Influence of pre-sowing treatments on seed quality parameters in Elaeocarpus floribundus.

Introduction

The olive species, Elaeocarpus floribundus (E. floribundus) (family: Elaeocarpaceae), is an evergreen tree of modest stature with a wide crown and a clean trunk ranging from 12 to 16 meters, celebrated for its fruit and recognized as a valuable medicinal plant native to India.

Elaeocarpus floribundus exhibits significant therapeutic potential, with various plant partsincluding the leaves, fruit, bark and roots-traditionally utilized in disease management. The species demonstrates notable biological activities, particularly antioxidant and antibacterial properties, which are attributed to a rich profile of bioactive phytochemicals, predominantly phenolic compounds. Morphologically, the fruit contains a hard, three-celled stone housing spindle-shaped seeds that exhibit recalcitrant behavior despite their rigid endocarp. E. floribundus typically thrives on hill slopes and ridges across sandy-to-clay soil compositions. Commercially, the edible fruits of the Indian olive (Elaeocarpus floribundus) are primarily processed into pickles, while its timber serves as a suitable raw material for light interior construction, plywood, particle board, fiberboard and paper pulp production. The species is primarily propagated via seeds; however, cultivation is significantly hindered by extended seed dormancy, which results in low germination rates and delayed seedling development. Species within the genus Elaeocarpus are predominantly evergreen trees or shrubs, though a minority exhibit briefly deciduous, epiphytic, or lianescent growth habits. Leaves are simple (strictly unifoliolate) and arranged alternately, featuring prominent venation, frequently serrated margins and a distinct pulvinus at the petiole base, characteristically turning red prior to abscission. The inflorescences are typically racemose, bearing mostly bisexual flowers with 4-5 sepals and petals, alongside numerous stamens; the petals are notably distinguished by finely divided, linear lobes. The resulting fruit is a spherical-to-oval drupe, typically blue or black in color, enclosing a characteristically sculptured endocarp. Given the paucity of data regarding seed biology in Elaeocarpus floribundus, this study investigates the efficacy of various pre-sowing treatments on the species' seeds. Although seeds represent the primary natural mechanism for plant reproduction, genetic conservation, transport and species propagation, viable seeds frequently fail to germinate under optimal environmental conditions due to seed dormancy [5] . This physiological inhibition is governed by internal mechanisms-such as seed coat impermeability, embryo immaturity, or endogenous biochemical inhibitors-which directly restrict germination rates [7] . Overcoming these physiological barriers requires species-and lot-specific pre-treatment protocols tailored to the precise mechanism of dormancy involved. Seed dormancy presents a primary barrier to successful germination, which can be mitigated through targeted presowing treatments tailored to the specific dormancy mechanism of the seed. Standard methods include mechanical or acid scarification, hot or cold water soaking, chemical application, plant growth regulators, and alternate wetting and drying cycles. Recent technological advancements have expanded traditional seed processing to include innovative techniques such as seed priming, gamma irradiation, and magnetic field exposure to enhance germination rates. Furthermore, nanotechnology has emerged as a promising frontier in accelerating seed germination. Given that a key bottleneck in mass propagation and species conservation is the lack of standardized protocols for many tree species, this review highlights both traditional and advanced seed pre-treatment methodologies along with their functional significance.

Material and Methods

Study site and growing media: The present study was carried out during 2023 in laboratory of Department of Forest Biology and Tree improvement, College of Forestry, Ponnampet, Kodagu district of India. Mature seeds of Elaeocarpus floribundus were harvested from a sacred grove (Devarakadu) located near Ponnampet which lies between 12° 11' 60'' N latitude and 75° 33' 36'' E, elevation of 867 above MSL. To minimize non-treatment variation, uniform-sized seeds were selected for the study.

Experimental design and treatment combinations:

