Can the new boron-fertilization method improve the system productivity of rice (Oryza sativa L.) – mustard (Brassica juncea L.) cropping system under upland calcareous soils?


 Background

Calcareous soils are highly deficient in boron (B) which has become one of the most important deficient micronutrients in Indian soil after zinc (Zn). For various rice-based cropping systems, B-fertilization is essential for increasing crop productivity and the biofortification of the crop, thus suitable soil application protocol for B-fertilization are required for B-deficient soils.
Results

In a six years-long experiment, different rates of B application viz. 0.5, 1.0, 1.5, and 2.0 kg ha− 1 y− 1 were evaluated to determine the effects of three different modes of B fertilization viz. applied only in the first year, in alternating years, and every year in rice ( Oryza sativa L.) – Indian mustard (Brassica juncea L.) cropping system. It was observed that the application of B at 1.5 kg ha− 1 in every year or 2 kg ha− 1 in alternate years resulted in the highest yield of rice and mustard as well as the system productivity of the rice–mustard cropping system. Application of 2 kg ha− 1 B in the initial year showed the maximum B-uptake by rice, while, application of 1.5–2.0 kg ha− 1 B in every year resulted in the maximum B-uptake by the mustard crop.
Conclusion

Application of B at 2 kg ha− 1 in alternate years or 1.5 kg ha− 1 in every year was the best B-application protocol under rice–mustard cropping system in B-deficient calcareous soils for ensuring the best system productivity of rice–mustard cropping system and B-availability in soil.

above msl). The eld observation was initiated in 2012. The soil of the experimental eld is sandy loam soil (typic calciorthent in the soil taxonomy by USDA).

Climatic condition
The climate of the site comprises mainly three seasons, i.e., rainy (June to September), winter (October to  4 ].8H 2 O} to the rice-mustard cropping system. All treatments were arranged in a randomized complete block design (RCBD) and repeated three times.
During the six years of study, B was applied only once in the initial year applied plot; thrice in the alternate years applied plots; and six times in every year mode of applications. The present study was undertaken during the sixth year (2017-2018) in both rice and mustard crops. Rice crop of locally popular variety, Rajendra Bhagwati was transplanted during late June and was harvested in late October while the mustard (variety, Rajendra Sufalam) was sown in early November and was harvested in late February in each year of the study. Recommended package of practices was followed to grow both of these crops.
The fertilizers urea at 211 kg ha − 1 , diammonium phosphate (DAP) at 130 kg ha − 1 , and muriate of potash (MOP) at 100 kg ha − 1 were applied as sources of N, P, and K, respectively to rice in each experimental plot. For mustard, the rates of urea, DAP and MOP were 96 kg ha − 1 , 87 kg ha − 1 , and 100 kg ha − 1 in each plot.

Extraction procedures
Available B and extraction of B fractions in soil Finely ground and 2 mm sieved air-dried soil samples were used for B fractions extraction. The sequential extraction procedure as proposed by [23], [24] and modi ed by Datta et al. [25] was followed to determine the different sequential B fractions in the soil after the harvest of mustard i.e. at the end of the six years of experimentation.
Readily soluble fraction (Solution plus Non-speci cally adsorbed) The supernatant solution was ltered using Whatman 42 No. lter paper, which was aggregated with 5 gm of soil adding 10 ml of 0.01 M CaCl 2 (1:2 soil: solvent) in 50 ml centrifuges of polyethylene shaken for 16 h and centrifuged for 30 min at 10,000 rpm. Clear extracts with azomethine-H were determined in B [26].

Speci cally adsorbed
The residue from the above step was then extracted with 10 ml of 0.05 M KH 2 PO 4 by shaking for 1 h.
After centrifugation, B was measured in the clear extract as described in the previous step.

Oxide bound
With 0.2 M NH 4 -oxalate (1:4 ground: solution) of 20 ml, the residue from the previous step has been removed by shaking for 4 h at the pH of 3.25. A 14 ml aliquot from the extract was taken into a 50 ml beaker to remove a slightly yellowish to a reddish colour because of Fe and the minor dissolution of organic matter colour. The content of the dissolved Fe as Fe(OH) 3 was held on a hot plate and 2 ml of 5 N NaOH solution had been applied. After the aliquot in the beaker had been weighed, weights were decreased by adding distilled water. The suspension has been ltered by the ltering substance Whatman No. 42 and Fe has been removed. A 9 ml aliquot was taken from the ltrate and heated on a hotplate at 135 ± 5 o C to kill the organic material by adding 50 ml Te on beaker. 4 ml concentrated H 2 SO 4 and 1 ml HClO 4 (60%) was applied. Once the volume has been decreased to about 6 ml, the HClO 4 has been additionally applied to the solution by an increase of 0.5 ml. The material was then converted into a polythene tube of 15 ml and the nal volume was 6 ml. B in the clear extract was calculated with a carmine method after centrifugation at 10,000 rpm for 15 min [26].

