Fulvic Acid, Chitosan, and Amino Acids are a common biostimulant. Biostimulants are increasingly recognized in modern agriculture as eco-friendly inputs that enhance plant growth, improve stress tolerance, and promote product quality. This study investigated the effects of fulvic acid, amino acids, and chitosan on the growth, yield, and nutritional quality of parsley grown under hydroponic greenhouse conditions. The research was conducted in two stages. These findings suggest that fulvic acid and chitosan, applied individually and particularly in combination, may serve as effective biostimulant strategies for improving yield and nutritional quality while reducing nitrate accumulation in hydroponically grown parsley.
Introduction
Hydroponic agriculture has emerged as a controlled cultivation method that plays a significant role in the modern greenhouse industry of developed countries. These hydroponic greenhouse systems provide a controlled environment that leads to substantial improvements in plant growth and yield, while also enhancing the efficiency of agricultural water management and enabling year-round production. This advanced agricultural technique, which has rapidly developed over the past 30-40 years, is widely applied particularly in greenhouse environments, where the majority of hydroponic crops are cultivated in high-technology greenhouse facilities equipped with fully automated climate control systems. This method, in which plants are grown without soil using a nutrient-rich solution dissolved in water, is considered both environmentally friendly and space-efficient, making it an ideal option for urban areas or regions with limited agricultural land.
Biostimulants are substances, either natural or synthetic, that promote plant growth, increase yield, and improve stress resistance by triggering physiological processes, without functioning as traditional nutrients or pesticides. Agricultural advancements, including the use of chemical fertilizers, have played a crucial role in ensuring food production meets the demands of the growing global population. However, these developments have resulted in considerable environmental costs. Enhancing the efficiency of plant nutrient utilization is essential for addressing greenhouse gas emissions, supporting biodiversity, and improving agricultural productivity to satisfy the needs of the expanding world population. Biostimulants, which are compounds derived from various sources, have the ability to regulate plant physiological functions and enhance nutrient absorption and utilization. As a result, incorporating biostimulants into conventional farming practices could help reduce the need for fertilizers while sustaining crop yields. Plant biostimulation is a practical approach that involves biological processes triggered by environmental factors, including physical, chemical, and biological stimuli. These processes lead to adaptive modifications in the plant’s metabolism, allowing it to optimize the use of environmental resources and enhance its ability to cope with stress conditions.
Humic substances are organic compounds that naturally form through the decomposition of plant and animal residues and are classified into three groups: humin, humic acid, and fulvic acid (FA). Fulvic acid is distinguished by its low molecular weight and oxygen-rich functional groups, which enable it to pass through biological membrane micropores. Its high total acidity, abundant carboxyl groups, and superior adsorption and cation-exchange capacity allow fulvic acid to function as a natural chelating agent, facilitating the transport and uptake of micronutrients across cell membranes. Additionally, it promotes plant growth by enhancing photosynthesis, stimulating the secretion of growth hormones, improving nutrient absorption, and increasing resistance to both biotic and abiotic stresses. As a water-soluble, carbon-based chelating agent that remains stable across a wide pH range (acidic, neutral, and alkaline), fulvic acid plays a significant role in agricultural production due to its versatile and dynamic nature.
Amino acids (AAs) are nitrogen-containing compounds that serve as the essential building blocks of proteins and play a fundamental role in protein synthesis. They are crucial in metabolic processes by supplying key enzymes that promote cell growth. Recognized as potent biostimulants, AAs contribute significantly to plant growth and development. Extensive studies have highlighted their diverse benefits across various plant species. Research indicates that AAs enhance fertilizer uptake, improve nutrient and water absorption, and boost photosynthetic efficiency in many vegetable crops. These combined effects lead to increased flower production, better fruit set, and higher yields, emphasizing their importance in enhancing agricultural productivity.
Chitin is the second most significant natural polymer globally. The primary sources used are two types of marine crustaceans: shrimp and crabs. Through partial deacetylation in alkaline conditions, chitosan is produced, which is considered the most significant derivative of chitin for various applications. Due to their cationic polymer structure, biodegradability, ability to be absorbed by living organisms, and antimicrobial effects, chitosan nanoparticles (ChNPs) are considered highly suitable for developing slow-release pesticide and fertilizer formulations. The antimicrobial activity of chitosan is influenced by various factors such as the microbial species, degree of acetylation, and molecular weight, along with conditions like inoculum density, temperature, type of culture medium, and pH level. Chitosan enables the removal of heavy metals such as lead, uranium, copper, and mercury from soil and agricultural waste, thereby allowing the treated water to be reused for irrigation purposes. Utilizing chitosan as a plant biostimulant provides protection against soil-borne pathogens and enhances the population of beneficial soil microorganisms. This, in turn, supports better nutrient absorption by plants and contributes to improved growth and overall productivity.
