii, I. A., Bystrova, N. A. & Kochetkov, K. A. The effect of a biostimulant based on a protein hydrolysate of rainbow trout (oncorhynchus mykiss) on the growth and yield of wheat (triticum aestivum L.), Molecules, vol. 27, no. 19, p. 6663, Oct. (2022). 10.3390/molecules27196663 [DOI] [PMC free article] [PubMed]
28.Popko, M. et al. Effect of the new plant growth biostimulants based on amino acids on yield and grain quality of winter wheat. Molecules23 (2), 470. 10.3390/molecules23020470 (Feb. 2018). [DOI] [PMC free article] [PubMed]
29.Yaldiz, G. & Camlica, M. Assessing the effect of various doses of organic manures on herbage yield, essential oil, and compositions of sage grown under climate chamber and field conditions, Org. Agric., vol. 13, no. 3, pp. 377–397, Sep. (2023). 10.1007/s13165-023-00434-5
30.Ali, Q. et al. Aug., Low doses of cuscuta reflexa extract act as natural biostimulants to improve the germination vigor, growth, and grain yield of wheat grown under water stress: Photosynthetic pigments, antioxidative defense mechanisms, and nutrient acquisition, Biomolecules, vol. 10, no. 9, pp. 1–30, (2020). 10.3390/biom10091212 [DOI] [PMC free article] [PubMed]
31.Schmidt, R. E., Ervin, E. H. & Zhang, X. Questions and answers about biostimulants. Golf Course Manag. 71 (6), 91–94 (2003). [Google Scholar]
32.Lopes, W. O., Varão, L. C. & Buso, W. H. D. Foliar application of biostimulant doses in two phenological stages in common bean culture. Rev. Agric. Neotrop.9 (3), e6981–e6981. 10.32404/rean.v9i3.6981 (Sep. 2022).
33.Seciu, A. M. et al. Water use efficiency on cabbage and cauliflower treated with a new biostimulant composition. Agric. Agric. Sci. Procedia. 10, 475–484. 10.1016/j.aaspro.2016.09.019 (Jan. 2016).
34.Chen, C. H., Lin, K. H., Sen Chang, Y. & Chang, Y. J. Application of water-saving irrigation and biostimulants on the agronomic performance of maize (Zea mays), Process Saf. Environ. Prot., vol. 177, pp. 1377–1386, Sep. (2023). 10.1016/j.psep.2023.08.008
35.Gitau, M. M., Farkas, A., Ördög, V. & Maróti, G. Evaluation of the biostimulant effects of two chlorophyta microalgae on tomato (Solanum lycopersicum). J. Clean. Prod.364, 132689. 10.1016/j.jclepro.2022.132689 (Sep. 2022).
36.4-Field. Capacity | PDF | Soil | Water. Accessed: Sep. 03, 2025. [Online]. Available: https://www.scribd.com/document/711011056/4-Field-Capacity
37.Arnon, D. I. Copper enzymes in isolated chloroplasts. Polyphenoloxidase in beta vulgaris. Plant. Physiol.24 (1), 1–15. 10.1104/pp.24.1.1 (Jan. 1949). [DOI] [PMC free article] [PubMed]
38.Feung, C. S., Hamilton, R. H. & Witham, F. H. Metabolism of 2,4-dichlorophenoxyacetic acid by soybean cotyledon callus tissue cultures. J. Agric. Food Chem.19 (3), 475–479. 10.1021/jf60175a032 (May 1971).
39.Armstrong, R. A. & Hilton, A. C. Split-plot analysis of variance, Stat. Anal. Microbiol. Statnotes, pp. 71–75, Nov. (2010). 10.1002/9780470905173.CH13

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

Supplementary Material 1^{(21.8KB, docx)}

Data Availability Statement

The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.

[CR1] 1.Jardin, P. Plant biostimulants: Definition, concept, main categories and regulation. Sci. Hortic. (Amsterdam). 196, 3–14. 10.1016/j.scienta.2015.09.021 (Nov. 2015).

[CR2] 2.Bulgari, R., Franzoni, G. & Ferrante, A. Biostimulants application in horticultural crops under abiotic stress conditions. Agronomy9 (6), 306. 10.3390/agronomy9060306 (Jun. 2019).

