Effect of Nitrobenzene granules and Seaweed extracts on biochemical contents of Arachis hypogaea callus culture.
Abstract
The present study is aimed to evaluate the effect of organic extracts (benzene, diethyl ether and water) of seaweeds (Halimeda gracilis, Ceramium rubrum and Cystophyllum muricatum) and nitrobenzene granuleson biochemical contents of Arachishypogea L. callusunder in vitro conditions. The callus of Arachishypogea L. was obtained from the leaf explants on MSmedium containing 2, 4-D (1 mgL-1) and BAP (0.5 mg L-1). The mass multiplication of callus was achieved at 1mg L-1 of 2, 4-D and 0.5 mg L-1 of GA . The calli were then treated with different concentrations (0.5, 1.0 and 1.5 mg L-1) of 3 seaweed extracts and Nitrobenzene granules. Total carbohydrate, total protein and total chlorophyll contents were analyzed at 5, 10 and 15 days intervals. The total carbohydrate content was high (3.7mg/100mg) in callus treated with Benzene extract of Ceramium rubrumat 1.5 mg L-1 on 15th day. The total protein content was increased (6.9mg/100 mg) in callus treated with Benzene extract of Cystophyllum muricatum at 0.5 mg L-1 on 5th day and the total chlorophyll content was lower (0.36mg/100mg) in Nitrobenzene granules at 0.5mg L-1 in 5th day when compare to control. The present study reveals the positive role of different extracts of seaweeds on increasing the biochemical contents of callus culture of A.hypogea. The extracts can be further evaluated for their role on enhanced regeneration of plants from callus culture.
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Introduction
Seaweed extracts are well-known biostimulants. The seaweed extract consist of trace elements and particularly plant growth regulators such as cytokinin, amino acids, antibiotics, and Vitamins. In modern trends the seaweeds were used along with the farmland as a soil conditioner in some European countries. There are different extraction methods can be used for seaweed extracts preparation i.e. water extraction under high pressure, alcohol extraction, alkaline extraction, microwave-assisted extraction (MAE) and supercritical CO extraction. Cytokinins can be extracted using chilled 70% ethanol. Deuterium is 2 used as co solvent in this process (Yokoyaet al.,2010, Stirket al., 2009). Extraction in 85% methanol leads to obtainment of algae extract rich in gibberellins (Hytonen.et al., 2009). Algal extracts improve plant resistance to frost and drought and increase crop yields. Plants sprayed with the use of seaweed extracts are also characterized by higher resistance to pests and pathogens and more efficient consumption of nutrients from soil (Matysiak K et al.,2010).
Groundnut (Arachishypogaea L.,)is one of the most important oil crops. The Groundnut provides major source of edible oil and vegetable protein. It contains 47-53% oil and 25-36% of protein. Groundnut is a self pollinated crop whereas flowers are produced aboveground and after fertilization, pegs move towards the soil, and seed-contain pods are formed. The cultivated groundnut (Arachishypogaea L.,) originated in South America. China is the first country in cultivate groundnut and India is in second place. Peanut oil is often used in cooking, because it has a mild flavor and a relatively high smoke point. Due to its high monounsaturated content, it is considered healthier than saturated oils, and is resistant to rancidity. There are several types of peanut oil including: aromatic roasted peanut oil, refined peanut oil, extra virgin or cold pressed peanut oil and peanut extract. In the United States, refined peanut oil is exempt from allergen labeling laws. The top countries of peanut cultivars around the world in 2012.
Nitrobenzene is a greenish yellow crystal or yellow oily liquid with the odor of bitter. The nitrobenzene is soluble in water, acetone, benzene, diethyl ether, and ethanol. Nitrobenzene is applied with nitrogen for the enhancement of flowering and growth of agricultural crops. The nitrobenzene from seaweed extract has the capacity to increase flowering in plant (flower stimulant) and also prevent flower shedding. 1.1 Structure and Molecular formula: NO2 C H NO 6 5 2 The science of tissue culture is historically linked to the discovery of the cell and subsequent propounding of the cell theory. Plant tissue culture is a collection of techniques used to maintain or grow plant cells, tissues or organs under sterile conditions on a nutrient culture medium of known composition. More than 234 years ago, Henri-Louis Duhamel du Monceau’s(1756) pioneering experiments on wound healing in plants demonstrated spontaneous callus formation on the decorticated region of elm plants. The development of the multicellular or multiorganed body of a higher organism from a single-celled zygote supports the totipotent behaviour of a cell. . Haberlandt is regarded as the father of tissue culture. German botanist Gottlieb Haberlandt (1902) developed the concept of invitro cell culture. He was the first to culture isolated, fully differentiated cell in a nutrient medium containing glucose, peptone, and Knop’ssalt solution. Callus, which shown stable characteristics under specific conditions after subculture through many successive passages, is a suitable material for cyto differentiation. The advantage of using such callus is that it is composed of fairly homogeneous mass cells and can be proliferated in large amounts under known culture conditions. Wetmore and Sorokin (1955) induced vascular strands in syringa callus derived from the cambial region of the stem or graft apices of shoots. Since then (1950) vascular or tracheary element differentiation has been induced in callus derived from tissues of many species. To Biostimulate the callus of Arachishypogaea L., using different seaweed extracts and it’sbiochemical assay. To collect seaweeds from Mandapam, Rameshwaram coastal Islands, Tamilnadu, India. To prepare seaweed extract by using three different organic solvents.To collect seed of Arachis hypogaea L. from Tamil Nadu Agriculture University Trichy,Tamilnadu,India.To prepare explants in half-strength MS Media.To induce callus in MSmedia with growth hormones.To Mass multiple the callus using growth hormones.To biostimulate callus by using crude seaweed extracts. To biostimulate callus by using nitrobenzene granules (Standard). To estimate the amount of total chlorophyll by Mackinney method. To estimate the amount of total carbohydrate by Anthrone method. To estimate the amount of total Protein by Lowery’smethod.
Conclusion
In present study totally three seaweeds namely Halimeda gracilus, Ceramium rubrum and Cystophyllum muricatum were used. The seaweeds were extracted with three different organic solvents such as benzene, diethyl ether and water. Totally nine extracts were obtained from three seaweeds. Among the nine extracts, Cystophyllum muricatum aqueous extract gave high amount of crude extract 3.150grams, Callus Bioassay was done at different concentration (0.5,1.0,1.5 mg L-1) at different incubation (5, 10, 15 days). Ceramium rubrum benzene extract showed increased level of total carbohydrate content, while the total protein level increased in Cystophyllum muricatum benzene extract and there was no increase in total chlorophyll content with all the extracts tested. This is the first report on application of seaweed extract and nitrobenzene granule, a flower enhancing hormone in A.hypogaea callus culture. GC – MSand LC-MShave to be done for further characterization of the bio molecules present in the three seaweed different solvent, water extracts and TLC for confirmation of Nitrobenzene compound.
