Growth and Polyphenol Content of Ashwagandha (Withania somnifera L.Dunal) under Combined Moisture Stress and Salicylic Acid Treatment

Growth and Polyphenol Content of Ashwagandha (Withania somnifera L.Dunal) under Combined Moisture Stress and Salicylic Acid Treatment | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Growth and Polyphenol Content of Ashwagandha (Withania somnifera L.Dunal) under Combined Moisture Stress and Salicylic Acid Treatment M. S. Jayathilaka, Siripala Subasinghe, KMW Rajawatta, K. H.T. Karunarathna This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4689316/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 10 You are reading this latest preprint version Abstract Ashwagandha is a medicinal herb that has very high demand in the global market. Root is the main part harvested from this plant because of its rejuvenating properties. This study investigates the effects of moisture stress and salicylic acid on the growth yield and polyphenol content of Ashwagandha. A polyhouse experiment was conducted where Ashwagandha plants were grown under four levels of soil moisture that included 50%, 60%, 80%, and 100% of field capacity (FC) and four Salicylic acid (SA) levels as 10 mM,1 mM,0.1 mM and control was applied as a foliar spray as treatments to induce the secondary metabolite production. Plant height, number of leaves, number of fruits, number of fallen leaves, SPAD value and leaf area, root dry of the root, free proline content and total polyphenol content were measured. The highest root dry weight (7.62g) and total polyphenol content (3.78 GAE mg/g d.w.b) was reported with the interaction effect of 50% of FC and 10 − 3 M SA application. Under the combined application of 50% of FC and 1 mM SA the dry weight and total polyphenol content was increased by 2.7 folds and 12.49 folds respectively compared to control (100% FC and without SA). These findings suggest that a moisture stress level of 50% field capacity combined with a 1 mM SA foliar spray can significantly enhance the root yield and polyphenol content of Ashwagandha. Drought stress Moisture stress Polytunnel Root yield Salicylic acid 1 Introduction Ashawagandha ( Withania Somnifera L. Dunal) is a medicinal plant, also known as Indian Ginseng and Amukkara, belonging to family Solanaceae. This woody shrub, which grows to a maximum height of 150 cm [ 1 ] thrives in semi tropical with annual rainfall of 500 to 750 mm and the temperature is between 20°C to 38°C, is cultivated as a late rainy season crop [ 2 ] .The plant is widely used in ayurveda, Yunani medicine due to medicinal properties including anti- inflammatory, anti-microbial, anti-tumor, anti-stress, and anti-diabetic properties [ 3 ].The extract of Ashwagandha is a complex mixture of phenolic compounds and flavonoids. Withanolides are the active pharmacological compounds of Ashwagandha which is a series of naturally occurring steroids containing a lactone with a side chain of nine carbons, attached to C-17 [ 4 ]. Further, Withaferin A, an active ingredient with the anticancer activity is well distributed in leaves, bark and stems in addition to the roots of Ashwagandha [ 5 ].The global market for ashwagandha is growing rapidly due to the rapid increase in use of Ashwagandha as supplements, functional food and beverages and pharmaceuticals. According to market research, the market for ashwagandha is forecasted to grow with a Compound annual growth rate (CAGR) of 11.6% in the period of 2023 to 2030 and expected to reach USD 115502.48 by 2030. Ashwagandha root product consisting of 5% withanolides (in dry weight basis) is expected to dominate the global market as it is widely used tonic to reduce stress and boost [ 6 ]. The plant has been categorized under ‘threatened category’ of IUCN due to over exploitation caused by higher demand [ 7 ]. Plant secondary metabolites are sources of medicines, food additives and many other industrially important biochemicals. The biosynthesis of these secondary metabolites can be enhanced by deliberately exposing medicinal plants to drought stress. This can be achieved by manipulating the moisture levels, light intensities and by exogenous application of plant growth regulators including Salicylic acid, Methyl Jasmonate [ 8 ]. For example, severe water stress has been shown to significantly increase secondary metabolite production in Satureja hortensis and Hypericum Brasiliense [ 9 ]. Under drought stress conditions, plants close the stomata to prevent water loss by evapotranspiration reducing CO 2 intake through stomata. Correspondingly, CO 2 fixation by Calvin cycle decreases, leading to extensive decrease in consumption of reduction equivalents (NADPH + H + ). The oversupply of NADPH + H + induce the production of reduced compounds such as alkaloids, isoprenoids and phenols which are active compounds in plants [ 10 ].Salicylic acid as a plant growth regulator, induces the plant response to many biotic and abiotic stresses and elicit the production of secondary metabolites in plants [ 11 ]. Similarly, in Mint ( M.spicata) , total phenol and flavonoid content was significantly increased when treated with 200 µg ml − 1 of salicylic acid (SA) [ 12 ]. In most studies, the treatment of SA to stimulate secondary metabolite production is only limited to in vitro studies of Ashwagandha. Though in vitro studies have proven their potentials inducing secondary metabolite production upon treatments of elicitors, real world application is limited due requirement of skilled labor, higher maintenance cost and the genetic stability of in vitro cultures [ 13 ]. The open field cultivation results in larger variability in yields, as both biomass production and production of secondary metabolites are affected by many factors including genotype, climate, soil type, management practices and pests and diseases. Inhouse Hydroponic technology can be used as a strategy to overcome this problem of inconsistent yields. Hydroponic technologies provide optimum growing conditions and can be used to stimulate secondary metabolism by appropriate manipulation of mineral nutrition [ 14 ]. In this study, the individual effects and interaction effect of moisture stress and SA on the growth dynamics and secondary metabolite production in Ashwagandha was investigated. It was hypothesized that different moisture levels would have a significant effect on the plant's growth, yield, and total polyphenolic content (TPC), as certain degree of moisture stress could stimulate an adaptive mechanism in plants leading to increased synthesis of polyphenols which are the active compounds in Ashwagandha. Further, it was speculated that the foliar application of SA, a plant growth regulator, would not only influence the plant’s stress response mechanisms but also enhance its growth and secondary metabolite production, particularly under varying moisture conditions. A significant interaction between the levels of moisture stress and SA treatments was expected, proposing that this interaction might affect the plant's growth parameters and secondary metabolite production in a synergistic manner. The most severe level of moisture stress, when optimally managed with SA application, is expected to yield the highest production of polyphenols, illustrating the complex balance between stress induction and metabolic enhancement in Ashwagandha. This research can contribute to optimizing growth and yield of Ashwagandha which will ensure consistent supply of high-quality products in global market. The increased production of Ashwagandha can meet the growing global demand for pharmaceuticals, herbal health supplements and functional foods. In addition, this study provides strategies which can be applied to enhance growth and yield other medicinal plants, thereby protecting biodiversity. Through this study, we aimed to identify effective agronomic strategies that can be employed to optimize the medicinal value of Ashwagandha, catering to the growing demand for high-quality medicinal plants. Therefore, the objectives of the study were to investigate the growth, yield and polyphenols content of Ashwagandha as affected by different moisture levels and foliar application of different concentrations of Salicylic acid. 2 Materials and methods 2.1 Experimental site The study was conducted in a 139.35 m 2 protected house in Faculty of Technology, University of Ruhuna, Sri Lanka. The experimental area was in the WL2B climate zone, which is a low-lying wet zone (60 06 'N 80022 'E). 2.2 Experimental treatments Ashwagandha plants were transplanted in coir dust bags after one month of nursery. Recommended Fertilizer schedule was applied for all plants [15]. There were three replicates for each treatment combination and 16 treatment combinations (4*4), arranged in split plot design. One coir dust bag with two plants was considered as one experimental unit. Two-month-old plants (one month after transplanting) were subjected to moisture stress and salicylic acid stress. Four moisture stress levels as 100% field capacity (FC) (control), 80% of FC, 60 % of FC and 50 % of FC were maintained using automated drip irrigation system with XH-M214 humidity controller soil sensor modules. Foliar spray of Salicylic acid was applied to the plants in concentrations of 10 mM, 1 mM, 0.1 mM and 0 mM (control) once in two weeks. 2.3 Data collection 2.3.1 Growth and yield parameters The plant height (cm) from the pot media surface to the plant tip was measured using a meter scale, while the number of leaves per plant was counted manually [16]. The plants were uprooted for root harvesting when the berries turn red to orange and leaves become dry after 150 -180 days of transplanting [17].The SPAD value was measured using Konica Minolta, SPAD 502 Plus Chlorophyll Meter after 60 days of treatment application from 3 rd fully grown leaf from the apex of each plant. Individual leaf area (mm 2 ) was measured by using ADC BioScientific Leaf Area Meter AM350 Ver.1.00 from top three fully grown leaves of the plants at the end of harvesting. At the harvesting, the number of fruits and nodes from fallen leaves were counted. After root harvesting dry weights were measured by oven drying at 72 °C until a constant weight was obtained [16]. 2.3.2 Free proline content The third fully grown leaf was used to extract free proline following the acid ninhydrin method. First the fresh leaf samples were grind using liquid nitrogen and 0.1 g of sample was weighed. Then the leaf sample was homogenized with 3% aqueous sulfosalicylic acid using vortex mixture and 200 µL supernatant was harvested. Then the supernatant solution was vortexed with 200 μL acid ninhydrin and 200 μL glacial acetic acid. The solution was incubated in a 100°C water bath for a period of one hour and immediately transferred to an ice bath to stop the reaction. After cooling, 400 μL toluene was added, and vortexed the samples. After the mixture cooled to room temperature 100 μL chromophore was diluted with 900 μL toluene. Consequently, the absorbance of the separated upper layer was measured at 520 nm wavelength using toluene as blank. A standard curve was plotted to calculate the final concentration of free proline (mg proline g –1 fresh weight) [18] . 2.3.3 Total poly phenolic content - Folin-Ciocalteu method (modified ISO 14502-1) First, 5.00 ml of 70% methanol (at 70 °C) was added to 0.2 g of dried and finely ground root sample. The mixed sample was heated at 70°C for 10 min, while mixing in 5 min intervals. The samples were centrifuged at 3500 rpm for 10 min after reaching room temperature. The supernatant was decanted to a 10.00 ml volumetric flask. The extraction was repeated for residue with fresh 5.00 ml of 70% methanol. The extracts were combined and top up to 10 .00 ml by adding cold 70% methanol. The sample (5.00 ml) was diluted up to 25.00 ml with distilled water for polyphenol analysis. Anhydrous gallic acid standard series (10 µg/ml, 20 µg/ml, 30 µg/ml, 40 µg/ml and 50 µg/ml) was prepared. 10% of Folin-Ciocalteu phenol reagent (5.00 ml) was added into 1.00 ml of gallic acid standard series, and 1.00 ml of sample. 7.5 % Sodium carbonate solution (4.00 ml) was added and mixed within 3 min to 8 min after the addition of the Folin-Ciocalteu phenol reagent. All mixtures were allowed to stand at room temperature for 60 min. The absorbance was measured in 10mm path length cells against blank using the spectrophotometer at 765 nm. The total polyphenol content (TPC), expressed as a percentage by mass on a sample dry matter basis, is given by the formula: 2.4 Data analysis Collected data were analyzed using two-way analysis of variance (ANOVA) and the mean values were compared using Duncan test significant difference test at P≤0.05. The test was carried out using SAS© for academics software and IBM SPSS software. 