Growth, Yield, Nodulation and Amino Acid Content of Bambara Groundnut (Vigna subterranea) under Inorganic and Organic Fertilizer Application

Bambara groundnut (Vigna subterranea) plays a major role in the diet of West African people and positioned it as the third most significant legume crop following groundnut and cowpea. Ripe seeds of bambara groundnut contain protein 16 to 21%, fat 4.5 to 6.5% and carbohydrate 50 to 60% (Nyau et al., 2015).
 
There is an inadequate study concerning the effect of inorganic and organic fertilizer on the growth, nitrogen yield and nutritional component in the seed of bambara groundnut, especially in Malaysia. Organic material is broken down and decomposed in the natural and biological process and form compost. Although urea is the most suitable source of nitrogen (N), the plant does not use more than 50% (Uddin et al., 2017 and Hayat et al., 2012) and through volatilization, denitrification and leaching the remaining N is lost. Phosphorus (P) furnishes superior root and soil contact which outcome the advanced getting of phosphorus and other vital and low availability of nutrients and attractions of higher absorption of mineral nutrients (Gharpinde and Wankhade, 2017; Kamble et al., 2016).
 
Biofertilizer is cost-effective and also the renewable basis during which plant acquires essential nutrients to complement inorganic fertilizers (Hasan et al., 2019). Biofertilizers supply nutrients like nitrogen and phosphorus and their actions in the soil or rhizosphere and formulate them obtainable for plants in soil (Zi et al., 2019). A legume lives in association with rhizobium bacteria and helps to improve the soil status (Haque et al., 2020 and Uddin et al., 2020). Soil bacteria are one of the important biotic components that influence decomposition and nutrient mineralization in the terrestrial ecosystem (Khan et al., 2018).
 
Although some reports on the relationship between fertilizers and growth, yield and seed quality of bambara groundnut are available, relatively less attention has been paid on the application of chemical fertilizer and compost in Malaysia. Thus, the main objective of the present study was to find out the effect of inorganic and organic fertilizer on the growth, yield, nodulation and amino acid content of bambara groundnut.

Experimental site, design and treatments

A glasshouse experiment was conducted during the period of 2018, at the Faculty of Agriculture in Ladang 15, Universiti Putra Malaysia, Malaysia. The size of the pot was 65.94 cm². There were 40 experimental units (pot) and in each pot, one seed was sowed. Sandy clay loam soil was used to fill the pot and the amount was 10 kg/plot. Ten different levels of inorganic and organic fertilizers (T₁: N₃₀ + P₆₀ kg/ha, T₂: N₂₀ + P₆₀ kg/ha, T₃: N₂₅ + P₅₀ + K₇₅ kg/ha, T₄: compost 2.5 t/ha, T₅: compost 5.00 t/ha, T₆: compost 7.5 t/ha, T₇: compost 10 t/ha, T₈: B-green biofertilizer 15 kg/ha, T₉: gypsum 400 kg/ha, T₁₀: B-green biofertilizer 15 kg/ha + gypsum 400 kg/ha) were applied. N, P and K were used in the form of urea, triple superphosphate (TSP) and MOP and the inorganic and organic fertilizer were mixed with the soil before sowing the seeds. The experiment was desingned with RCBD with four replication.
 
Growth and yield components

The observations on the growth and yield related data were recorded manually from each pot of each replication separately as well as harvest index was determined as the weight of pod divided by total plant weight with pod and then multiplied by 100. Leaf area and chlorophyll content of the seed was measured by Leaf Area meter (LI-3100C) and Spad meter respectively.
 
Estimation of nitrogen yield

Bambara groundnut plants were dug carefully around the plants to a depth of 50 cm without disturbing the roots for nodule number, nodule weight, root and shoot dry weight. Nitrogen content in the root, shoot and pods with seed was determined by dry combustion technique using a LECO CR-412 carbon analyzer (LECO, Corporation, St. joseph, USA).

Amino acids content in seed

Concentrations of amino acids in grains was measured by HPLC described by (Strydom and Cohen, 1994) following pre-column derivatization with the 6-aminoquinolyl-N-hydroxysuccinimdly carbonate (Waters, USA).
 
