Adaptive physiological response, carbon partitioning, and biomass production of Withania somnifera (L.) Dunal grown under elevated CO₂ regimes

Abstract

Winter cherry or Ashwagandha (Withania somnifera) is an important medicinal plant used in traditional and herbal medicine system. Yet, there is no information available on response of this plant to changing climatic conditions particularly elevated atmospheric CO₂ concentrations. Therefore, we conducted an experiment to examine the effect of elevated CO₂ concentrations (ECs) on Withania somnifera. The variations in traits of physiological adaptation, net primary productivity, carbon partitioning, morphology, and biomass in response to elevated CO₂ concentrations (ambient, 600 and 800 µmol mol⁻¹) during one growth cycle were investigated within the open top chamber (OTC) facility in the foothill of the Himalayas, Dehardun, India. ECs significantly increased photosynthetic rate, transpiration rate, stomatal conductance, water use efficiency, soil respiration, net primary productivity and the carbon content of plant tissues (leaf, stem, and root), and soil carbon. Furthermore, ECs significantly enhanced biomass production (root and shoot), although declined night leaf respiration. Overall, it was summarized that photosynthesis, stomatal conductance, water use efficiency, leaf, and soil carbon and biomass increased under ECs rendering the physiological adaptation to the plant. Increased net primary productivity might facilitate mitigation effects by sequestering elevated levels of carbon dioxide. We advocate further studies to investigate the effects of ECs on the accumulation of secondary metabolites and health-promoting substances of this as well as other medicinal plants.

Introduction

The atmospheric CO₂ has changed from its preindustrial concentration of 280 µmol CO₂ mol⁻¹ to the current level of approximately 400 µmol CO₂ mol⁻¹ largely due to anthropogenic activities. If atmospheric CO₂ levels continue to increase at the current level (2.11 µmol CO₂ mol⁻¹), it is projected to reach 720–1000 µmol CO₂ mol⁻¹ causing increased air temperature (2.6–5.4 °C) before the end of this century (IPCC 2007; Dlugokencky and Tans 2017). Increasing atmospheric CO₂ concentration is also increasing the uncertainty in precipitation which could decline growth and biomass of crops and influence the performance, adaptation, and survival of vegetation system globally (Thompson et al. 2005). Elevated CO₂ (EC) directly affects the species growth performance by shifting their physiological tolerance limits to cope with modified climatic conditions. EC is already forcing living organisms and ecosystems to adapt either through shifting habitat, changing life cycles and the development of new physical and/or physiological traits (CBD 2014; Zari 2014; Tilman and Lehman 2001). Species that are unable to adapt might face extinction (Thomas et al. 2004; Urban 2015). According to predictions, approximately one million species may extinct due to climate change (Gopalakrishna et al. 2011). However, impacts of climate change are expected to vary across regions (Parry et al. 2007). Studies indicate that lately in addition to shifting phenology, plant species have started to conform to latest climate adjustments through alteration at the metabolic and physiological levels of the species (Buckley et al. 2013) or through shifting species ranges (Parmesan 2006). The impact of increased CO₂ concentrations on the adaptation of agricultural species has been understood widely (Bunn et al. 2015; Saadi et al. 2015). However, there is dearth of knowledge regarding the physiological response and adaptations of medicinal plants to increasing CO₂ concentrations. This information is urgently required for understanding the adaptive behavior, ultimately biomass, and yield in future conditions.

