Study On Microbiologically Influenced Corrosion Of 316L Stainless Steel And Titanium By Oral Microbiota

Objective:Microbiologically influenced corrosion(MIC)is a widespread phenomenon in natural environment,which can cause material degradation and affect material properties.The corrosion of biomedical materials in human environment can induce inflammatory or allergic reactions,leading to serious complications such as host tissue damage and implant loosening.At present,the vast majority of MIC research is carried out in pure medium,and a few in double strain medium.Although this research method is conducive to the repetition and analysis of the results,it is far from the corrosion in the actual environment.Microbial corrosion in the actual environment(including natural environment and human environment)is the result of a variety of microorganisms.Oral cavity has the most complex micro ecosystem of human body,from which more than 700kinds of microorganisms have been isolated.At present,there are few reports on oral microbiological corrosion,and almost all focus on the evaluation of corrosion behavior of oral common bacteria and single bacteria.Therefore,the study of oral microbiota corrosion behavior has a better reference value for evaluating the degree of microbial corrosion of biomaterials in oral environment,and exploring the corrosion mechanism of oral flora has a greater guiding significance for the improvement of oral biomaterials.Therefore,this study is expected to establish a multi-species biofilm of oral flora in vitro to evaluate its microbiologically influenced corrosion behavior on commonly used oral metal materials,so as to provide a theoretical basis for elucidating the corrosion mechanism of oral flora and improving biomedical materials.Methods:Part I:First,establish multi-species biofilms(subject1 and subject2)of supragingival plaque in vitro in aerobic environment.316L stainless steel was taken as the research object.Scanning electron microscopy and laser confocal microscopy were used to characterize the biofilm and corrosion morphology.The corrosion products were analyzed by X-ray photoelectron spectroscopy and atomic absorption spectrometry.The corrosion rate was evaluated by linear polarization resistance,electrochemical impedance spectroscopy and polarization curve.16S r DNA sequencing technology was used to evaluate the composition characteristics of the multi-species biofilms.Part II:The ability of oral multi-species biofilms to secrete riboflavin was detected by HPLC.The effect of riboflavin on the corrosion ability of oral microbiota was analyzed by electrochemical methods,including linear polarization resistance and electrochemical impedance spectroscopy.Part III:First,establish subgingival plaque biofilms(subject1 and subject2)in vitro in anaerobic environment.Titanium alloy(TC4)and pure titanium(TA2)were studied.Scanning electron microscopy and laser confocal microscopy were used to characterize the biofilm and corrosion morphology.The corrosion products were analyzed by X-ray photoelectron spectroscopy.The corrosion rate was evaluated by electrochemical methods,including linear polarization resistance and polarization curve.Results:Part I:The results of scanning electron microscope showed that biofilms could be formed on the surface of 316L stainless steel.On the third day,the biofilm structures were relatively simple,and on the seventh day,grew into more complex multilayer network structures;The results of fluorescence staining showed that the density of biofilms in the two groups on the 7th day was significantly higher than that on the 3rd day,yellow fluorescence was detected on the 7th day,indicating that the proportion of dead bacteria increased;The results of laser scanning confocal microscopy showed that the average maximum pit depths of subject1 and subject2 groups were significantly higher than that of the sterile group,the average maximum pit diameter of subject1 group was significantly larger than that of sterile group;The electrochemical results showed that compared with the sterile group,the open circuit potential,polarization resistance,corrosion potential and charge transfer resistance of the experimental group were significantly reduced,and the corrosion current density was significantly increased;The X-ray photoelectron spectroscopy results show that the oxide ratio of Fe,Cr and Ni on the surface of the two experimental groups decreases significantly,the proportion of pure metals(Fe⁰,Cr⁰and Ni⁰)increased;The results of p H measurement showed that the p H value of sterile group was relatively stable,the two experimental groups showed similar changes,which decreased significantly on the first day,and then increased gradually;the results of atomic absorption