Boron Application in Yermosols Improves Grain Yield and Quality of Chickpea (Cicer arietinum L.)

1 Introduction

Chickpea (Cicer arietinum L.) is one of the prominent pulses and widely cultivated in the world after beans and soybean. In Pakistan, it’s area under cultivation is 940 thousand hectares having annual seed production is 545 thousand tones (Govt. of Pakistan, 2019–2020). Chickpea average yield is lower in Pakistan than other chickpea growing countries. It is a well-adjusted crop in arid to semi-arid regions and is commonly cultivated on dry conditions (Kagan and Kayan, 2014). It is common and cheap protein source particularly for poor peoples of unindustrialized countries and thus can help to decrease malnutrition and improves human health. Chickpea has ability to bear low-risk in semi-arid environment due to its intensive rooting features (Rehman et al., 2014). Among different factors causing low yield, lack of balanced fertilization is the most important factor, especially deficiency of micronutrients (Borie et al., 2006). Boron (B) has significant microelement which is essential for crop growth, but quickly becoming scarce in soils (Tahir et al., 2009). However, Pakistani calcareous soils are B-deficient, that results in lower crop production and economic benefits (Atique-ur-Rehman et al., 2020). It has dominant role in cell growth, pollen tube development, membranes stability, grain development and fertilization (Zhao et al., 2001).

Boron deficiency and surplus leads to physiological and morphological disorder in crop plants (Kastori et al., 2008). Boron deficiency symptoms are poor root proliferation, inferior development of apical meristem, weak leaves growth, lower chlorophyll and photosynthetic rate, disturbance in ion channel; higher phenolic and lignin concentrations, and lower crop production (Wang et al., 2015). Boron deficiency leads to higher enzymatic and non-enzymatic oxidation reactions by using phenolic contents as substrate, outcomes in higher polyphenolic and quinine concentrations, which are hazardous for crop production (Hajiboland et al., 2013). Boron application is reported to have significant improvement in chickpea development and its scarcity cause destructive effects on chickpea production. Boron deficiency may be under alkaline soil, and accessibility lower at pH 6.5–7.0 or above on sandy soils or having lower organic content soils (Sims 2000). Hence, chickpea shows more response to the application of B as compared to other legumes, even though dissimilarity among chickpea cultivars correlated to B deficiency (Ahlawat et al., 2007). Boron application in soil is necessary when its concentration is less than 0.3 mg kg per ha (Ahlawat et al., 2007).

Deficiency of B may be corrected in a number of ways, however, soil and exogenous application strategy are more suitable way for effectiveness of microelements especially B (Rehman et al., 2019). Agronomic biofortification of micronutrients is easier and can be applied by exogenous spray, soil incorporation and seed coating methods (priming and coating) (Rehman et al. 2018a, 2018b). It has been observed that soil applied B showed significant role on chickpea dry matter production while foliar application has prominent effect on grain production (Padbhushan and Kumar, 2014).

In Pakistan, limited research was done to evaluate the consequence of B application in calcareous condition. Adjusting the B concentration in soil may be helpful for improving growth and production on calcareous Yermosols. The objective of present study was to surmise the response of chickpea to different concentrations of soil applied B on Yermosols. It was hypothesized that chickpea cultivars will exhibit changing behavior to soil applied B with improvement in grain yield and quality.

2 Materials and methods

The experiment was performed in a wire-house at Bahauddin Zakariya University, Multan (30.10 ^oN, 71.25 ^oE and 421 ft. altitude above sea level) in winter during 2017–18. Calcareous alluvial soil of 20 kg weight and 1.04 mg m⁻³ bulk density were added in earthen pots (25 cm × 40 cm), firstly the inner side of pots were enclosed with polyethylene sheet. Water holding and field capacity were kept with 33% and 70% through using de-ionized water respectively. Before performing study, different soil physico-chemical characteristics were determined. Soil texture was silty clay loam calculated through Hydrometer and fits to Sindhlianwali series (hyperthermic, sodic haplocambids) in USDA Haplic Yermosols classification. Soil pH was 8.3 recorded through pH meter (Beckman 45 Modal, US) and EC was 12 dS m⁻¹ noted by EC meter (VWR Conductivity Meter DIG2052). Soil organic content was 0.78% (Walkely-Block method), N was 4.16% (Kjeldahl Method), extractable-P was 7.65% (Olsen’s Method) and potassium was 270%. Soil analysis showed 0.24 mg kg⁻¹ of B that it was deficient which determined through procedure modified by Wolf (1974). Environmental data of crop growing period during 2017–2018 is given in Table 1.