Winter cherry or Ashwagandha (Withania somnifera) is one of the important ancient medicinal plants utilized for curing various diseases. It is an important plant species used since ancient era and has been mentioned in various pieces of literature particularly Ayurvedic, Unani and African traditional medicines (Patwardhan et al. 1998; Singh et al. 2015). Due to it is medicinal properties leading to huge demand in the domestic and international markets, World Health Organization (WHO) and National Medicinal Plant Board (NMPB) of India are continuously focusing on cultivation, conservation, sustainable exploration and its management for future demand (Mirjalili et al. 2009; Prajapati et al. 2003; NMPB; WMO). Effects of climate change particularly EC on medicinal plants could be detrimental for the economy of local communities and medical industries. Studies on medicinal plant’s response to EC have been carried out on species such as Digitalis lanata (Stuhlfauth and Fock 1990), Mentha spicata, Thymus vulgaris, Mentha aquatica (Tisserat 2002), Hymenocallis littoralis (Idso et al. 2000), Ocimum basilicum (Singh et al. 2016), Hizikia fusiforme (Zou and Gao 2005), Podophyllum hexandrum (Chaturvedi et al. 2009), Papaver setigerum (Ziska et al. 2008), Scutellaria barbata and Scutellaria lateriflora (Stutt et al. 2008), Thymus vulgaris (Vurro et al. 2009), Halia bentong, Halia bara (Ghasemzadeh et al. 2010), Vernonia herbacea (Oliveira et al. 2010), and Catharanthus roseus (Saravanan and Karthi 2014). The near tripling of EC increased the dry biomass production of the woolly foxglove (Digitalis purpurea) plant by 63 and 83% under dry and well-watered conditions, respectively. Stuhlfauth et al. (1987) also obtained similar results when near tripling of EC enhanced plant dry weight biomass by 75% with 15% increase in digoxin per unit dry weight of plant material. Later, Idso et al. (2000) investigated the response of spider lily plants grew at EC of 400 or 700 ppm and reported 48 and 56% increment in above and below ground plant biomass, respectively. Zobayed et al. (2003) reported an increase of net photosynthetic rates of herb Hypericum perforatum by 124 and 107% greater dry weight under EC (1000 ppm). Above-ground biomass of the plants at elevated CO₂ increased too (Ziska et al. 2008). Ibrahim et al. (2012) grew oil palm Elaeis guineensis at ECs 400, 800, and 1200 µmol CO₂ mol⁻¹ and reported increase in flavonoid contents by 86 and 132%, and total phenolics by 52 and 91% at 800 and 1200 µmol CO₂ mol⁻¹.

However, we do not have sufficient knowledge of Withania somnifera in respect of physiological adaptation, carbon partitioning, net primary productivity, and biomass production to EC. The magnitude of the direct and indirect effect of EC on stimulation of traits related to physiological adaptation especially photosynthesis, water use efficiency, growth, and biomass production of medicinal plants has significant economic and social bearing. Thus, the study was performed to understand the adaptive physiological response, carbon partitioning, net primary productivity, growth, morphology, and biomass production of Withania somnifera grew at various levels of EC, i.e., ambient, 600 and 800 µmol CO₂ mol⁻¹.

Materials and methods

Description of open top chamber (OTC) facility

The experiment was conducted in the open top chamber (OTC) facility of Ecology, Climate Change and Forest Influence Division, Forest Research Institute—Dehradun, Uttarakhand—India (32°20′44.2172″N, 78°0′41.6185″E and 668 m.a.s.l.). Each OTC (3.0 m, width × 3.0 m, length × 4.0 m height) was constructed using high-quality multilayer polycarbonate sheet of 3.0–4.0 mm thick and 80–85% transmittance (Singh et al. 2018). CO₂ gas of commercial grade (100%) was released into the chambers at the required concentration. One OTC was used as control chamber. Another two OTCs, EC (600 and 800 µmol mol⁻¹) was maintained during whole growth cycle of Withania somnifera, from seedling to harvest stage. The CO₂ concentration was measured by sensors installed in each chamber. The EC in each chamber was maintained using the solenoid valves of the corresponding chamber.

Seedlings preparation of Withania somnifera

The seeds of Withania somnifera acquired from Non-wood Forest Product Division, Forest Research Institute—Dehradun, Uttarakhand, India. Seeds were sown in plastic trays with growing media (2:1:1; soil:sand:manure). One-month-old seedlings were transferred to pots and kept for 4 days outside of OTC for acclimatization in pot conditions. Subsequently, seedlings were placed in OTCs and left for 4 days to acclimatize in OTCs conditions. Finally, the required concentration of CO₂ was supplied into each OTC, except control. CO₂ treatment was given during whole one growth cycle of plants. Regular watering and weeding were carried to maintain the uniform soil moisture.

