VOL. 3, NO. 4, JULY 2008 ISSN 1990-6145 ARPN Journal of Agricultural and Biological Science © 2006-2008 Asian Research Publishing Network (ARPN). All rights reserved. www.arpnjournals.com VERMIWASH: BIOCHEMICAL AND MICROBIOLOGICAL APPROACH AS ECOFRIENDLY SOIL CONDITIONER Zambare V. P., Padul M. V., Yadav A. A. and Shete T. B. Post Graduate Department of Biochemistry, New Arts, Commerce and Science College, Ahmednagar (Maharashtra), India E-mail: vasuzambare@gmail.com ABSTRACT Vermiwash was found to contain enzyme cocktail of proteases, amylases, urease and phosphatase. Microbiological study of vermiwash revealed that it contains nitrogen-fixing bacteria like Azotobactrer sp., Agrobacterium sp. and Rhizobium sp. and some phosphate solublizing bacteria. Laboratory scale trial showed effectiveness of vermiwash on Cowpea plant growth. Keywords: Vermiwash, soil conditioner, enzymes, nitrogen, bacteria, cowpea. INTRODUCTION Vermiculture is a mixed culture containing soil bacteria mixed and an effective strain of earth worms (NIIR Board, 2008). Earthworm has efficiency to consume all types of organic rich waste material including vegetable waste, industrial and other organic waste. Vermicroposting refers to the production of plant nutrient rich excreta of worms. Earthworms play a vital role in plant growth. It is a quite possible to effect quick change over for sustainable agriculture by harnessing brand new vermicompost technology to the soil. In recent times, the commercial vermin culturists have started promoting a product called vermiwash. This vermiwash would have enzymes, secretions of earthworms which would stimulate the growth and yield of crops and even develop resistance in crops receiving this spray. Such a preparation would certainly have the soluble plant nutrients apart from some organic acids and mucus of earthworms and microbes (Shivsubramanian and Ganeshkumar, 2004). But so far there are no experimental evidences to quantify the effect of such spray. Microbes in the environment significantly influence the biogeochemical cycle of phosphorus. The organic phosphorous compounds are decomposed and mineralized by enzymatic complexes like posphatases produced by microbes. In the ecosystem, a mixed population of microbes is essential to promote enzymatic degradation of naturally occurring phosphorous compounds (Trivedi and Bhatt, 2006). Shweta and Singh (2007) reported that presence of plant growth promoting substance in vermicompost and in an article published in The Hindu Newspaper by Subasashri (2004), vermiwash was reported for an effective biopesticide. The present study was carried out to evaluate composition of vermiwash by considering biochemical and microbiological approaches for sustainable development of plant growth promoting factor or as a soil conditioning agent on cowpea plant. MATERIALS AND METHODS Earthworms Earthworms were collected from vermicompost soil. Preparation of extract The earthworms (10g) were immersed in 25ml warm water and kept for 30 minutes at room temperature. Secreted enzyme extracts was centrifuged to remove the insoluble materials at 3000 rpm for 10 minutes. The filtrate was made cell free using 0.2μ membrane filtration. Preparation of soil extract agar Soil (10%) was taken in water and filtered through simple filter paper. The filtrate with 2.5% agar was used for preparation of soil extract agar medium and sterilized. To the sterilized extract added 5% (v/v) of filter-sterilized vermiwash. Extracellular enzymes Exracellular enzymes secretion from earthworms was screened by qualitative and quantitative methods. Qualitatively, protease, amylase and lipase secretions were tested on gelatin, starch and oil emulsion agar plate medium respectively. The zone of clearance was measured. Quantitatively, Protease (caseinase and gelatinase) assay was performed as per Zambare et al. (2007) with 1% casein and gelatin substrates. Amylase assay was performed as per Raghuramula et al. (1988) with 1% starch substrate. In addition to these enzymes, urease, phosphatases and nitrate reductases were also studied. Protein content of vermiwash was estimated by method of Lowery et al. (1951). Microbiological studies Microflora of Vermiwash for Azotobactor, Agrobacterium, Rhizobium and Phosphate sollublizing microbes was isolated on different media like Johnson’s medium, Rhizobium medium and soil extract agar medium, respectively. RESULTS AND DISCUSSIONS Vermicomposting contains mainly earthworm excreta and decomposed matter. As the main substrates presented in the waste is of rich source of macromolecules. Resultant complex materials can easily broken by secretary enzymes of earthworms. Soil with simpler substances is the best suitable media to growth of 1