3 Results Table 1 Significance levels in two-way ANOVA of the effect of moisture stress and salicylic acid on growth, yield and total polyphenol content of Ashwagandha Source of variance Moisture stress effect (main factor) Salicylic acid stress (main factor) Interaction effect (Moisture stress* Salicylic acid stress) Growth parameters Plant height * * NS Number of leaves per plant * * NS Number of fallen leaves per plant * * NS Number of fruits per plant * * * SPAD value * * * Leaf area * * Yield parameter Root dry weight * * * Biochemical parameters Total polyphenol content * * * Free proline content * * * *, and ns represent significant (P ≤ 0.05), and not significant, respectively The of the effect of moisture stress and salicylic acid on growth, yield and biochemical parameters (free proline content and total polyphenol content) of Ashwagandha is presented in Table 1. Moisture stress and SA eac individually influenced the growth parameters: plant height, number of leaves per plant and number of fruits per plant. The interaction effect of SA and moisture stress treatments were significant for the parameters including number of fallen leaves, number of fruits, SPAD value, root dry weight and total polyphenol content and free proline content. As shown in Fig 1, the highest plant height, number of leaves per plant and leaf area was achieved under 100% FC and 1 mM of SA. Upon application of moisture stress the plant height was decreased by 12.02%, 14.3% and 24.11% in 80% FC, 60% FC and 50% FC respectively compared to 100% FC. The number of leaves decreased with a maximum reduction observed at 50% FC by 47.85% relative to 100% FC. When the plants were undergoing moisture stress level 50% FC the leaf area was reduced by 42.63% compared to 100% FC (Fig 1). Upon receiving the treatment 1 mM SA plant height, number of leaves per plants and leaf area was increased by 21.08 %, 16.5% and 47.99 % respectively relative to control (0 mM SA). The lowest plant height, number of leaves per plant and leaf area was reported in the salicylic acid level 10 mM (Fig 1). The highest number of leaves fell under the interaction effect of treatments 50% FC and 10 mM SA while the lowest was reported with interaction effect of 100% FC and 1 mM SA (Fig 2). The highest SPAD value was observed in the Ashwagandha plants received combined application of 100 % FC and 1mM SA which was 17.14% higher than control (100%FC and 0 mM SA). The plants treated with 10 mM SA had had significantly higher SPAD values than control with dark green, shrunken which was evident with lowest leaf area given in Fig 1. The lowest SPAD value was observed in the plants treated with 50 % FC and without SA application. The significantly highest number of fruits per plant was observed under the combined treatment 60% FC and 1mM SA while the lowest was recorded with the application of 80% FC and 10 mM SA. The significantly highest root dry weight (7.62g) and total polyphenol content (3.78 (GAE mg/g d.w.b) of the plant was recorded with the interaction effect of treatments 50% FC and 1 mM SA. The root dry weight was increased by 2.7 folds under the interaction effect of treatment 50% FC and 1 mM SA relative to 100% FC and 0 mM SA (control). The lowest root dry weight reported at 100% FC and 10 mM SA was 1.33 folds lesser than the control. The TPC of plants undergone the treatment 50% FC and 1 mM SA showed an increment of 12.49 folds relative to that of control. Highest free content of 150.72 µg/g FW was reported under the combined application of 50% FC and 1mM SA while the lowest was reported in the control (100 % FC and 0 mM SA) (Fig 3). The morphological and biochemical parameters, number of fruits per plant, root dry weight and free proline showed a positive correlation with total polyphenol content while leaf area had a negative relationship to total polyphenol content (Table 02). The root dry weight was positively influenced by total polyphenol content, free proline content and number of fruits per plant. Table 02 Correlation between morphological and biochemical parameters of Ashwagandha under moisture stress and salicylic acid treatment Plant height Number of leaves per plant Number of fallen leaves per plant Number of fruits per plant SPAD value Leaf Area Root dry weight Free proline content Total polyphenol content Plant height 1 0.48 * 0.08ns 0.23 * 0.27 * 0.19ns 0.05ns -0.23* -0.12ns Number of leaves per plant 0.48 * 1 -0.10ns 0.11ns 0.26 * 0.21 * -0.12ns -0.33* -0.17ns Number of fallen leaves per plant 0.08ns -0.10ns 1 0.08ns 0.10ns -0.26 * 0.09ns 0.38* 0.01ns Number of fruits per plant 0.23 * 0.11ns 0.08ns 1 -0.06ns 0.13ns 0.43 * 0.17ns 0.39 * SPAD value 0.27 * 0.26 * 0.10ns -0.06ns 1 -0.009ns -0.36 * -0.34* -0.17ns Leaf Area 0.19ns 0.21 * -0.26 * 0.13ns -0.009ns 1 0.10ns -0.50* -0.28 * Root dry weight 0.05ns -0.12ns 0.09ns 0.43 * -0.36 * 0.10ns 1 0.23* 0.39 * Free proline content -0.23 * -0.33 * 0.38 * 0.17ns -0.34 * -0.50 * 0.23 * 1 0.33 * Total polyphenol content -0.12ns -0.17ns 0.01ns 0.39 * -0.17ns -0.28 * 0.39 * 0.33* 1 *, and ns represent significant (P ≤ 0.05), and not significant, respective 4 Discussion Water stress is the most significant abiotic elicitor, which affects medicinal plant growth and active ingredients by altering biochemical and physiological characteristics. Many studies revealed that moisture stress has a positive role in augmenting plant secondary metabolite production in various medicinal plants [ 19 ].In present study, the moisture stress had a negative effect on plant growth (plant height, number of leaves, SPAD value and leaf area) which is in fair agreement with reduced plant height with the application of drought in Rosmarinus officinalis L [ 20 ], Satureja hortensis L [ 21 ] and Trachyspermum ammi l. [ 22 ]. In Portulaca oleracea L , the plant height was decreased by 18.5% and 45% under 60% FC and 90% FC, respectively and the number of leaves decreased with a maximum reduction observed at 30% FC by 51.3% compared to the control plants. In the same study the chlorophyll A content was reduced by 23.6% and 43.8% under 60 FC and 30% FC [ 23 ].The decrease in number of leaves per plant and leaf area observed in plants is a strategic adaptation manifested by plants to reduce water loss by transpiration. The physiological damage occurred in plants due to drought was evident by the degraded morphological characteristics of plant including plant height, number of leaves and SPAD value [ 24 ]. During the present study, the higher number of leaves (Fig. 1 ) recorded in 100% FC and higher number of fallen leaves (Fig. 2 ) recorded in 50% FC were also indicators of drought stress undergone by the plant. Similarly, leaf area Ashwagandha was reduced by 27.4% and 34.4% when subjected to mild and severe water stress in a study conducted in Gujarat, India. In the same study. Chlorophyll content was also reduced by 60% in the severe water stress conditions which was in line with present study [ 25 ]. The increased production of Reactive Oxygen Species due to drought conditions, leads to oxidative stress in plant cells. Oxidative stress cause disintegration and reduction of chlorophyll content in plants thereby reducing photosynthesis rate[ 26 ] . With the increase of SA level, the leaf area was decreased while increasing the number of fallen leaves in Ashwagandha in our study. Though the number of fallen leaves increased with increasing moisture stress and SA concentration, new leaves were developed under the stress conditions. The leaf senescence was higher with the increase of substrate moisture stress. The reduction of shoot parts (plant height) with the increase of drought stress level could be due to the decrease in number and area photosynthetic organs specifically the leaves[ 27 ]. Under stressful conditions, plants promote leaf senescence to increase the translocation of essential nutrients to growing tissues of the plant [ 28 ]. It seems that with increase in the drought stress level, the plant height and number of leaves decreased in response to the reduction in the leaf area and photosynthetic pigments [ 20 ]. During our study, a significant increment of SPAD value was observed in plants which received 1mM SA treatment in both 60% FC and 50% FC (Fig. 2 ). This result agrees with those reported [ 29 ] who found that foliar application of 1 mM SA increased plant height, chlorophyll content in purslane plant under drought conditions. Moreover, [ 30 ] showed that foliar application of 1 mM SA increased Chlorophyll a, b and c content significantly in Conocarpus erectus and Populus deltoides plants under 60% FC and 30% FC moisture stress conditions. One of the mechanisms plants responses to any stress condition, including drought, is ensuring survival of species by fruiting and flowering. This fact was proven during the study as the number of fruits increased under treatments 60% FC and 1 mM SA[ 31 ]. SA is a plant hormone that can ameliorate drought stress by stimulating the production of antioxidants and other defense compounds in plants, as well as improving the photosynthesis mechanism [ 32 ]. Many studies have demonstrated that SA can efficiently stimulate secondary metabolite production in plants [ 33 ].From the results of the present study, it was evident that SA has a significant positive impact growth of Ashwagandha. The present results agree with [ 12 ] who observed significantly enhanced plant growth, biochemical properties and antioxidant activity compared to control plants of Mentha spicata with the application of 200 µgml − 1 (1.44 mM) SA. Low concentrations of SA decrease the negative impacts of drought stress on plant growth as it enhances the drought stress tolerance of plants—however, high concentrations of SA result in detrimental effects on plants [ 34 ]. The SA concentration of 10 mM cannot be recommended for foliar spray as the plants treated and it did not show any significant difference with that of control. SA increases plant growth under drought stress by preventing reduction of cytokinin and auxin, which induce cell division of apical root meristem and plant growth [ 35 ]. Root is the most common plant part harvested from Ashwagandha because of its rejuvenated properties. The results of increased root dry weight were consistent with findings of [ 25 ],where authors reported increased root weight under moderate drought stress. Similarly, in Rosemary plants, root yield was highest under 60% FC. In the same study, root yield was reported to be highest in SA treatment 2 mM [ 20 ]. In a similar study where root dry weight was increased when SA was foliar sprayed to purslane ( Portulaca oleracea L .) seedlings under drought stress, there was no significant effect on root dry weight when SA was applied under non drought stress conditions[ 23 ]. Plants increase the root biomass and growth in response with early drought stress signals to penetrate deeper soils to explore more water to keep shoot in hydrated condition. These can be the reasons for the increase of dry weight observed in our study. Higher branching and more fibrous roots were observed in plants which received the moisture stress treatments 100% FC and 80% FC while the plants under treatment 50%FC had good quality roots with higher biomass similar to [ 25 ]. In the Stellaria dichotoma L.var laceolata Bge increase in root biomass was reported until a water stress level of 60–70% FC and then decreased with afterwards [ 19 ].The increased root dry mass due to the undergone drought stress was observed in many plants including Ashwagandha and tall Fescue Cultivars [ 25 ],[ 36 ]. This observation was in accordance with the theory of functional balance which states “that plants will respond to a limited water availability by increasing the flow of assimilates to the root leading to an increased root dry mass ratio”[ 25 ]. Plants treated with Salicylic acid levels 1mM and 0.1mM) under the moisture stress level 50% FC showed significantly more root dry weight when compared to combined application of 50% FC and without SA (Fig. 3 ). This data showed that SA application in low concentration could effectively improve plant tolerance and overcome the adverse conditions induced by the limited water availability. Many similar observations have been reported in sweet basil plants [ 26 ], Conocarpus erectus and Populus deltoides [ 30 ] under drought conditions. SA can stimulate root growth by promoting cell division and elongation when plants undergo stress conditions. Foliar application of SA performs the same function when plant is under moisture stress. This is important for drought-stressed plants, as a larger root mass can aid in absorption of more water and nutrients from the soil [ 30 ].The metabolic changes triggered by drought stress results in accumulation of higher amount of secondary metabolites in medicinal plants which also can been a reason for the increased root dry weight[ 10 ]. SA has a positive regulation of plant responses for drought stress as SA induces stomatal closure to maintain water content in plant leaves [ 34 ]. This proline buildup is particularly enhanced by higher concentrations of SA (1 mM and 10 mM), particularly in situations of acute water stress (60% and 50% FC). This suggests that SA may increase the plant's resistance to drought by activating biochemical pathways that are involved in the synthesis of proline. As field capacity (FC) drops from 100–50%, proline concentration in plants increases, confirming its function as an osmoprotectant that aids in maintaining turgor and stabilizing cellular structures under stressful situations. These results are consistent with earlier studies, including one by [ 37 ], which showed that SA efficiently stimulates proline synthesis and contributes to stress signalling and defensive responses in plants exposed to different degrees of water stress. Particularly under some circumstances, higher SA concentrations are particularly more effective in eliciting these reactions. In the present study, there was a significant increment in TPC under interaction effect of application of 10mM Salicylic acid under moisture stress of 50% FC. Similarly, Portulaca oleracea L . showed a significant increase in total phenol when 1mM SA was foliar sprayed under drought stress, in comparison to plants treated with drought stress alone [ 23 ].