[CR3] 3.Colla, G., Rouphael, Y., Canaguier, R., Svecova, E. & Cardarelli, M. Biostimulant action of a plant-derived protein hydrolysate produced through enzymatic hydrolysis, Front. Plant Sci., vol. 5, no. SEP, p. 110917, Sep. (2014). 10.3389/fpls.2014.00448 [DOI] [PMC free article] [PubMed]

[CR4] 4.Savarese, C. et al. Combination of humic biostimulants with a microbial inoculum improves lettuce productivity, nutrient uptake, and primary and secondary metabolism. Plant. Soil.481, 1–2. 10.1007/s11104-022-05634-8 (Aug. 2022).

[CR5] 5.PACCI, S. & AGBOR, D. O. DENGIZ, and Impact of biostimulants on soil quality, in International soil science symposium on soil science & plant nutrition (8th International Scientific Meeting), Samsun, Turkey: Federation of eurasian soil science societies, pp. 153–161. Accessed: May 05, 2025. [Online]. (2023). Available: https://www.researchgate.net/profile/SenaPacci/publication/377627483_Impact_of_biostimulants_on_soil_quality/links/65affba37fe0d83cb562a344/Impact-of-biostimulants-on-soil-quality.pdf

[CR6] 6.Vashi, J., Saravaiya, S., Patel, A. & Chaudhari, B. Silicon – The most under-appreciated element for vegetables. Int. J. Chem. Stud.8 (4), 2122–2127. 10.22271/chemi.2020.v8.i4w.9941 (2020). [Google Scholar]

[CR7] 7.Swoboda, P., Döring, T. F. & Hamer, M. Remineralizing soils? The agricultural usage of silicate rock powders: A review. Sci. Total Environ.807, 150976. 10.1016/j.scitotenv.2021.150976 (Feb. 2022). [DOI] [PubMed]

[CR8] 8.Elnahal, A. S. M. et al. Correction: The use of microbial inoculants for biological control, plant growth promotion, and sustainable agriculture: A review (European Journal of Plant Pathology, 162, 4, (759–792), (2022). 10.1007/s10658-021-02393-7), Eur. J. Plant Pathol., vol. 162, no. 4, p. 1007, Apr. 2022, doi: 10.1007/s10658-022-02472-3.

[CR9] 9.Battacharyya, D., Babgohari, M. Z., Rathor, P. & Prithiviraj, B. Seaweed extracts as biostimulants in horticulture, Sci. Hortic. (Amsterdam)., vol. 196, pp. 39–48, Nov. (2015). 10.1016/j.scienta.2015.09.012

[CR10] 10.Johnson, R., Joel, J. M. & Puthur, J. T. Biostimulants: The futuristic sustainable approach for alleviating crop productivity and abiotic stress tolerance, J. Plant Growth Regul., vol. 43, no. 3, pp. 659–674, Mar. (2024). 10.1007/s00344-023-11144-3

[CR11] 11.Agbor, D., Štyriaková, D. & Eedings, O. D. U. Agroecological significance of ekofertile™ plant biostimulant on tropical soils and crop improvement, in International soil science symposium on soil science & plant nutrition (7th International Scientific Meeting), Samsun, Turkey: Federation of Eurasian Soil Science Societies, 2022, pp. 63–68. Accessed: May 05, 2025. [Online]. (2022). Available: https://www.fesss.org/uploadfile/1671012304.pdf#page=80

[CR12] 12.Mazhar, M. W. et al. Optimizing water relations, gas exchange parameters, biochemical attributes and yield of water-stressed maize plants through seed priming with iron oxide nanoparticles. BMC Plant. Biol.24 (1), 1–17. 10.1186/S12870-024-05324-W/TABLES/4 (Dec. 2024). [DOI] [PMC free article] [PubMed]

[CR13] 13.Ahmed, N. et al. Micronutrients and their effects on horticultural crop quality, productivity and sustainability. Sci. Hortic. (Amsterdam). 323, 112512. 10.1016/J.SCIENTA.2023.112512 (Jan. 2024).

[CR14] 14.Haghaninia, M., Javanmard, A., Rasouli, F., Radicetti, E. & Memarzadeh Mashhouri, S. Optimizing camelina performance under drought conditions through the combined action of nanochitosan-encapsulated rosemary oil and arbuscular mycorrhizal fungi, Biocatal. Agric. Biotechnol., vol. 67, p. 103682, Jul. (2025). 10.1016/J.BCAB.2025.103682

[CR15] 15.Sellami, M. H., Di Mola, I. & Mori, M. Evaluating wheat response to biostimulants: a 25-year review of field-based research (2000–2024). Front. Sustain. Food Syst.9, 1543981. 10.3389/FSUFS.2025.1543981/BIBTEX (Mar. 2025).