Measurement of adaptive physiological traits

The adaptive physiological traits, i.e., photosynthetic rate (μmol CO₂ m⁻² s⁻¹), transpiration rate (mmol H₂O m⁻² s⁻¹), stomatal conductance (mol H₂O m⁻² s⁻¹), and intercellular CO₂ concentration (µmol CO₂ mol⁻¹), were measured during vegetative phase using portable photosynthetic system (Model 6400 XT- LICOR, Incl, USA). The youngest and fully expended leaves were preferred for measuring physiological parameters between 09:30 am and 12:30 pm in clear sky condition. Forty observations for each parameter were recorded from eight plants (five replicates from each plant). The instantaneous water use efficiency (A/E) was computed as the ratio of CO₂ photosynthetic rate (A) and transpiration (E). Intrinsic water use efficiency (A/gs) was estimated as the ratio of photosynthetic rate (A) to stomatal conductance (gs), whereas intrinsic carboxylation efficiency (A/Ci) was calculated using ratio of photosynthetic rate (A) to intercellular CO₂ concentration (Ci) furthermore ratio of intercellular CO₂ concentration (Ci) to stomatal conductance (gs) was calculated for obtaining the intrinsic mesophyll efficiency (Ci/gs) (Warrier et al. 2013; Singh et al. 2015, 2018).

Estimation of leaf and soil respiration, and net primary productivity

Night leaf respiration (μmol CO₂ m⁻² s⁻¹) was measured from fully developed leaf during vegetative stage between 4:30 to 5:00 am and soil respiration was measured between 09:30 am and 12:30 pm with help of portable photosynthesis system (Model 6400 XT- LICOR, Incl, USA). For measuring soil respiration from pot soil, a chamber of 10 cm × 10 cm × 10 cm was constructed using polyvinyl chloride sheet with 80–85% transmission level of natural light. The chamber was placed on the top of the pot soil, made air tight by pressing it into soil about 2 cm deep and sealing it with soil from outside to break the contact of inside condition to the outer atmosphere. The packed chamber was connected to reference pipe of portable photosynthesis system to visualize variations in CO₂ concentration on a digital screen. Before measuring the soil CO₂ respiration, we recorded the initial CO₂ concentration and left for 1 h interval to observe the change in CO₂ concentration. After 1 h interval, the final CO₂ concentration was recorded. The soil CO₂ respiration (mg CO₂ m⁻² h⁻¹) was then calculated by subtracting the initial value of CO₂ from the final value using following equation (Singh et al. 2017):

$$\text{C}{\text{O}}_2\text{flux}=\frac{\mathrm{\Delta }X\times \text{ECV}\left(\text{STP}\right)\times \text{MW}\times 1000\times 60}{1000000\times 22400\times T\times A}$$

where ∆X = Difference in flux value between 60 and 0 min (converted to ppm based on the standard CO₂ values), ECV (STP) = effective chamber volume at standard temperature and pressure, MW = molecular weight of CO₂, T flux time (min), and A area of the chamber.

Moreover, net primary productivity (NPP; μmol CO₂ m⁻² s⁻¹ = GPP − respiration) was calculated by subtracting the carbon lost through plant respiration from the carbon assimilated during the photosynthesis (Gross Primary Production or GPP) (Singh et al. 2017).

Biomass and carbon estimation

Biomass allocation in plant parts was calculated by carefully uprooting the plants and segregating into leaves, stems, and root parts. The roots were cleaned with distilled water to remove soil particles sticking to root hairs. The fresh plant parts were weighed and subsequently oven dried at 65 °C until a constant weight reached and weighed again (Wu et al. 2013; Singh et al. 2018). Organic carbon was estimated following Walkley and Black (1934) method. In 500 ml conical flask, the plant tissue (0.1 gm) was first mixed with 30 ml of 1N potassium dichromate solution followed by 20 ml of concentrated sulphuric acid. After swirling for few minutes, the solution was incubated at room temperature for 40 min. Then, 200 ml of distilled and 10 ml orthophosphoric acid was added followed by five drops of diphenylamine indicator. At last, the solution was titrated against ferrous ammonium sulphate solution to reach an end point of brilliant bright green color solution. The following equation was used to calculate organic carbon in plant parts:

$$\%C=\frac{(B-T)\times S\times 0.003\times 1.33\times 100}{W}$$

where B amount of ferrous ammonium sulphate required in blank titration, T amount of ferrous ammonium sulphate required in sample titration, S strength of ferrous ammonium sulphate (from blank titration), and W weight of plant sample.

Statistical analysis

We used the SPSS 16.0 software to carry out the multivariate general linear model to observe the existence of significant mean difference in response of physiological parameters, biomass, and carbon accumulation at different CO₂ levels. Further post hoc Tukey test was performed to identify the homogeneous subsets.