[ 38 ] stated that the interaction effects of 1 mM SA application and increased irrigation interval significantly enhanced TPC of milk thistle leaves. These phenolic compounds make the plants resistant to drought stress by reducing ROS production. The increase TPC is considered to be a result of increasing enzymes involve in shikimate pathway due to water stress and SA application. Besides, SA induces phenylalanine ammonia-lyase activity, which is a major enzyme involved in the early stages of biosynthesis phenolic compounds [ 39 ]. According to[ 38 ] the levels of sucrose and total soluble carbohydrate (TSC) content increased significantly in Silybum marianum L . in response to the increase in drought stress. Sucrose and TSC decrease plant water potential in order to increase the gradient of water potential between soil and plant tissue which facilitated water flow from soil to root via osmosis. On the other hand, TSC and sucrose can stimulate production of secondary metabolites like phenols. In the shikimic acid pathway phenolic compounds are synthesized using soluble carbohydrates during drought stress [ 38 ]. 5 Conclusion This study on Ashwagandha under different moisture stress levels and Salicylic Acid treatments has demonstrated a complex interaction between abiotic stress and plant physiological responses, particularly in relation to growth and secondary metabolite production. The findings showed that while moisture stress negatively impacts plant growth in terms of height, leaf number, SPAD values, and leaf area, it simultaneously triggers an adaptive mechanism enhancing secondary metabolite production, as evidenced by the significant increase in total polyphenolic content. The application of SA further modulates this response, optimizing plant growth and secondary metabolite production, particularly under severe moisture stress conditions. The Salicylic acid application gradually reduced the decremental effects of drought conditions. From the results of the present study foliar spray of 1mM SA can be recommended to enhance the root yield, plant growth and secondary metabolite production of Ashwagandha plants under the moisture stress of 50% field capacity. This research highlights the potential of using agronomic strategy, which consists with controlled moisture stress and SA application, to enhance the medicinal value of Ashwagandha. By controlling environmental stressors and growth regulators, it is possible to optimize conditions for maximizing the yield and quality of medicinal compounds, to address the growing global demand for high-quality medicinal plants without expanding cultivated land area. Moreover, these findings contribute to the understanding of plant stress physiology, offering insights into how plants can be engineered to thrive under stress while enhancing their pharmacological potential. This balance between stress management and metabolic enhancement opens new avenues for agricultural practices aimed at sustainable cultivation of medicinal plants, ensuring the availability of key pharmacological compounds and supporting the health and wellness industry. Declarations Funding No funding was received for conducting this study. Competing interests The authors declare no competing interests Data availability (data transparency) All the data are presented in tables and figures in the manuscript and further inquiries can be directed to the corresponding authors. Code availability Not applicable Ethical Statement The seeds used in this study was obtained from Prof. Siripla Subasinghe (one of authors) Faculty of Agriculture, University of Ruhuna, Sri Lanka. Therefore, the plants used in the study comply with national guidelines without further affirmation. Contribution M.S.Jayathilaka : Conceptualization, Methodology, Investigation, Review and editing , Data analysis, Data interpretation, Writing-original draft Siripala Subasinghe, KMW Rajawatta , K.H.T. Karunarathna : Conceptualization, Methodology, Review and editing,supervision, validation All authors read and approved the final manuscript. 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Konrad, “Increased biomass and salicylic acid elicitor activity in fennel (Foeniculum vulgare Miller),” Brazilian Journal of Food Technology, vol. 20, 2017, doi: 10.1590/1981-6723.17216. M. Kundu, S. Halder, and A. Bhattacharjee, “Salicylic acid-induced modulation of growth and metabolism of a medicinal plant mentha spicata l,” Int J Pharm Sci Res, vol. 9, no. 12, pp. 5294–5300, 2018, doi: 10.13040/IJPSR.0975-8232.9(12).5294-00. V. Tomar, N. Das, H. Chauhan, P. Roy, and D. Sircar, “Closed polybag foliar methyl-jasmonate treatment: New technology for rapid enhancement of bioactive withanolide biosynthesis in field-grown plants of Withania somnifera,” Ind Crops Prod, vol. 162, Apr. 2021, doi: 10.1016/j.indcrop.2021.113262. R. Maggini, C. Kiferle, L. Guidi, A. Pardossi, and A. Raffaelli, “Growing medicinal plants in hydroponic culture,” Acta Hortic, vol. 952, pp. 697–704, 2012, doi: 10.17660/ActaHortic.2012.952.88. B. Dmapr, R. S. Jat, R. N. Reddy, R. Bansal, and P. Manivel, “Good Agricultural Practices for Ashwagandha Good Agricultural Practices for Ashwagandha Compiled by,” 2015. [Online]. Available: www.dmapr.org.in S. Udpuay, H. Ullah, S. K. Himanshu, R. Tisarum, S. Cha–um, and A. Datta, “Drought tolerance screening of okra genotypes in relation to growth and physio–biochemical traits at the vegetative stage,” Genet Resour Crop Evol, 2023, doi: 10.1007/s10722-023-01689-3. H. T. Chandranath and V. Naik, “Impact of Planting Dates and Stage of Harvesting on Yield and Economics of Ashwagandha (Withania sominifera Dunal.),” 2012. [Online]. Available: www.rjas.info A. U. Alam et al., “Seed Priming and Foliar Application of Salicylic Acid is Equally Beneficial in Mitigating Drought Stress in Cucumber,” J Soil Sci Plant Nutr, 2023, doi: 10.1007/s42729-023-01485-z. W. Zhang et al., “Effect of water stress on roots biomass and secondary metabolites in the medicinal plant Stellaria dichotoma L. var. lanceolata Bge,” Sci Hortic, vol. 224, pp. 280–285, Oct. 2017, doi: 10.1016/j.scienta.2017.06.030. B. Abbaszadeh, M. Layeghhaghighi, R. Azimi, and N. Hadi, “Improving water use efficiency through drought stress and using salicylic acid for proper production of Rosmarinus officinalis L.,” Ind Crops Prod, vol. 144, Feb. 2020, doi: 10.1016/j.indcrop.2019.111893. Z. F. Baher, M. Mirza, M. Ghorbanli, and M. B. Rezaii, “The influence of water stress on plant height, herbal and essential oil yield and composition in Satureja hortensis L,” Flavour Fragr J, vol. 17, no. 4, pp. 275–277, 2002, doi: 10.1002/ffj.1097. N. Azhar, B. Hussain, M. Yasin Ashraf, and K. Yar Abbasi, “Medicinal Plants: Conservation & Sustainable use) Water stress mediated changes in growth, physiology and secondary metabolites of desi ajwain (trachyspermum ammi l.,” 2011. F. Saheri, G. Barzin, L. Pishkar, M. M. A. Boojar, and L. Babaeekhou, “Foliar spray of salicylic acid induces physiological and biochemical changes in purslane (Portulaca oleracea L.) under drought stress,” Biologia (Bratisl), vol. 75, no. 12, pp. 2189–2200, Dec. 2020, doi: 10.2478/s11756-020-00571-2. J. Pradhan, S. K. Sahoo, S. Lalotra, and R. S. Sarma, “Positive impact of abiotic stress on medicinal and aromatic plants,” Int J Plant Sci, vol. 12, no. 2, pp. 309–313, Jul. 2017, doi: 10.15740/has/ijps/12.2/309-313. S. Shah, R. Saravanan, and N. A. Gajbhiye, “Phytochemical and physiological changes in ashwagandha (withania somnifera dunal) under soil moisture stress,” Brazilian Journal of Plant Physiology, vol. 22, no. 4, pp. 255–261, 2010, doi: 10.1590/S1677-04202010000400005. S. Kordi, M. Saidi, and F. Ghanbari, “Induction of Drought Tolerance in Sweet Basil (Ocimum basilicum L) by Salicylic Acid,” International Journal of Agricultural and Food Research, vol. 2, no. 2, Apr. 2013, doi: 10.24102/ijafr.v2i2.149. S. Nogué and N. R. Baker2, “Effects of drought on photosynthesis in Mediterranean plants grown under enhanced UV-B radiation,” 2000. S. Munné-Bosch and L. Alegre, “Die and let live: Leaf senescence contributes to plant survival under drought stress,” Functional Plant Biology, vol. 31, no. 3. pp. 203–216, 2004. doi: 10.1071/FP03236. F. Saheri, G. Barzin, L. Pishkar, M. Mashhadi, A. Boojar, and L. Babaeekhou, Foliar spray of salicylic acid induces physiological and biochemical changes in purslane (Portulaca oleracea L.) under drought stress, doi: 10.2478/s11756-020-00571-2/Published. Z. Zafar, F. Rasheed, R. M. Atif, M. A. Javed, M. Maqsood, and O. Gailing, Foliar application of salicylic acid improves water stress tolerance in conocarpus erectus l. And populus deltoides l. saplings: Evidence from morphological, physiological and biochemical changes, Plants, vol. 10, no. 6, Jun. 2021, doi: 10.3390/plants10061242. Y. Nakajima, S. Susanto, and K. Hasegawa, Influence of Water Stress in Autumn on Flower Induction and Fruiting in Young Pomelo Trees (Citrus grandis (L.) Osbeck), 1993. J. González-Villagra, M. M. Reyes-Díaz, R. Tighe-Neira, C. Inostroza-Blancheteau, A. L. Escobar, and L. A. Bravo, Salicylic Acid Improves Antioxidant Defense System and Photosynthetic Performance in Aristotelia chilensis Plants Subjected to Moderate Drought Stress, Plants, vol. 11, no. 5, Mar. 2022, doi: 10.3390/plants11050639. B. Ali, Salicylic acid: An efficient elicitor of secondary metabolite production in plants,Biocatalysis and Agricultural Biotechnology, vol. 31. Elsevier Ltd, Jan. 01, 2021. doi: 10.1016/j.bcab.2020.101884. S. Yuan and H.-H. Lin, Role of Salicylic Acid in Plant Abiotic Stress, 2008. [Online]. Available: http://www.znaturforsch.com E. Sánchez-Rodríguez, D. A. Moreno, F. Ferreres, M. D. M. Rubio-Wilhelmi, and J. M. Ruiz, Differential responses of five cherry tomato varieties to water stress: Changes on phenolic metabolites and related enzymes, Phytochemistry, vol. 72, no. 8, pp. 723–729, Jun. 2011, doi: 10.1016/j.phytochem.2011.02.011. S. Gao et al., Effects of drought stress on growth, physiology and secondary metabolites of Two Adonis species in Northeast China, Sci Hortic, vol. 259, Jan. 2020, doi: 10.1016/j.scienta.2019.108795. M. I. R. Khan, M. Fatma, T. S. Per, N. A. Anjum, and N. A. Khan, Salicylic acid-induced abiotic stress tolerance and underlying mechanisms in plants, Front Plant Sci, vol. 6, no. JUNE, Jun. 2015, doi: 10.3389/fpls.2015.00462. A. Estaji and F. Niknam, Foliar salicylic acid spraying effect’ on growth, seed oil content, and physiology of drought-stressed Silybum marianum L. plant, Agric Water Manag, vol. 234, May 2020, doi: 10.1016/j.agwat.2020.106116. M. I. R. Khan, M. Fatma, T. S. Per, N. A. Anjum, and N. A. Khan, Salicylic acid-induced abiotic stress tolerance and underlying mechanisms in plants, Front Plant Sci, vol. 6, no. JUNE, Jun. 2015, doi: 10.3389/fpls.2015.00462. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Editorial decision: Revision requested 04 Sep, 2024 Reviews received at journal 28 Jul, 2024 Reviews received at journal 25 Jul, 2024 Reviewers agreed at journal 24 Jul, 2024 Reviewers agreed at journal 22 Jul, 2024 Reviewers agreed at journal 22 Jul, 2024 Reviewers invited by journal 22 Jul, 2024 Editor assigned by journal 12 Jul, 2024 Submission checks completed at journal 11 Jul, 2024 First submitted to journal 04 Jul, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Jayathilaka","email":"data:image/png;base64,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","orcid":"","institution":"Department of Biosystems Technology, Faculty of Technology,University of Ruhuna","correspondingAuthor":true,"prefix":"","firstName":"M.","middleName":"S.","lastName":"Jayathilaka","suffix":""},{"id":333239409,"identity":"179ea78e-7bca-4af0-b4b9-233d94f41431","order_by":1,"name":"Siripala Subasinghe","email":"","orcid":"","institution":"Department of Crop Science, Faculty of Agriculture, University of Ruhuna","correspondingAuthor":false,"prefix":"","firstName":"Siripala","middleName":"","lastName":"Subasinghe","suffix":""},{"id":333239410,"identity":"81281fdb-c7f3-43a6-8f92-569bcc645fd4","order_by":2,"name":"KMW Rajawatta","email":"","orcid":"","institution":"Department of Biosystems Technology, Faculty of Technology,University of Ruhuna","correspondingAuthor":false,"prefix":"","firstName":"KMW","middleName":"","lastName":"Rajawatta","suffix":""},{"id":333239411,"identity":"e8b41206-1059-47b2-96d9-8c15cf6b575d","order_by":3,"name":"K. H.T. Karunarathna","email":"","orcid":"","institution":"University of Ruhuna, Karagoda-Uyangoda","correspondingAuthor":false,"prefix":"","firstName":"K.","middleName":"H.T.","lastName":"Karunarathna","suffix":""}],"badges":[],"createdAt":"2024-07-05 03:21:24","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4689316/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4689316/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":61762070,"identity":"9c269c02-d1f3-427d-bc46-a0ee6da986ec","added_by":"auto","created_at":"2024-08-05 09:37:13","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":475044,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4689316/v1/8db61482-2a8b-4bfe-b387-9b4f6b43ddcc.