[CR16] 16.Antón-Herrero, R. et al. Biostimulant Effects of Micro Carbon Technology (MCT^®)-Based Fertilizers on Soil and Capsicum annuum Culture in Growth Chamber and Field, Agronomy, vol. 12, no. 1, p. 70, Dec. (2022). 10.3390/agronomy12010070

[CR17] 17.Irani, H., ValizadehKaji, B. & Naeini, M. R. Biostimulant-induced drought tolerance in grapevine is associated with physiological and biochemical changes. Chem. Biol. Technol. Agric.8 (1), 1–13. 10.1186/s40538-020-00200-9 (Dec. 2021).

[CR18] 18.Giordano, M. et al. Jul., Stand-alone and combinatorial effects of plant-based biostimulants on the production and leaf quality of perennial wall rocket, Plants, vol. 9, no. 7, pp. 1–15, (2020). 10.3390/plants9070922 [DOI] [PMC free article] [PubMed]

[CR19] 19.Ahmed, N. et al. The power of magnesium: unlocking the potential for increased yield, quality, and stress tolerance of horticultural crops. Front. Plant. Sci.14, 1285512. 10.3389/FPLS.2023.1285512/FULL (Oct. 2023). [DOI] [PMC free article] [PubMed]

[CR20] 20.Adriaje, L. Role of magnesium in the process of photosynthesis. J. Plant. Biochem. Physiol.13 (1), 1–2. 10.35248/2329-9029.25.13.342 (Feb. 2025).

[CR21] 21.Rissler, H. M., Collakova, E., DellaPenna, D., Whelan, J. & Pogson, B. J. Chlorophyll biosynthesis. Expression of a second Chl i gene of magnesium chelatase in Arabidopsis supports only limited Chlorophyll synthesis. Plant. Physiol.128 (2), 770. 10.1104/PP.010625 (2002). [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR22] 22.Nair, P. et al. Transcriptional and metabolomic analysis of ascophyllum nodosum mediated freezing tolerance in Arabidopsis Thaliana. BMC Genom.13 (1), 1–23. 10.1186/1471-2164-13-643 (Nov. 2012). [DOI] [PMC free article] [PubMed]

[CR23] 23.Jannin, L. et al. Mar., Brassica napus growth is promoted by ascophyllum nodosum (L.) Le Jol. seaweed extract: microarray analysis and physiological characterization of N, C, and S metabolisms, J. Plant Growth Regul., vol. 32, no. 1, pp. 31–52, (2013). 10.1007/s00344-012-9273-9

[CR24] 24.microfertile^® plant EN – ekolive. Accessed: Sep. 12, 2025. [Online]. Available: https://ekolive.eu/agriculture-en/microfertile-plant/

[CR25] 25.Chrysargyris, A. et al. Assessing the biostimulant effects of a novel plant-based formulation on tomato crop, Sustain., vol. 12, no. 20, pp. 1–15, Oct. (2020). 10.3390/su12208432

[CR26] 26.Hidangmayum, A. & Sharma, R. Effect of different concentrations of commercial seaweed liquid extract of Ascophyllum nodosum as a plant bio stimulant on growth, yield and biochemical constituents of onion (Allium cepa L.), J. Pharmacogn. Phytochem., vol. 6, no. 4, pp. 658–663, [Online]. (2017). Available: https://www.phytojournal.com/archives/2017.v6.i4.1409/effect-of-different-concentrations-of-commercial-seaweed-liquid-extract-of-ascophyllum-nodosum-as-a-plant-bio-stimulant-on-growth-yield-and-biochemical-constituents-of-onion-allium-cepa-l

[CR27] 27.Mironenko, G. A., Zagorskii, I. A., Bystrova, N. A. & Kochetkov, K. A. The effect of a biostimulant based on a protein hydrolysate of rainbow trout (oncorhynchus mykiss) on the growth and yield of wheat (triticum aestivum L.), Molecules, vol. 27, no. 19, p. 6663, Oct. (2022). 10.3390/molecules27196663 [DOI] [PMC free article] [PubMed]

[CR28] 28.Popko, M. et al. Effect of the new plant growth biostimulants based on amino acids on yield and grain quality of winter wheat. Molecules23 (2), 470. 10.3390/molecules23020470 (Feb. 2018). [DOI] [PMC free article] [PubMed]