Parsley (Petroselinum crispum), a member of the Apiaceae family (formerly known as Umbelliferae), is part of a plant group that has long been utilized as medicinal, aromatic, and edible vegetable species. Parsley is high in iron, vitamins A, B, and C, as well as in apiol, an oil extracted from the seeds, which is helpful to treat several health issues. Parsley leaves are rich in an essential oil known as myristicin, which exhibits anti-inflammatory, analgesic, and antiproliferative properties, and also demonstrates strong antibiotic activity against certain fungi and Gram-negative bacteria.
In addition to being a rich source of vitamins, minerals, and bioactive compounds, parsley is a widely cultivated leafy vegetable, making it a suitable model crop for evaluating the impact of biostimulant applications under hydroponic conditions. The primary objective of this research is to investigate how different biostimulant treatments influence yield and quality parameters in hydroponically grown parsley. By doing so, this study aims to provide valuable insights into the potential of biostimulants as sustainable tools to enhance crop productivity and nutritional quality, while reducing the dependency on synthetic inputs in modern agricultural systems.
Materials and Methods
Plant Material
This study was conducted in a designated section of a 500 m2 glasshouse allocated for soilless cultivation at the Department of Horticulture, Faculty of Agriculture, Çukurova University. Two separate trials were established. The objective of the first trial was to determine the optimal doses of the biostimulants. Based on the outcomes of the first trial, the second trial employed the most effective doses in combination with different biostimulants. The plant material used in the experiments was parsley cultivar procured, characterized by large, flat, dark-green leaves with broad lamina and slightly pointed tips.
Experimental Conditions
Parsley seeds were subjected to a priming treatment prior to sowing in order to promote rapid and uniform germination and to obtain homogeneous, high-quality seedlings. As the seeds used in this study were not commercially pre-primed by the supplier, the priming treatment was applied equally to all seeds, including the control, to ensure uniform emergence and experimental consistency. For priming, 15 g of seeds were soaked in a solution containing 0.5 g L-1 humic acid at 15 ℃ for 24 h under continuous aeration and then dried. This treatment promoted synchronized germination and improved seed vigor, leading to more consistent experimental outcomes. Following priming, the seeds were sown in plug trays filled with a peat-perlite mixture (2:1, v/v). Germination began approximately 10 days later, after which the seedlings were transferred to the greenhouse and grown under daytime temperatures of 20 ℃ and nighttime temperatures of 15 ℃. The seedlings were subsequently transferred to the hydroponic system. The plants were grown in rigid PVC containers with a capacity of 50 L and dimensions of 105 × 55 cm, filled with aerated nutrient solution. A floating culture system was used, in which the roots remained submerged in the nutrient solution, and air stones were installed to ensure continuous aeration. During the first experiment, conducted in winter, daytime temperatures inside the glasshouse ranged between 20 and 23 ℃ and nighttime temperatures between 13 and 15 ℃. In the second experiment, carried out in spring, the daytime temperature was maintained at 26-28 ℃, while nighttime conditions ranged from 17 to 19 ℃. Relative humidity was maintained between 50 and 60% under natural sunlight conditions. The plants were arranged in a randomized block design with a spacing of 15 × 15 cm, placing three plants in each position, corresponding to a planting density of 133 plants m-2. For plant nutrition, a fertilizer program consisting of two separate stock solutions (Stock A and Stock B) was prepared and diluted prior to use. Electrical conductivity (EC) was regularly adjusted, initially set at 1.5 dS m-1 for the first 20 days, increased to 1.8 dS m-1 between days 20 and 40, and subsequently maintained at 2.2 dS m-1 for the remainder of the cultivation period. The pH of the nutrient solution was consistently adjusted to 6.0 throughout the experiment.