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Growth and Polyphenol Content of Ashwagandha (Withania somnifera L.Dunal) under Combined Moisture Stress and Salicylic Acid Treatment","fulltext":[{"header":"1 Introduction","content":"\u003cp\u003eAshawagandha (\u003cem\u003eWithania Somnifera\u003c/em\u003e L. Dunal) is a medicinal plant, also known as Indian Ginseng and Amukkara, belonging to family Solanaceae. This woody shrub, which grows to a maximum height of 150 cm [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e] thrives in semi tropical with annual rainfall of 500 to 750 mm and the temperature is between 20\u0026deg;C to 38\u0026deg;C, is cultivated as a late rainy season crop [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e] .The plant is widely used in ayurveda, Yunani medicine due to medicinal properties including anti- inflammatory, anti-microbial, anti-tumor, anti-stress, and anti-diabetic properties [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e].The extract of Ashwagandha is a complex mixture of phenolic compounds and flavonoids. Withanolides are the active pharmacological compounds of Ashwagandha which is a series of naturally occurring steroids containing a lactone with a side chain of nine carbons, attached to C-17 [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. Further, Withaferin A, an active ingredient with the anticancer activity is well distributed in leaves, bark and stems in addition to the roots of Ashwagandha [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e].The global market for ashwagandha is growing rapidly due to the rapid increase in use of Ashwagandha as supplements, functional food and beverages and pharmaceuticals. According to market research, the market for ashwagandha is forecasted to grow with a Compound annual growth rate (CAGR) of 11.6% in the period of 2023 to 2030 and expected to reach USD 115502.48 by 2030. Ashwagandha root product consisting of 5% withanolides (in dry weight basis) is expected to dominate the global market as it is widely used tonic to reduce stress and boost [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. The plant has been categorized under \u0026lsquo;threatened category\u0026rsquo; of IUCN due to over exploitation caused by higher demand [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e].\u003c/p\u003e \u003cp\u003ePlant secondary metabolites are sources of medicines, food additives and many other industrially important biochemicals. The biosynthesis of these secondary metabolites can be enhanced by deliberately exposing medicinal plants to drought stress. This can be achieved by manipulating the moisture levels, light intensities and by exogenous application of plant growth regulators including Salicylic acid, Methyl Jasmonate [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. For example, severe water stress has been shown to significantly increase secondary metabolite production in \u003cem\u003eSatureja hortensis and Hypericum Brasiliense\u003c/em\u003e [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Under drought stress conditions, plants close the stomata to prevent water loss by evapotranspiration reducing CO\u003csub\u003e2\u003c/sub\u003e intake through stomata. Correspondingly, CO\u003csub\u003e2\u003c/sub\u003e fixation by Calvin cycle decreases, leading to extensive decrease in consumption of reduction equivalents (NADPH\u0026thinsp;+\u0026thinsp;H\u003csup\u003e+\u003c/sup\u003e). The oversupply of NADPH\u0026thinsp;+\u0026thinsp;H\u003csup\u003e+\u003c/sup\u003e induce the production of reduced compounds such as alkaloids, isoprenoids and phenols which are active compounds in plants [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e].Salicylic acid as a plant growth regulator, induces the plant response to many biotic and abiotic stresses and elicit the production of secondary metabolites in plants [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. Similarly, in Mint (\u003cem\u003eM.spicata)\u003c/em\u003e, total phenol and flavonoid content was significantly increased when treated with 200 \u0026micro;g ml\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e of salicylic acid (SA) [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. In most studies, the treatment of SA to stimulate secondary metabolite production is only limited to \u003cem\u003ein vitro\u003c/em\u003e studies of Ashwagandha. Though \u003cem\u003ein vitro\u003c/em\u003e studies have proven their potentials inducing secondary metabolite production upon treatments of elicitors, real world application is limited due requirement of skilled labor, higher maintenance cost and the genetic stability of in vitro cultures [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe open field cultivation results in larger variability in yields, as both biomass production and production of secondary metabolites are affected by many factors including genotype, climate, soil type, management practices and pests and diseases. Inhouse Hydroponic technology can be used as a strategy to overcome this problem of inconsistent yields. Hydroponic technologies provide optimum growing conditions and can be used to stimulate secondary metabolism by appropriate manipulation of mineral nutrition [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eIn this study, the individual effects and interaction effect of moisture stress and SA on the growth dynamics and secondary metabolite production in Ashwagandha was investigated. It was hypothesized that different moisture levels would have a significant effect on the plant's growth, yield, and total polyphenolic content (TPC), as certain degree of moisture stress could stimulate an adaptive mechanism in plants leading to increased synthesis of polyphenols which are the active compounds in Ashwagandha. Further, it was speculated that the foliar application of SA, a plant growth regulator, would not only influence the plant\u0026rsquo;s stress response mechanisms but also enhance its growth and secondary metabolite production, particularly under varying moisture conditions. A significant interaction between the levels of moisture stress and SA treatments was expected, proposing that this interaction might affect the plant's growth parameters and secondary metabolite production in a synergistic manner. The most severe level of moisture stress, when optimally managed with SA application, is expected to yield the highest production of polyphenols, illustrating the complex balance between stress induction and metabolic enhancement in Ashwagandha.\u003c/p\u003e \u003cp\u003eThis research can contribute to optimizing growth and yield of Ashwagandha which will ensure consistent supply of high-quality products in global market. The increased production of Ashwagandha can meet the growing global demand for pharmaceuticals, herbal health supplements and functional foods. In addition, this study provides strategies which can be applied to enhance growth and yield other medicinal plants, thereby protecting biodiversity.\u003c/p\u003e \u003cp\u003eThrough this study, we aimed to identify effective agronomic strategies that can be employed to optimize the medicinal value of Ashwagandha, catering to the growing demand for high-quality medicinal plants. Therefore, the objectives of the study were to investigate the growth, yield and polyphenols content of Ashwagandha as affected by different moisture levels and foliar application of different concentrations of Salicylic acid.\u003c/p\u003e"},{"header":"2 Materials and methods","content":"\u003ch2\u003e\u003cstrong\u003e2.1 Experimental site\u003c/strong\u003e\u003c/h2\u003e\n\u003cp\u003eThe study was conducted in a 139.35 m\u003csup\u003e2\u003c/sup\u003e protected house in Faculty of Technology, University of Ruhuna, Sri Lanka. The experimental area was in the WL2B climate zone, which is a low-lying wet zone (60 06 \u0026apos;N 80022 \u0026apos;E).\u0026nbsp;\u003c/p\u003e\n\u003ch2\u003e\u003cstrong\u003e2.2 Experimental treatments\u003c/strong\u003e\u003c/h2\u003e\n\u003cp\u003eAshwagandha\u0026nbsp;plants were transplanted in coir dust bags after one month of nursery.\u0026nbsp;Recommended Fertilizer schedule was applied for all plants [15]. There were three replicates for each treatment combination and 16 treatment combinations (4*4), arranged in split plot design. One coir dust bag with two plants was considered as one experimental unit. Two-month-old plants (one month after transplanting) were subjected to moisture stress and salicylic acid stress.\u003c/p\u003e\n\u003cp\u003eFour moisture stress levels as 100% field capacity (FC) (control), 80% of FC, 60 % of FC and 50 % of FC were maintained using automated drip irrigation system with XH-M214 humidity controller soil sensor modules. Foliar spray of Salicylic acid was applied to the plants in concentrations of 10 mM, 1 mM, 0.1 mM and 0 mM (control) once in two weeks.\u0026nbsp;\u003c/p\u003e\n\u003ch2\u003e\u003cstrong\u003e2.3 Data collection\u003c/strong\u003e\u003c/h2\u003e\n\u003ch3\u003e\u003cstrong\u003e2.3.1 Growth and yield parameters\u003c/strong\u003e\u003c/h3\u003e\n\u003cp\u003eThe plant height (cm) from the pot media surface to the plant tip was measured using a meter scale, while the number of leaves per plant was counted manually [16]. The plants were uprooted for root harvesting when the berries turn red to orange and leaves become dry after 150 -180 days of transplanting [17].The SPAD value was measured using Konica Minolta, SPAD 502 Plus Chlorophyll Meter after 60 days of treatment application from 3\u003csup\u003erd\u003c/sup\u003e fully grown leaf from the apex of each plant. Individual leaf area (mm\u003csup\u003e2\u003c/sup\u003e) was measured by using ADC BioScientific Leaf Area Meter AM350 Ver.1.00 from top three fully grown leaves of the plants at the end of harvesting. At the harvesting, the number of fruits and nodes from fallen leaves were counted. After root harvesting dry weights were measured by oven drying at 72 \u0026deg;C until a constant weight was obtained [16].\u003c/p\u003e\n\u003ch3\u003e\u003cstrong\u003e2.3.2 Free proline content\u003c/strong\u003e\u003c/h3\u003e\n\u003cp\u003eThe third fully grown leaf was used to extract free proline following the acid ninhydrin method. First the fresh leaf samples were grind using liquid nitrogen and 0.1 g of sample was weighed. Then the leaf sample was homogenized with 3% aqueous sulfosalicylic acid using vortex mixture and 200 \u0026micro;L supernatant was harvested. Then the supernatant solution was vortexed with 200 \u0026mu;L acid ninhydrin and 200 \u0026mu;L glacial acetic acid. The solution was incubated in a 100\u0026deg;C water bath for a period of one hour and immediately transferred to an ice bath to stop the reaction. After cooling, 400 \u0026mu;L toluene was added, and vortexed the samples. After the mixture cooled to room temperature 100 \u0026mu;L \u0026nbsp; chromophore was diluted with 900 \u0026mu;L toluene. Consequently, the absorbance of the separated upper layer was measured at 520 nm wavelength using toluene as blank. A standard curve was plotted to calculate the final concentration of free proline (mg proline g\u003csup\u003e\u0026ndash;1\u003c/sup\u003e fresh weight) \u003cspan lang=\"EN-US\"\u003e[18]\u003c/span\u003e.\u003c/p\u003e\n\u003ch3\u003e\u003cstrong\u003e2.3.3 Total poly phenolic content - Folin-Ciocalteu method (modified ISO 14502-1)\u003c/strong\u003e\u003c/h3\u003e\n\u003cp\u003eFirst, 5.00 ml of 70% methanol (at 70 \u0026deg;C) was added to 0.2 g of dried and finely ground root sample. The mixed sample was heated at 70\u0026deg;C for 10 min, while mixing in 5 min intervals. The samples were centrifuged at 3500 rpm for 10 min after reaching room temperature. The supernatant was decanted to a 10.00 ml volumetric flask. The extraction was repeated for residue with fresh 5.00 ml of 70% methanol. The extracts were combined and top up to 10 .00 ml by adding cold 70% methanol. \u0026nbsp;The sample (5.00 ml) was diluted up to 25.00 ml with distilled water for polyphenol analysis. Anhydrous gallic acid standard series (10 \u0026micro;g/ml, 20 \u0026micro;g/ml, 30 \u0026micro;g/ml, 40 \u0026micro;g/ml and 50 \u0026micro;g/ml) was prepared. 10% of Folin-Ciocalteu phenol reagent (5.00 ml) was added into 1.00 ml of gallic acid standard series, and 1.00 ml of sample. 7.5 % Sodium carbonate solution (4.00 ml) was added and mixed within 3 min to 8 min after the addition of the Folin-Ciocalteu phenol reagent. All mixtures were allowed to stand at room temperature for 60 min. The absorbance was measured in 10mm path length cells against blank using the spectrophotometer at 765 nm.