[CR29] 29.Yaldiz, G. & Camlica, M. Assessing the effect of various doses of organic manures on herbage yield, essential oil, and compositions of sage grown under climate chamber and field conditions, Org. Agric., vol. 13, no. 3, pp. 377–397, Sep. (2023). 10.1007/s13165-023-00434-5

[CR30] 30.Ali, Q. et al. Aug., Low doses of cuscuta reflexa extract act as natural biostimulants to improve the germination vigor, growth, and grain yield of wheat grown under water stress: Photosynthetic pigments, antioxidative defense mechanisms, and nutrient acquisition, Biomolecules, vol. 10, no. 9, pp. 1–30, (2020). 10.3390/biom10091212 [DOI] [PMC free article] [PubMed]

[CR31] 31.Schmidt, R. E., Ervin, E. H. & Zhang, X. Questions and answers about biostimulants. Golf Course Manag. 71 (6), 91–94 (2003). [Google Scholar]

[CR32] 32.Lopes, W. O., Varão, L. C. & Buso, W. H. D. Foliar application of biostimulant doses in two phenological stages in common bean culture. Rev. Agric. Neotrop.9 (3), e6981–e6981. 10.32404/rean.v9i3.6981 (Sep. 2022).

[CR33] 33.Seciu, A. M. et al. Water use efficiency on cabbage and cauliflower treated with a new biostimulant composition. Agric. Agric. Sci. Procedia. 10, 475–484. 10.1016/j.aaspro.2016.09.019 (Jan. 2016).

[CR34] 34.Chen, C. H., Lin, K. H., Sen Chang, Y. & Chang, Y. J. Application of water-saving irrigation and biostimulants on the agronomic performance of maize (Zea mays), Process Saf. Environ. Prot., vol. 177, pp. 1377–1386, Sep. (2023). 10.1016/j.psep.2023.08.008

[CR35] 35.Gitau, M. M., Farkas, A., Ördög, V. & Maróti, G. Evaluation of the biostimulant effects of two chlorophyta microalgae on tomato (Solanum lycopersicum). J. Clean. Prod.364, 132689. 10.1016/j.jclepro.2022.132689 (Sep. 2022).

[CR36] 36.4-Field. Capacity | PDF | Soil | Water. Accessed: Sep. 03, 2025. [Online]. Available: https://www.scribd.com/document/711011056/4-Field-Capacity

[CR37] 37.Arnon, D. I. Copper enzymes in isolated chloroplasts. Polyphenoloxidase in beta vulgaris. Plant. Physiol.24 (1), 1–15. 10.1104/pp.24.1.1 (Jan. 1949). [DOI] [PMC free article] [PubMed]

[CR38] 38.Feung, C. S., Hamilton, R. H. & Witham, F. H. Metabolism of 2,4-dichlorophenoxyacetic acid by soybean cotyledon callus tissue cultures. J. Agric. Food Chem.19 (3), 475–479. 10.1021/jf60175a032 (May 1971).

[CR39] 39.Armstrong, R. A. & Hilton, A. C. Split-plot analysis of variance, Stat. Anal. Microbiol. Statnotes, pp. 71–75, Nov. (2010). 10.1002/9780470905173.CH13

PERMALINK

ekofertile and microfertile plant biostimulants enhanced wheat productivity and water use efficiency

David Tavi Agbor

Darina Štyriaková

Sena Pacci

Desmond Kwayela Sama

Salih Demirkaya

Abdurrahman Ay

Elis-Bright Iteke Molua

Endalamaw Dessie Alebachew

Orhan Dengiz

Abstract

Supplementary Information

Introduction

Result

Sole and interactive effect of biostimulant type and treatment level on wheat chlorophyll content

Table 1.

Table 2.

Sole and interactive effect of biostimulant type and treatment level on wheat yield and yield components

Correlation

Table 3.

Discussion

Materials and methods

Description of the site

Biostimulants

Experimental design

Table 4.

Greenhouse management

Collection of data

Collection of plant data

Plant samples were collected at the wheat tillering and stem elongation stage to analyze chlorophyll a, b, and carotenoids

Water use efficiency (WUE)

Analysis of data

Conclusion

Supplementary Information

Acknowledgements

Author contributions

Funding

Data availability

Declarations

Competing interests

Footnotes

References

Associated Data

Supplementary Materials

Data Availability Statement

ACTIONS

PERMALINK

RESOURCES

Similar articles

Cited by other articles

Links to NCBI Databases