The pre-sowing treatment experiments for Elaeocarpus floribundus were carried out at the College of Forestry, Ponnampet. Germination assays were conducted in polythene trays using a 1:1 (v/v) sand and topsoil substrate. The study followed a Completely Randomized Design (CRD) comprising six treatments, including a control, with four replications per treatment. Each replication utilized 100 seeds to systematically evaluate the influence of the presowing treatments on germination performance. The presowing experiment comprised six distinct treatment protocols. In T 0 (control), seeds received no chemical or physical treatment. In 𝑇 1 , seeds were soaked in a 4% sulfuric acid (H 2 SO 4 ) solution for 24 hours. Seeds in T 2 were immersed in boiling water for 2 minutes followed by a 24hour soak in ambient-temperature water. T 3 involved soaking seeds in ambient-temperature water for 24 hours. For 𝑇 4 , seeds were immersed in a 4% hydrogen peroxide (H 2 0 2 ) solution for 24 hours. In T 5 , seeds were subjected to boiling water immersion for 2 minutes only. For T 6 , seeds were soaked in a 350ppm gibberellic acid (GA 3 ) solution for 24 hours. Following their respective pre-treatments, seeds across all groups were sown in a sand medium for germination evaluation. The experimental treatment details are summarized below:  T0: Control  T1: H 2 SO 4 4% for 24 hrs  T2: Soaking in boiling water for 2 min and then in normal water for 24 hrs  T3: Soaking in water for 24 hrs  T4: H 2 O 2 4% for 24 hrs  T5: Soaking in boiling water for 2 min  T6: GA 3 350ppm for 24 hrs

Germination percentage

Pre-treated seeds were placed in a seed germinator maintained at 30 ∘ C and 90% relative humidity. The count of normal seedlings was recorded on the 30th day postsowing under controlled laboratory conditions. Germination percentage was subsequently calculated using the following equation:

Shoot length

Ten normal seedlings were randomly selected from each of the four replications per treatment. Shoot length was measured using a standard graduated scale and the mean shoot length was computed and expressed in centimeters (cm).

Root length

At the time of the final germination count, ten normal seedlings were randomly sampled from each of the four replications per treatment. Root length was measured using a standard graduated scale and the mean root length was calculated and expressed in centimeters (cm).

Seedling dry weight

Seedling dry weight was evaluated following the final germination count on day 30. Seedlings were dried in a hot air oven, after which the dried biomass for each replication was weighed to determine the mean seedling dry weight, expressed in grams.

Vigour Indices

Seed vigour reflects the total properties that determine the potential for rapid, uniform emergence and robust seedling development under diverse environmental conditions. The seed vigour index was calculated according to the formula proposed by Abdul-Baki and Anderson (1973) and expressed as a whole number.

Vigour Index-I It was calculated by using following formula:

Standard germination (%) × Average seedling length (cm) Vigour index-II It was calculated by using following formula:

Standard germination (%) × Average seedling dry weight (g) Statistical Analysis: Data were subjected to statistical analysis to assess treatment variations using Microsoft Excel and AGRISTAT software. Analysis of Variance (ANOVA) was performed in accordance with the experimental design, following the methodology described by Panse and Sukhatme (1967) [13] .

Results and Discussion: Germination period: Seed germination initiated on day 22 post-sowing and extended through day 34. Pre-sowing treatments significantly influenced the germination timeframe. The shortest imbibition period to germination onset (22 days) was recorded in treatmentsT 3 , T 4 , T 5 , andT 6 , whereas the control group (T 0 ) exhibited significantly delayed germination, requiring the longest imbibition period of 26 days (Table 1).

Germination percentage:

The perusal of results presented reveals that germination was significantly affected by all the treatments except control, soaking in water for 24 hours, soaking in H 2 O 2 4% for 24 hours and soaking in GA 3 350ppm for 24 hours. The highest germination percentage was recorded in seeds treated with 4% H 2 0 2 for 24 hours, followed by those soaked in water for 24 hours. Conversely, complete germination failure occurred in seeds exposed to 4% concentrated H 2 SO 4 , boiling water for 2 minutes, and boiling water (2 minutes) combined with a 24-hour water soak. The detrimental effect of H 2 SO 4 likely stems from acid penetration through the exposed micropyle, which damages internal embryonic tissues (Ells, 1963) [5] . The enhanced germination observed under 4% H 2 0 2 , 24-hour water soaking, and 350 ppm GA 3 can be attributed to seed coat softening, scarification and increased coat permeability. Structural modifications to the seed coat facilitate water imbibition into the embryo and cotyledons, thereby promoting radicle emergence.

Germination value and germination energy:

Germination values varied significantly across treatments, ranging from 0.6 to 1.9. The highest germination value was recorded in treatment T 4 (1.9), followed by T 3 and T 6 (both 1.2), while the lowest was observed in the control group (T 0 , 0.6). Similarly, germination energy peaked in T 3 (25%), followed by T 6 (23.25%) and was lowest in T 0 (13.75%) (Table 1).