\u003c/p\u003e\n\u003cp\u003eThe total polyphenol content (TPC), expressed as a percentage by mass on a sample dry matter basis, is given by the formula:\u003c/p\u003e\n\u003cp\u003e\u003cimg 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xpaOWuMt+w5tYSf55hibqc2pR83GEck25HeS1XHpaByX3qDWX66n/TE+Psde6n9N9iEUE7KuQXbNi13pI+fDkCzZVmPigPqdx0FEMo0ZE/IxtYXt0TfXcmT9Sg81+2uM5hij/tT0WqNVTw/kVjvoaGxbGa7THED/cz1zfBJ0Ltu01VfqH2qn5gPkc0R6fiidjY07LSRvzWboM+arTdrKtPTR86FW2+i6JVNrPAP5NdtrTESmjmHF1jE3e8tau/Tsn5kin+Aa9Rc9jPUZ+XHNx1t9AflIzf+H+or9OJ+RrjNjdCed9cZXthW0fF1+K58S2dfG0tNlpGcPnYt1YGfy8pho0RvDoLhTG68tv5Suxvj2FJCROjXma/YbC/JxtJAftMacro8y1PLEUH3L4J9WGjBbiKeffrqkJ5xwQklr/PnPf1785je/Ka8//OEPlzTy1FNPlfTjH/94SQXXwcpAKKl4+OGHS3raaaeVtMYvf/nLkuYyv/jFL0p6+eWXl7SH2jn//PNLGuEccr3nPe9ZzXmNZ555pqTnnntuSYf49a9/XdKPfOQjJY088cQTJb3gggtKKqI+VyaQxYknnrj41re+tXjppZcWK8G66DPb49FHHy3pxRdfXFLxq1/9qqQf+tCHShpBRyuBrry+6qqrSp/+8Y9/lP8jjz/+eEnPPPPMkkaefPLJko7Rh3S5svAoKdx1112lj1Ps9f73v7+kEcn43ve+t6SR3//+90V373vf+1ZzpoPsjzzySJH15ptvXrzrXe8q9UawG32r+Y3O1fxA51YmssXJJ5+8mvsa+A9t7tmzZzXnNaSL3A72o66VIP0mH/nb3/5W0re+9a0lFb/97W9LmsfSz372s5Lefvvti2OOOWZx6aWXLu6+++7FF77whdLGGJsh90pcn3xcc801qzXUcVxyXNpqcWmKvdT/muxD/PznPy+y1Xxfdl1ZnJdUSB81ew/JMicOSCd5bEWmjIkf/ehHJd27d2+R4atf/WqJx7fddtvi2WeffZN+//SnP5X005/+dEkjf/nLX0p63HHHlRTwL3y7pde1Qv3XX399kftrX/taifX4xY033vh6vJkCMt53331FXsbf/fffv3rmNeb4JOi6HC9/97vflfRTn/pUSQU2gHe/+90ljUyNr4rp2Q/nxJ0Wr7zySknf+c53llTQBn3//Oc/X/6XP0Cee0Fx/owzziip6PmQdJXXL3/4wx9KmuuC1njW2oOxE9tqrXWWMZfXQG9cv4y1S8v+a2WtY2/uWGqtBaDXV9mQNVyEfkjXmTHzSWu+nOvrc+b2Hi2/yEy1x5R5RrrozQNqf8p9iHT1sY99rKTL4LHHHlucd955ZR5jXv/6179e4iLzwmZy7LHHrr4aR74fWSbeUNpgGDhr5T/+4z9Kum/fvjdNKhqgOTiCJss80DWgc10RLapzmb/+9a8lPemkk0raQzcQOYAxUDlX24BQwBgbLCXP2WefXdLIT3/605Lmxdjf//73krKI0A32Qw89VBbPrQW6Fm8EmIiCX2szBd2zIKct9H7RRRetnnkDTSi1OjQxjtUHfqDJC9/49re/vfjOd75T/h9C1/UmvQ984AMljehm+8ILLyzpXPCT5557brF///4y4XPDHRcksltNT1rw1/xANz/5Zk8Li7yAaC04QDeOtb5qUvvkJz9ZUtEaS+rb4cOHiw/Sd/zwyiuv7I7NrYLjUhvHpaNZS1yaYy9t4rb634I6obYxBLo5zXGwt8gfkmVqHEB/9Lt24xOZOia4UZDf4VcsnPGT2sJf/a0t+uV7cdNEc0RLrz3y5kQN4jIxG7294x3vKK9vuummxWc/+9ni4+hsDl/84hdLqo0SMccno92yTnWjlu0tXeY5eU58bY3nOXGnhdrABpGrr7661Kd+yx9q8gNxHmJcGxqbLV1pszTHSGiNZ236Z/sqP8//6zWXL3Pt0ovna2GtY683lqirJW9rLQC9vrZsqI2Rmn+NmU9abc7xdZg7t9fAP1t+kZka26bMM2PmAa0TptyHSFenn356SZcBm0dsCvPm68svv1zmoR/+8IclZhw4cGC11PLQhuLQeIl20fh/9dVXS1qjtgZdFt5Q2mBOPfXUkhLoa8R38WpwHe96sNj4t3/7t9XcN9AArX1yhMEABHahSVly1VCZ3oK1tpAcCwMVajflChhrDQz0QYG6Fphg7EDT4o3JKy8Eya8tBCNcw+IcfVI++wJytuqQPvICrYXqoA0+BUDgbvV/LExGkrG2wNRGSu0dwDkQwK+77royAWqS74F9uEGFml+23l3S2Ml+0FpwgG6kc1+RARujo3gjNWYs5Yl7K+C4dDSOS0ezFeJSz17Sca//NbRx2BqTWsznd8V1w1Hzj7GyjI0DvU9D1Rg7JqbcoLT6i39z08GNS+yv9DrmpkPoUztjfIx5SRsWfFJUN0X4GDcbeXxEuDmobahGxmxqQc8nZbf8qT7d7GX9M1ZaN4FT4yv0xjMs88Yj+sUdd9xR2swbXC30CQA2I6LdemOzpSvaxhdqMRJa41mxMG/6tz4FJcaO4be//e2rr/osc+0yZP8aY8quZez16I2lobVAr681GzKX3nDDDeV17VNIY2L4HP22fH2tc3tGm8y1eDGWnj1gzDwzZx4QrfsQ6QpfmOtrNdgg1RqFDWI2zqifzdKhT/jPQTGIvtCniPwdP4loLlGciGiNNmYTcS7eUNpgLrnkkpJqAsjo5hOHzeBUfGQWCNi1AKIBmm8AH3zwwTL4cMChhTuTJx9ZFfrqTm2BoXb++Mc/llQgK4uyPBAyTPD6CF4tACnwreUmGxkIfgSY2mDSRwa1YI0Q4KMuQLbLC3gN8rwQJL/2VZBant7ZmrKY590+XZeR3lhYsMnC18fGwuQF2YZs7sAtt9xS0ozeHah99HgMfEQ8t9n7aHHmS1/60uu2qfl6690lbYRlP9RisuafCtJvectbSipuvfXWsqBFR3FS0zipjSWNA7UnNJZaNt4IHJeOxnFpmPWKSzWG7JVB92NuaPV1rdpXjECL/OzzuuEY4x9ZlqlxYOxXV6aMCRbp8vEIMlE2o/7mBTxfewE+ZToENiQetNCN01CcyKBbPiUyFmJ6Lc7Bj3/84yJD/opajSGfbH2qT3rPN2oa37UbuKnxVb5VG89T404P9UV+wfXf//73yzwR0YZK9EHgf93Yf+Mb3yipkHy1sfl///d/q6/egD7zaVDs13qjojWeW592ojzk/KljWO3VxlxkWWuXnv1rKGZMvfGfOvZaDI2l3lpgqK982iTD1/PkI2M2sLFrjOG9+XKOr0+d24fQJ/GmrKsjPXssY+0V54Gp9yHSVW0dsyyYH9ZjEylCTNC6ML+Rfs8995T0c5/7XEmF3jTQpzAF/sg8wFpzLWvWIbyhtMFoIaIJIIMTMVDY1WdBrkHEAGMwks/zLlqLYU2yd9555+vXMon/67/+a3Gm733veyVPaPHAAKE8AYrve3/mM58p+aCPd5PimLStQEZ/mKBZJCiIIisfy7ziiiuOWmBqNxV5gMBBsNckwMf0+L8WdPif8q1+Cw1AFn5cg0z/8i//Uha0caeXdyH1MUUGGLph0aB+cR3nCfBRF9B7Z0zQDgsvZKY8bfNaoCP0gLyxHi2EuJ4jygkK4gRrzuMjkBdAQostno1C4G2Vq6GP9zO5RnkIVnytiY9vZyijhf+UtoT0rzYBn/jP//zPUmd8pgSLOPJYhFFG+uArZlokkIeuo98QWGvyDX1MFv9EvuifWgB+97vfLSn56Igb5JaOQGOJutRnTQ74ierXuOedpDn6XBa07bjkuMT1HFFO2Mi4NMde+AKQr3GXv4pa48UXXyxpbezLJtioB/JJHzAky9Q4MPbTclPGBLGLDXH5BnKgb8YgZSP0Q6g89TMm8C/iQvZJ3XQoLnAdcuWvcwjqI8YMjbNlQMzJcY4+qj/69EVkjk+27NZ7xg/U4t3U+Cooy0FMmBt3esSbQa5n3cs7/Rl8mpgT4xFt8uaQ5pZWXNPY5AZSfa/NH9dee235igp+Deq32muNZ8ohQ2+cUwZ9yz/nzOX4ELLR7xbLXLsArzmi/WtojGseW0/mjKXeWkC0+lqbu9jkjRu3WXdDMbw3X87x9alz+xDaIB2aM2CqPabMM2Pmgan3IWN0tdmgD9GzF+tDdIlPyp+xA98GoO/5GyLcE5Ef5y8ObCibrCtHzIazEkzK09Z7v3qwElRef/o9x8rEXJ4uv+IoqyXqrDhNuW7FqV7/5QKuXXHM1RJvhnrVDtflNpBTspDmJ8jzKwNR1hXnrf5yAPWq78hEH4GykjVfq1/q4Dxy9nQmuEb1xf7wWm1nfVAvbXM+lqm1pz7UnqRf6x8pdZM3pv4oZ/5lBXSjemIbLaifssg1B2SMctMP7N0CnaylPa6n/9GfsGXNL4Hyko82ZRP0Eq+VnnvykU9dmZZ/kiov+g6yt3SEHJK3NpaQO+s7l9lMkC/aBlkdlxyX0I3qiW20oH7K1sbhEHPsRRmd49qxYyrXGX0QPZFf82Gd49qszzGyUEb6VNstmVVmDGPHBODXkpO0NgZBY4HyY+tGH7JhTUcZxYKxdlsLtIVsUf+9/sMcn1TdGdWT20I/Md7FOYY2yENGtYccNd8UyKNyeTzTls7Huno2qiG7ZXlrUHeUXzpv+RC+EHWb9aW2KcNryV6zE2jMZp3RDvnIkuF6tcH5qJ8pYxhUV5QpIjmQP0M+bWV6/tezf4YytfrXgzljCb3HsZH13Osr9ZCfr8Vf1R62jf6FTDqHrDW7ck2rzam+Ln3U2mn5cw/Kc4xhjj3GzjPoQXVTVyvGkC8bqb5WPOnparNBNuksH5yr6Yg8+S8HesgxKlPTV63uZeMNpU2AAYqRCSDLZL3qNdsXAgsBLE6G6wnt4YNDAW8nwMRKX8dO4rsVxyWT2ei4ZNYPxjXje71s6fjRx/rZGXAjTUzkRnwrwY07/rUb1nS7ATZasCcbGMYsE3/lbRPgI48rN6HlY2v6GOEy0NdvtvJH/czGgW/xMe8f/OAH1Y9Yrwcb8eC3rYK+tjrmY8O7GcclE9mMuGTWD74+sXIjvC621Mf1V26yF9/85jdXc03E8XVnwFeB+DolX4Va5n3BWvnKV75Svrq43s+MMRuDvhKm5/kasyyWuqHEdyh5oO7YI8L3A2tlOI4//vjynVi+p9mD72vyvUEephavRa74ncWtAP1hUwl5+d4rC6e1ou/etx4ganY++Dy+xPdy9+7dW557U/sltvWAdvmOP4uP3bC41XMLvJDv47hkNjMumfUDmz7/fP1n8dcKa0KepcGNNs/dyc8tMq/h+Lpz4L5g37595b5g6H5nI0AOniP3k5/8ZDXHbHd4jiPoeb7GLIulbijxTgnvVB06dIiv0pWFBnCDyf8chw8fLnn5Eww8TIoNFqAOlX/hhRfKQzt5ANcFF1zw+kO+Muzo0w43efxMqtonj5tc/aLGVoLJg3cjoPUwyinoQWtmd6KHtp144onlYbcsTPhJy42Ch+aBfqZ4J8ONlMau6eO4tLvZ7Lhk1g/99P2yYeORX0Dk8GZSH8fXnQWfUiJGcr/Te2DveqIHTeNb/Ky+3zTbGXCfzLp1//79/nSwWT5HlgQPfKK6+DwRXpPHA6EifE+Yh+Rl9N3OXB70UKraw+Na7YDObcUHdC0b+slxyimnbMgDuMzWQt91x/5DD1lcJmq39jDEnYpiFQffRa89SNC8hvTkuLQ72ay4ZNYfPTOPo7amM+uP4+vOhDUGz8Wq3desJ/iQYrXXNTsD7Og52Kw3x/BnxdHWDDuf/KQoP9Mp+CoXP5+94sBH/bQfO998Lzd/5J3deN7BzOVBdbGzGr/Ly8dCzzrrrPKppmefffZN72TpPJ908rtcxhhjjDHGGGOMMWtnaV954+tbcTMJ9FHc888/v6SCjzDXnp/ARyshlwfVdcYZZ5RU8HBP4GOitQ0jPqrJnpk3k4wxxhhjjDHGGGOWw9I+oZThGSM8MwHGNsEDtIGHwEV4oPYnPvGJ8jykuGkVP52Ur5kLDw+dCs+DYpPMGGOMMcYYY4wxZjew1IdyR5555pmSjv35cB4C99JLLy0+8pGPrOa8lnfgwIGymcSmUf6lgf/5n/8pKZ+OWhZsfk09vJlkjDHGGGOMMcaY3cS6bSg9/vjjJb3wwgtLOgS/EAf8mpt+8p9PJPEThzw3iecj5V8a0NfgLr744pIaY4wxxhhjjDHGmPVn3b7yxoO3+bn+Q4cOLfbs2bOa2+bqq69e3H777aPLg76e9vzzz2/5n0Cc81U6Y4wxxhhjjDHGmGWw7O2fddtQ0gbK2OpPPfXUsjE0tvxjjz22+OAHP1heL7MLczZ+/AwlY4wxxhhjjDHG7CbW5StvPCwbzjnnnJIOwbOS2EwaW349YXNq6uHNJGOMMcYYY4wxxuwm1mVD6YknnihpfMB2Dz0/aWx5OPbYY1df1WGTik8b8WtzxhhjjDHGGGOMMWZ5rMuG0iOPPFLSM888s6RDPProoyU97rjjSjoGHtDNL78Bm0cZnsnEQ71POOGE1RxjjDHGGGOMMcYYswyWvqHE1934pTZ45ZVXStqD8vfdd195PaZ85Lbbbispm0faVCK9/PLLywPBt8JX6IwxxhhjjDHGGGN2GkvdUOJB2Wedddbqf4vFVVddtThw4MDqf3Uo/9JLL5XXN99882D5CBtHDzzwQPlaG59G4itupC+//PJi3759iy9/+curJc0y0NcIdTz44IOrZ3YX9JtfMYy6qH1KLhPLaxN1u8N4Pffcc1/vFw/XX2/shxsHfhp1PeUrxHGMsOlvzFaHN7hYVxx//PGv++5OidVzIaZLF9dff/1q7vpAfLnjjjtKm3oW5xyog/iz2223G2ANIv9k3Bpj1s7c+5ydylrWwruGI2bX8/zzzx+57rrrjpxyyin8XF453va2tx356Ec/euTee+9dLfUaL7zwwutluG63gZ7o+8GDB8v/pOhqDOhOOt4JujvnnHNK3w8dOlT+v+yyy8qR2b9//1E6WwZDfojvcu7w4cOrOWYuDzzwQNElvlujp2v8gXM5jmxl7Du7E/n5vn37SnzB/vaD1yDWr/c4Rs/EGMaf5pS1gKyqD3tuJvQHWYb6hZz4H/Mq+iaVP7ZAb4qzHLTDnNuD85TTNVzf8/M5cm0k2Bq5sLUxZm2s5T5nJzO0Ft7teENpl8MigkDBgjEudrQJwKIhs1sHFPpp6WQs6G0r6A57c8xF/jHmBqN3g66Ja87CtOeHnPMEuByG/L6na67h/FbZQB3j9/ad3QfxB5t7oVhHMXy9xrHWIWuZW2tgV435zdj8QF9xs6e3oSRZ8UHNldzIcV1LfulNm2YcirktXeo8N0eATNTBUZuj58i10ehGb2gjbbcgfcjGNcaU2c3g1+iHNepmwhjj2CiWcZ+zU7Fu+qzLQ7nN9uHSSy8tXznkuVd79uxZzV0srrnmmpJefPHFJRX6yONKgCvpbuKee+4padbJWPiI5MoCc7GywFzN2Ryw4W9+85vyVbW53HnnnSX98Ic/XNIeDz30EBvX5UH6GfxuZaE6+eH5Q35Iey+++OLqf2Yt/O1vfytpzX7Q0/VTTz1VbHTyySev5mweY/3evrP7+MUvflHmwc2OzVuV5557rvwIynqN4y984QtlHtBzMZcF8wpzDPPurbfeupq7MfBVLP1yMbob4q677irx6b/+679ej7VXXnlleXwD+ffff3/Ji1x77bXFb3/4wx+WvnKgQ3R5++23v+lrgzyWgvyVm+TFxz/+8ZLHuo9rqOeWW24peZE5cm00v/rVr0r6/ve/v6S7nZ/97GclPe2000paY0yZ3QxzAoz9can1YBlr9ams9T5nJzO0Ft71rCyezS6Fd89wgSkfE9ZHi5f59aXtgHamOea+I7cV3hHC5nq3ea4N9emktbxrgg716STewZ3KbvXDzUDvaNfeve6xVd7hg2X4vdmZ4Bt8+gLf8Lv1b0brhDlxegyaT9ZT95prep8QWia0g760VlDs6bXPJ4Qok9G6Ic+3xGPya+s39TfHXmSqyaFYzZHXN1Pl2gykX/PG2gi7tRhTZjfD+NCcMHXdsyw2Y81Cv2mPI8cBM38tvFtwBN7FaEHCpDI2eGhAtRZG5GvRwkFQzh9DpgyLEM6Tqm3KKYgTSFsyEWiRQ2WRf+pNKxOqZJAcrQWtyuRj6gJbizz6FXVAP1oBirLoJcoaF6pC5aQTDvRCWdWt9msH+hwiTjb5qE14tK/z0Qe0EK0dyD+Gmh/Kn3XEc2v1ubF2APKQT/2nfq5FR3HxP0amsT5Wk4/2ycug/9wP5UWdCckS6elayM6kjFna4X/kYvyJfE7+GuUkP0M5XcdBWepF97HfY/x+TH+A/NimbBsZY9eery0L+YT8UDqPeou26NkoE/tCWfSIvWp2khwqr2toS7YWWb8c6DDruMcYGwkt2PNR60cPzUlcpzpol7yancfoT3WqTG1cUIfyo9/FfBgzxvI1Aj/gfM8GlIm6pM5aXTVol2Ms1BvtS1tqv+Wz9J+y6HMzkG6wUQ3yOU+5jGTniGAP8mp2kc1yfbV6BDbgXJRxjlzrCWMpjjNklh7wA0E54r7KyS+y78hHqUNlsz6lAw7KtNjseEv7nG8dtDemTAa55L8ccWxP1fMQ6JrrVJ/ay3JJ19IJB9fFdsbIxrmsd9pTnfmo2SjLjEyxHOs+yan4g2zRj5EzMmbNMsQYHUViG/HgminQ96xD/Ad/zWQ/4Tqup49xHJI3VofZ7lm3c+whdF0L/JR6VI76WnXtRDZuJjBbEjk+A5lBNURvQDFwOMeAUl28Jk/BQcGXgafy5PFaAUf5MaAIAguDVHXQjiY76hkiltcmCHkKgK06kI3zawkOtMuhgEm7BFTqRa8ZzpGPbJKrpRvqRS8qh24UqPMEoqA5F8lQmyAiQ3qVfGPslmn5If0mP56j/prPYQP5gGTBNpkpdlBZdCz90K50TptQk4ny2JFz6K4lT0Z6jm1yHderf0JtIQdtxzwO5Qn+Jx+5MjrHUQPdcA6dcD3lkVW20//qs8YY10Xb0DfpTSgOcE7+res5anob8vuh/kTdITfwmjz5Qc2u5MX+9Hyth2w6FtrhGvkSekZHkh+9kY9eoo2kB/Izshfn6Q//xzxsm+EcdaEHoH7pQLYDzpMX9SubUn4MY2yUoRzn8aU5cL18kb4B/VIciERd9fSnvDwudADXky87q8/yE+msVRflVJfyKZeR/mS/iOqmn9QHvboy8kHaGIP6Rt3StfTQklHU7DEGZBvTlx5c35NPfWi1U+ufxpBsGKGcrhHSdcyLSMZY3xy5hpirzzim1B4+wP/IwNgXGhP4p+Tnf40Rymqska+xqLp4HYljrUWtTcaC2pT+aYPXnEd+DuVnuI5ynJeMMU99iEjWml+IMWXUDnLRL8hje4qeh+A66uBadML1WtdEan5AHv9zcB3UZIt6lz1q8umc+tlC8w16pJ3oj0C+ztEX8nlNvcjOa9onn2szsa4pjNVRRvaNY2kK9JXr8S+gnZwnsA2yDK3hajpEV9iX66IO6WvUrcqr/lpdvMbnVBfXq64IbZLf8gn1B7ko26trpzLdU82OQkEGpydlQLdggLQGlAIBgymia2pBW4GG+mK7yFRrh0FZa0P1jBm0mgBycJP8BJYaakNBbg5cTxDLbcQgKdAb9kBvvBbSAfKIll0UACNDQXEMXJvlbaG+1dAkMJWeH/bOQfS5aEvlR73CFDtAnsCEdFbzH7VNO3Mmnlab5MX6WmNUfamNUV2T+wlDuiafOkmj7jQGsy4kB3qotSfwu5pNgOs5sm+O8ftef9Ya38b4Wg3apU3aQT9qh/qw+5gxCLRH+dw38nUu6pJ2yM+04qf6lGNbS6eyR0R15zFAnWN1xfVTbATyu1bsH6I1RrBR7vdU/YHkoy6uF3G8xD73fL1VF1AP5zKKLzWoh3O12FNrP6N+Z33U0LoAeTLkc/RAR5TJ/lUD3ciepNgFP+L1mH5lpKdW29JDq271L16vOiVnRHbmELW8SK2+OXLVWIY+5YdT5lbgHL49ph3kojxjOoJ/jumnoOxmxFtQH3qyjikjvY4d2+SP1XOmFbtr/tHyA+mq5gfkyx5R7+RzZKiDfPlsDdkg20Z6y5Av/UTZVT7bYsyapcUcHYH61Do/hNrN4N+xf/QNXZAf7QFcz1Fb20QdxvPSIedin3t+3qpLYz3bvjXPA/Vzrua/rfZ3IvWZxewqGEwakK0BAxpQBKWMJsRaEFC9GQ32XB8BpnYNbRAAcgCS7LW2I5ynHPVkFBCoq4bayAF6LKq/FkBVdww6rUBYC/jSV63uzJiJcgiuxw5joCyTTEa2aOm7R88PpefaOdBNBfqNtCbaKXaQbnPd0PNRydSSuUevzUxrjEpntTqkl7zAhiFdc44jjxnpLvugJvKav0Rkk2wrIL82vqWnnt/3+tPSHZDPkZnqay2Qi2vQD3LgS6TZJ1ugf8mY5Vd+rou8PMY1Zmv6rY0HmBKbpJe8QB/LHBuBZJ/bbutmKDNHf6Bxka+Tv+Y5Ufk1eTQOamNMNwPRF2S/WvnWmFL7Y2KSxnL2vxq1eRIk49Bc0tNxDcrhkxwac6Q1/xqiJbuQbK0+cC5frzqz/kE24BC1vEitvjlytViLPuVrNZ+S3LV6FPsYI1PayTql3dq4rbGZ8RYoz7keQ2VaepAPZTtM1XNG8gxd2/MD6STL3LJHnJsyQ3FJtsnjgjrRQa1O8rkm20xxN/e9ZYMh5uhIaCyhszmoL0PXS79ZF0B+TX/Q0qHkzv3q9Uc+l9e2ms+zbbU+qa2FqQvZ4jwMan/OmNiO9KOO2VUo2HDUAmlrQLUGoFCdGQ3oPAg1acX61IaCJAGCoKIAVgtMGclfC6a1NiMEC465SLc1OXMQzhMdwYj+t25QQQGaa1qTIPSC4hiQkeuHbp5AOqXNjOxZOzdErw/Sc6t/8rkc4GuT+lQ7qO7W5NXyH12Xx8EYxk7g+B3lejeZtRtq1V+Tradr2b7mq7Jfbk8+3LIdYAPK1HSpc7U+jvH7Vn/4n/y58W2Mrw2BPnUdepriK7JvtkUr5rXGuOxT8xOdq/mhzqGPXmziWuxKWew1pY9zbQQaz0NjqAVyyja037LrWvWX5w75K2lEG1C9dmrjgHyOiPRai9PRj9EBttU449wY+6Evyvf8AjS28aGM/LgmY6Slrx7oUOO4ps+xDPVTsg35b7xeddb6I51wiFpepFbfHLl6zNWnrqmNj97aTLFvyDcE9VM+xj78mPrz+GuxmfEWWcln/LUYU2bq2J6q54hiTG3ezkiumr/JV/N4aNlD7eZ8UDu5n0K6kE8gD3aSL2a7tOKXbFGLa2qjFqt7zNGR6I2lMUjX1NEa39JFrR2dq/lCbw5Qfdlerf6oLnSVaemo5RPZj7A9elB5+chu4J9WOmxM4Zprrik/BQvf/e53Sxr57W9/W9L8M6P6ydYLL7ywpBF+phZWBldJhX5Cf2VyfdNPxuunGd/61reWFPQTp/zk7THHHLO49NJLF3fffXf5uWHqufzyy8v5Hg8//HBJaz8t+/jjj5f0ve99b0kj/PQuP6n7vve9bzVnOr/85S9LWvuZfX4WFPRTlPq5UvpFX9EdP9m7EkgXK8Gq+tPK5K1MQOWaD37wg4urr7569czRSAdzfyr2iSeeKOnZZ59d0h7Sae1nV+Uzc36SteWHID3XzvETrOgHfcafv8YXsQH6jflT7KC6VyavN/2kqM7pJ6QjOlcbB0PoJ2WRZ+hnTB999NGSXnDBBSWN6Nz5559f0ojqr8nW07VsX/vpWdnv3e9+d0mFfFM/Z13j17/+dUlrutS5Wh/H+H2rP3Pi21Rfa0H566+/fvHVr3518bWvfa3Uh4/deOONpY0xyL7ZFn/84x9LesUVV5RUtMa4dFjzE87VfB/GxiaufeSRR4pubr755sW73vWuN/3seYs5NhLq19AYasHYeOihh8oY/vnPf17kv++++1bPvsFc/em6PHf87ne/K+mnPvWpkgp0CHl8wVNPPVXSPMbkS8gQefrpp0ua47RiD9DfE088cfGtb32rzJMrC+zSztR41kOxeOVGo6QRxZoPfOADJV0G9O+iiy4quvziF79Y1kW8PnDgQBmTm8Wxxx67+moc2Z5jOO6441ZfjWdIrrXok2tbc6vWZrX5ABT7Pvaxj5V0CNVPe+Kuu+4qPj5mjQmbGW+ffPLJkvZ+Zn6ozJyxPVXPEa3tP/nJT5a0heZO2LNnT0kjmr/POOOMkoqWPTRnfOhDHyqpiHN0K4b96Ec/KunevXvL2pD5GX9mrnv22WffZBetTXL8ki1q/iv799Ysmbk6gmXc5zBG7r333vL6qquuKj6Wx/fcNVxLh1oj0Ha0V68/f/rTn0r66U9/uqSRv/zlLyXNcbDlE8u4N90peEPJHMVnP/vZkr788ssljWixnG+EdIPY26jJwUOBtHYDoAkgTjBMcsCN/JEjRxbPPfdcWcRfeeWVo27MYEygrS1Kn3nmmZLWZB0LwZAbmhyMHnzwwZLGIPnXv/61pAcPHix9ffHFF0tfb7rppuoNh+A8+qEdghs3ohkFxbE6yyh412ydkU5rAV0+M2fyavkh9M5pEmn5Ys6fYoe///3vJa315/777y9pbRNOMtU2ModQm6eeempJe2j85JtMJnvd/OY+6Ua8NvFDT9e62T3vvPNKKmiP61gMx3FIvjZgesgmNV0yiUPtRnqM3y8zvk31tRaUZ1GE373jHe8or/E/4jQxIy/WamgTIduCBTCcfvrpJRXKz/3VTVbNTzjXG8tjYhNQN7F9//79pa8sLOW7PebYCKibdtgMWgvEdWzEDRe+zY2GYruYo784LvLcoZvK7K+6EcnzHHUxDmp91WI9L35bMVyxBx8kNnLQf26oehvCc9G4f+c731nSyJ133lnSKTdeQ7CJcMkllxy1MchrbjK+9KUvreYsD93gDY3n6DuaM1555ZWS1oh2i+O8106Mn3PkqrEWffbmVmI2tN7gUuzLMa4H40NrRfr97W9/e/Gd73yn/D+GzYy32gDOmySRoTJzxvYcPQvFd+a3HlqHt2K1fCHrvWUPxcmsR7XTm6PRP7FX+qEN/Dlvaog//OEPJc33F61NLZizVp+rI1jGfQ6gAzbV8B/6wEZyZO4arqVDbcjmcaL8Wn+k99p6QX4R38DprYWXcW+6U/CG0i6EwcFOao88EDSghm74MlrsfeYznymp0KSWd8qZwAnMBOvap3mGFi5zICDoJrc2YWqDq7arPwbtlDM5ZLS7/eUvf7mkkVpQHQL9EMyAG7eIbDhmA6KFgu2YhYN0WguqmtimBtyeHw75qCaR/G67fLHlW3PsILQghdrkJZnW8u76WhYALPRbN9S6Ea/pZUjXutnNiyt90uDrX/96ScXUjZYM8sin8o30GL8f6k+LVnyb62sZ4hELefTIJzd1k0EfWcTWFq8R/E8L01xWN01ZltaGRAvevYYhP+zFpgyfluVTTfimfGYuLRuBNlLmbOjWwF4/+MEPyuv//u//LukQPf1pXORPE2gjLN8Maa6pjWfdMNT6qoW83lASvRgOrRv5sYyN/61PNbJW0M3d2Lpqm1IZNkC5IQDesdZNA3lxU2RZaDOMMZk3bxSb4ptOoNii8RpR7GYTRzD+tQaRX0Vk6zju58hVYz30OTS3KvYR04fiZER+xFhifmROGhsLNzveagO4tzkzpgyMHdtz9ZwZ2/8a8iF8McowZI9a3NDYOemkk0raorah10I2zpveimt5PT1mzTKVlo7EWu9zItRPe/gEetYb0T3oc2sNBy0dqu48N7Q2aqGnd+YT5s/oF721sBi7ptvJeENpF6LBoUk98r//+78lzQtLfVy3Nslo4L366qslFXfccUcZnASwPNg0qb3lLW8pqbj11lvLgviWW245Kujp6285MDFhsANe60tGiymuiXDjArRZQ+9wzH0HVDvlal8QvLixInjFAKqPWkrnEb4uEt/55iPj+d1+9JbbaoE+W58WyKA37FmbnDOyx9hJl7rHfDS054f6qmRrIaRJJMskX8wT0hQ7tODdV22S1CZJyTTHt/Q1A30aKIJ8WpS0wPa6trY41bnahlpP19TLGM4+iI2Ri0Xc5z//+dXc19BGy5yvQFIvi2wWL6135jLZ73v9mRPfpvraGBgfvAM2hda7rYxPLZwi5Lc2JGqgA8XmvHCbEpt4gyPH5Sm+MMdGoHc95/gdMPfkODBlLPf0B61PE2gjLI/b3rvMmvfzR/mRn4U8OooxSnNtLYb3Yg83EmPnFN201WJsRJtgGqeAr2ojbMw8I1mHbqQzbOhqI3S9wH815vIG6j333FPSz33ucyUVvOFGLOWGLa9/2ETlXH5Tjq+bgd7IErrZ1CMPxBy5hliWPofmVo2F3te/aihOsG5jw4qv345lM+NtC+bc7B+ZWGbq2J6rZ9Fa2/N/XBO+/e1vL2meJ/j/hhtuKK+/8Y1vlFS07KG10dj1KXqPcZ6xpRgcQeb8iRzQppbmKaENlDGbEXnNUmOOjsRa7nPQZ83+Y30C+YbWcC0dtjZkKQ+12CC95/sYvroIfEI60lsLL+PedMewskA1uwweHobpVwZveTgZkOphZJzP8IA1zvGgOMquDPrXH/pGmuvjQWQrQbfk1R5sxzmuWVnElv8p02s/PvhM9ZFH/cg0hlg/dQy1CZThPPLOhT5Sx0owHKUfyuicyh8+fLjUg95jef6nrGzBOdmKvkWoQ3JQjmt4Tf4YZAPZrIfaUv+QJcoj+1OOPlJuzMPren6YzyGnzgHnanakDs5RlkM6nmIHoCwHZTiHT9En1U8eOoz6a8k0FmTjevWTtmkv+7N0o3z1UzZFZ8gXdcZrzlGGPNkSerrWOQ7l6XrpJyMd1c5FOE85ySIfiPkQ/U3nen7f6w+p6pYfDMU3ytfsqn5SJwf/165fFuiA9uhfJNo9Qv/UV+SKepQ/aJxSJ/6kNqgTnapPpNE3yZOeVSdIv9QlXVAPMnD9GP3MsRFwjuuiL0wht0lKP8hXv8VU/YH8JcuH3XRNhLrJr/mx2tc4gJ6OaJPyOked0W7kR/vSHuenzClqQ3Gphewr3fC/+kZ+lKsFciHvRoNeaBc5kbkFuqBc1Kns2dKPzkcflP/JzyLojrKcV0xQuzUfgDlyLRu1jyzISLvIoPFBHmMh