Biostimulants were applied via the root zone by incorporation into the nutrient solution. The nutrient solution was completely renewed at 7-day intervals, and at each renewal, the respective biostimulants were freshly added to the solution at the designated concentrations. No partial replenishment or top-up was performed between solution renewals. Following biostimulant addition, the pH of the nutrient solution was adjusted to 6.0, and electrical conductivity (EC) values were measured and recorded to ensure consistency among treatments throughout the experimental period.
The nutrient solution was formulated to supply the following concentrations of essential mineral elements: 212 mg L-1 nitrogen (N), 50 mg L-1 phosphorus (P), 305 mg L-1 potassium (K), 205 mg L-1 calcium (Ca), and 60 mg L-1 magnesium (Mg). Micronutrients were provided at the following concentrations: 3.0 mg L-1 iron (Fe), 0.78 mg L-1 manganese (Mn), 0.51 mg L-1 boron (B), 0.50 mg L-1 zinc (Zn), 0.23 mg L-1 copper (Cu), and 0.18 mg L-1 molybdenum (Mo). These concentrations were optimized to meet the nutritional requirements of parsley under hydroponic cultivation.
The same plants were harvested repeatedly throughout each trial. Total leaf yield (g m-2) represents the cumulative yield obtained by summing all harvests per treatment. Yield was expressed on an area basis (g m-2) to ensure comparability across treatments and seasons and was calculated using plant spacing (row × plant distance). Plant weight was calculated at the end of each trial as the total fresh weight of all leaves harvested per plant across all harvests. Dry matter content represents the mean dry matter value determined from leaf samples collected at each harvest.
Biostimulants
In this study, three root-applied biostimulants were used. The amino acid-based biostimulant Amino and fulvic acid. The compositions of the amino acid and fulvic acid biostimulants were as follows: the fulvic acid contained 80% total organic matter, including 70% humic-fulvic substances, 70% fulvic acid, and a maximum moisture content of 7%, with a pH ranging from 2.0 to 4.0. The amino acid product contained 70% total organic matter, 14% organic carbon, 3% organic nitrogen, 29% free amino acids, and a maximum moisture content of 20%, with a pH between 2.5 and 4.5. In addition, a chitosan-based biostimulant, was used as the chitosan source, with a guaranteed content of 2.5% N-acetyl-d-glucosamine (w/w).
A two-stage experimental design was adopted (Table 1). In Trial 1, different single doses of fulvic acid (FA), amino acids (AA), and chitosan (C) were tested to identify the most effective dose of each biostimulant. Dose selection was based on total yield (g m-2), calculated from cumulative leaf harvests, as shown in Table 2. Accordingly, FA 80 ppm, AA 40 ppm, and C 0.3 mL L-1 were identified as the most favorable doses.
| Trial 1 | Trial 2 |
| Control | Control |
| Fulvic acid 80 ppm | Fulvic acid 80 ppm + Amino acid 40 ppm |
| Fulvic acid 120 ppm | Fulvic acid 80 ppm + Chitosan 0.3 mL L−1 |
| Amino acid 40 ppm | Amino acid 40 ppm + Chitosan 0.3 mL L−1 |
| Amino acid 80 ppm | Amino acid 40 ppm + Fulvic acid 80 ppm + Chitosan 0.3 mL L−1 |
| Chitosan 0.3 mL L−1 | |
| Chitosan 0.6 mL L−1 |
| Treatments | Plant Height (cm) | Plant Weight (g Plant−1) | Total Leaf Yield (g m−2) | Dry Matter (%) | Chlorophyll (SPAD) |
| Control | 23.4 a | 7.3 abc | 1590 c | 16.07 | 39.67 |
| Fulvic acid 80 | 25.0 a | 8.8 a | 1966 ab | 17.53 | 39.32 |
| Fulvic acid 120 | 22.0 ab | 6.8 bc | 1603 c | 18.42 | 43.54 |
| Amino acid 40 | 19.0 bc | 6.1 bc | 1438 cd | 17.69 | 46.49 |
| Amino acid 80 | 18.0 c | 5.7 c | 1283 d | 17.13 | 45.82 |
| Chitosan 0.3 | 23.0 a | 8.5 a | 2068 a | 16.87 | 41.77 |
| Chitosan 0.6 | 18.0 c | 7.6 ab | 1699 bc | 17.58 | 43.34 |
| LSD0.05 | 5.51 | 1.569 | 300.7 | NS | NS |
| p | 0.0037 | 0.0073 | 0.0015 | 0.079 | 0.617 |
Measurements of Plant Growth Parameters
For growth measurements, 10 plants were randomly selected and evaluated per replicate. Plant height (cm) was measured using a meter with 1 cm precision. The number of leaves per plant was recorded by manual counting. Leaf area (cm2 plant-1) was determined using a leaf area meter (Li-3100, LICOR, Lincoln, NE, USA). Fresh plant weight was measured using a digital balance with 0.1 g accuracy, while dry weight was obtained with a scale precise to 0.01 g. Leaf chlorophyll content was assessed non-destructively using a SPAD chlorophyll meter (Minolta SPAD-502, Tokyo, Japan). Dry matter content was determined from 100 g of fresh parsley samples, which were oven-dried until a constant weight was achieved. Dry matter percentage was then calculated using the following formula:
% Dry Matter = Dry Weight (g) ÷ Fresh Weight (g) × 100.
