Study On Intracellular Synthesis Of Selenium Nanoparticles By Comamonas Testosteroni S44 Using Single-cell ICP-MS

Cells are relatively independent basic units of life,and the same type of cell metabolism is basically the same.Generally speaking,to study the basic growth law of cells,the main way is to observe the growth state of the whole cell populations.Because of the difference of microenvironment,the phenomenon of"cell heterogeneity"exists especially.For Single-Cell analysis,which advantage is that it can obtain more comprehensive information about the physiological state of cells.Under the existing technology,the method of Single-Cell analysis with inductively coupled plasma mass spectrometer(ICP-MS)as one of the main means that provides the method for detecting elements in a single cell.In fact,Single-Cell analysis with ICP-MS has demonstrated its detecting ability in drug,toxicology and so on.The content and distribution of selenium in single C.testosteroni S44 cell were analyzed by ICP-MS.Cells of Strain S44 could reduce sodium selenite,and produce selenium nanoparticles(BioSeNPs),by adding sodium selenite to culture and optimize strain S44.The Single-Cell analysis of ICP-MS was carried out by using the obtained cells containing nanoparticles.The main contents and results are as follows:(1)Preparation of intracellular biological selenium nanoparticles.By exploring the cultured conditions of strain S44,we found that the spherical selenium nanoparticles were obtained under the conditions of 28?,shaking speed 150 r/min,inoculation ratio 1%,volume of culture solution 100 mL and Se(?)concentration was 0.2mmol/L.The reducted efficiency of Se(?)was the highest(51.3%),when the size distribution was 50-300 nm.It was confirmed that the majority of BioSeNPs was intracellular under the concentration of Se(?)with 2mmol/L,and these cells were used as Single-Cell materias of ICP-MS.In addition,FT-IR spectra showed that strain S44 produced proteins and polysaccharides during the reduction of Se(?).(2)Optimized preparation of intracellular selenium nanoparticles.After determining the single factor culture conditions of strain S44,we optimized the culture medium.The effects of different additives on the reduction reaction were investigated,including bovine serum albumin(BSA),chitosan(CTS),polyvinylpyrrolidone(PVP),and the optimum ratio to the medium was 1:4,1:10,1:8.After adding BSA,the maximum reduction rate of Se(?)reached 62.3%,the amount of products increased obviously,and the secretion of proteins and other substances increased equally.Under the condition of 2mmol/L concentration of Se(?).The additives were cultured to obtain intracellular BioSeNPs but only PVP was found.(3)Single strain S44 cell was analyzed by ICP-MS.We used the natural materials containing intracellular selenium nanoparticles to explore the Single-Cell experiment of ICP-MS.The optimal resolution time of 0.1ms was determined.The cells were treated with cation exchange resin(CER).The suitable injecting concentration of cells were5×10~5cells/mL,combined with mathematical statistical analysis and integral analysis.The time-resolved data of ICP-MS showed the reducted products in details,and single BioSeNPs were used to qualitatively analyze the intracellular BioSeNPs.It is proved that the simulation degree is better by the aid of transmission electron microscope(TEM).