Organically bound
The above residue was removed by shaking for 24 h and then ltering through Whatman No. 42 using 20 ml of 0.5 M NaOH. The method described by Datta et al. [25] was used to eliminate the colour from the ltrate. Carmine [26] determined the B in the simple extract.

Residual fraction
The residue was dried and ground from the previous phase. 1 g sub-sample has been taken into a 50 ml of Te on beaker with a small volume of H 2 SO 4 , HF of 5 ml (40%), and HClO 4 of 0.5 ml (60%) [27]. At 135 ± 5 o C, the beaker was put on a thermal plate and the ability was reduced to approximately 3 ml. The heating was then applied and proceeded by adding 5 ml concentrated H 2 SO 4 and 5 ml of HF (40%). In increments of 2-5 ml, additional HF was applied before the soil was completely digested. Depending on color intensity in the extract, a clear extract was added 3 to 5 ml of HClO 4 (60%) after digestion. The heating and removal of HF und HClO 4 reduced the volume to 3-4 ml. The volume was upto 25 ml and the material moved into centrifuge tubes. The obvious supernatant was calculated with carmine after centrifugation of 10,000 rpm B.
Analysis of plant samples for the estimation of B content and uptake During harvest, grain or seed and straw or stover yields for each experimental plot were reported and composite grain/ seed and straw/stover samples from each plot were collected. The samples were washed with deionized (DI) water with 0.1 M HCl. Additional moisture was removed. They were placed in new bags of paper and dried at 70 o C in the oven. Wiley Millground weight samples have been diluted by adding distilled water and the material is ltered through Whatman no. 42 lter papers and the nal volume is provided at 50 ml by the combination of concentrated HNO 3 and HClO 4 (ratio 9:4).
A 20 ml B free tube and a vortexed tube were added with 5 ml sample aliquot, 2 ml ammonium acetate buffer (pH 5.5), and 2 ml 0.02 M EDTA. The tube was once more vortexed and assisted for 1 h at 25 o C, and vortexed again after 1 ml of azomethine-H reagent had been applied (0.9% azomethine-H plus 2% ascorbic acid solution) and the reading was taken at 420 nm with a spectrophotometer (Systronics 2203) [28]. The current concentration was determined using the standard curve which consists of the observed x-axis boron concentration and y-axis absorption.
The uptake of boron by rice and mustard crop was estimated as: Boron uptake (g ha − 1 ) = The Rice equivalent yield (t ha − 1 ) of the rice-mustard system was the average rice yield and rice equivalent yield of mustard. Rice equivalent yield of mustard was determined as follows: Rice equivalent yield (Mg ha − 1 ) = where, MSP of rice and mustard denotes the minimum support price.

Statistical analysis
The data are analyzed statistically using the randomized complete block design process [29]. All the data are subjected to the analysis of variance (ANOVA

Yields of rice and mustard
Compared with control, all the B fertilization practices signi cantly increased the rice grain and straw yields. However, there was no such signi cant impact of different rates and methods of B application on grain yield of rice, while the straw yield of rice was signi cantly varied with the different B-application protocols (Table 1). during alternate years. Application of B at 0.5 kg ha − 1 during alternate years in rice resulted in the improvement in the grain and straw yields to the tune of about 24% and 51%, respectively than no application of B to this crop. Seed and stover yield of mustard was signi cantly improved when B applied at 0.5-2.0 kg ha − 1 of the alternate year or each year. B applied at 1.5 kg ha − 1 during each year signi cantly improved the seed and stover yield of mustard. This treatment resulted in increment of seed and stover yield to the tune of almost 124% and 31%, respectively, over no application of B. Concerning the system productivity of rice-mustard cropping system, application of B at 2.0 kg ha − 1 in the alternate year resulted in the maximum yield being closely followed by the application of B at 1.5 kg ha − 1 in every year (Fig. 1).

