Effect of organic manures and biofertilizers on herbage and essential oil yield of holy basil (Ocimum tenuiflorum L.)

Introduction

Revered as the "Queen of Herbs," holy basil scientifically known as Ocimum tenuiflorum L. It is a diploid (2n= 32) perennial shrub belonging to the Lamiaceae family. Beyond its profound spiritual and religious status, the plant is a cornerstone of traditional Eastern medicinal systems, including Ayurveda and Unani. Its therapeutic utility has been documented for centuries, most notably in the "Charaka Samhita". The species exhibits remarkable ecological adaptability, with a broad distribution spanning from the Andaman and Nicobar Islands to the high-altitude reaches of the Himalayas (up to 1800 m).Within the Indian subcontinent, commercial cultivation primarily focuses on two distinct morphotypes: Sri Tulsi: The more prevalent green-leaved variety and Krishna Tulsi: Distinguished by its purple foliage, this variety is highly prized in the pharmaceutical market for its greater concentration of bioactive compounds. Holy Basil serves as a versatile medicinal agent. Traditionally, it has been employed to address a diverse range of health concerns, including renal dysfunction, respiratory distress, digestive issues and parasitic infections. Modern scientific inquiry corroborates these traditional practices, identifying the plant's essential oils as potent sources of antioxidant, antimicrobial (antibacterial/antifungal) and insecticidal agents (Baranauskiene et al., 2003) [1] . The pharmaceutical industry's increasing preference for chemical-free raw materials has created a clear imperative to scale up organic production. Historically, the reliance on indiscriminate synthetic fertilization has led to soil nutrient imbalances and degradation, ultimately undermining crop productivity and medicinal quality. To ensure the long-term viability of holy basil cultivation, there is a strategic shift toward the use of organic amendments and biofertilizers, such as Azotobacter, Azospirillum, PSB and VAM. By enhancing the soil's physicochemical and biological properties, these inputs not only serve as a nutrient source but also optimize the efficiency of the soil-plant system, ensuring a sustainable, high-quality yield that meets rigorous industry standards.

Material and methods

A field trial was conducted over two consecutive seasons (2023-24 and 2024-25) at Nursery No. 4, College of Horticulture, DBSKKV, Dapoli. The study employed in FRBD with 16 distinct treatment combinations, replicated three times. The experimental treatments comprised a factorial arrangement of four organic manures and four biofertilizers as follows: Organic Manures (M): absolute control (M 1 ), FYM at 10 t/ha (M 2 ), vermicompost at 5 t/ha (M 3 ) and biogas slurry at 5 t/ha (M 4 ). Biofertilizers (B): PSB at 6 kg/ha (B 1 ), Azospirillum at 6 kg/ha (B 2 ), Azotobacter at 6 kg/ha (B 3 ) and VAM at 6 kg/ha (B 4 ). Standard agronomic practices for the cultivation of holy basil were strictly followed throughout the trial. Yield parameters, specifically fresh herbage yield (g/plant and kg/hectare) and essential oil yield (g/plant and kg/hectare), were measured at two specific harvest intervals: 90 DAT and 150 DAT. All collected data were subjected to statistical analysis using SAS software, with the analysis of variance (ANOVA) performed in accordance with the methodology outlined by Panse and Sukhatme (1965) [2] .

Results and discussion

1 Fresh herbage yield per plant (g) 1.1 Fresh herbage yield per plant at first harvest Data presented in Table 1 indicated that the organic manures, biofertilizers and their synergistic application significantly influence the fresh herbage yield per plant at first harvest, across two consecutive seasons (2023-24 and 2024-25) along with pooled data.

Treatment M₃ consistently proved to be the most effective and recorded significantly the highest fresh herbage (262.54 g, 274.89 g and 268.72 g) during the years 2023-24, 2024-25 and pooled analysis, respectively and superior over rest of the treatments. In contrast, treatment M 1 exhibited significant treatment variations and recorded the lowest fresh herbage yields across the same periods, pulling in just 72.17 g, 82.26 g and 77.22 g.

Effect of biofertilizers on fresh herbage yield per plant at first harvest of holy basil (B)

At first harvest, the treatment B₂, resulted in the highest fresh herbage yield (197.06 g, 207.88 g and 202.47 g), demonstrating superiority over all other treatments. Conversely, treatment B 1 exhibited the lowest fresh herbage yield (174.16 g, 185.35 g and 179.76 g) during 2023-24, 2024-25 and in pooled analysis, respectively.