Results
The Effects of Biostimulant Applications on Growth Parameters of Parsley in the First and Second Trials
The effects of biostimulants applied to parsley plants are presented in the findings of the first and second trials in Table 2 and Table 3. Evaluation of the data obtained from the first trial revealed that among the applied biostimulant doses, Fulvic acid 80 and Chitosan 0.3 treatments were particularly prominent. The Fulvic acid 80 treatment resulted in marked increases in plant height, plant weight, and total leaf yield compared with the control. In contrast, the Chitosan 0.3 treatment led to increases in plant weight and total leaf yield relative to the control. Overall, the findings indicate that these doses exerted positive effects on plant growth and yield parameters. However, no statistically significant differences were observed among the applied biostimulant doses with respect to dry matter and chlorophyll (SPAD) values (Table 2). In the second trial, examination of the applied biostimulant combinations revealed that the Fulvic acid 80 + Chitosan 0.3 combination resulted in statistically significant increases in plant height, plant weight, total leaf yield, and dry matter compared to the control group. Furthermore, the evaluations indicated that none of the applied biostimulant combinations had a statistically significant effect on chlorophyll (SPAD) values (Table 3). Total leaf yield (g m-2) was higher in the winter (first experiment) than in the spring (second experiment), which can be attributed to the more favorable day and night temperatures observed during the first experiment.
| Treatments | Plant Height (cm) | Plant Weight (g Plant−1) | Total Leaf Yield (g m−2) | Dry Matter (%) | Chlorophyll (SPAD) |
| Control | 17.82 ab | 6.5 d | 493 d | 16.3 d | 37.38 |
| Fulvic acid 80 + Amino acid 40 | 16.07 b | 6.4 d | 487 d | 19.5 b | 42.38 |
| Fulvic acid 80 +Chitosan 0.3 | 22.00 a | 11.0 a | 856 a | 22.3 a | 41.00 |
| Amino acid 40 + Chitosan 0.3 | 18.81 b | 10.3 b | 777 b | 18.1 c | 41.50 |
| Amino acid 40 + Chitosan 0.3 + Fulvic acid 80 | 20.00 ab | 9.2 c | 694 c | 18.2 c | 40.02 |
| LSD0.05 | 2.44 | 0.722 | 54.18 | 0.233 | NS |
| p | 0.0478 | <0.0001 | <0.0001 | <0.0001 | 3.684 |
The Effects of Biostimulant Applications on Nitrate Content and Antioxidant Activity in Parsley During the First Trial
The effects of different biostimulant applications on nitrate content were examined. Compared to the control group (1048 mg/kg), the Fulvic acid 120 (405 mg/kg) and Amino acid 40 (423 mg/kg) treatments significantly reduced nitrate accumulation, making them the most effective applications in lowering nitrate content (Figure 1).
Vitamin C analysis revealed notable differences among the biostimulant applications. Compared to the control group (68.57 mg 100 g FW-1), the Fulvic Acid 120 treatment showed the highest vitamin C content (75.35 mg 100 g FW-1), indicating a positive effect on vitamin C accumulation. In contrast, the Fulvic Acid 80 application resulted in the lowest value (57.35 mg 100 g FW-1), suggesting a negative impact on vitamin C content (Figure 1).
In this study, DPPH analysis revealed that the Amino Acids 40 biostimulant application consistently resulted in the lowest antioxidant activity in parsley plants, showing the lowest inhibition and radical scavenging percentages compared to the control and other treatments (Figure 1).
