Study On The Oxidation Of Cyclohexane/Cyclohexanol To ?-Caprolactone By Cascading Visible-light-driven Photocatalysis Combined With Whole Cell Biocatalysis

?-caprolactone is an important raw material for the production of nontoxic and degradable plastics.Because of its good biocompatibility,permeability and biodegradability,?-caprolactone is widely used in food,medicine and other industries.At present,the main method for preparing?-caprolactone is chemical method,but it has the disadvantages of high risk,high energy consumption,and producing many by-products during the process.Compared with chemical method,visible-light-driven photocatalysis and biological enzyme catalysis have the advantages of mild reaction conditions and environmental friendliness.The photo-enzymatic cascade reaction has shown unique advantages in reducing pollution,improving reaction safety and product selectivity,which is one of the most promising research directions in the field of green chemistry.However,the current research on producing lactone by this technology needs further exploration.In this study,the photo-enzyme catalytic reaction system is to be applied to the synthesis of?-caprolactone.The purpose of synthesizing?-caprolactone directly from cyclohexane or cyclohexanol is realized by constructing a combined catalytic reaction system of visible light photocatalytic reaction?Au-Ti O₂,g-C₃N₄ as photocatalyst?and enzyme?Ac CHMO@E.coli?,which provides a new thought for green synthesis of?-caprolactone.The specific research contents and results are as follows:1.Preparation,characterization and catalytic capability of visible light photocatalyst.Two kinds of semiconductor photocatalysts were prepared:titanium dioxide loaded with gold nanoparticles on the surface?Au-Ti O₂?and graphite phase carbon nitride?g-C₃N₄?.The effects of reaction time,light intensity,substrate concentration and photocatalyst concentration on the photocatalytic cyclohexane oxidation reaction were investigated respectively.The experimental results show that Au-Ti O₂ nanocomposites have the same crystal form and structure as Ti O_2.Both photocatalysts have absorption peaks in visible light wavelength range,and can catalyze cyclohexane oxidation reaction in visible light.Under the conditions of light intensity of 149?M/cm²,100 m M cyclohexane was used as the substrate to react for 72 h,1.75±0.12 m M cyclohexanone was obtained from Au-Ti O₂ system,which is 3 times to the g-C₃N₄ system,indicating that the photocatalytic efficiency of Au-Ti O₂ system for this reaction was higher than that of g-C₃N₄ system.2.Preparation and catalytic properties of cyclohexanone monooxygenase Ac CHMO.Firstly,cyclohexanone monooxygenase?Ac CHMO?derived from Acinetobacter calcoaceticus was expressed in E.coli BL21?DE3?,and the target protein was purified by affinity chromatography.The experimental results showed that the molecular weight of Ac CHMO recombinant enzymes was 62.0 k Da and the total cell yield was 30.00 g/L?wet weight?.Secondly,the efficiency of cyclohexanone to?-caprolactone catalyzed by whole cell biocatalysis and pure enzyme was compared.The results show that under the conditions of p H8.0,temperature 25?and reaction time 24 h,the product concentration catalyzed by Ac CHMO@E.coli can reach 4.27±1.28 m M,which is higher than that catalyzed by Ac CHMO purified enzyme?0.55±0.11 m M?,indicating that Ac CHMO@E.coli has higher catalytic efficiency.3.Study on synthesis of?-caprolactone from cyclohexane or cyclohexanol catalyzed by combination of visible light and whole cell biocatalysis.The synthesis of?-caprolactone from cyclohexane or cyclohexanol catalyzed by combination of visible light and Ac CHMO@E.coli was investigated.The experimental results show that cyclohexanone can be generated in situ by photocatalytic oxidation of cyclohexane or cyclohexanol with Au-Ti O₂ and g-C₃N₄ as photocatalysts under visible light irradiation.Cyclohexanone produced in situ was used as the catalytic substrate,Ac CHMO@E.coli,NADPH and glucose were added,and?-caprolactone was finally obtained through Baeyer-Villiger reaction.The influencing factors of cyclohexanol synthesis route are further studied and optimized.The experimental results show that when cyclohexanol is 5.0 m M,Ac CHMO@E.coli concentration is 100.0 g/L,NADPH concentration is 2.5 m M,glucose concentration is 1.0%,the conversion rate reaches 23.70±1.41%and1.18±0.07 m M?-caprolactone can be produced.In summary,the photocatalyst Au-Ti O₂ and g-C₃N₄ were successfully prepared and characterized in this study.The prepared photocatalyst is combined with cyclohexanone monooxygenase?Ac CHMO?to construct a visible-light-driven photoenzymatic cascade reaction system and successfully synthesized?-caprolactone from cyclohexane or cyclohexanol through direct oxidation,thus realizing the goal of green and safe preparation of?-caprolactone.The results of this study provide innovative thought for the green synthesis of?-caprolactone and have reference significance for the green synthesis of other lactone compounds.