The highest fresh herbage yield per plant (274.56 g, 286.60 g and 280.58 g) was recorded under M₃B₂ which was significantly higher than all other treatments, while the lowest fresh herbage yield (67.64 g, 77.66 g and 72.66 g) was noticed in M₁B₁ during 2023-24, 2024-25 and in pooled analysis.

Fresh herbage yield per plant at second harvest

Table 2 presents a comprehensive dataset illustrating the impact of organic manures, biofertilizers and their combined application on fresh herbage yield per plant at second harvest over two successive seasons (2023-24 and 2024-25), along with pooled data analysis.

At second harvest, treatment M₃ was the most effective treatment, achieving a highest fresh herbage yield (138.74 g, 150.84 g and 144.79 g) which significantly outperformed the remaining treatments during 2023-24, 2024-25 and in pooled analysis. In contrast, treatment M₁ recorded the lowest herbage yield of 35.81 g, 47.61 and 41.71 g highlighting significant treatment variations. This finding underscores the significant treatment variations observed in the study.

Effect of biofertilizers on fresh herbage yield per plant at second harvest (B)

At second harvest, the treatment B 2 achieved the highest fresh herbage yield (103.13 g, 115.20 g and 109.16 g) this was statistically significant, with a superior outcome compared to all other treatments. In contrast, treatment B 1 resulted in the lowest fresh herbage yield of 90.99 g, 102.59 g and 96.79 g during 2023-24, 2024-25 and in pooled analysis.

At second harvest, the highest fresh herbage yield (144.68 g, 157.51 g and 151.10 g) was recorded under M₃B₂ which was significantly superior over other treatments, while the lowest fresh herbage yield (33.71 g, 44.53 g and 39.13 g) was noticed in M₁B₁ during 2023-24, 2024-25 and in pooled analysis.

2 Fresh herbage yield per hectare (kg) 2.1 Fresh herbage yield per hectare at first harvest The data on the impact of organic manures, biofertilizers and their combined application on fresh herbage yield per hectare at first harvest including results from two consecutive seasons (2023-24 and 2024-25) and pooled data are presented in Table 3.

At first harvest, treatment M₃ emerging as the most effective treatment. This treatment achieved a highest fresh herbage yield per hectare of 7295.14 kg, 7637.73 kg and 7466.44 kg during 2023-24, 2024-25 and in pooled analysis, respectively. In contrast, treatment M₁ recorded the lowest fresh herbage yield per hectare of 2004.63 kg, 2284.72 kg and 2144.68 kg highlighting significant variations among treatments during 2023-24, 2024-25 and in pooled analysis, respectively.

Effect of bio fertilizers on fresh herbage yield per hectare at first harvest of holy basil (B)

The findings of the experimental trial indicated that the treatment B₂ was the most efficacious in enhancing fresh herbage yield per hectare, achieving a highest value (5473.38 kg, 5775.46 kg and 5624.43 kg) during 2023-24, 2024-25 and in pooled analysis, respectively. This value was statistically highest over other treatments. Conversely, treatment B 1 exhibited the lowest fresh herbage yield per hectare (4839.12 kg, 5149.31 kg and 4994.22 kg) during 2023-24, 2024-25 and in pooled analysis, respectively highlighting the significance of biofertilizer application.

The data collected during 2023-24, 2024-25 and pooled analysis indicated that highest fresh herbage yield per hectare (7629.63 kg/ha, 7962.97 kg and 7796.30 kg, respectively) was recorded under M₃B₂ while the lowest fresh herbage yield per hectare (1879.63 kg/ha, 2157.41 kg/ha and 2018.52 kg/ha, respectively) was noticed in M₁B₁.

Fresh herbage yield per hectare at second harvest

The data on the impact of organic manures, biofertilizers and their combined application on fresh herbage yield per hectare at second harvest including results from two consecutive seasons (2023-24 and 2024-25) and pooled data are presented in Table 4.

At second harvest, treatment M₃ emerging as the most effective treatment. This treatment achieved the highest fresh herbage yield per hectare (3854.17 kg, 4188.66 kg and 4021.41 kg) during 2023-24, 2024-25 and in pooled analysis, respectively. In contrast, treatment M1 recorded the lowest fresh herbage yield per hectare (994.21 kg, 1321.76 kg and 1157.99 kg) highlighting significant variations among treatments during 2023-24, 2024-25 and in pooled analysis, respectively. (Table 4).