Two chickpea cultivars viz. Noor-2013 and Bittle-98 with five soil applied B concentration viz. B₀ (control), B₁: 0.18; B₂: 0.27; B₃: 0.36 and B₄: 0.45 mg B kg⁻¹ were tested and each treatment was replicated five times. Borax (Na₂B₄O₇·10H₂O) was used as source of B. Solution of different B concentrations were applied on dried soil than mixed into soil manually. Ten seeds were sown into each pot and kept five after 20 days after sowing. Soil moisture contents checked regularly and maintained up to 70%. Recommend doses of N and P @ 34 and 85 kg ha⁻¹ were equally incorporated into the pots on soil weight basis. At maturity, pods were separated and air dried to detach seed and seed dried weight was recorded. Standard agronomic production practices were applied during growing period.

At maturity plant height (cm) and number of pods were measured. Pods were separated manually to find out seeds per pod and plant. Weight 100-grain was measured by using an analytical balance (Model Number: HC2204) after oven-drying at 70 °C for 24 h. Chlorophyll contents were determined after 45 days of sowing by ‘SPAD-502′ chlorophyll meter. For measurements of transpiration and stomatal conductance, portable photosynthesis system (LI-6200, LI-COR, Inc., Lincoln, NE) was used. Water use efficiency (WUE) was calculated by dividing photosynthesis with transpiration rate. For determination of B contents in plants, mature plants were uprooted and separated into roots, shoots, leaves and seeds. Separated parts were cleaned with distilled water and dried for constant weight in a thermo-ventilated oven at 65 ± 5 °C. Material was crushed into a John Wiley mill and sieved through a 40 mesh screen. The powdered material was converted into ash by using muffle furnace at 550 °C for six hours. After that 0.36 N H₂SO₄ was added and the B concentration was determined by spectrophotometer at 420 nm wavelength using azomethine-H method (Bingham, 1982). Soluble protein (mg g⁻¹ dry weight) content in the seed sample was estimated by Folin-Ciocalteau reagent method Lowry et al. (1951). For determination of B content in seed, 500 mg seed was milled in a pestle and mortor using 5–10 ml of potassium sodium tartarate buffer, subjected to centrifugation and the supernatant was collected. From extract, 0.2 ml was diluted up to 1 ml by addition of distilled water and allowed rest for ten minutes. Folin-Ciocalteau reagent (0.5 ml) was mixed into it and covered in dark at room temperature for 30 min. The concentration of blue color established was measured at 660 nm. Blank was prepared without plant sample and the absorbance was measured. Starch content in grains was estimated by Anthrone Reagent method (Sadasivam and Manickam, 1996). For this purpose, 0.5 g sample was standardized then centrifuged in 80% ethanol to eliminate sugars and continued to wash until green color of anthrone reagent developed. Filtrate was desiccated well into a water bath, consisted of 5 ml water and 6.5 ml of 52% perchloric acid, centrifuged at 0 °C for 20 min and supernatant was pooled repeatedly using fresh perchloric acid and made up to 100 ml. Distilled water was added in 0.1 ml supernatant and volume was made up to 1 ml, then 4 ml anthrone reagent was added and warmed into a water bath for eight minutes. It was air-cooled and green color intensity was read in spectrophotometer at 630 nm. Standard curve was prepared with standard glucose solution. Glucose content in sample was obtained from standard graph then multiplied by 0.9 factor to attain starch content. Starch contents were calculated by applying the procedure of Rong et al. (1996). The termination and gradation of sugar compounds from seeds were carried out by an Agilent 1100 series HPLC system (Agilent, USA), (Zeng et al., 2007).

Experimental treatments were set in completely randomized design with factorial arrangement. SAS software (Version 9.1; SAS Institute, Cary, NC, USA) (SAS Institute, 2008) was used for ANOVA on all traits and their means were marked by applying Duncan’s multiple range test at significance (p less than 0.05) level of 5% (Steel, 1997). Analysis of correlation and scatter plot with linear regression were prepared by applying Minitab (Version 12, State College, PA, USA) (Minitab, Inc. 1998) to evaluate the effect of studied traits particular on the achieved results.

3 Results

Soil applied 0.45 mg B kg⁻¹ produced 13.5% taller plant height than control (Table 2). Similarly, no. of pods of Bittle-98 was measured 38.7% maximum than Noor-2013. Boron application of 0.45 mg kg⁻¹ gave 15% more no. of pods. Among different treatments, Bittle-98 with 0.45 mg kg⁻¹ had 47.9% higher no. of pods than Noor-2013 with B₀ (Table 2). Likewise, 0.45 mg B kg⁻¹ had 34.8% significantly maximum no. of grains per pod than control. Cultivar Noor-2013 produced 24.6% more no. of grains per pod than Bittle-98 in different concentrations of B (Table 2). Cultivar Noor-2013 produced 41.5% more no. of grains per plant than Bittle-98 under different concentrations of B (Table 2). Regarding 100-grain weight, 0.45 mg kg⁻¹ had 11.6% more grain weight than control. The cultivar Noor-2013 produced 43% heavier grains than Bittle-98 under different concentrations of soil applied B. Likewise, 0.45 mg B kg⁻¹ produced 51.8% and 28.5% higher grain and biological yield than control, respectively. The cultivar Noor-2013 produced 31.2% and 3.4% higher grain and biological yield per plant than Bittle-98 under different concentrations of B, respectively (Table 3).