6lfjKse+IeRTc65ttbkR8RZ0DWWlp2yjMWWmjO1Wn8ci3SAD8nBQV60t2VttIVfWU6Qlm2ws3WEXlSHlnHyJ//PYUPzVNZyjfWTOcqgt1RchnwOoS3ag3+RTH3VTR00fNabqCGiD89h8DjU9Y1fqQ56I+iadIh9lYj5QJwe0dCi5dU0EfXFOttG1aodjrI50jj4hS/SH7L/Ko0we7zsdbyjtQhgwDGAcXgOLg0HDYKnBQNEA5bpcTgNIdfE6BpeIBjTtaaDqGuppgdySQde35G1BgMp19NpUIMtBcQrSBQGHAEt9yIAsCkAZdJT1WQt01IFsqrdXFjQRciAPgXQstMV1CvJDUD/l6Wv2BeSTzMg/1o49P+ScdJbP9eyI/SVL1slYOwBtSLbYJ8qTRxtx0unJNBbkGzuGZA/kkO9lncVrkU96oY2ol56u8THaog3pQ31v+RtlOMaAL9Xspf5pbEWG/L7XH0AvOq8yrfjWs2vP19YDZKCt3B+N5Zovt/SIrKqPc7oWH4x+wv/AtZTXOQ70lttENtqM+h3ylxpTbCRUdg7Ihl9JJzpa89JU/YHqzKierJ/eeKYN8tCr2quNlYhiC+WyLmlL52Nd5E+B6ziG0BiWb9COYmvsZw30RDmu2yiQl35JP/HADrWxl+O5dNqj5vc9faC36ANRny3myLVMGE/SJb6v8SX71/qgMVIbiz2oQ+1MpdUmuiK/ZnPk5lzW6Zx4wWvpKV4TGVMGHYwd23P1HKFuydTzR/Jqvht1EOnJVtMt0IbOUX+r34zvLEeOxyDbR9sKnau1M7RmaTFVR4B+KEu/54D+8BfZkKPnM1PXcNJTzAPJzXUZZOKc+i85aJt89BvjZh5LEdqRvJTLtqCt2HfKrGU8bFe8oWQ2HA30GMS3Kq1AthtZxsLBGGO2AmtdRG8nWADT19rCe7PRHFvbBOEmAPu05hzdgNRuWiK6OfPcZYbAH7l5HHsDb8x2Zzfd52jO8PhePn6Gktlw9PC3Ob8IsdHomScri9qS7mZqD+00xpjtSO3BxTsVPe9jzLM6NhoewH7KKacsvvKVr6zmvAHPyWHeqa0VeEYFzyC8rPGQWcEzLHgOzsqNhOcu04VnBF177bXlwfr5WS3G7FR2030Oz1ziPsbje/l4Q8lsOHr421Zc3EZYsLKYZbG71WVdb/RAwjEPzjbGmK3Oj3/845LWfk10p6GHj895GPxG8JOf/GTx4osvlocDR/TA1dqG0Y033ljS2i+kCuZwNpzOOeecxTe/+c3VXGPeQD8IwIOX9+7dW36kpfXz+MbsNHbTfQ59ff7550c/jN1MwxtKZkNhQOvp+1sdLVj1SzK7FWz27//+72VX34tyY8x2h08iMA/t379/V7xTqY2ZrQo3MvzMM3LGX8bRGxjc7DMPAefYeOLTSdiv9akjfnnovPPOK5tR/KpY71NMZneiX/s68cQTF1dddVX5FNtNN91U8ozZDeym+5wnn3xy9ZVZD7yhZDYU/YwnsHDUInErwadxeNeKT1IdOnRoV39Mnp8pZbHFuxfcgHlRbozZrjDfENv5CeWDBw+Wr1vtBvQmzhVXXPGmnzfeKrCpxIKfryDyM9/IyRsYbBp9//vfL/MQtuPTRnyaibm5ZT82oPgZag5vJpkWr776aklZ3xAPbrvttvK/MTud3Xif8/TTT5eUT2Rxb2OWyzE8SGn1tTHGGGOMMcYYY4wxg/gTSsYYY4wxxhhjjDFmEt5QMsYYY4wxxhhjjDGT8IaSMcYYY4wxxhhjjJmEN5SMMcYYY4wxxhhjzCS8oWSMMcYYY4wxxhhjJuENJWOMMcYYY4wxxhgzCW8oGWOMMcYYY4wxxphJeEPJGGOMMcYYY4wxxkzCG0rGGGOMMcYYY4wxZhLeUDLGGGOMMcYYY4wxk/CGkjHGGGOMMcYYY4yZhDeUjDHGGGOMMcYYY8wkvKFkjDHGGGOMMcYYYybhDSVjjDHGGGOMMcYYMwlvKBljjDHGGGOMMcaYSXhDyRhjjDHGGGOMMcZMwhtKxhhjjDHGGGOMMWYCi8X/A0JPqp4sCePeAAAAAElFTkSuQmCC\" style=\"width: 1166px;\" width=\"1166\" height=\"94\"\u003e\u003c/p\u003e\n\u003ch2\u003e\u003cstrong\u003e2.4 Data analysis\u003c/strong\u003e\u003c/h2\u003e\n\u003cp\u003eCollected data were analyzed using two-way analysis of variance (ANOVA) and the mean values were compared using Duncan test significant difference test at P\u0026le;0.05. The test was carried out using SAS\u0026copy; for academics software and IBM SPSS software.\u0026nbsp;\u003c/p\u003e"},{"header":"3 Results ","content":"\u003cp\u003eTable 1\u0026nbsp;\u0026nbsp;Significance levels in two-way ANOVA of the effect of moisture stress and salicylic acid on growth, yield and total polyphenol content of Ashwagandha\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003eSource of variance\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003eMoisture stress effect (main factor)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003eSalicylic acid stress (main factor)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003eInteraction effect (Moisture stress* Salicylic acid stress)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003eGrowth parameters\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003ePlant height\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003eNS\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003eNumber of leaves per plant\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003eNS\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003eNumber of fallen leaves per plant\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003eNS\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003eNumber of fruits per plant\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003eSPAD value\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003eLeaf area\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003eYield parameter\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003eRoot dry weight\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003eBiochemical parameters\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003eTotal polyphenol content\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"37.770382695507486%\" valign=\"top\"\u003e\n \u003cp\u003eFree proline content\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.80199667221298%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.299500831946755%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"23.12811980033278%\" valign=\"top\"\u003e\n \u003cp\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e*, and ns represent significant (P\u0026thinsp;\u0026le;\u0026thinsp;0.05), and not significant, respectively\u003c/p\u003e\n\u003cp\u003eThe of the effect of moisture stress and salicylic acid on growth, yield and biochemical parameters (free proline content and total polyphenol content) of Ashwagandha is presented in Table 1. Moisture stress and SA eac individually influenced the growth parameters: plant height, number of leaves per plant and number of fruits per plant. The interaction effect of SA and moisture stress treatments were significant for the parameters including number of fallen leaves, number of fruits, SPAD value, root dry weight and total polyphenol content and free proline content.\u003c/p\u003e\n\u003cp\u003eAs shown in Fig 1, the highest plant height, number of leaves per plant and leaf area was achieved under 100% FC and 1 mM of SA. Upon application of moisture stress the plant height was decreased by 12.02%, 14.3% and 24.11% in 80% FC, 60% FC and 50% FC respectively compared to 100% FC. The number of leaves decreased with a maximum reduction observed at 50% FC by 47.85% relative to 100% FC. When the plants were undergoing moisture stress level 50% FC the leaf area was reduced by 42.63% compared to 100% FC (Fig 1). Upon receiving the treatment 1 mM SA plant height, number of leaves per plants and leaf area was increased by 21.08 %, 16.5% and 47.99 % respectively relative to control (0 mM SA). The lowest plant height, number of leaves per plant and leaf area was reported in the salicylic acid level 10 mM (Fig 1).\u003c/p\u003e\n\u003cp\u003eThe highest number of leaves fell under the interaction effect of treatments 50% FC and 10 mM SA while the lowest was reported with interaction effect of 100% FC and 1 mM SA (Fig 2). The highest SPAD value was observed in the Ashwagandha plants received combined application of 100 % FC and 1mM SA which was 17.14% higher than control (100%FC and 0 mM SA). The plants treated with 10 mM SA had had significantly higher SPAD values than control with dark green, shrunken which was evident with lowest leaf area given in Fig 1. The lowest SPAD value was observed in the plants treated with 50 % FC and without SA application.\u003c/p\u003e\n\u003cp\u003eThe significantly highest number of fruits per plant was observed under the combined treatment 60% FC and 1mM SA while the lowest was recorded with the application of 80% FC and 10 mM SA. The significantly highest root dry weight (7.62g) and total polyphenol content (3.78 (GAE mg/g d.w.b) of the plant was recorded with the interaction effect of treatments 50% FC and 1 mM SA. The root dry weight was increased by 2.7 folds under the interaction effect of treatment 50% FC and 1 mM SA relative to 100% FC and 0 mM SA (control). The lowest root dry weight reported at 100% FC and 10 mM SA was 1.33 folds lesser than the control. The TPC of plants undergone the treatment 50% FC and 1 mM SA showed an increment of 12.49 folds relative to that of control. Highest free content of 150.72 \u0026micro;g/g FW was reported under the combined application of 50% FC and 1mM SA while the lowest was reported in the control (100 % FC and 0 mM SA) (Fig 3). \u0026nbsp;\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe morphological and biochemical parameters, number of fruits per plant, root dry weight and free proline showed a positive correlation with total polyphenol content while leaf area had a negative relationship to total polyphenol content (Table 02). \u0026nbsp;The root dry weight was positively influenced by total polyphenol content, free proline content and number of fruits per plant.\u003c/p\u003e\n\u003cp\u003eTable 02 Correlation between morphological and biochemical parameters of Ashwagandha under moisture stress and salicylic acid treatment\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" width=\"836\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.335329341317365%\" valign=\"bottom\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.622754491017965%\"\u003e\n \u003cp\u003ePlant height\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.341317365269461%\"\u003e\n \u003cp\u003eNumber of leaves per plant\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.377245508982035%\"\u003e\n \u003cp\u003eNumber of fallen leaves per plant\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.538922155688622%\"\u003e\n \u003cp\u003eNumber of fruits per plant\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.461077844311378%\"\u003e\n \u003cp\u003eSPAD value\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.425149700598802%\"\u003e\n \u003cp\u003eLeaf Area\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.221556886227544%\"\u003e\n \u003cp\u003eRoot dry weight\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.059880239520957%\"\u003e\n \u003cp\u003eFree proline content\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.616766467065869%\"\u003e\n \u003cp\u003eTotal polyphenol content\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.335329341317365%\" valign=\"bottom\"\u003e\n \u003cp\u003ePlant height\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.622754491017965%\" valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.341317365269461%\" valign=\"top\"\u003e\n \u003cp\u003e0.48\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.377245508982035%\" valign=\"top\"\u003e\n \u003cp\u003e0.08ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.538922155688622%\" valign=\"top\"\u003e\n \u003cp\u003e0.23\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.461077844311378%\"\u003e\n \u003cp\u003e0.27\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.425149700598802%\" valign=\"top\"\u003e\n \u003cp\u003e0.19ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.221556886227544%\" valign=\"top\"\u003e\n \u003cp\u003e0.05ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.059880239520957%\" valign=\"top\"\u003e\n \u003cp\u003e-0.23*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.616766467065869%\" valign=\"top\"\u003e\n \u003cp\u003e-0.12ns\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.335329341317365%\" valign=\"bottom\"\u003e\n \u003cp\u003eNumber of leaves per plant\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.622754491017965%\" valign=\"top\"\u003e\n \u003cp\u003e0.48\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.341317365269461%\" valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.377245508982035%\" valign=\"top\"\u003e\n \u003cp\u003e-0.10ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.538922155688622%\" valign=\"top\"\u003e\n \u003cp\u003e0.11ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.461077844311378%\"\u003e\n \u003cp\u003e0.26\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.425149700598802%\" valign=\"top\"\u003e\n \u003cp\u003e0.21\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.221556886227544%\" valign=\"top\"\u003e\n \u003cp\u003e-0.12ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.059880239520957%\" valign=\"top\"\u003e\n \u003cp\u003e-0.33*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.616766467065869%\" valign=\"top\"\u003e\n \u003cp\u003e-0.17ns\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.335329341317365%\" valign=\"bottom\"\u003e\n \u003cp\u003e\u0026nbsp;Number of fallen leaves per plant\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.622754491017965%\" valign=\"top\"\u003e\n \u003cp\u003e0.08ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.341317365269461%\" valign=\"top\"\u003e\n \u003cp\u003e-0.10ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.377245508982035%\" valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.538922155688622%\" valign=\"top\"\u003e\n \u003cp\u003e0.08ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.461077844311378%\"\u003e\n \u003cp\u003e0.10ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.425149700598802%\" valign=\"top\"\u003e\n \u003cp\u003e-0.26\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.221556886227544%\" valign=\"top\"\u003e\n \u003cp\u003e0.09ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.059880239520957%\" valign=\"top\"\u003e\n \u003cp\u003e0.38*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.616766467065869%\" valign=\"top\"\u003e\n \u003cp\u003e0.01ns\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.335329341317365%\" valign=\"bottom\"\u003e\n \u003cp\u003eNumber of fruits per plant\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.622754491017965%\" valign=\"top\"\u003e\n \u003cp\u003e0.23\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.341317365269461%\" valign=\"top\"\u003e\n \u003cp\u003e0.11ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.377245508982035%\" valign=\"top\"\u003e\n \u003cp\u003e0.08ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.538922155688622%\" valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.461077844311378%\"\u003e\n \u003cp\u003e-0.06ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.425149700598802%\" valign=\"top\"\u003e\n \u003cp\u003e0.13ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.221556886227544%\" valign=\"top\"\u003e\n \u003cp\u003e0.43\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.059880239520957%\" valign=\"top\"\u003e\n \u003cp\u003e0.17ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.616766467065869%\" valign=\"top\"\u003e\n \u003cp\u003e0.39\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.335329341317365%\" valign=\"bottom\"\u003e\n \u003cp\u003eSPAD value\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.622754491017965%\" valign=\"top\"\u003e\n \u003cp\u003e0.27\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.341317365269461%\" valign=\"top\"\u003e\n \u003cp\u003e0.26\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.377245508982035%\" valign=\"top\"\u003e\n \u003cp\u003e0.10ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.538922155688622%\" valign=\"top\"\u003e\n \u003cp\u003e-0.06ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.461077844311378%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.425149700598802%\" valign=\"top\"\u003e\n \u003cp\u003e-0.009ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.221556886227544%\" valign=\"top\"\u003e\n \u003cp\u003e-0.36\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.059880239520957%\" valign=\"top\"\u003e\n \u003cp\u003e-0.34*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.616766467065869%\" valign=\"top\"\u003e\n \u003cp\u003e-0.17ns\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.335329341317365%\" valign=\"bottom\"\u003e\n \u003cp\u003eLeaf Area\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.622754491017965%\" valign=\"top\"\u003e\n \u003cp\u003e0.19ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.341317365269461%\" valign=\"top\"\u003e\n \u003cp\u003e0.21\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.377245508982035%\" valign=\"top\"\u003e\n \u003cp\u003e-0.26\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.538922155688622%\" valign=\"top\"\u003e\n \u003cp\u003e0.13ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.461077844311378%\"\u003e\n \u003cp\u003e-0.009ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.425149700598802%\" valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.221556886227544%\" valign=\"top\"\u003e\n \u003cp\u003e0.10ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.059880239520957%\" valign=\"top\"\u003e\n \u003cp\u003e-0.50*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.616766467065869%\" valign=\"top\"\u003e\n \u003cp\u003e-0.28\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.335329341317365%\" valign=\"bottom\"\u003e\n \u003cp\u003eRoot dry weight\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.622754491017965%\" valign=\"top\"\u003e\n \u003cp\u003e0.05ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.341317365269461%\" valign=\"top\"\u003e\n \u003cp\u003e-0.12ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.377245508982035%\" valign=\"top\"\u003e\n \u003cp\u003e0.09ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.538922155688622%\" valign=\"top\"\u003e\n \u003cp\u003e0.43\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.461077844311378%\"\u003e\n \u003cp\u003e-0.36\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.425149700598802%\" valign=\"top\"\u003e\n \u003cp\u003e0.10ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.221556886227544%\" valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.059880239520957%\" valign=\"top\"\u003e\n \u003cp\u003e0.23*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.616766467065869%\" valign=\"top\"\u003e\n \u003cp\u003e0.39\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.335329341317365%\" valign=\"bottom\"\u003e\n \u003cp\u003eFree proline content\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.622754491017965%\" valign=\"top\"\u003e\n \u003cp\u003e-0.23\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.341317365269461%\" valign=\"top\"\u003e\n \u003cp\u003e-0.33\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.377245508982035%\" valign=\"top\"\u003e\n \u003cp\u003e0.38\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.538922155688622%\" valign=\"top\"\u003e\n \u003cp\u003e0.17ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.461077844311378%\"\u003e\n \u003cp\u003e-0.34\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.425149700598802%\" valign=\"top\"\u003e\n \u003cp\u003e-0.50\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.221556886227544%\" valign=\"top\"\u003e\n \u003cp\u003e0.23\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.059880239520957%\" valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.616766467065869%\" valign=\"top\"\u003e\n \u003cp\u003e0.33\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.335329341317365%\" valign=\"bottom\"\u003e\n \u003cp\u003eTotal polyphenol content\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.622754491017965%\" valign=\"top\"\u003e\n \u003cp\u003e-0.12ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.341317365269461%\" valign=\"top\"\u003e\n \u003cp\u003e-0.17ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.377245508982035%\" valign=\"top\"\u003e\n \u003cp\u003e0.01ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.538922155688622%\" valign=\"top\"\u003e\n \u003cp\u003e0.39\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.461077844311378%\"\u003e\n \u003cp\u003e-0.17ns\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.425149700598802%\" valign=\"top\"\u003e\n \u003cp\u003e-0.28\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.221556886227544%\" valign=\"top\"\u003e\n \u003cp\u003e0.39\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.059880239520957%\" valign=\"top\"\u003e\n \u003cp\u003e0.33*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.616766467065869%\" valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e*, and ns represent significant (P\u0026thinsp;\u0026le;\u0026thinsp;0.05), and not significant, respective\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e"},{"header":"4 Discussion","content":"\u003cp\u003eWater stress is the most significant abiotic elicitor, which affects medicinal plant growth and active ingredients by altering biochemical and physiological characteristics. Many studies revealed that moisture stress has a positive role in augmenting plant secondary metabolite production in various medicinal plants [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e].In present study, the moisture stress had a negative effect on plant growth (plant height, number of leaves, SPAD value and leaf area) which is in fair agreement with reduced plant height with the application of drought in \u003cem\u003eRosmarinus officinalis L\u003c/em\u003e[\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e], \u003cem\u003eSatureja hortensis L\u003c/em\u003e[\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e] and \u003cem\u003eTrachyspermum ammi l.\u003c/em\u003e[\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. In \u003cem\u003ePortulaca oleracea L\u003c/em\u003e, the plant height was decreased by 18.5% and 45% under 60% FC and 90% FC, respectively and the number of leaves decreased with a maximum reduction observed at 30% FC by 51.3% compared to the control plants. In the same study the chlorophyll A content was reduced by 23.6% and 43.8% under 60 FC and 30% FC [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e].The decrease in number of leaves per plant and leaf area observed in plants is a strategic adaptation manifested by plants to reduce water loss by transpiration. The physiological damage occurred in plants due to drought was evident by the degraded morphological characteristics of plant including plant height, number of leaves and SPAD value [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]. During the present study, the higher number of leaves (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e1\u003c/span\u003e) recorded in 100% FC and higher number of fallen leaves (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e2\u003c/span\u003e) recorded in 50% FC were also indicators of drought stress undergone by the plant. Similarly, leaf area Ashwagandha was reduced by 27.4% and 34.4% when subjected to mild and severe water stress in a study conducted in Gujarat, India. In the same study. Chlorophyll content was also reduced by 60% in the severe water stress conditions which was in line with present study [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. The increased production of Reactive Oxygen Species due to drought conditions, leads to oxidative stress in plant cells. Oxidative stress cause disintegration and reduction of chlorophyll content in plants thereby reducing photosynthesis rate[\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e] .\u003c/p\u003e \u003cp\u003eWith the increase of SA level, the leaf area was decreased while increasing the number of fallen leaves in Ashwagandha in our study. Though the number of fallen leaves increased with increasing moisture stress and SA concentration, new leaves were developed under the stress conditions. The leaf senescence was higher with the increase of substrate moisture stress. The reduction of shoot parts (plant height) with the increase of drought stress level could be due to the decrease in number and area photosynthetic organs specifically the leaves[\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. Under stressful conditions, plants promote leaf senescence to increase the translocation of essential nutrients to growing tissues of the plant [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]. It seems that with increase in the drought stress level, the plant height and number of leaves decreased in response to the reduction in the leaf area and photosynthetic pigments [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eDuring our study, a significant increment of SPAD value was observed in plants which received 1mM SA treatment in both 60% FC and 50% FC (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e2\u003c/span\u003e). This result agrees with those reported [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e] who found that foliar application of 1 mM SA increased plant height, chlorophyll content in purslane plant under drought conditions. Moreover, [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e] showed that foliar application of 1 mM SA increased Chlorophyll a, b and c content significantly in \u003cem\u003eConocarpus erectus\u003c/em\u003e and \u003cem\u003ePopulus deltoides\u003c/em\u003e plants under 60% FC and 30% FC moisture stress conditions. One of the mechanisms plants responses to any stress condition, including drought, is ensuring survival of species by fruiting and flowering. This fact was proven during the study as the number of fruits increased under treatments 60% FC and 1 mM SA[\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eSA is a plant hormone that can ameliorate drought stress by stimulating the production of antioxidants and other defense compounds in plants, as well as improving the photosynthesis mechanism [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e]. Many studies have demonstrated that SA can efficiently stimulate secondary metabolite production in plants [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e].From the results of the present study, it was evident that SA has a significant positive impact growth of Ashwagandha. The present results agree with [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e] who observed significantly enhanced plant growth, biochemical properties and antioxidant activity compared to control plants of \u003cem\u003eMentha spicata\u003c/em\u003e with the application of 200 \u0026micro;gml\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e (1.44 mM) SA. Low concentrations of SA decrease the negative impacts of drought stress on plant growth as it enhances the drought stress tolerance of plants\u0026mdash;however, high concentrations of SA result in detrimental effects on plants [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e]. The SA concentration of 10 mM cannot be recommended for foliar spray as the plants treated and it did not show any significant difference with that of control. SA increases plant growth under drought stress by preventing reduction of cytokinin and auxin, which induce cell division of apical root meristem and plant growth [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eRoot is the most common plant part harvested from Ashwagandha because of its rejuvenated properties. The results of increased root dry weight were consistent with findings of [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e],where authors reported increased root weight under moderate drought stress. Similarly, in Rosemary plants, root yield was highest under 60% FC. In the same study, root yield was reported to be highest in SA treatment 2 mM [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e]. In a similar study where root dry weight was increased when SA was foliar sprayed to purslane (\u003cem\u003ePortulaca oleracea L\u003c/em\u003e.) seedlings under drought stress, there was no significant effect on root dry weight when SA was applied under non drought stress conditions[\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]. Plants increase the root biomass and growth in response with early drought stress signals to penetrate deeper soils to explore more water to keep shoot in hydrated condition. These can be the reasons for the increase of dry weight observed in our study. Higher branching and more fibrous roots were observed in plants which received the moisture stress treatments 