Results

Adaptive physiological response to EC

The traits of adaptive physiological response showed significant difference across control and elevated CO₂ concentrations (Table 1). The photosynthetic rate increased to 38.63 and 86.77% at EC of 600 and 800 µmol mol⁻¹, respectively, to ambient CO₂ level. It was reported that the increased CO₂ concentration induced higher water lost form leaf through the process of transpiration. Approximately, transpiration rate was 20.16 and 36.18% at EC (600 and 800 µmol mol⁻¹) level when compared to control. The stomatal opening and closing in terms of stomatal conductance was found to be stimulated significantly under EC (800 µmol mol⁻¹), but not for 600 µmol mol⁻¹ compared to ambient (Table 1). Stomatal conductance was 45.19% higher in EC 800 µmol mol⁻¹ condition as compared to ambient. The study also reported a significant increase in intercellular CO₂ concentration (Table 1) and the increase was 69% higher at 800 µmol mol⁻¹ and 26.52% higher at 600 µmol mol⁻¹, whereas lowest in ambient conditions. Instantaneous (A/E) and intrinsic (A/gs) water use efficiency had the same trend as seen in stomatal conductance (Table 1). The percentage increase of A/E was 36.31% higher and for A/gs around 28.5% at EC 800 µmol mol⁻¹ condition than ambient. The carboxylation efficiency showed significant increment at elevated CO₂ concentrations than ambient (Table 1). Furthermore, a tendency of higher value was observed for the intrinsic mesophyll efficiency under 800 µmol mol⁻¹ CO₂ level when compared to the plant grown at ambient condition (Table 1). Mesophyll efficiency increased to 16.29% at 800 µmol mol⁻¹ CO₂ compared to ambient. It can be seen from the results that there was an enhancement of mesophyll efficiency around 10% at 600 µmol mol⁻¹ and 16.20% at 800 µmol mol⁻¹ CO₂ level (Table 1).

Table 1.

Effects of elevated CO₂ concentrations (EC) on physiological parameters of Withania somnifera

Physiological traits	CO2 concentration (μmol CO2 mol−1)			P value
	Ambient	EC (600)	EC (800)
A (µmol CO2 m−2 s−1)	15.03a ± 0.23	20.83b ± 0.08	28.06c ± 0.31	0.000
E (mmol H2O m−2 s−1)	7.04a ± 0.27	8.45b ± 0.34	9.581c ± 0.22	0.000
Gs (mol H2O m−2 s−1)	0.35a ± 0.01	0.40a ± 0.02	0.51b ± 0.02	0.000
A/E	2.16a ± 0.09	2.49a ± 0.10	2.94b ± 0.09	0.000
A/gs	43.29a ± 1.90	52.17b ± 2.26	55.65b ± 2.38	0.002
Ci (µmol CO2 mol−1)	284.17a ± 7.52	359.52b ± 8.76	480.25c ± 6.22	0.000
A/Ci	0.053a ± 0.001	0.058b ± 0.001	0.0585b ± 0.001	0.011
Ci/gs	816.63a ± 34.93	897.82ab ± 34.31	949.62b ± 30.31	0.032
NLR (μmol CO2 m−2 s−1)	5.65c ± 0.10	4.70b ± 0.04	3.99a ± 0.11	0.000
SR (mg CO2 m−2 h−1)	3.13a ± 0.19	4.64b ± 0.37	7.79c ± 0.36	0.000
NPP (μmol CO2 m−2 s−1)	11.89a ± 0.35	16.19b ± 0.35	20.27c ± 0.62	0.000

Means not sharing a common letter are significantly different at p ≤ 0.05

A photosynthetic rate, E transpiration rate, gs stomatal conductance, A/E instantaneous water use efficiency, A/gs intrinsic water use efficiency, Ci intercellular CO₂ concentration, A/Ci intrinsic carboxylation efficiency, Ci/gs intrinsic mesophyll efficiency, NLR night leaf respiration, SR soil respiration, NPP net primary productivity, ± the standard error of the mean (SEM). N = 8.

