Bioinsecticides Production by Bacillus thuringiensis Using Agroindustrial By Product in Solid Fermentation

Abstract

Bacillus thuringiensis is one of well-known bioinsectiside resources, and utilized in organic farming for the replacement of chemical insecticide. Substrate composition affect the bioinsecticides production. In fermentation process, microorganisms require water, carbon, nitrogen, and minerals for the growth. Substrate composition also affect the characteristics of bioinsecticides products, especially in growth, toxicity, and potential products. Agroindustrial by product contains the necessary complex nutrients components as a substrate for the growth of microorganisms. This research aimed to assess the ability of Bacillus thuringiensis using substrate combination of carbon source (onggok, pulp of coffee, starch fractions of iles-iles, and sago’s waste) and nitrogen source (tofu’s waste, peanut meal, palm kernel cake, and corn hominy) in bioinsecticides production using solid media cultivation. The main parameters to select the carbon and nitrogen sources are LC50 value and potential bioinsecticides products. The research consisted of three stages, there were substrates characterization, microorganisms characterization, and bioinsecticides production. Substrates characterization was conducted to analyze proximates content and physical characteristics such as aw, bulk density, and water holding capacity. Microorganisms that was used consisted of three strains, there were Bacillus thuringiensis subsp. aizawai, Bacillus thuringiensis subsp. berliner dan Bacillus thuringiensis subsp. israelensis. Microorganisms characteristics includes the ability of cellulolytic, amylolytic, and ability of the cell production. Bioinsecticides production stages was divided into three stages, there were influence of carbon source substrates, and nitrogen source substrates for bioinsecticides production, and the influence of bioinsecticides production using carbon and nitrogen sources that were selected on a scale of 2 kg. Carbon source substrates have main component contents like carbohydrate (50.7 – 65.9 %), total simple sugars (23.5 – 30.5 %), crude fiber (11.3 – 31.4 %), and protein (0.5 – 11.5 %). Nitrogen source substrates have carbohydrate (25.1 – 59.4 %), total simple sugars (10.5 – 25.1 %), crude fiber (6.5 – 37.4 %), and high protein (7.1 – 24.9 %). Nitrogen source substrates has a higher protein content than the carbon source substrates. The results showed that all strains of Bacillus thuringiensis that were determined has the ability to produce cellulase and amylase enzymes so they can use all substrates as a carbon and nitrogen sources in solid media cultivation. Pulp of coffee has an advantage because it contains simple sugars higher than other substrates so in the carbon source substrate selection, cultivation using pulp of coffee as a substrate for Bacillus thuringiensis subsp. berliner and Bacillus thuringiensis subsp. israelensis can produce the highest potential product. Cultivation of Bacillus thuringiensis subsp. berliner using pulp of coffee as a substrate can produce the highest toxicity with LC50 value of 0.07 μg/ml and potency of 11429 IU/mg, while cultivation of Bacillus thuringiensis subsp. israelensis using pulp of coffee as a substrate can produce the highest toxicity with LC50 value of 0.05 μg/ml and potency of 16000 IU/mg. Bacillus thuringiensis subsp. aizawai has high amylolytic capability thus in the carbon source selection, cultivation of Bacillus thuringiensis subsp. aizawai can produce the highest toxicity with LC50 value of 0.09 μg/ml and potency of 8889 IU/mg. Suitable substrates for the cultivation of Bacillus thuringiensis subsp. aizawai is combination of onggok (carbon source) and palm kernel cake (nitrogen source) that produced highest toxixity (LC50 value) of 0.04 μg/ml and potency of 20000 IU/mg. Suitable substrates for the cultivation of Bacillus thuringiensis subsp. berliner is combination of pulp of coffee (carbon source) and palm kernel cake (nitrogen source) that produced highest toxixity (LC50 value) of 0.02 μg/ml and potency of 40000 IU/mg. Suitable substrates for the cultivation of Bacillus thuringiensis subsp. israelensis is combination of pulp of coffee (carbon source) and peanut meal (nitrogen source) that produced highest toxixity (LC50 value) of 0.02 μg/ml and potency of 40000 IU/mg. Bioinsecticides production using selected carbon and nitrogen substrates on a scale of 2 kg/batch, LC50 values increased which means a decrease in toxicity. LC50 values for Bacillus thuringiensis subsp. aizawai cultivation using combination of onggok and palm kernel cake is 2.63 μg/ml, LC50 values for Bacillus thuringiensis subsp. berliner cultivation using combination of pulp of coffee and palm kernel cake is 1.28 μg/ml, and LC50 values for Bacillus thuringiensis subsp. israelensis cultivation using combination of pulp of coffee and peanut meal is 0.67 μg/ml