B concentration and uptake
The Concerning B content in mustard seed, it was observed that initial application of B at 1.5 or 2.0 kg ha − 1 was better than other B application protocols, while, B application at 1.5 or 2.0 kg ha − 1 in every year showed higher B content in mustard stover than other B application protocols. Values followed by different letters in columns are signi cantly different at P = 0.05 by LSD In several B treatments, the B uptake by rice grain and straw was increased signi cantly than no application of B (Table 3). B uptake by rice grain was much higher when B application rate was increased i.e. >1.0 kg ha − 1 in the initial year or alternate years or every year. However, application of B at 1.0 kg ha − 1 in every year showed at par result with the application of B at 1.5-2.0 kg ha − 1 during the initial year or alternate years or every year. Concerning total B uptake by rice crop, the best result was estimated with B application at 2.0 kg ha − 1 in the rst year. This treatment showed improvement in B uptake by the crop about 114% over no B application. All the B fertilization protocols resulted in signi cant improvement in B uptake by mustard seed than no B application. The highest amount of B uptake by mustard seed was recorded with the application of B at 1.5 kg ha − 1 in every year. B application at 1.5-2.0 kg ha − 1 in every year showed better uptake of B by the mustard crop. Concerning B uptake by stover almost similar trends was recorded. The highest amount of total B uptake by mustard was recorded with the application of B at 2.0 kg ha − 1 in every year and this treatment was narrowly followed by application of B at 1.5 kg ha − 1 in every year. These two treatments showed morte than 200% B uptake by the crop compared to no B application.   Relationship between readily soluble B and grain yield of rice and seed yield of mustard Readily soluble B was related to both the yield of rice grain aw well as a mustard seed and B uptake by these two crops. Figure 2a and 2b represent the relationship between readily soluble B and yields of rice and mustard, respectively. R 2 values of Fig. 2a and 2b were 0.46 and 0.88, respectively. Thus, a strong relationship between soluble B and mustard yield was evaluated. However, the relationship between rice yield and soluble B was not so high as evaluated in the case of mustard yield. Likewise, Fig. 3a and 3b represent the relationship between soluble B with b uptake by rice and mustard, respectively. Here also the R 2 value was a higher increase of uptake of B by mustard than of in rice. Thus, a comparatively stronger relationship between soluble B and B-uptake by mustard was evaluated than that the relationship between B-uptake by rice and soluble B.

Discussion
Boron de ciency is the second most important micronutrient constraint of soils in India after that of zinc (Zn). Owing to this acute problem in Indian soil, considerable yield reduction in many crops was previously reported [30], [31]. The availability of boron is mostly related to soil pH and is widely available at low pH but is frequently leached down from acid, sandy soils. Therefore, de ciency of boron prevails due to the low availability of the boron. Calcareous soils with low organic matter content are more prone to B de ciency [32]. The application of B increased rice yields might be due to its favourable effect on cell-dividing metabolic pathways [33]. Khan et al. [34] found that the application of B at 2 kg ha − 1 to rice along with recommended basal dose of N, P and K fertilizers resulted in the highest yield of the crop compared to other B application protocols. Remesh and Rani [35] reported the signi cant improvement of number of spikelet panicle − 1 , grain weight and the number of lled grains panicle − 1 through the application of B at 1 kg ha − 1 than no application. Katyal and Singh [36] reported higher B-uptake by rice plant with soil applied B. They also reported that only less than 40% uptaken B was accumulated in ricegrain, and the remaining portion was accumulated in rice straws. Soil application of B at10 kg ha − 1 enhanced the B content of rice grain [37]. They opined that the concentration of B in rice-grain and Buptake by the grain were increased dramatically as B in the soil solution was increased through the B application in soil.
Boron (B) is an important micronutrient that contributes to mustard production and growth [31]. Many studies have shown that yields and yield attributes of the crop such as the number of silique plants − 1 , 1000 seed weight etc. were signi cantly higher with the application of B [38], [39]. The B fractional data showed that the majority of B-fractions, except speci cally adsorbed B, were improved with the application of B at 1.5-2.0 kg ha − 1 in alternate years or every year. The chances of losses of speci cally adsorbed B from the upper soil might be higher compared with other fractions. The supply of speci cally adsorbed B decreased due to the high pH level of calcareous soils [32]. B might be used during humi cation to bind to organic matter. In most agricultural lands, B-pool is highly correlated with humic colloids [40]. Gürel et al. [41] recorded the development of the B-buffer zone on organic-effect complexes, B-bound fractions leading to B-labile and making it less available for plant uptake. They also reported that the majority of B was present in the residual form (85-88%), followed by organically bound B (2.84-4.50%), adsorbed directly on the colloid soil surfaces (0.93-1.31%), oxide-bound B (7.27-8.31%), and readily soluble B which was the smallest ranging from 0.40-0.50% only.

Conclusions
From this experiment, it was observed that the application of B at 1.5 kg ha − 1 in every year or 2 kg ha − 1 in alternate years resulted in the highest yield of rice and mustard. Hence, the maximum system productivity of rice-mustard cropping system was also attained under these two B application protocols. This protocol of B application (1.5-2.0 kg ha − 1 B in every year) also showed the maximum amount of readily soluble B. It was also observed that the relationships between readily soluble B with mustard yield as well as B-uptake by mustard were much stronger than rice yield as well as B-uptake by rice. So, from this experiment, it can be concluded that application of B at 2 kg ha − 1 in alternate years or 1.5 kg ha − 1 in every year was better concerning the improvement of yield as well as soil available B under rice-mustard cropping system.