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Regulatory Mechanism of Enhancing Fertilization of Mango Flower in Response to Urea Application

Uttam Kumar Roy
Laboratory of Protein and Enzyme Research, Department of Biochemistry and Molecular Biology, University of Rajshahi, Rajshahi 6205, Bangladesh

Md. Shahidul Haque
Laboratory of Protein and Enzyme Research, Department of Biochemistry and Molecular Biology, University of Rajshahi, Rajshahi 6205, Bangladesh

Sohel Hasan
Laboratory of Protein and Enzyme Research, Department of Biochemistry and Molecular Biology, University of Rajshahi, Rajshahi 6205, Bangladesh

Swapan Kumar Roy
Laboratory of Protein and Enzyme Research, Department of Biochemistry and Molecular Biology, University of Rajshahi, Rajshahi 6205, Bangladesh

Narayan Roy
Laboratory of Protein and Enzyme Research, Department of Biochemistry and Molecular Biology, University of Rajshahi, Rajshahi 6205, Bangladesh

Abstract/ Executive Summary:

Urea is a potent N fertilizer inducing biological process of plants. However the regulatory mechanism of enhancing fertilization in mango flower in response to urea is not clarified. In this respect, flower of Langra variety of mango were used and the enzyme urease in flower extract when treated with 100 mM urea were maximally increased. For the regulation of urease activity, role of heavy elements like arsenic (As) and mercury (Hg) and light elements like zinc (Zn) and calcium (Ca) was investigated. 10 mM Na₂HAsO₄ potentially prohibited the urea induced urease activity in flower than the effect of 1 mM concentration. Similar inhibitory effects were observed whenever the extract was treated with different doses of mercuric chloride (1, 10, 50, and 100 mM) and the effective concentration was 10 mM which reduces 36% of activity. On the other hand, the effects of 1, 10 and 100 mM Zn(NO₃)₂ were examined on urea induced urease activity where 1 and 10 mM concentrations were found to enhance 30 and 58.4% enzyme activity respectively. We also examined the effect of different doses of CaCl₂ (1, 10 and 100 mM) on urea induced urease activity and declined by increasing concentration of CaCl₂. However, the effective concentration of CaCl₂ was 1 mM showing 103.2% increased activity. The findings indicate that calcium is more potent than zinc on increasing urease activity. To clarify the regulatory mechanism of urea induced primary amine synthesis, the flower was treated with 10 mM Na₂HAsO₄ which potentially reduced color pigmentation for primary amine demonstrating clearly that arsenic is involved in prevention of primary monoamine during flowering.

Keywords: Mango flower, Primary monoamine, Regulation of urease

Location: Laboratory of Protein and Enzyme Research, Department of Biochemistry and Molecular Biology, University of Rajshahi, Rajshahi

Start Date:

End Date:

Program Area: Crop-Soil-Water Management

Commodity: Mango

Objectives:

1. To involve the effect of urea on the synthesis of primary monoamine in mango flower, and

2. To examine whether urea induces the synthesis of primary monoamine.

Methodology:

Plant material - Mango flower (Magnifera indica) (Langra) was collected from the mango garden located to the northern side of the University Campus during February-March. The flowers were quickly stored at –80^oC refrigerator. About 5–6 g of flower were homogenized with mortar and pestle with 10 mM phosphate buffer (pH 7.0) and centrifuged at 6000 rpm for 10 min. After centrifugation, the supernatant was collected and used as crude extract while the sediment was discarded. Assay of urease activity - 5.0 g of mango flower were placed into a mortar with pestle and were homogenized with 30 ml of 0.2 M phosphate buffer (pH 7.0) and centrifuged at 6000 rpm for 10 min. The supernatant was collected and used as crude extract. The urease activity in crude extract of flower was assayed. For assay of urease activity, 0.4 ml of the crude extract was used. To examine the effect of urea on urease activity, 50, 100 and 200 mM of urea were used as substrate of the enzyme. The enzyme activity was expressed as μmole/min/mg of protein. Treatment with Na₂HAsO₄, HgCl₂, Zn(NO₃)₂ and CaCl₂ - To examine the role of different modulators (heavy and light elements) on the activity of urease in mango flower extract, the effect of arsenic compound (1 and 10 mM Na₂HAsO₄) was performed. Different concentrations of mercuric chloride (1, 10, 50 and 100 mM) were used to examine the role of Hg on urease activity. Similarly Zn(NO₃)₂ solutions (1, 10 and 100 mM) was used to find the role of Zn on the activity of urease in the flower extract. Another compound CaCl₂ (1, 10 and 100 mM) was also used to examine its effects on the enzyme activity in the extract. The flower sample extracts obtained by the homogenization procedure were treated with the different concentrations of the above modulators and the urease enzyme activity was determined accordingly by the conventional procedure as described above. Test of primary monoamine - 400 μL samples (5 g of flower in 30 mL solution) in a test tube was taken, mixed well with 0.5 mL urea (100 mM) and incubated at 55^oC for 15 min. After incubation, 1.0 mL diluted HCl and 4 drops of 10% NaNO₂were added and cooled. In another test tube, a few drops of alkaline β-napthanol were taken and the mixture of the previous test tube was added slowly to the alkaline β-napthanol. An orange deep color was appeared showing the synthesis of primary monoamine in the flower sample. Control tube was similarly used for identification of primary amine where no urea was used and 1.5 mL diluted HCl were used and the color pigmentation was different from the urea induced sample tube. To examine the role of arsenic on the regulation of primary amine synthesis in flower, similar procedure was done however the flower extract was treated with 0.5 mL Na₂HAsO₄ (10 mM) instead of urea. Deep orange color was disappeared in response to arsenic. Statistical analysis - Results of the experiments were expressed as mean and standard error of different groups. The differences between the mean values were evaluated by ANOVA followed by paired t-test using SPSS software.

Source of Information: International Journal of Biosciences | IJB |; ISSN: 2220-6655 (Print) 2222-5234 (Online); Vol. 6, No. 1, p. 308-317, 2015

Article Link (Online Journal): http://www.innspub.net; http://dx.doi.org/10.12692/ijb/6.1.308-317

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

The results revealed that 100 mM concentration plays the critical role on enhancing urease activity in the mango flower extract. The increased activity of urease showed the higher uptake of urea in the flower because higher concentration of NH₄+ is formed in response to urea.

Knowledge Management: Journal