Phenolic content analysis revealed that the Amino Acids 40 (391.1 mg GA 100 g FW-1), Amino Acids 80 (367.9 mg GA 100 g FW-1), and Chitosan 0.3 (387.7 mg GA 100 g FW-1) treatments resulted in the highest phenol levels compared to the control group (237.9 mg GA 100 g FW-1), indicating a strong enhancing effect of these applications on phenolic accumulation. In contrast, flavonoid content analysis showed no statistically significant differences between the control and the treatment groups, suggesting that biostimulant applications did not markedly influence flavonoid synthesis in parsley plants (Figure 1).
The Effects of Biostimulant Applications on Nitrate Content and Antioxidant Activity in Parsley During the Second Trial
In the second trial, the nitrate content in the control group (937 mg/kg) was higher than in all biostimulant-treated groups, with the lowest nitrate content recorded in the Fulvic Acid 80 + Chitosan 0.3 treatment (460 mg/kg). The results indicated that nitrate content was lower in all biostimulant-treated groups compared to the control, demonstrating that the applications effectively reduced nitrate accumulation in parsley (Figure 2).
In the control group, the vitamin C content (63.53 mg 100 g FW-1) was lower than that of the Fulvic Acid 80 + Chitosan 0.3 treatment, where the highest value (66.63 mg 100 g FW-1) was obtained (Figure 2).
In the control group, %DPPH inhibition (77.06%) and radical scavenging activity (64.42%) were measured, while the highest %DPPH inhibition was obtained in the Fulvic Acid 80 + Amino Acids 40 treatment (83.71%), and the highest radical scavenging activity was observed in the Amino Acids 40 + Chitosan 0.3 + Fulvic Acid 80 treatment (80.22%), indicating that these combinations most effectively enhanced the antioxidant capacity of parsley (Figure 2).
In the second trial, the phenolic content in the control group (414.5 mg GA 100 g FW-1) was lower than that of the Fulvic Acid 80 + Chitosan 0.3 and Amino Acids 40 + Chitosan 0.3 treatments, which both recorded the highest value (560.3 mg GA 100 g FW-1) (Figure 2).
The control group exhibited a flavonoid content of (523 mg RU 100 g FW-1), while the highest value was recorded in the Fulvic Acid 80 + Chitosan 0.3 treatment (539.9 mg RU 100 g FW-1); however, this difference was not statistically significant (Figure 2).
The Effects of Biostimulant Applications on the Mineral Profile of Parsley During the First Trial
The results of the mineral analyses conducted on parsley plants during the first trial are presented in Table 4. According to the findings, biostimulant applications had no statistically significant effect on the nitrogen (N) and potassium (K) contents of parsley plants. Nitrogen levels ranged between (2.25 and 3.25%), with the highest value recorded in the control group, while potassium concentrations varied from (3.08 to 4.39%), reaching the maximum in the Chitosan 0.6 treatment. Phosphorus (P) content, on the other hand, was significantly affected by the treatments, showing a decreasing trend in the biostimulant treatments compared to the control. The highest phosphorus level (0.34%) was found in the control group, while the lowest (0.17%) was observed in the Amino Acids 40 treatment. Calcium (Ca) and magnesium (Mg) contents were also significantly influenced by the biostimulant treatments. The highest Ca concentration (3.00%) was obtained from the Amino Acids 80 ppm application, whereas the maximum Mg content (0.86%) was detected in the Amino Acids 40 ppm treatment. Copper (Cu) concentration reached its highest values in the Fulvic Acid 80 and Amino Acids 40 applications, while manganese (Mn) and zinc (Zn) contents were highest in the control group. Iron (Fe) accumulation, however, was most pronounced in the Fulvic Acid 80 and Amino Acids 40 treatments. Statistically significant differences were observed among treatments for P, Ca, Mg, Cu, Mn, Zn, and Fe contents (Table 4).