Response of leaf and soil respiration, and net primary productivity to EC

The response of leaf and soil respiration and net primary productivity (NPP) against EC is presented in Table 1. Night leaf respiration significantly reduced in plants treated with EC than the one in ambient. The EC had negative impact on night leaf respiration by suppressing 16.80 and 29.38% significantly for the plant grown at 600 and 800 µmol mol⁻¹ compared to ambient. Soil respiration, a major component in the global carbon cycle, is affected by elevated atmospheric CO₂ concentrations. The soil respiration was higher by 48.24 and 148.88% under 600 and 800 µmol CO₂ mol⁻¹, respectively, compared to ambient CO₂. The NPP showed a positive response to rising CO₂ by showing elevated values by 36.16 and 70.47% for the plants exposed to 600 and 800 µmol mol⁻¹ CO₂ to ambient condition.

Response of carbon and biomass partitioning to EC

Interestingly, the EC showed substantial effects on leaf, stem, and root organic carbon (Table 2). The leaf organic carbon of the plant increased by 8.3% at 600 µmol mol⁻¹ and 14.2% at 800 µmol mol⁻¹ than the ambient. The parameters investigated after harvesting of the plant are presented in Table 2. The stem organic carbon increased by 5.63% at 600 µmol mol⁻¹ and 8.90% at 800 µmol mol⁻¹ to the ambient. Accumulation of root organic carbon also increased under EC both at 600 and 800 µmol mol⁻¹ concentrations by 10.4 and 15.8% (Table 2). There was a uniform accumulation of dry matter of W. somnifera grown under elevated CO₂ concentration recorded for all plants (Table 2). Dry weight of leaves was too increased by 53.2 and 90.7% at 600 and 800 µmol mol⁻¹, respectively. Stem dry weight was also enhanced when plants were exposed to elevated CO₂ with maximum value at 800 µmol mol⁻¹ and lowest under controlled condition. Simultaneously, root dry weight was also maximum at 800 µmol mol⁻¹ (Table 2). The magnitude of percent increase in leaves dry biomass accumulation was 90.66 and 53.19% at 800 and 600 µmol mol⁻¹, respectively. Similar trend was observed in stem and root dry biomass accumulation, i.e., 93.06 and 32.59% in stems and 14.24 and 8.75% in roots when exposed to 800 and 600 µmol mol⁻¹ CO₂ concentrations.

Table 2.

Effects of elevated CO₂ concentrations (EC) on carbon/biomass partitioning parameters of Withania somnifera

Carbon/biomass partitioning	CO2 concentration (µmol CO2 mol−1)			P value
	Ambient	EC (600)	EC (800)
Organic carbon leaves (%)	49.59a ± 0.30	53.62b ± 0.34	56.61c ± 0.32	0.000
Organic carbon stem (%)	54.60a ± 0.29	57.67b ± 0.28	59.46c ± 0.24	0.000
Organic carbon root (%)	57.46a ± 0.29	63.44b ± 0.34	66.56c ± 0.44	0.000
Leaves biomass (gm plant−1)	7.74a ± 0.32	11.86b ± 0.26	14.76c ± 0.26	0.000
Stem biomass (gm plant−1)	14.03a ± 0.40	18.60b ± 0.24	27.08c ± 0.64	0.000
Root biomass (gm plant−1)	7.88a ± 0.07	8.57b ± 0.08	9.02c ± 0.14	0.000

± Indicates standard error of the mean (SEM). N = 40. Means not sharing a common letter are significantly different at p ≤ 0.05

Relationship between functional traits

Variation in selected adaptive physiological traits (viz. photosynthetic rate, transpiration rate, leaf respiration, soil respiration, net primary productivity, and biomass production is depicted graphically (Fig. 1). The relationship between functional traits is illustrated in Figs. 2 and 3. As per our expectations, we observed a strong correlation between photosynthetic rate (r = 0.99), stomatal conductance (r = 0.81), transpiration rate (r = 0.81), soil respiration (r = 0.90), leaf respiration (r = − 0.94), and net primary productivity (r = 0.94). Root organic carbon and root biomass production were also strongly correlated (r = 0.84).

Fig. 1.

Response of important traits lined to physiological adaptation, carbon partitioning, and biomass production of Withania Somnifera grown under elevated CO₂ concentration

Fig. 2.

Correlation between various physiological parameters. Where, A photosynthetic rate, E transpiration rate, gs stomatal conductance, WUEins instantaneous water use efficiency, WUEintr intrinsic water use efficiency, Ci intercellular CO₂ concentration, A/ci carboxylation efficiency, Ci/gs mesophyll efficiency, SR soil respiration, LR leaf respiration, NPP net primary productivity, NEE net ecosystem exchange (at pot conditions)

Fig. 3.

