Preparation and Application of Indoxacarb Degrading Bacteria Immobilized Sodium Alginate Microspheres

The objective of this study is to prepare microsphere of degrading bacterium Pseudomonas stutzeri (ACCC 02521) with sodium alginate as carrier, and to establish the application conditions for degrading indoxacarb in farmland.

Through the micro drop embedding ball forming method, bacteria were suspended and added into sodium alginate solution. After mixing, it was dropped into CaCl₂ for granulation. It was washed with 0.9% NaCl after fixing at low temperature. The mass transfer performance and mechanical strength of degrading bacteria microspheres were measured to determine the optimal concentration of sodium alginate. The best formulation was obtained by single factor optimization. 1.0%-5.0% CaCl₂, 40-200 g·L^-1 bacteria or 20-100 g·L^-1 preparation were dropped into 3.0% sodium alginate, respectively. The CaCl₂ concentration, embedded bacteria and preparation dosage in the degrading bacteria microspheres were determined according to the degradation rate of indoxacarb. The morphology of microspheres, cell morphology and distribution of degrading bacteria were observed by scanning electron microscope. Quantitative degrading bacteria microspheres were put into different types of soil suspension, temperature, pH or treatment time. The effects of environmental factors on the release capacity, degradation effect and stability of degrading bacteria microspheres were evaluated by calculating the amount of released bacteria (CFU/mL) or the degradation rate of indoxacarb. Two days after the routine application of pesticides, the degrading bacteria microsphere preparation was applied. Topsoil was collected to detect the residue of indoxacarb, the field application of dose was determined, and the field application conditions of the preparation were determined. The residual concentrations of indoxacarb were detected and tracked by HPLC.

The preparation was composed of 3.0% sodium alginate, 2.0% CaCl₂ and 80 g·L^-1 degrading bacteria. The particle size was about 3.0 mm. The degrading bacteria microspheres had uniform particle size, moderate mechanical strength, good mass transfer performance, degradation activity and storage stability. Scanning electron microscopy showed that the degrading bacteria were evenly distributed in sodium alginate microspheres and their morphology was normal. In the soil with 10-30℃ and pH of 6.0-8.0, the degrading bacteria were released stably, and the degradation rate of indoxacarb was more than 85%. The release performance was not affected by soil type, the stability was good, and was less affected by environmental conditions. When 90-900 kg·hm^-2 of degrading bacteria microspheres were applied 2 d after field spraying of 150 g·L^-1 of EC active ingredient 20 mg·L^-1, the residual half-life (T_1/2) of indoxacarb in soil was shortened to 2.49-3.32 d (7.53 d for blank control area); furthermore when 450 kg·hm^-2 of degrading bacteria microspheres were applied 2 d after spraying indoxacarb with 5, 20 and 50 mg·L^-1, the values of T_1/2 was shortened from 6.03-7.45 d to 2.34-3.59 d.

The preparation of degrading bacteria P. stutzeri microspheres with sodium alginate as carrier has stable performance, can significantly degrade indoxacarb residues in farmland and shorten T_1/2 time, provides technical and product support for bioremediation of soil pesticide residue pollution, and has the potential for further optimization and application.