| Treatments | N (%) | P (%) | K (%) | Ca (%) | Mg (%) | Cu (ppm) | Mn (ppm) | Zn (ppm) | Fe (ppm) |
| Control | 3.25 | 0.34 a | 3.08 | 2.15 c | 0.51 de | 8.25 c | 59.12 a | 83.62 a | 78.87 b |
| Fulvic Acid 80 | 3.21 | 0.19 cd | 3.37 | 2.33 bc | 0.66 bc | 14.00 a | 57.37 a | 76.37 ab | 126.87 a |
| Fulvic Acid 120 | 2.88 | 0.18 cd | 3.88 | 2.61 b | 0.70 bc | 8.00 c | 43.37 b | 35.12 c | 83.00 b |
| Amino Acids 40 | 2.53 | 0.17 d | 3.54 | 2.49 b | 0.86 a | 14.00 a | 57.37 a | 76.37 ab | 126.87 a |
| Amino Acids 80 | 2.25 | 0.23 b | 3.7 | 3.00 a | 0.79 ab | 12.12 ab | 58.00 a | 53.37 bc | 82.37 b |
| Chitosan 0.3 | 2.66 | 0.18 d | 3.47 | 2.47 b | 0.25 cd | 8.37 bc | 51.12 ab | 69.87 ab | 93.00 a b |
| Chitosan 0.6 | 2.76 | 0.22 bc | 4.39 | 2.53 b | 0.45 e | 8.37 bc | 53.75 ab | 67.12 ab | 86.62 b |
| LSD0.05 | NS | 0.41 | NS | 0.282 | 0.148 | 3.871 | 11.126 | 27.799 | 35.591 |
| p | 0.9133 | <0.0001 | 0.5029 | 0.0003 | 0.0003 | 0.0058 | 0.0337 | 0.02 | 0.007 |
Effects of Biostimulant Applications on the Mineral Profile of Parsley During the Second Trial
The mineral analyses of parsley plants in the second trial with biostimulant combinations are presented in Table 5. Nitrogen (N) content was significantly affected by the treatments. The highest nitrogen content was observed in the Amino Acids 40 + Fulvic Acid 80 + Chitosan 0.3 treatment (3.61%), while the lowest was found in the control treatment (1.70%). Magnesium (Mg) content was significantly influenced by the treatments, with the highest concentration (0.43%) recorded in the Fulvic Acid 80 + Amino Acids 40 treatment, and the lowest (0.13%) observed in the Amino Acids 40 + Fulvic Acid 80 + Chitosan 0.3 treatment. Manganese (Mn) content in parsley plants was not significantly affected by the biostimulant treatments. The Mn levels ranged from 56.50 mg kg-1 to 63.66 mg kg-1, with the highest value observed in the control group (63.66 mg kg-1). Other treatments, including Fulvic Acid 80 + Amino Acids 40, Fulvic Acid 80 + Chitosan 0.3, Amino Acids 40 + Chitosan 0.3, and Amino Acids 40 + Fulvic Acid 80 + Chitosan 0.3, showed similar manganese concentrations, with no significant differences. Overall, certain elements such as Nitrogen (N), Manganese (Mn), and Magnesium (Mg) were significantly influenced by the biostimulant treatments, whereas elements including Phosphorus (P), Potassium (K), Calcium (Ca), Iron (Fe), Copper (Cu), and Zinc (Zn) did not exhibit statistically significant differences.
| Treatments | N (%) | P (%) | K (%) | Ca (%) | Mg (%) | Cu (ppm) | Mn (ppm) | Zn (ppm) | Fe (ppm) |
| Control | 1.70 d | 0.21 | 3.32 | 2.34 | 0.29 bc | 8.0 | 63.66 a | 77.25 | 60.33 |
| Fulvic Acid 80 + Amino Acids 40 | 2.29 c | 0.18 | 2.78 | 3.05 | 0.43 a | 6.0 | 56.50 b | 77.25 | 52.5 |
| Fulvic Acid 80 + Chitosan 0.3 | 2.87 b | 0.18 | 3.31 | 2.52 | 0.41 ab | 7.6 | 59.25 ab | 87.75 | 49.25 |
| Amino Acids 40 + Chitosan 0.3 | 2.80 b | 0.2 | 3.22 | 2.64 | 0.21 c | 9.5 | 57.25 ab | 104.75 | 58.5 |
| Amino Acids 40 + Chitosan 0.3 + Fulvic Acid 80 | 3.61 a | 0.19 | 3.1 | 3.04 | 0.13 c | 8.0 | 60.75 ab | 95.5 | 45.25 |
| LSD0.05 | 0.297 | NS | NS | NS | 0.132 | NS | 7.045 | NS | NS |
| p | <0.0001 | 0.4154 | 0.685 | 0.528 | 0.0124 | 0.2677 | 0.3805 | 0.6073 | 0.7685 |
Discussion
Biostimulants represent an innovative and eco-friendly agricultural approach that enhances plant productivity, promotes growth, and improves product quality. These applications support sustainability in agricultural production by enabling the efficient use of natural resources. Additionally, by reducing the use of chemical fertilizers, they contribute to the protection of ecosystems and allow for the production of healthier products in the long term.