Correlation between plant dry biomass and plant organic carbon (where OC.Lvs organic carbon leaves, OC.Stem organic carbon stem, OC.Root organic carbon root, Lvs.Bms leaves biomass, Stem.bms stem biomass, Root.Bms root biomass, Dry.Bms dry biomass)

Discussion

Adaptive physiological response to EC

Our study showed that all the physiological parameters of Withania somnifera increased at EC except leaf respiration. The increase in photosynthetic rate potentially related to rising in the activity of ribulose 1, 5-bisphosphate carboxylase enzyme (Rubisco) that can change the growth and even secondary metabolites (Ibrahim and Jaafar 2011). With an increase in CO₂ concentration, there is an increase in the rate at which carbon is subsumed into carbohydrate and the rate continues to grow until one of the factors becomes limiting. Increase in carbon dioxide cause rapid development of leaf area, which increases availability of transpiring surface subsequently enhancing transpiration rate (Morison and Gifford 1984). Stomatal conductance increased at EC which could be attributed to rapid opening of stomatal guard cells to the elevated CO₂ concentration. Water use efficiency increased by improving plant water relationship, i.e., by increasing plant turgor pressure which is important for the growth processes. Mechanism for the observed response could potentially be as follows. CO₂ helps in accumulation of more carbohydrate concentration that enhances the leaf osmotic potential and ultimately helps to maintain adequate leaf water contents for other phenomena of photosynthesis and growth. Similarly, the growing concentration of climatic carbon dioxide influences the mechanisms of plant photosynthesis, metabolism, and development (Nowak et al. 2004; Ainsworth and Long 2005). The other adaptive physiological process of W. somnifera such as intrinsic carboxylation efficiency (A/Ci) and intrinsic mesophyll efficiency (Ci/gs) exhibited increased trends at EC compared to plants grown at ambient condition. These physiological observations were in congruence to earlier studies (Singh et al. 2018; Warrier et al. 2013; Saravanan 2014). The response of physiological processes of W. somnifera to EC (600 and 800 µmol mol⁻¹) is represented in Table 1 and Fig. 1.

Response of leaf and soil respiration, and net primary productivity to EC

The CO₂ efflux (net respiratory CO₂ evolution originating from respiratory substrates minus the CO₂ being re-fixed by carboxylases) reduced in leaves of C₃ plants (Pinelli and Loreto 2003). This can be explained by stimulation of dark CO₂ fixation catalyzed by phosphoenolpyruvate carboxylase (PEPC), evolved in an evident reduction of CO₂ efflux (Amthor 1997; Drake et al. 1999). Griffin et al. (1996) investigated a reduction of 20–4% in leaf respiration of plant grown under increased CO₂ concentration. The response of leaf respiration of study was also reported in other studies (Hamilton et al. 2001; Tan et al. 2013).

Soil respiration, a major component in the global carbon cycle, is affected by elevated atmospheric CO₂ concentrations. The soil respiration increased linearly with increased CO₂ concentration under the present study. However, effect of elevated CO₂ on soil respiration has rarely been studied. This study reported an increase in soil respiration under elevated CO₂ condition both at 600 and 800 µmol mol⁻¹ than ambient. The maximum return of CO₂ flux from terrestrial ecosystems back to the atmosphere comes from the soil (Raich and Schlesinger 1992). Similar results of stimulation of soil respiration were seen in the wheat field under FACE experiment (Pendall et al. 2001). The increased rate of soil respiration under elevated CO₂ might be due to greater carbon availability for microbial activity through CO₂ induced carbon inputs (Adair et al. 2011). An increment of 20% in soil respiration was reported in a temperate grassland ecosystem by Keidel et al. (2015).

In terrestrial ecosystem, doubling of CO₂ concentrations without change in temperature is predicted increase in Global NPP by 16.3% (Melillo et al. 1993). NPP is a sensitive indicator of climatic and other forms of environmental change (Beer et al. 2010). The exchange of carbon from the atmosphere to the plant is through the stomata. The increase of CO₂ may influence the response of stomata which in turn may affect the plant’s NPP. The result was similar to the study that showed elevated CO₂ may increase rates of photosynthesis, increase productivity, and increase biomass in most C₃ Plants (Houghton 2007). Elevated CO₂ concentrations increase the net exchange of CO₂ between the biosphere and the atmosphere, through stimulation of photosynthesis, increased water and nitrogen use efficiency and increase in biomass (Drake et al. 1997).