100% FC and 80% FC while the plants under treatment 50%FC had good quality roots with higher biomass similar to [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. In the \u003cem\u003eStellaria dichotoma L.var laceolata Bge\u003c/em\u003e increase in root biomass was reported until a water stress level of 60\u0026ndash;70% FC and then decreased with afterwards [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e].The increased root dry mass due to the undergone drought stress was observed in many plants including Ashwagandha and tall Fescue Cultivars [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e],[\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e]. This observation was in accordance with the theory of functional balance which states \u0026ldquo;that plants will respond to a limited water availability by increasing the flow of assimilates to the root leading to an increased root dry mass ratio\u0026rdquo;[\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. Plants treated with Salicylic acid levels 1mM and 0.1mM) under the moisture stress level 50% FC showed significantly more root dry weight when compared to combined application of 50% FC and without SA (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e3\u003c/span\u003e). This data showed that SA application in low concentration could effectively improve plant tolerance and overcome the adverse conditions induced by the limited water availability. Many similar observations have been reported in sweet basil plants [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e],\u003cem\u003eConocarpus erectus\u003c/em\u003e and \u003cem\u003ePopulus deltoides\u003c/em\u003e[\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e] under drought conditions. SA can stimulate root growth by promoting cell division and elongation when plants undergo stress conditions. Foliar application of SA performs the same function when plant is under moisture stress. This is important for drought-stressed plants, as a larger root mass can aid in absorption of more water and nutrients from the soil [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e].The metabolic changes triggered by drought stress results in accumulation of higher amount of secondary metabolites in medicinal plants which also can been a reason for the increased root dry weight[\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. SA has a positive regulation of plant responses for drought stress as SA induces stomatal closure to maintain water content in plant leaves [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThis proline buildup is particularly enhanced by higher concentrations of SA (1 mM and 10 mM), particularly in situations of acute water stress (60% and 50% FC). This suggests that SA may increase the plant's resistance to drought by activating biochemical pathways that are involved in the synthesis of proline. As field capacity (FC) drops from 100\u0026ndash;50%, proline concentration in plants increases, confirming its function as an osmoprotectant that aids in maintaining turgor and stabilizing cellular structures under stressful situations. These results are consistent with earlier studies, including one by [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e], which showed that SA efficiently stimulates proline synthesis and contributes to stress signalling and defensive responses in plants exposed to different degrees of water stress. Particularly under some circumstances, higher SA concentrations are particularly more effective in eliciting these reactions.\u003c/p\u003e \u003cp\u003eIn the present study, there was a significant increment in TPC under interaction effect of application of 10mM Salicylic acid under moisture stress of 50% FC. Similarly, \u003cem\u003ePortulaca oleracea L\u003c/em\u003e. showed a significant increase in total phenol when 1mM SA was foliar sprayed under drought stress, in comparison to plants treated with drought stress alone [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e].[\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e] stated that the interaction effects of 1 mM SA application and increased irrigation interval significantly enhanced TPC of milk thistle leaves. These phenolic compounds make the plants resistant to drought stress by reducing ROS production. The increase TPC is considered to be a result of increasing enzymes involve in shikimate pathway due to water stress and SA application. Besides, SA induces phenylalanine ammonia-lyase activity, which is a major enzyme involved in the early stages of biosynthesis phenolic compounds [\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e]. According to[\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e] the levels of sucrose and total soluble carbohydrate (TSC) content increased significantly in \u003cem\u003eSilybum marianum L\u003c/em\u003e. in response to the increase in drought stress. Sucrose and TSC decrease plant water potential in order to increase the gradient of water potential between soil and plant tissue which facilitated water flow from soil to root via osmosis. On the other hand, TSC and sucrose can stimulate production of secondary metabolites like phenols. In the shikimic acid pathway phenolic compounds are synthesized using soluble carbohydrates during drought stress [\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e].\u003c/p\u003e"},{"header":"5 Conclusion","content":"\u003cp\u003eThis study on Ashwagandha under different moisture stress levels and Salicylic Acid treatments has demonstrated a complex interaction between abiotic stress and plant physiological responses, particularly in relation to growth and secondary metabolite production. The findings showed that while moisture stress negatively impacts plant growth in terms of height, leaf number, SPAD values, and leaf area, it simultaneously triggers an adaptive mechanism enhancing secondary metabolite production, as evidenced by the significant increase in total polyphenolic content. The application of SA further modulates this response, optimizing plant growth and secondary metabolite production, particularly under severe moisture stress conditions. The Salicylic acid application gradually reduced the decremental effects of drought conditions. From the results of the present study foliar spray of 1mM SA can be recommended to enhance the root yield, plant growth and secondary metabolite production of Ashwagandha plants under the moisture stress of 50% field capacity.\u003c/p\u003e \u003cp\u003eThis research highlights the potential of using agronomic strategy, which consists with controlled moisture stress and SA application, to enhance the medicinal value of Ashwagandha. By controlling environmental stressors and growth regulators, it is possible to optimize conditions for maximizing the yield and quality of medicinal compounds, to address the growing global demand for high-quality medicinal plants without expanding cultivated land area. Moreover, these findings contribute to the understanding of plant stress physiology, offering insights into how plants can be engineered to thrive under stress while enhancing their pharmacological potential. This balance between stress management and metabolic enhancement opens new avenues for agricultural practices aimed at sustainable cultivation of medicinal plants, ensuring the availability of key pharmacological compounds and supporting the health and wellness industry.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNo funding was received for conducting this study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability (data transparency)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll the data are presented in tables and figures in the manuscript and further inquiries can be directed to the corresponding authors.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCode availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthical Statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe seeds used in this study was obtained from Prof. Siripla Subasinghe (one of authors) Faculty of Agriculture, University of Ruhuna, Sri Lanka. Therefore, the plants used in the study comply with national guidelines without further affirmation.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eContribution\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eM.S.Jayathilaka : Conceptualization, Methodology, Investigation, Review and editing , Data analysis, \u0026nbsp;Data interpretation, Writing-original draft\u003c/p\u003e\n\u003cp\u003eSiripala Subasinghe, KMW Rajawatta , K.H.T. Karunarathna : Conceptualization, Methodology, Review and editing,supervision, validation\u003c/p\u003e\n\u003cp\u003eAll authors read and approved the final manuscript.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eSapra NC, Kalyanrao P, Sasidharan N, Das A, and Susmitha P, \u0026ldquo;Effect of Mechanical, Chemical, Growth Hormone and Biofertilizer Treatments on Seed Quality Enhancement in Ashwagandha (Withania somnifera Dunal),\u0026rdquo; Med Aromat Plants (Los Angel), vol. 9, no. 3, pp. 1\u0026ndash;4, 2020, doi: 10.35248/2167-0412.20.9.350.\u003c/li\u003e\n\u003cli\u003eTNAU, \u0026ldquo;TNAU Agritech Portal:: Sustainable Agriculture.\u0026rdquo; Accessed: Aug. 21, 2023. [Online]. Available: https://agritech.tnau.ac.in/farm_enterprises/Farm%20enterprises_%20Ashwagantha.html\u003c/li\u003e\n\u003cli\u003eS. Chandra, P. Chatterjee, P. Dey, and S. 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JUNE, Jun. 2015, doi: 10.3389/fpls.2015.00462.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"discover-plants","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"Learn more about [Discover Plants](https://link.springer.com/journal/44372)","snPcode":"44372","submissionUrl":"https://submission.springernature.com/new-submission/44372/3","title":"Discover Plants","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Discover Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Drought stress, Moisture stress, Polytunnel, Root yield, Salicylic acid","lastPublishedDoi":"10.21203/rs.3.rs-4689316/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4689316/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eAshwagandha is a medicinal herb that has very high demand in the global market. Root is the main part harvested from this plant because of its rejuvenating properties. This study investigates the effects of moisture stress and salicylic acid on the growth yield and polyphenol content of Ashwagandha. A polyhouse experiment was conducted where Ashwagandha plants were grown under four levels of soil moisture that included 50%, 60%, 80%, and 100% of field capacity (FC) and four Salicylic acid (SA) levels as 10 mM,1 mM,0.1 mM and control was applied as a foliar spray as treatments to induce the secondary metabolite production. Plant height, number of leaves, number of fruits, number of fallen leaves, SPAD value and leaf area, root dry of the root, free proline content and total polyphenol content were measured. The highest root dry weight (7.62g) and total polyphenol content (3.78 GAE mg/g d.w.b) was reported with the interaction effect of 50% of FC and 10\u003csup\u003e\u0026minus;\u0026thinsp;3\u003c/sup\u003eM SA application. Under the combined application of 50% of FC and 1 mM SA the dry weight and total polyphenol content was increased by 2.7 folds and 12.49 folds respectively compared to control (100% FC and without SA). These findings suggest that a moisture stress level of 50% field capacity combined with a 1 mM SA foliar spray can significantly enhance the root yield and polyphenol content of Ashwagandha.\u003c/p\u003e","manuscriptTitle":"Growth and Polyphenol Content of Ashwagandha (Withania somnifera L.Dunal) under Combined Moisture Stress and Salicylic Acid Treatment","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-08-05 09:29:06","doi":"10.21203/rs.3.rs-4689316/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2024-09-04T05:58:41+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-07-28T12:55:20+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-07-25T09:25:06+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"292550494385634437167893113635075611447","date":"2024-07-24T13:49:41+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"68406511731940316087315745664167994103","date":"2024-07-22T21:27:17+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"216371523966878414655108657347829368500","date":"2024-07-22T13:04:16+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2024-07-22T11:28:13+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-07-12T08:29:47+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-07-11T05:02:26+00:00","index":"","fulltext":""},{"type":"submitted","content":"Discover Plants","date":"2024-07-05T03:19:38+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"discover-plants","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"Learn more about [Discover Plants](https://link.springer.com/journal/44372)","snPcode":"44372","submissionUrl":"https://submission.springernature.com/new-submission/44372/3","title":"Discover Plants","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Discover Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"90773b0c-aba9-4c1b-85d8-10168ee19f94","owner":[],"postedDate":"August 5th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2025-02-27T03:23:16+00:00","versionOfRecord":[],"versionCreatedAt":"2024-08-05 09:29:06","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-4689316","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4689316","identity":"rs-4689316","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}