The uniform application of humic acid seed priming across all treatments was intended solely to improve germination uniformity and seedling quality and, therefore, is not considered to have influenced the comparative effects of the post-emergence biostimulant treatments.
The Effects of Biostimulant Applications on Growth Parameters of Parsley
When evaluating the results obtained from the first trial in terms of plant growth parameters, it was found that the Fulvic Acid 80 and Chitosan 0.3 biostimulant applications demonstrated more favorable effects compared to other treatments. Notably, the Fulvic Acid 80 + Chitosan 0.3 combination stood out in the second trial as well. These findings suggest that the combination of fulvic acid and chitosan may represent an effective biostimulant strategy to support the growth and development of parsley plants. Overall, based on the results of the growth parameters assessed in this study, it can be concluded that Fulvic Acid 80 and Chitosan 0.3 biostimulant applications are effective in promoting the development of parsley. These findings highlight the potential of biostimulant combinations to improve the total leaf yield of parsley. Pepper, radish, and cucumber plants exhibited increased growth and produced higher-quality fruit following the application of foliar chitosan. Chitosan is a naturally derived biopolymer that functions as an effective elicitor by activating plant defense mechanisms, thereby contributing to sustainable agricultural practices. Its beneficial effects are generally influenced by factors such as its concentration, the growth conditions, the method of preparation, and environmental variables.
The Effects of Biostimulant Applications on Nitrate Content and Antioxidant Activity in Parsley
The effects of different biostimulant applications on the nitrate levels, vitamin C content, antioxidant activities, and total phenolic and total flavonoid contents in parsley were evaluated. The results revealed significant variation in the responses to different biostimulant combinations. Biostimulants have a significant impact on secondary metabolite synthesis. Compounds such as alkaloids, phenylpropanoids, terpenoids, and phenolics contribute significantly to enhancing the plant’s resilience to both biotic and abiotic stress factors.
In terms of nitrate content, the control group exhibited the highest nitrate level, while certain biostimulant applications, particularly Fulvic Acid 120 and Amino Acids 40, significantly reduced nitrate content. Additionally, the Fulvic Acid 80 + Chitosan 0.3 combination was also found to notably decrease nitrate content. Among the applied biostimulants, some treatments and their respective dosages were capable of significantly reducing nitrate levels. Compared to the control plants, nitrate content was lower in all biostimulant applications during the first trial, except for the Fulvic Acid 80 treatment. The observed differences in nitrate content between the first and second trials in the control plants can be attributed to the seasonal timing of the experiments, with the first trial conducted in winter and the second trial in spring. The decrease in nitrate content with increasing daylight supports the close relationship between nitrate metabolism and light intensity.
In terms of vitamin C content, the control group exhibited a moderate level. However, the Fulvic Acid 80 application resulted in a decrease in vitamin C content, while the Fulvic Acid 120 application displayed the highest vitamin C content, suggesting that Fulvic Acid 120 may enhance vitamin C levels in parsley. Additionally, the Fulvic Acid 80 + Chitosan 0.3 combination showed higher vitamin C content compared to other combinations. Previous studies have also reported that different biostimulant applications can enhance vitamin C content, and the findings of the present study are consistent with those reported in the literature.
In this study, the Fulvic Acid 80 and Fulvic Acid 120 treatments exhibited slightly higher antioxidant activity, as measured by DPPH inhibition and radical scavenging assays, compared to the other applications. This finding suggests that these treatments may stimulate antioxidant defense mechanisms in parsley. Moreover, the Fulvic Acid 80 + Amino Acids 40 and Amino Acids 40 + Chitosan 0.3 + Fulvic Acid 80 combinations also displayed enhanced antioxidant activity, whereas the Amino Acids 40 treatment showed the lowest inhibition value, indicating that amino acid applications alone may have a limited effect on promoting antioxidant activity.