Response of carbon and biomass partitioning to EC

Despite the fact that analyzing plant development by surveying leaves, stem and root biomass might be an important beginning stage in deciding plant reaction to elevated CO₂ it might turn out to be fairly a heartless pointer of what is really happening to plant development regarding structure and capacity (Stulen and den Hertog 1993; Taylor et al. 1994; Sattler and Rutishauser 1997). The present study exhibited an enhancement of biomass production under elevated CO₂ condition. Carbon accumulation in roots was higher followed by stem and leaf (Singh et al. 2018; Lin and Wang 1998). Dry weight of leaved increased by 90.66% under 800 µmol mol⁻¹ and 53.19% at 600 µmol CO₂ mol⁻¹ than ambient. Similarly, shoot dry weight increased by 93.06 and 32.59%, root dry weight by 14.24 and 8.75% under 800 and 600 µmol mol⁻¹, respectively. More biomass was allocated to the stem tissues while minimum to the root tissues for the plant under elevated CO₂ concentration. Besides above, the carbon allocation to leaves, stem, and root parts was also enhanced under an elevated CO₂ condition. The plants become taller and produce maximum biomass under elevated CO₂ condition. It was also observed that rising of atmospheric CO₂ could significantly stimulate leaf photosynthesis and above-ground dry weight biomass production of a C3 species, Xanthium strumarium (Ziska 2001). Similar results were reported for biomass and photosynthesis for Cirsium arvense L. (Ziska 2002). The biomass production and water use efficiency of the plants exposed to elevated CO₂ condition too increased. The corresponding results were reported by Singh et al. (2018).

Relationship between functional traits

The relationship between functional traits in presented in Figs. 2 and 3. The increased CO₂ concentration had stimulatory effect on photosynthesis rate that positively correlated to stomatal conductance and transpiration rate. More stomatal conductance leads to exchange higher rate of CO₂ and H₂O from the leaf and extended CO₂ atmospheric condition. Stomatal conductance was positively correlated to water use efficiency which produces more carbohydrate and ultimately higher biomass production. Similar relationship was observed by Singh et al. (2018). Night leaf respiration (NLR) suppressed under EC. Amthor et al. (1992) stated that there was an immediate and fully reversible decrease in leaf respiration rates when atmospheric CO₂ partial pressures are increased due to increased CO₂ concentration. This reduction mechanism of respiration has not been elucidated, but may be related to dark CO₂ fixation by phosphoenolpyruvate carboxylase (PEPc), altered cytosolic pH, or direct effects on membranes or enzymes (Amthor et al. 1992). Similar relationship was also reported by other researcher (Griffin et al. 1996; Hamilton et al. 2001; Tan et al. 2005). The photosynthesis was enhanced under EC which eventually increased NPP and total biomass production. The NPP, photosynthesis, and biomass in plant tissues were also correlated with each other. Increasing rate of photosynthesis enhances NPP and biomass under elevated CO₂ condition in this study. Total daily photosynthesis (GPP) and net primary production (NPP) correlated positively and were consistently higher under the elevated CO₂ treatment than the ambient CO₂ treatment (Cheng et al. 2000). This study reported positive correlation between elevated CO₂ and soil respiration. The increased rate of soil respiration under elevated CO₂ might be due to greater carbon availability for microbial activity through CO₂-induced carbon inputs (Adair et al. 2011). A relationship was also observed by Keidel et al. (2015) in a temperate grassland ecosystem, where soil respiration increased by 20%.

Conclusion

The present study concludes that the W. somnifera could adapt to changing climatic conditions especially rising of atmospheric CO₂ concentration. In such conditions of elevated CO₂ concentration, the physiological processes likely photosynthetic rate, water use efficiency, transpiration, and stomatal conductance might be enhanced contributing to plant growth and ultimately more biomass production particularly the root biomass in W. somnifera. Therefore, this species could be one of the most important medicinal species which will be more suitable to mitigate climate change in the future. Consequently, this species might be more adaptable in near future and can be used in suitable production of W. Somnifera and meet the increasing demand of pharmaceutical industries/medical to produce ayurvedic medicine in the country.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest in the publication.