Boron applied @ 0.45 mg kg⁻¹ produced 11.8% higher chlorophyll contents than control which was also significantly lower than other B concentrations. Between cultivars, Noor-2013 resulted 3.4% more chlorophyll contents than Bittle-98 (Table 4). Plants applied with 0.45 mg B kg⁻¹ recorded 4.7% more transpiration rate than all other B concentrations. From both cultivars, transpiration rate of Noor-2013 had 0.73% higher than Bittle-98. Likewise, Noor-2013 with 0.45 mg kg⁻¹ gave 5.7% higher transpiration rate than Bittle-98 with control and other treatments. Boron applied @ 0.45 mg kg⁻¹ produced 36.9% higher stomatal conductance than control and other B concentrations. Moreover, Noor-2013 recorded 1.9% higher stomatal conductance than Bittle-98 (Table 4). A 7.7% higher WUE was recorded with 0.45 mg kg⁻¹ than control and other treatments. Moreover, Noor-2013 with 0.45 mg kg⁻¹ showed 10.9% higher WUE than Bittle-98 with control. Among both cultivars, 2.5% higher WUE was noted in Noor-2013 than Bittle-98 (Table 4).

Boron application @ 0.45 mg kg⁻¹ recorded 74.3% more B content of roots than control. Among treatments, Noor-2013 with @ 0.45 mg kg⁻¹ produced 76.1% higher B in roots than Bittle-98 with control (Table 5). Among various concentrations of B, 0.45 mg B kg⁻¹ gave 40.1% more grain B contents than control. Noor-2013 and Bittle-98 with 0.45 mg kg⁻¹ produced 41.5% more grain B contents than Bittle-98 and Noor-2013 with control (Table 5). Nonetheless, 0.45 mg B kg⁻¹ produced 36.6% more B contents of leaves in Noor-2013 than Bittle-98. The cultivar Noor-2013 recorded with 10.8% more B in leaves than Bittle-98 under various concentrations of B. Likewise, higher concentration of B in stalk was measured with 0.45 mg kg⁻¹ that was 9% higher than control (Table 5).

Grain protein was significantly improved by B application and 0.45 mg kg⁻¹ produced 2.9% higher grain protein than control. Moreover, 4.1% higher grain protein was recorded in Noor-2013 with 0.45 mg kg⁻¹ than Bittle-98 with control (Table 6). Likewise, applied B significantly affected grain starch and 0.45 mg B kg⁻¹ recorded 1.4% more grain starch than control. Overall, the cultivar Noor-2013 presented 0.3% higher grain starch than Bittle-98 (Table 6). Regarding grain total sugar, 0.45 mg kg⁻¹ exhibited 4.5% higher grain total sugar than control. Contrarily, Bittle-98 without B produced 10% higher grain crude fiber than Noor-2013 and Bittle-98 with 0.45 mg B kg⁻¹ (Table 6).

Significant and positive correlation was recorded between grain yield per plant and plant height (r = 0.99), no. of pods (r = 0.98); no. of grains per pod (r = 0.99); no. of grains per plant (r = 0.99); 100-grain weight (r = 0.99); biological yield per plant (r = 0.99); chlorophyll contents (r = 0.98); transpiration rate (r = 0.99); stomatal conductance (r = 0.98); WUE (r = 0.97); B uptake by roots (r = 0.91); B uptake by seed (r = 1.00); B uptake by leaves (r = 0.97); B uptake by stalk (r = 0.99); grain protein (r = 0.99); grain starch (r = 0.98) and grain total sugar (r = 0.98) (Table 7). Scatterplot with regression line was drawn among grain yield with all other parameters (Fig. 1). The regression line among grain yield with all parameters showed positive relationship except grain crude fiber. Scatterplot dots showed the variation of parameters with grain yield.

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Fig. 1

Scatterplot with regression line of grain yield vs plant height, number of pods per plant, number of seeds per pod, number of seeds per plant, 100 grain weight, biological yield, chlorophyll contents, transpiration rate, stomatal conductance, water use efficiency, boron uptake by roots, boron uptake by seeds, boron uptake by leaves, boron uptake by shoot, grain protein, grain crud fiber, grain starch, grain total sugar.