Effect of biofertilizers on fresh herbage yield per hectare at second harvest of holy basil (B)

The findings of the first trial (2023-24), second trial (2024-25) and pooled analysis indicated that treatment B2 was the most efficacious in enhancing fresh herbage yield per hectare at second harvest, achieving a highest value of 2863.43 kg/ha, 3199.07 kg/ha and 3031.25 kg/ha, respectively. This value was statistically highest over other treatments. Conversely, treatment B₁ exhibited the lowest fresh herbage yield per hectare (2527.78 kg, 2848.38 kg and 2688.08 kg) during 2023-24, 2024-25 and in pooled analysis, respectively highlighting the significance of biofertilizer application.

At second harvest, collected data indicated that highest fresh herbage yield per hectare (4018.52 kg, 4375.00 kg and 4196.76 kg) was recorded under M₃B₂ during 2023-24, 2024-25 and in pooled analysis, respectively. This treatment was found significantly superior over other treatments, while the lowest fresh herbage yield per hectare (935.19 kg, 1236.11 kg and 1085.65 kg) was noticed in M₁B₁ during 2023-24, 2024-25 and in pooled analysis, respectively.

Table 5 provides a data of the influence of organic manures, biofertilizers and their synergistic application on essential oil yield per plant at first harvest, across two consecutive seasons (2023-24 and 2024-25) along with pooled data.

At first harvest, the treatment M₃ emerged as the most effective treatment among other treatments, achieving a highest essential oil yield (1.445 g, 1.493 g and 1.469 g) during 2023-24, 2024-25 and in pooled analysis, respectively which was significantly superior over other treatments. Conversely, treatment M1 recorded the lowest essential oil yield of 0.177 g, 0.231 g and 0.204 g highlighting significant treatment variations during 2023-24, 2024-25 and in pooled analysis, respectively.

Effect of biofertilizers on essential oil yield per plant at first harvest of holy basil (B)

At first harvest, the both experimental trial and pooled data demonstrated that treatment B₂ resulted in the highest essential oil yield of 0.950 g, 1.001 g and 0.976 g respectively, demonstrating superiority over all other treatments. Conversely, treatment B 1 exhibited the lowest essential oil yield of 0.719 g, 0.779 g and 0.749 g during 2023-24, 2024-25 and in pooled analysis, respectively.

The data collected during the first trial, second trial and pooled analysis indicated that highest essential oil yield (1.660 g, 1,710 g and 1.686 g, respectively) was recorded under M 3 B 2 which was significantly higher than all other treatments while the lowest essential oil yield (0.169 g, 0.217 g and 0.193 g) during 2023-24, 2024-25 and in pooled analysis, respectively was noticed in M₁B₁.

Essential oil yield per plant at second harvest

Table 6: represents a comprehensive dataset illustrating the impact of organic manures, biofertilizers and their combined application on essential oil yield per plant at second harvest over two successive seasons (2023-24 and 2024-25), along with pooled data analysis.

At second harvest, the treatment M₃ was the most effective treatment, achieving a highest essential oil yield (0.776 g, 0.805 g and 0.791 g) during 2023-24, 2024-25 and in pooled analysis, respectively which significantly outperformed the remaining treatments. In contrast, treatment M₁ recorded the lowest essential oil yield of 0.095 g, 0.124 g and 0.110 g, highlighting significant treatment variations during 2023-24, 2024-25 and in pooled analysis, respectively.

Effect of biofertilizers on essential oil yield per plant at second harvest (B)

The examination of first trial, second trial and pooled data revealed that treatment B₂, which included Azospirillum at 6 kg/ha achieved the highest essential oil yield (0.515 g, 0.542 g and 0.529 g, respectively). This was statistically significant, with a superior outcome compared to all other treatments. In contrast, treatment B₁ resulted in the lowest essential oil yield of 0.385 g, 0.413 g and 0.399 g during 2023-24, 2024-25 and in pooled analysis, respectively.

The data collected during 2023-24, 2024-25 and pooled data indicated that highest essential oil yield (0.892 g, 0.922 g and 0.907 g, respectively) was recorded under M₃B₂ which was significantly superior over other treatments while the lowest essential oil yield (0.084 g, 0.112 and 0.099 g) was noticed in M₁B₁ during 2023-24, 2024-25 and in pooled analysis, respectively. (Table 6).

