Study On Isolation And Characterization Of Norfloxacin-degrading Bacterium Strain NOR-36

As a representative of the quinolones antibiotics,norfloxacin was ubiquitously detected in the soil and aquatic environments,causing potential harm to human health.A strain that could degrade norfloxacin was isolated from pharmaceutical wastewater treatment pool and designated as NOR-36.According to the phenotype,physiological and biochemical characteristics,compared with 16 s rRNA gene sequence homology,it was preliminary identified as Staphylococcus caprae.The stain NOR-36 entered logarithmic growth period after 3 h when cultured in LB liquid medium.After 10h,the NOR-36 entered into stable growth period.The optimum temperature for the growth of strain NOR-36 was 30?,the optimal pH was 7.0.More dissolved oxygen in the water was benefit for the growth of NOR-36.The concentration of NaCI had great impact on the growth of stain NOR-36.When the concentration of NaCl was<15 g/L,the strains growed well.Whereas,the growth was significantly inhibited when the concentration of NaCl was over 15 g/L.NOR-36 could utilize peptone as the nitrogen source and fructose as the carbon source.Moreover,NOR-36 was resistant to anpicillin and chloramphenicol,and sensitive to carbenicillin,kanamycin,genta-micin and streptomycin.The growth characteristics and the co-metabolism degradation characteristics of NOR-36 were further investigated.The results showed that piperazine could be used as a co-metabolic substrate which can effectively increase the degradation rate.The strain was subjected to chemical treatment under following conditions with temperature of 30 ?,pH 7.0,norfloxacin concentration of 5mg/L,and piperazine to norfloxacin concentration ratio of 5:1.Under the above optimum conditions,the maximum degradation rate of norfloxacin can reach 92.6%.When piperazine/norfloxacin concentration ratio was larger than 5,a key enzyme binding site was occupied by piperazine and the degradation efficiency was decreased.The optimum temperature was 30 ?,the optimum pH was 7.0,and the optimum concentration of NaCl was below 15 g/L.The results of HPLC-MS/MS analysis indicated that three main intermediate metabolites were generated.The formulas were C16H18FN3O4,C14H16FN3O3,C12H11FN2O3,respectively.The C-N on the piperazine ring of norfloxacin was broken and oxidized.Afterwards,-COCH3 and-CH2CH2NH2 group were removed subsequently.Used sodium alginate embedding the norfloxacin degrading strain NOR-36,results showed that mechanical strength of immobilized beads best and the degradation of norfloxacin effect was best when the concentration of sodium alginate was 0.6%,the concentration of CaCl2 was 4%and the concentration of PVA were 5%,immobilized time was 9 d.The degradation rate of immobilized bacteria was slower than the free bacteria,but immobilized bacteria could tolerance pH and temperature in a wider range.