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Fig. 1

Scatterplot with regression line of grain yield vs plant height, number of pods per plant, number of seeds per pod, number of seeds per plant, 100 grain weight, biological yield, chlorophyll contents, transpiration rate, stomatal conductance, water use efficiency, boron uptake by roots, boron uptake by seeds, boron uptake by leaves, boron uptake by shoot, grain protein, grain crud fiber, grain starch, grain total sugar.

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Fig. 1

Scatterplot with regression line of grain yield vs plant height, number of pods per plant, number of seeds per pod, number of seeds per plant, 100 grain weight, biological yield, chlorophyll contents, transpiration rate, stomatal conductance, water use efficiency, boron uptake by roots, boron uptake by seeds, boron uptake by leaves, boron uptake by shoot, grain protein, grain crud fiber, grain starch, grain total sugar.

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4 Discussion

Boron shortage is increasing and becoming a serious issue on calcareous soils (hyperthermic, sodic haplocambids) (Atique-ur-Rehman et al., 2020). Regardless of different concentrations and cultivars, considerable change was recorded in plant height of chickpea due to soil applied B (Table 2). Significant improvement in chickpea plant height of soil applied at 0.45 mg B kg⁻¹ (Table 2) could be possible due to appropriate dose of boron, which is used as a part of many plant body functions. Deficiency of B reduces plant height as a result of depression in various physiological and growth parameters (Shrestha et al., 2019). Current findings are in line with Rehman et al. (2019) that crop with B incorporation supported in producing higher number of pods per plant. Suitable concentrations of B had prominent role in movement of photo assimilates from roots to other body parts and it also brings improvement in pollen tube development that leads to enhance number of seeds per pod (Silva et al., 2011). Significantly, higher 100-grain weight was recorded in 0.45 mg B kg⁻¹ might be improvement in plant growth and less flower abortion. Experimental findings confirm the finding of Khatun et al. (2016) who found significant higher 100-grain weight in chickpea due to improvements of nutrients uptake that leads to healthy plant growth and flower development. Similarly, B had prominent role in fertilization that leads to enhance the grain filling rate, which encouraged to increase grain weight (Islam et al., 2017) (Table 3). Boron application brings improvement in flower formation and development, pollen formation, fertilization and seed development. Therefore, these improvements reduced the rate of flower and fruit shedding and ultimately resulted in higher number of pods and grain weight (Rehman et al., 2019).

Chlorophyll contents significantly improved with the increasing B concentration in the soil and maximum value was recorded with 0.45 mg B kg⁻¹. Moreover, some researchers reported that B had indirect relationship with photosynthesis (Liu et al., 2005). Our study agreed with the outcomes of Liu et al. (2005) who reported that soil applied B had higher photosynthetic rate of crop by support of cell membrane and photosynthate movement as well as larger leaf area. The smaller size of leaf is primarily cause for reduction in photosynthetic rate of chickpea under B deficiency. Present study showed minimum rate of transpiration, stomatal conductance and WUE in B deficiency than 0.45 mg B kg⁻¹ which might be due to damage of vascular bundle by B₀ (Li et al., 2017). Statistically lower rate of transpiration rate and stomatal conductance in B₀ also lowered the WUE (Table 6). Pinho et al. (2010) supported our results and concluded that linear correlation exists in stomatal conductance and B concentrations. Han et al. (2008) reported that 0.45 mg B kg⁻¹ improved stomatal conductance and decreased intercellular CO₂ absorption and resulting a considerable improvement in physiological characteristics. Boron contents were increased as increased concentration of B from 0.18, 0.27, 0.36 and 0.45 mg B kg⁻¹ (Rehman et al., 2012). Boron deficiency quickly degenerated the appearance of genes in Arabidopsis of roots, hence, delayed the root development (Martín-Rejano et al., 2011). The division of B contents among plant parts had considerable change with increasing B concentration. Boron was absorbed in the direction of leaf > shoot > root (Table 7) (Reid et al., 2004).

Improved grain protein, crude fiber, starch and total sugar supported by the results of Rehman et al., (2019), that soil applied B in chickpea cultivars improves the grain quality traits due to considerable role of B on enzymes which are responsible for the buildup and alteration of assimilates into grains. Rehman et al. (2019) concluded that B application enhanced seed quality in form of storage, protein, fiber and starch. Genetic characteristics of cultivars is also responsible for improving grain quality traits (Bellaloui et al., 2009b). The improving development in quality traits by soil applied B (Rehman et al., 2019) could be the positive role of B on enzymes activities which are responsible for buildup and alteration of fatty acids (Bellaloui et al., 2009a).

5 Conclusion

Boron application to chickpea cultivars improved growth and grain production under semiarid region Yermosols. Application of B @ 0.45 mg B kg⁻¹ remained better than other treatments.