4 Essential oil yield per hectare (kg) 4.1 Essential oil yield per hectare at first harvest The data on the impact of organic manures, biofertilizers and their combined application on essential oil yield per hectare at first harvest including results from two consecutive seasons (2023-24 and 2024-25) and pooled data are presented in Table 7.

The first trial (2023-24), second trial (2024-25) and pooled data yielded notable results, with treatment M₃ emerging as the most effective treatment. This treatment achieved a highest essential oil yield per hectare of 40.14 kg, 41.48 kg and 40.81 kg, respectively. In contrast, treatment M₁ recorded the lowest essential oil yield per hectare of 4.93 kg, 6.41 kg and 5.67 kg during 2023-24, 2024-25 and pooled data highlighting significant variations among treatments.

Effect of biofertilizers on essential oil yield per hectare at first harvest of holy basil (B)

The findings of the both experimental trials (2023-24 and 2024-25) and pooled data indicated that treatment B₂ was the most efficacious in enhancing essential oil yield per hectare, recorded the highest value of 26.38 kg, 27.81 kg and 27.10 kg, respectively. This value was statistically highest over other treatments. Conversely, treatment B₁ exhibited the lowest essential oil yield per hectare 19.97 kg, 21.63 kg and 20.80 kg during 2023-24, 2024-25 and in pooled analysis, respectively highlighting the significance of biofertilizer application.

The data collected during 2023-24, 2024-25 and pooled analysis indicated that the highest essential oil yield per hectare (46.11 kg, 47.50 kg and 46.81 kg, respectively) was recorded under M 3 B 2 while the lowest essential oil yield per hectare (4.70 kg, 6.02 kg and 5.36 kg) was noticed in M₁B₁ during 2023-24, 2024-25 and in pooled analysis, respectively.

Essential oil yield per hectare at second harvest

The data on the impact of organic manures, biofertilizers and their combined application on essential oil yield per hectare at second harvest including results from two consecutive seasons (2023-24 and 2024-25) and pooled data are presented in Table 8.

At second harvest, the treatment M₃ emerging as the most effective treatment and recorded the highest essential oil yield per hectare of 21.56 kg, 22.37 kg and 21.97 kg during 2023-24, 2024-25 and in pooled analysis, respectively. In contrast, treatment M₁ exhibited the lowest essential oil yield per hectare of 2.66 kg, 3.44 kg and 3.05 kg during 2023-24, 2024-25 and in pooled analysis, respectively highlighting significant variations among treatments.

Effect of biofertilizers on essential oil yield per hectare at second harvest of holy basil (B)

The findings of the first trial (2023-24), second trial (2024-25) and pooled data indicated that treatment B₂ was the most efficacious in enhancing essential oil yield per hectare, achieving a highest value of 14.32 kg, 15.06 kg 14.69 kg, respectively and found significantly superior over other treatments. Conversely, treatment B1 exhibited the lowest essential oil yield per hectare 10.71 kg, 11.47 kg and 11.09 kg during 2023-24, 2024-25 and in pooled analysis, respectively.

At second harvest, the data collected during the first trial, second trial and pooled data indicated that the highest essential oil yield per hectare (24.77 kg, 25.61 kg and 25.19 kg, respectively) was recorded under M₃B₂, while the lowest essential oil yield per hectare (2.36 kg, 3.10 kg and 2.74 kg) was noticed in M 1 B 1 during 2023-24, 2024-25 and in pooled analysis, respectively. (Table 8).

Discussion

Vermicompost significantly boosts the fresh herbage yield by improving soil structure, aeration and water-holding capacity, creating an ideal environment for vegetative growth. Its rich profile of essential macro and micronutrients, combined with beneficial microorganisms and plant growth-promoting substances like phytohormones and humic acids, enhances soil fertility and enzymatic activity. Furthermore, vermicompost stimulates root development and increases the efficiency of nutrient uptake; notably, it promotes the accumulation of IAA by inhibiting IAA oxidase, which directly drives superior biomass production and sustained crop yield. The results of the present study are consistent with those reported by Kumar (2007) [3] in lemon tulsi, Prabu and Arumugam (2013) [4] in Japanese mint, Raina et al. (2013) [5] in Ocimum sanctum L., Abaszadeh et al. (2016) [6] in Lavandula angustifolia, Cheena et al. (2021) [7] in Ocimum basilicum L. and Beleri et al. (2024) [8] in tulsi.