Maximum shoot and root lengths (8.1 cm and 2.0 cm, respectively) were recorded in seeds treated with 4% hydrogen peroxide for 24 hours (T 4 ), which performed significantly better than all other treatments (Table 2). This was closely followed by water soaking for 24 hours (T 3 ; 7.6 cm shoot / 1.3 cm root) and 350 ppm GA 3 for 24 hours (T 6 ; 7.3 cm shoot / 1.4 cm root). The superior performance under H 2 O 2 and GA 3 treatments may be attributed to enhanced nutrient availability and the physiological activity of endogenous growth promoters during early seedling development. These findings align with those of Krishna et al. (2011) [9] and Ajay et al. (2022), who observed that presowing organic and growth regulator treatments significantly enhanced seedling vigor, shoot height and biomass accumulation in hard-coated seed species like Melia dubia.

The information (Table 2) demonstrated that various presowing treatments on seed had a substantial impact on the dry weight of Elaeocarpus floribundus seedlings. The highest seedling dry weight (0.13 g/10 seedling) was recorded in seeds treated with 4% hydrogen peroxide for 24 hours (T 4 ), performing significantly higher than all other treatments. This was followed by 24-hour water soaking (T 3 ) and 350 ppm GA 3 for 24 hours (T 6 ), both yielding 0.12 g/10 seedlings.

Seedling Vigour Index-I and Vigour Index-II

The Vigour Index-I of Elaeocarpus floribundus seedlings was strongly impacted by several pre-sowing treatments, (Table 2). The highest Vigour Index I (505.43) was recorded in seeds treated with 4% hydrogen peroxide for 24 hours (T 4 ), which was significantly superior to all other treatments. This was followed by seeds soaked in water for 24 hours (T 3 ; 406.2), 350 ppm GA 3 for 24 hours (T 6 ; 235.23) and the untreated control (T 0 ; 92.1). A similar trend was observed for Vigour Index II in Elaeocarpus floribundus across the pre-sowing treatments. Treatment T 4 (4% H 2 O 2 for 24 hours) recorded the highest Vigour Index II (6.60), significantly outperforming all other treatments. This was followed by 24-hour water soaking (T 3 ;5.83), 350 ppm GA 3 for 24 hours (T 6 ;3.29) and the untreated control (T 0 ;1.60).

Discussion:

Pre-sowing treatments are widely recognized for their capacity to enhance seed germination parameters (Hossain, 2005). The natural germinability of Elaeocarpus floribundus seeds is typically low (<50%), with hydrogen peroxide soaking often proving to be the most effective treatment for breaking dormancy (Halder et al., 2023) [6] . Consistent with these observations, the present study demonstrates that soaking E. floribundus seeds in 4% H 2 O 2 for 24 hours substantially increased germination speed, final germination percentage, seedling growth, and biomass production relative to the control. These results closely mirror the findings of Halder et al. (2023) [6] , who reported peak germination in E. floribundus using 5% H 2 O 2 , as well as Niamjit Das (2014) [12] , who noted enhanced germination following acid scarification with H 2 SO 4 .

Conclusion

Exploring potential indigenous flora as alternative industrial raw materials directly supports their conservation by preserving genetic diversity and safeguarding native species against endangerment, thereby enhancing regional ecological sustainability. Standardization of mass propagation protocols for native species like Elaeocarpus floribundus represents a crucial initial step toward sustainable laboratory and commercial production.

Ultimately, this study demonstrates that pre-sowing seed treatments are a key determining factor in significantly increasing the germination percentage and early seedling performance of E. floribundus.

Even though seed propagation is simpler, we might not always have an abundance of seeds (Hussain et al., 2018) [7] .

The second biggest obstacle to seed germination is its viability, which indicates the extent to which a seed is alive, metabolically active and possesses enzymes that can catalyze the metabolic reactions required for germination. Seed viability is likely at its peak during physiological maturity, after which it gradually decreases (Copeland & McDonald, 2012) [4] . Additionally, some seeds display seed dormancy, which is one of the major obstacle to seed germination. A physiological state known as "seed dormancy" occurs when a fertile seed fails to germinate even under ideal circumstances. It prevents an intact, viable seed from fully germinating. Based on the pre-sowing seed treatment evaluation in Elaeocarpus floribundus, it can be concluded that a 24-hour soak in 4% hydrogen peroxide (H 2 O 2 ) yields superior germination performance under laboratory conditions. Furthermore, key seedling growth metrics-including shoot length, root length, dry weight, and vigour indices-were significantly maximized by this treatment. Consequently, pre-treating seeds with 4% H 2 O 2 for 24 hours is recommended for nursery managers and growers to enhance the germination efficiency and early growth performance of E. floribundus.

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