Regarding phenolic content, all biostimulant treatments led to an increase compared with the control group. In particular, the Amino Acids 40, Amino Acids 80, Chitosan 0.3, Fulvic Acid 80 + Chitosan 0.3, and Amino Acids 40 + Chitosan 0.3 treatments significantly enhanced phenolic accumulation. This outcome aligns with the widely accepted understanding that phenolic compounds play a crucial role in determining antioxidant capacity. In terms of flavonoid content, the Fulvic Acid 80 + Chitosan 0.3 combination resulted in the highest flavonoid level; however, the differences among treatments were not statistically significant, suggesting that biostimulant applications may have a limited effect on flavonoid accumulation.
Effects of Biostimulant Applications on the Mineral Profile of Parsley
In this study, the effects of different biostimulant applications on the mineral content of parsley were evaluated. Among the treatments, several biostimulants had significant effects on different minerals, while others showed no substantial impact.
The mineral analysis results demonstrated that biostimulant applications had varying effects on nutrient accumulation in parsley plants. While nitrogen and potassium contents were not significantly influenced, notable improvements were observed in other elements. Amino acid treatments, particularly Amino Acids 40 and Amino Acids 80, led to significant increases in calcium, magnesium, copper, and iron contents, highlighting their effectiveness in enhancing mineral uptake. Similarly, Fulvic Acid 80 improved copper and iron levels. Overall, amino acid-based treatments, especially at moderate doses, showed the most consistent positive impact on mineral enrichment in parsley. The results of the second trial demonstrated that biostimulant combinations had selective effects on the mineral content of parsley plants. Notably, the Amino Acids 40 + Fulvic Acid 80 + Chitosan 0.3 combination resulted in the highest nitrogen accumulation, suggesting a synergistic effect of these biostimulants on nitrogen uptake and assimilation. Conversely, the Fulvic Acid 80 + Amino Acids 40 and Fulvic Acid 80 + Chitosan 0.3 treatments showed the highest magnesium concentration, indicating treatment-specific responses in mineral absorption. However, other macro- and micronutrients such as phosphorus (P), potassium (K), calcium (Ca), copper (Cu), manganese (Mn), zinc (Zn), and iron (Fe) did not exhibit statistically significant changes across treatments. These findings indicate that although certain biostimulant combinations can enhance the accumulation of specific nutrients, their overall effect on the mineral profile may be limited, depending on the type and combination used, and generally tends to be negative. Previous studies conducted on various plant species have demonstrated that biostimulant applications can modify the dynamics of nutrient uptake and accumulation, generally leading to increased mineral content and improved metabolic efficiency. However, the findings of the present study indicate that these effects are not always positive and may vary depending on the type, dosage, and combination of biostimulants applied.
Overall, the findings of this study indicate that biostimulant applications can influence mineral uptake and accumulation in parsley. However, the magnitude and direction of these effects appear to vary depending on the type, concentration, and combination of biostimulants used. The absence of significant changes in certain mineral elements further suggests that these treatments may have selective rather than uniform impacts on the mineral composition of parsley.
Conclusions
This study clearly demonstrates that fulvic acid, amino acids, and chitosan can effectively improve the growth, productivity, and nutritional quality of parsley grown in hydroponic greenhouse conditions. Among the tested treatments, fulvic acid (80 ppm) and chitosan (0.3 mL L-1) stood out individually and in combination for their strong positive effects on leaf yield and overall plant vigor. Biostimulant application not only enhanced biomass accumulation but also enriched the nutritional profile of parsley by increasing total phenolic compounds and antioxidant capacity, while reducing leaf nitrate levels. These outcomes highlight the value of biostimulants as sustainable and environmentally friendly tools that can replace or complement conventional inputs, particularly in modern soilless cultivation systems. Overall, the consistent improvements observed in yield and nutritional quality indicate a promising role for biostimulant-based strategies in advancing hydroponic vegetable production.
Future research should focus on elucidating the physiological mechanisms underlying the observed responses to biostimulant applications, with particular emphasis on photosynthetic performance. In addition, evaluating the effectiveness of biostimulant combinations across different parsley cultivars and soilless growing systems would help to broaden their practical applicability. Further studies are also warranted to assess potential biostimulant residue dynamics in edible plant tissues, as well as their effects on key postharvest traits such as shelf life and freshness, in order to support both food safety and commercial relevance in hydroponic production systems.
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