不同光源对Photofrin联合碘化钾抗大肠杆菌光动力效果的研究

Impact of Different Types of Light on Joint Treatment of Eschenrichia Coli with Photofrin and Potassium Iodide

目的比较两种波长光源对光敏剂(Photofrin,PF)联合碘化钾(Potassium Iodide,KI)介导下的光动力疗法(photodynamic therapy,PDT)抗大肠杆菌(Eschenrichia coli,E.coli)效果。方法检测PF吸收光谱后,以大肠杆菌标准株K-12(ATCC 33780)为实验对象,进行PDT处理。将实验分为PDT组(A组)和对照组(B组)后分别处理,A组处理分别为:(1)相同光剂量下两种波长对PF介导的a PDT抗菌能力;(2)相同光剂量下两种波长对PF联合KI介导的a PDT抗菌能力;(3)不同光剂量下,两种波长对相同浓度PF联合KI介导下的a PDT抗菌能力。其中PDT组分别将1μM、10μM与108菌落数(Colony forming units,CFU)/ml大肠杆菌各1 ml混匀,避光孵育30 min后,在光剂量10 J/cm^(2)的蓝光或红光下进行光照;PF联合KI组,将终浓度为10μM PF与108CFU/ml大肠杆菌各1 ml混合,避光孵育30 min后添加1ml KI(终浓度为100 m M),在蓝光或红光的不同光能量密度下进行光照。对照组分为3个小组,B1组:仅加入PF或仅加入PF+KI,不进行光照;B2组:不加入PF、KI,仅进行光照;B3组:空白对照组,仅加入菌悬液,不加入PF、KI也不进行光照。实验后的菌悬液经连续梯度稀释接种于培养基,孵育24 h后进行菌落计数。结果PF在(366±15)nm波长范围内有高吸收峰,在415 nm蓝光处吸光度值较635 nm红光处高。不同浓度的PF在10 J/cm^(2)的蓝光或红光作用下均无抗大肠杆菌作用(P>0.05)。PF联合100 m M KI后,PF在10μM时具有抗菌作用,在能量密度10 J/cm^(2)的蓝光或红光作用下大肠杆菌存活率分别为0.00003865%、28.7%,在该条件下蓝光抗大肠杆菌作用强于红光(P<0.001)。在10μM PF联合100 m M KI条件下,0 J/cm^(2)的蓝光或红光与空白对照组比较,差异均无统计学意义(P>0.05);在0.5 J/cm^(2)的蓝光作用下大肠杆菌存活率为0.0001209%,在20 J/cm^(2)的红光作用下其存活率为0.00002783%。结论PF联合KI,调节照射光的能量密度,可增强蓝光或红光对光敏剂PF介导下光动力抗大肠杆菌效果。

Objective To compare the photodynamic therapy(PDT)effect of photofrin(PF)plus Potassium Iodide(KI)on Eschenrichia coli(E.coli)with blue or red light.Methods The spectrum of PF was detected with spectrophotometer.E.coli standard strain K-12(ATCC 33780)was prepared to form a concentration of 108colony forming unit(CFU)/ml.The experiments were conducted in PDT groups(groups A)and control groups(groups B)respectively.For groups A firstly,E.coli was mixed with different concentrations of PF(0,1 and 10μM)with or without addition of KI(100 m M),then the suspension was illuminated with same radiant exposure of 10 J/cm^(2).Secondly,10μM PF were mixed with E.coli and incubated in dark for 30 min,then the suspension was illuminated with different light fluence of blue or red light after adding KI(100 m M).The control groups were further divided into 3 groups(group B1,B2and B3).For group B1,the E.coli was only mixed with different concentrations of PF(0,1 and 10μM)with or without addition of KI(100 m M).For group B2,the E.coli was only exposed to the same light adopted in the PDT groups.For the blank control groups(group B3):neither PF nor light was given to the E.coli suspension.The survival fraction of E.coli after treatment was evaluated via counting CFU.Results PF was efficiently excited at 366±15 nm,and the absorption of PF at 415 nm was higher than at 635 nm.With 10 J/cm^(2)of blue or red light,it was effective for PF to kill E.coli regardless of the concentration(P>0.05).Upon the addition of 100 m M KI,the effect was weak until PF reached 10μM.The survival fraction of E.coli was 0.00003865%with 10 J/cm^(2)of blue light and 28.7%with 10 J/cm^(2)of red light.Under this condition,the antimicrobial effect of blue light was better than that of red light(P<0.001).With the combination 10μM PF and 100 m M KI,there was no significant difference between 0 J/cm^(2)of blue or red group and blank control group(P>0.05).The survival fraction of E.coli was 0.0001209%at 0.5 J/cm^(2)of blue light and 0.00002783%at 20 J/cm^(2)of red light.Conclusions With blue or red light,the photodynamic effect of PF on E.coli was enhanced by adding KI and adjusting light density.