spectrometry showed that the ion precipitation concentrations of Ni and Cr in the two experimental groups were significantly higher than those in the sterile group;16S r DNA sequencing results showed that oral microbiota could form various and multifunctional biofilms on the surface of samples,including aerobic bacteria,microaerobes,facultative anaerobes and strict anaerobes,as well as acid-producing bacteria,nitrate reducing bacteria and electrochemically active bacteria with extracellular electron transfer ability.Part II:The results of high performance liquid chromatography showed that,no riboflavin was detected in the culture system of aseptic group,very low concentration of riboflavin was detected on the 5th and 7th day in subject1 group,relatively high riboflavin concentration could be detected on the 3rd,5th and 7th day in subject2 group;The electrochemical results showed that the addition of 10?g/m L exogenous riboflavin had no effect on the polarization resistance and charge transfer resistance of the sterile group,indicating that riboflavin itself did not interfere with the experimental results;Compared with the sterile group,the polarization resistance and charge transfer resistance of 316L stainless steel in subject1 and subject2 groups were significantly reduced,and the decrease in subject2 group was more obvious.The addition of exogenous riboflavin did not promote or inhibit the corrosion rate of the two experimental groups;The results of p H measurement showed that the p H of the sterile group was relatively stable,and the addition of riboflavin had no effect on the results.There were similar changes in subject1 and subject2 groups,which decreased significantly on the first day and increased gradually from the second day,the addition of riboflavin had no effect on the p H of the two experimental groups.Part III:The results of scanning electron microscopy showed that the oral flora could form biofilm on the surface of TC4 and TA2 on the 3rd and 7th day,and the structure of the biofilm was more compact and complex on the 7th day;The results of fluorescence staining showed that the maximum thickness of biofilm on TC4 and TA2 increased with time,the cell density increased significantly and the proportion of dead cells began to increase on the 7th day;The results of laser scanning confocal microscopy showed that the average maximum pit depth of subject1 and subject2 groups was significantly higher than that of the sterile group;The electrochemical results showed that compared with the sterile group,the open circuit potential,polarization resistance and corrosion potential of the experimental group were significantly decreased,and the corrosion current density was significantly increased;X-ray photoelectron spectroscopy results showed that the contents of Ti O₂and Al₂O₃on TC4 surface in subject1 and subject2 groups decreased significantly,the content of Ti O₂on TA2 surface in subject1 group also decreased significantly;The results of p H measurement showed that the p H value of the sterile group was relatively stable,both experimental groups decreased on the first day,and then gradually increased.Conclusion:Part I:Oral microbiota can form stable multi-species biofilm on 316L stainless steel surface,and its composition has individual differences.The multi-species biofilm in this study can well simulate the biodiversity of oral plaque and reproduce its multi-species and multi-function characteristics in vivo.The non-uniformity of multi bacteria biofilm leads to the formation of a variety of corrosive micro cells between the biofilm and the metal surface,such as oxygen concentration cell;Oral microbiota can increase the pitting corrosion rate of 316L stainless steel,at the same time,the oxide content of Fe,Cr and Ni in the surface passivation film is reduced,which weakens its corrosion resistance;Oral microbiota contains a variety of microorganisms with different functions and has the potential to produce EET-MIC.Part II:The oral microbiota contains bacteria that can synthesize riboflavin through their own metabolic activities,the ability of riboflavin secretion was different in different subjects;The corrosion ability of different oral microbiota groups was different,and the electrochemical data of the two experimental groups showed some differences;Exogenous riboflavin had no significant effect on the corrosion behavior of oral microbiota.Part III:In anaerobic environment,oral microbiota can form multiple biofilms with complex structures on the surface of TC4 and TA2;Oral microbiota can aggravate the pitting corrosion and accelerate the corrosion rate of TC4 and TA2.Moreover,it can reduce the content of Ti O₂or Al₂O₃in the surface passivation film and affect its corrosion resistance.