Statistical analysis
All data were subjected to statistical analysis of variance (ANOVA) using SAS version 9.4 at the 5% significance level and the least significant difference was employed for mean separation (Gomez and Gomez, 1984).

Vegetative growth and yield

From the results presented, it appears that inorganic and organic fertilizer greatly increased the vegetative growth of the plant (Table 1). Plant height ranges from 20.43 to 21.73 cm in this experiment. The less plant height was recorded at treatment T₄ (compost 2.5 t/ha). At harvesting time, use of T₃ (N₂₅ + P₅₀ + K₇₅ kg/ha) and T₇ (compost 10 t/ha) treatment gave 1.61% and 2.76% respectively less advanced in plant height compare to T₁ (N₃₀ + P₆₀ kg/ha) (control). There was a decrease in plant height by decreasing the N and P levels. Similar result was also reported by Hasan et al., (2019) where plant height decreased with decreasing N and P fertilizer.
 

 
The leaf area of the plant influences CO₂ uptake by determining active radiation absorbed by the canopy and stomatal area. Leaf area ranges from 1899.0 to 2802.9 cm² and T₅ (compost 5.0 t/ha) gave 47.60% decrese in leaf area compare to T₁ (N₃₀ + P₆₀ kg/ha) (control). Application of N and P fertilizer leaf area, as well as accumulation of dry matter, was increased. The finding of this experiment is similar to Hossain et al., (2007), where N (0, 20, 40, 60 kg/ha and P (0, 30, 60, 90 kg/ha) dose were applied and found the better growth and yield at N₆₀ + P₆₀ kg/ha. Increase the vegetative growth of bambara groundnut with the application of N and P with the rate of 0, 100, 150 and 200 kg/ha was reported by Wamba et al., (2012). A similar result was also reported by Hasan et al., (2019). The plant containing more leaves is not helpful for more economic yield and also if the vegetative growth of the plant is more it is undesirable for seed production.

The result showed that there was a significant difference in the pod yield and yield-related traits indicating a high variation of fertilizer among these traits (Table 1). As soon as the vegetative stage finish pod are started to form until the harvest. The results suggested that the highest number of the pod recorded in the treatment of T₁. The highest number of the pod was obtained at T₁ (41) which was treated with N₃₀ + P₆₀ kg/ha. Treatment T₁ gave a 64% increased pod number compare to T₄.  Treatment T₁ (N₃₀ + P₆₀ kg/ha) (49.67 g) also produced the highest pod weight and T₄ (compost 2.5 t/ha) (34.95 g) was the lowest. These result was quite dissimilar with Ikenganyia et al., (2017) because they used the dose of single super phosphate (0, 25, 50, 75 kg/ha). The difference observed between the treatments was because, in the organic residues, nutrient availability depended on nutrient concentration and release with synchrony with crop needs. In the current study, the highest plant growth and yield reported at N₃₀P₆₀ kg/ha could be attributed to the nutrients being readily available from the source. Organic sources alone could not improve the growth and yield of this crop but Ram and Dhaliwal (2012) advocated the use of intergrated nutrient management through chemical and organic sources for getting maximum crop yields.
 
Correlation among agronomic and yield characteristics

Correlation studies have great significance between the agronomic and yield character of bambara groundnut (Table 2). The parameter analysis showed that plant height recorded positive correlation coefficient with leaf number (r = 0.908**), branch number (r = 0.908**), plant fresh weight (r = 0.833**), revealing that the more plant height the higher leaf number, leaf area and plant fresh weight. The table also showed that the weight of hundred seed weight recorded significant correlation coefficient with pod number (r = 0.794**), pod fresh weight (r = 0.894**), pod dry weight (r = 0.938**) days of flowering (r = 0.932**). From the result it clearly showed that the more weight of 100 seed the higher pod number, pod fresh weight, pod dry weight and days of flowering.
 

