新型大气压冷等离子体射流处理对牙本质胶原纤维交联化的影响

Effect of a novel radio-frequency atmospheric-pressure glow discharge plasma jet treatment on crosslinking of dentin collagen

目的:研究新型大气压射频辉光放电(radio-frequency atmospheric-pressure glow discharge,RF-APGD)等离子体处理对牙本质胶原纤维交联化的影响。方法:(1)收集20颗新鲜拔除的、完整的第三磨牙,采用低速水冷精密切割机制备平行于牙合面的中层牙本质片,厚度为(1.5±0.1)mm,浸泡于10%(质量分数)H_(3)PO_(4)溶液中16 h获得全脱矿牙本质胶原纤维。将20个牙本质胶原纤维片随机分为5组,对照组无处理,4个实验组采用气体温度为4℃的等离子体处理不同时间(20 s、30 s、40 s、50 s)。采用衰减全反射傅里叶变换红外光谱仪测定牙本质胶原纤维结构及交联度,采用扫描电镜观察牙本质胶原纤维表面形貌,采用透射电镜观察牙本质胶原纤维微观结构。(2)收集完整第三磨牙40颗,制备精细牙本质粉5 g,10%H_(3)PO_(4)溶液中完全脱矿,将牙本质胶原纤维粉平均分为5组。对照组无处理,实验组分别采用等离子体处理20 s、30 s、40 s、50 s,采用茚三酮比色法测定各组交联度。(3)收集40颗完整第三磨牙,制备中层牙本质条。将牙本质胶原纤维条随机均分为5组,对照组无处理,实验组采用等离子体处理牙本质胶原纤维条各个轴面20 s、30 s、40 s、50 s,采用万能力学机测定牙本质极限拉伸强度。结果:(1)扫描电镜观察脱矿牙本质表面形貌显示,等离子体处理20 s、30 s及40 s,脱矿牙本质表面胶原纤维网状结构均能维持蓬松;等离子体处理50 s会出现部分微结构的破坏;透射电镜结果显示等离子体处理20 s、30 s及40 s后,纤维结构蓬松,可见天然Ⅰ型胶原纤维典型的周期性横纹;红外光谱结果显示,等离子体处理后,胶原纤维的二级构象与对照组一致,且处理30 s及40 s后酰胺带强度明显增加。(2)茚三酮交联度测试结果显示等离子体处理30 s组及40 s组交联度最高,差异有统计学意义(P<0.05)。(3)牙本质极限拉伸强度结果显示,对照组为(1.67±0.24)MPa,等离子体处理20 s、30 s、40 s、50 s组分别为(4.21±0.15)MPa、(7.06±0.30)MPa、(7.32±0.27)MPa、(6.87±0.17)MPa,与对照组差异有统计学意义(P<0.05)。结论:新型RF-APGD等离子体处理可以促进脱矿牙本质胶原纤维交联,同时明显增强了牙本质胶原纤维的机械性能。

Objective:To investigate the effect of a noval radio-frequency atmospheric-pressure glow discharge(RF-APGD)plasma jet on crosslinking of dentin collagen.Methods:(1)Twenty intact third molars were collected.The middle dentin discs were prepared for each tooth by low-speed water-cooled Isomet saw,and then immersed in 10%(mass fraction)H_(3)PO_(4) solution for 16 h to obtain fully demine-ralized dentin collagen.The twenty dentin discs were then randomly divided into five groups.The control group was untreated while the four experimental groups were treated by plasma jet with gas temperature of 4℃for different times(20 s,30 s,40 s,and 50 s).The structure and crosslinking degree of dentin collagen were characterized by attenuated total reflection-Fourier transform infrared spectroscopy.The surface morphology of demineralized dentin was observed by scanning electron microscope,and the microstructure was observed by transmission electron microscope.(2)Fourty non-caries third molars were collected to prepare 5 g fine dentin powder,then completely demineralized with 10%H_(3)PO_(4) solution.The control group was untreated,while the four experimental groups were treated by plasma jet for 20 s,30 s,40 s and 50 s.The crosslinking degree of each group was determined by ninhydrin colorimetric method.(3)Forty intact third molars were collected to obtain dentin strips.Only two central symmetrical dentin strips(nasty 80)were taken from each tooth and immersed in 10%H_(3)PO_(4) solution for 16 h to obtain fully demineralized dentin collagen.Eighty dentine collagen fiber strips were randomly divided into five groups.The control group was untreated and the axial surfaces of dentin collagen fiber strips in the expe-rimental groups were treated with the plasma jet for 20 s,30 s,40 s and 50 s.The ultimate tensile strength of dentin was measured by universal mechanical machine.Results:(1)The surface morphology of demineralized dentin observed by scanning electron microscope showed that the network structure of collagen fibers on the surface of demineralized dentin treated with the plasma jet for 20 s,30 s and 40 s could maintain the uncollapsed three-dimensional structure,and part of the microstructure was destroyed after plasma jet treated for 50 s.After being treated by plasma jet for 20 s,30 s and 40 s,the three-dimensional structure was uncollapsed,and the typical periodic transverse pattern of natural typeⅠcollagen fiber could be seen.The results of infrared spectra showed that the secondary conformation of dentin collagen fibers was the same as that of the control group after being treated with the plasma jet,and the intensity of amide band was significantly increased after being treated for 30 s and 40 s.(2)The results of ninhydrin crosslinking test showed that the crosslinking ratio of dentin collagen treated by plasma jet for 30 s and 40 s was the highest,and the difference was statistically significant(P<0.05).(3)The results of dentin ultimate tensile strength showed that the control group was(1.67±0.24)MPa,and the plasma jet treated 20 s,30 s,40 s and 50 s groups were(4.21±0.15)MPa,(7.06±0.30)MPa,(7.32±0.27)MPa,and(6.87±0.17)MPa,which were significantly different from that of the control group(P<0.05).Conclusion:The novel RF-APGD plasma jet treatment can promote the crosslinking degree of demineralized dentin collagen and improve their ultimate tensile strength.