Effect of biofertilizers on total fresh herbage yield of holy basil (B)

The increase in total fresh herbage yield per plant might be attributed to the enhanced bioavailability and subsequent uptake of essential nutrients by the plants. Azospirillum stimulates plant growth by producing various phytohormones and facilitating biological nitrogen fixation; this increased nutrient availability enables the plant to produce a greater number of branches, leaves and inflorescences, ultimately resulting in a higher total fresh herbage yield per plant. These findings align with the results reported by Kandeel et al. (2002) [9] in sweet basil as well as Manjunatha et al. (2002) [10] in patchouli.

This superior performance is driven by the synergistic effect of the two inputs: vermicompost optimizes the rhizosphere's physical and chemical properties, while Azospirillum enhances biological nitrogen fixation. The resulting increase in nitrogen availability supports essential protein synthesis, cell division and enlargement, which together maximize key growth parameters like plant height, leaf count and branching, ultimately leading to significant increases in total biomass accumulation. These results are in conformity with the findings of Hemalatha and Suresh (2012) [11] as well as Sanjutha et al. (2008) [12] in kalmegh.

Essential oil biosynthesis is intrinsically linked to photosynthetic activity, as it requires a high expenditure of ATP and NADPH to generate terpenoid precursors like IPP and DMAPP. In this study, the application of vermicompost at 5 t/ha significantly increased essential oil content by boosting the net photosynthetic rate, thereby enhancing both precursor availability and downstream enzymatic activity. This process is further supported by the presence of humic acids, plant growth regulators and beneficial rhizospheric microorganisms in the vermicompost, which promote structural growth such as increased leaf count and inflorescence number that correlates directly with higher volatile oil accumulation across leaves, stems and inflorescences. These results align with the findings of Jayasri and Anuja (2010) [13] in sweet basil, Raina et al. (2013) [5] in Ocimum sanctum L. Abaszadeh et al. (2016) [6] in Lavandula angustifolia, Cheena et al. (2021) [7] in Ocimum basilicum L. and Beleri et al. (2024) [8] in tulsi.

Effect of biofertilizers on essential oil yield of holy basil (B)

This improvement stems from a combination of increased vegetative biomass and higher oil concentration in leaf tissues, driven by the biofertilizer's ability to stimulate the endogenous production of growth regulators like IAA, gibberellic acid and kinetin, which upregulate secondary metabolic pathways. Furthermore, Azospirillum improves the mobilization and uptake of nitrogen and phosphorus, fuelling profuse vegetative growth by accelerating the conversion of photosynthates into structural proteins and amino acids, ultimately creating a more robust foundation for essential oil accumulation. The present results are in conformity with the findings of Manjunatha et al. (2002) [10] in patchouli (Pogostemon cablin) and Aboud et al. (2021) [14] in lavender (Lavandula officinalis L.).

The superior essential oil yield achieved through the integrated application of vermicompost and Azospirillum stems from their synergistic ability to optimize the rhizosphere and enhance nutrient bioavailability. By improving soil physical properties, water-holding capacity and nutrient uptake efficiency, this combination supports vigorous vegetative growth including increased leaf area, plant height and branching which is tightly correlated with higher volatile oil accumulation in aromatic species like holy basil. Furthermore, the ample nitrogen supply fuelled by these inputs promotes chlorophyll synthesis and metabolic activity, facilitating the production of essential carbohydrates and amino acids that serve as precursors for phytohormones and secondary metabolites. Ultimately, this integrated approach creates an optimal physiological environment that maximizes both biomass production and the biosynthetic capacity for essential oils. The findings of this research align closely with the results Kumar (2009) [15] in stevia, Rajamanickam et al. (2011) [16] in mint, Abhilasha Srivastava (2017) [17] in kalmegh, Yashaswini et al. (2020) [18] in tulsi.

Conclusion

Based on the two-year experimental results, it could be concluded that the application of vermicompost at 5 t/ha significantly enhanced the herbage and essential oil yield of holy basil. With respect to biofertilizers, treatment B₂ (Azospirillum at 6 kg/ha) enhanced the herbage and essential oil yield of holy basil. The combined effect of vermicompost at 5 t/ha + Azospirillum at 6 kg/ha (M₃B₂) significantly enhanced the herbage and essential oil yield of holy basil, outperforming other treatment combinations.

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