 
Nodule and nitrogen yield

The effects of different levels of organic fertilizer on nodule number, nodule weight and nitrogen yield were varied significantly (Table 3). The number and weight of nodules were increased with the application of N and P fertilizer (N₃₀ + P₆₀ kg/ha) compare to other organic fertilizers. The highest nodule number of 35.50 was recorded at T₁ (N₃₀ + P₆₀ kg/ha) followed by 32.50 at T₂ (N₂₀ + P₆₀ kg/ha) and 27.00 at T₃ (N₂₅ + P₅₀ + K₇₅ kg/ha). Treatment T₄ (compost 2.5 t/ha) (20.75) contained the lowest nodule number. These findings are similar to other studies (Egbe and Egbo, 2011). Nodulation plays a critical role in N fixation by leguminous crops. Nodule growth and function require proper light photosynthate and fertilizer. In the treatment of T₁ (0.26 g), the nodule weight was higher than any other treatment. Nodule weight was illustrated 0.21 g with T₄ (compost 2.5 t/ha) which was the lowest among all the treatments. Nodule weight increase of 6.07% at T₁₀ (biofertilizer 15 kg/ha+ gypsum 400 kg/ha) and 5.90% at T₈ (biofertilizer 15 kg/ha) compared to T₁ (N₃₀ + P₆₀ kg/ha) (control) was also recorded. There was an increasing trend of nodule number, the weight of nodule and nitrogen yield with increasing N and P fertilizer (N₃₀ + P₆₀ kg/ha). Significant differences in number and biomass of nodulation were also reported by Egbe and Bar-Nyam, (2011). The N content of roots shoots and pods with the seed of bambara groundnut seemed to have affected by the inorganic and organic fertilizer. The nitrogen yield of pods with the seed of bambara groundnuts was consistently higher than that of the shoot, which in turn was higher than root in all the treatments. A similar result was also reported by Egbe et al., (2013).
 

 
Amino acids content in seed
 
Bambara groundnut is an edible legume which serves as one of the main sources of income for smallholder farmers. The seeds contain sufficient quantities of lysine, cysteine and methionine such kind of essential amino acids. Effect of inorganic and organic fertilizer on amino acid content of bambara groundnut is presented in (Table 4). Application of different levels of fertilizer and compost has non significant differences among the content of the amino acid of bambara groundnut seed. In seed, amino acid content was decreased at T₇ (compost 10 t ha^-1) such as lysine (14.92%), argenine (17.89%), cystine (33.59%), glumatic acid (4.83%), glysine (7.01%) and histidine (9.74%) in compare to T₁ (N₃₀ + P₆₀ kg/ha) (control). On the other hand alanine (4.50 mg/kg), arginine (4.75 mg/kg) and aspartic acid (10.40 mg/kg) was the highest from the T1 (N₃₀ + P₆₀ kg/ha) treatment (control). It is seen that aspartic acid, glumatic acid and leucine are the most abundant amino acids in all the samples. (Olaleye et al., 2013) reported the same result. However, the amino acids contained in the seed were lower than the result reported by Mazahib et al., (2013).


 
Bambara groundnut has numerous agronomic and nutritional attributes which make it an excellent crop to develop. The results indicate that the inorganic N and P application exerts a strong influence on the bambara groundnut on growth, yield, nitrogen content and seed quality. This is the clear indication that N₃₀ + P₆₀ kg/ha increase the growth, yield, nitrogen content and yield of bambara groundnut rather than other fertilizer.

Bambara groundnut is a low- priced commodity but a valuable source of all the nutrients. The level of nutrients present in organic fertilizer is often low and usaully complexed in organic chemical structure. The growth, yield, nitrogen yield and amino acid content of bambara groundnut was increased with the combination 30 kg nitrogen and 60 kg phosphorus fertilizer. The application of 30 N and 60 P kg/ha could be recommended for getting maximum growth, yield, nitrogen yield and amino acid content of bambara groundnut.

The authors are grateful to (GP-IPM/2016/9494700) for funding this research project. We also sincerely acknowledge the University Putra Malaysia for providing all land and lab facilities.