高分子脲醛肥料降解菌的筛选及降解酶性质初探

Screening of polymeric urea - formaldehyde fertilizer degrading microorganisms and preliminary study on its degrading enzyme properties

关键词:  脲醛基缓释肥料，生物降解，产红青霉，抗热解材料，从头降解

Key words:  urea - formaldehyde slow release fertilizer  biodegradation  Penicillium rubens  Anti pyrolysis fertilizer, Ab initio degradation

摘要:

目的：为了筛选能降解脲醛基高分子缓释肥料的微生物并初步研究其降解酶性质。方法：以含磷的脲醛高分子缓释肥(PSRF)为唯一碳源和氮源设计筛选培养基，从含材料的土壤分离筛选降解菌，并从形态学和ITS进行菌种鉴定。PSRF经100 ℃超声处理后制备得到不溶性抗热解部分作为从头降解试验材料（rPSRF），与菌种或无菌发酵液共培养，应用生物电镜、微米级和纳米级激光粒度观察、分析样品分解状况，通过化学方法检测溶液中磷、甲醛、电导率的含量及变化进行了研究。结果：从施用PSRF的土壤中分离出一株PSRF降解菌，经鉴定为产红青霉菌Penicillium rubens ，保藏号CGMCC 23229 (P. rubens 23229)。SEM与激光粒度分布结果表明，与rPSRF共培养期间，P. rubens 23229可以在不溶性抗热解肥料中附着生长并将其从致密大块结构降解为小块以及更小的碎屑；共培养7 d后溶液中磷含量从0 μg·mL-1增加到21.52 μg·mL-1，甲醛含量从0 μg·mL-1增加到0.213 mg·mL-1。P. rubens 23229在含PSRF的筛选培养基或无PSRF的察氏基中培养后离心去掉菌体，得到的无菌发酵液与rPSRF共培养15 d，前者溶液中磷含量持续增加至10057.143 μg·mL-1，后者几乎无任何增加，说明其降解酶可以分泌到发酵液中，且受PSRF诱导产生。结论：从土壤中筛选到一株PSRF降解菌，经鉴定为产红青霉菌，能将抗热解的致密脲醛高分子从头降解为小颗粒并进一步代谢利用，初步分析其降解酶为胞外诱导酶。

Abstract:

This study aimed to screen microorganisms that can degrade polymeric urea - formaldehyde macromolecule slow-release fertilizers and to research the properties of their enzymes. A screening medium was designed using PSRF as the only carbon source and nitrogen source and. A degrading strain was isolated from the material-bearing soil. Morphological and ITS methods were applied to identify the strain. PSRF was ultrasonically treated at 100℃ to obtain an insoluble pyrolysis resistant urea-formaldehyde ab initial degradation test material (rPSRF). rPSRF were co-cultured with the strain or sterile fermentation liquid for a certain time, then biological electron microscopy (SEM) and micrometer and nanometer laser particle size meter (LPSM) were applied to observe and analyze the decomposition conditions of samples, and the contents and changes of phosphorus, formaldehyde and electrical conductivity were detected. A PSRFdegrading microorganism was isolated from the PSRFfertilizing soil and it was identified as Penicillium rubens, with preservation number CGMCC 23229 (P. rubens 23229). Results of SEM and LPSM showed that?P. rubens 23229 could grow on the insoluble pyrolytic resistant fertilizer and degrade it from a dense bulk structure to small pieces and smaller debris during the co-culture time. After co-cultured for 7 d, the phosphorus content of the solution increased from 0 μg·mL-1 to 21.52 μg·mL-1; and the formaldehyde content increased from 0 μg·mL-1 to 0.213 mg·mL-1. P. rubens 23229 was cultured in the screening medium containing PSRF or Czapek without PSRF, then the respective sterile fermentation broth (SFB) was obtained by centrifuge filtration. Two kinds of SFB were co-cultured with rPSRF for 15 d, and the phosphorus content in the PSRF-SFB co-culture solution continued raising until 10057.143 μg·mL-1 while there was nearly no substantial increase in phosphorus content for the Czapek-SFB co-culture. It could be deduced that P. rubens 23229 could produce PSRF biodegradable enzyme with the induction of PSRFand secreted it into the fermentation broth. A PSRF-degrading strain was isolated from soil and identified as Penicillium rubens. It could ab initial biodegrade the dense urea-formaldehyde pyrolysis resistant macromolecule material into small particles and smaller metabolize. The degradation enzyme was preliminarily analyzed as an inducible extracellular enzyme.