Preparation Of Enzyme-mimicking Hydrogel Complexes And Their Applications In Colorimetric Biochemical Sensing

In recent years,the problem of environmental pollution has seriously threated to human henlth,and the incidence of various cancer diseases tends to be younger.The development detection of rapid,sensitive and simple methods for heavy metal ions and biomarkers is of great significance for environmental protection,early diagnosis and prevention of clinical diseases and monitoring during treatment.Compared with traditional detection methods,the colorimetric biochemical sensors based on enzyme mimic possesses the advantages of visualization,simple operation,response rapid,low cost,large instruments-free and even suitable for point-of-care testing（POCT）,they have considerable application prospect in the field of accurate and rapid detection of targets in complex matrix.In addition,due to their excellent biocompatibility,flexibility,and mechanical stability,hydrogels can be well combined with thermal stability enzyme mimic,which provides new ideas for designing a variety of highly sensitive and portable colorimetric biochemical sensors,and also helps to broaden the application range of enzyme mimic.Therefore,based on enhanced oxidase-mimicking enzyme mimic activity with excellent biocompatibility and stimuli-responsive hydrogels,this thesis construct the colorimetric biochemical sensors for the highly sensitive and portable detection of heavy metal ions and biomarker,which are as follows:1.Simultaneously colorimetric detection and effective removal of mercury ion based on preparation of a hydrogel analysis tube containing novel enzyme mimicBy combining the enhanced oxidase-mimicking activity of composite material and agarose hydrogel,this work successfully develops a new colorimetric sensing method for the rapid and simple detection of mercury ion(Hg²⁺).Firstly,an environmentally-friendly and cost-effective enzyme mimic was obtained by facile one-pot preparation of chitosan/Cu/Fe（CS/Cu/Fe）composite.The CS/Cu/Fe composite exhibited significantly enhanced oxidase-mimicking activity during catalyzing the oxidation of 3,3’,5,5’-tetramethylbenzidine（TMB）.Benefiting from the thermal stability and the compatibility with hydrogels,the CS/Cu/Fe composite was further integrated with agarose hydrogel to fabricate a portable analytical tube containing oxidase mimic.Based on the inhibition of the catalytic oxidation of TMB in the presence of cysteine,as well as the recovery of oxidase-mimicking activity of CS/Cu/Fe due to the specific complexation of cysteine and Hg²⁺,the rapid colorimetric detection of Hg²⁺was successfully carried out in the analytical tube by using the change of solution absorbance.Under optimal conditions,this method could accurately detect Hg²⁺in the linear range of 40 n M to 8.0μM with detection limit as low as 8.9 n M.The method also revealed high selectivity and satisfactory results in recovery experiments of Hg²⁺detection in tap water and lake water.Furthermore,it was found that the effective removal of Hg²⁺could be realized in the analytical tube based on efficient Hg²⁺adsorption by CS/Cu/Fe composite and agarose hydrogel.Therefore,this work not only prepared a robust and low-cost enzyme mimic,but also proposed an intelligent strategy to simultaneously monitor and remove toxic Hg²⁺from contaminated water,which has great application potential in the field of environmental protection.2.Colorimetric detection of ALP based on pyrophosphate-triggered controlled release and activity-enhanced Co OOH nanozymeA highly sensitive colorimetric method for alkaline phosphatase（ALP）detection was successfully constructed based on pyrophosphate ion（PPi）specific triggering dissociation of hydrogel to release cobalt oxyhydroxide（Co OOH）nanozyme with oxidase-mimicking activity.Firstly,Cu²⁺was used as crosslinking agent to react with alginate（Alg）,and the Co OOH nanozyme was wrapped in Cu/Alg matrix to prepare Co OOH@Cu/Alg hydrogel.Due to the strong affinity between PPi and Cu²⁺,the addition of PPi could competitively bind Cu²⁺to form the PPi-Cu²⁺complex,thus destroying the network structure of Co OOH@Cu/Alg hydrogel and releasing Co OOH nanozyme.The transformation process of the gel-sol could not only be observed with naked eyes,but also catalyzed TMB color rendering by using Co OOH oxidase-mimicking activity released.Moreover,it was found that PPi and PPi-Cu²⁺complex could further enhance the activity of Co OOH nanozyme.Since ALP can catalyze the hydrolysis of PPi to phosphate ion（Pi）,it cannot trigger Co OOH@Cu/Alg hydrogel dissociation to release Co OOH nanozyme.Therefore,based on the above-mentioned controllable release strategy of Co OOH nanozyme,a new colorimetric detection method for ALP can be established.Under optimal conditions,the as-constructed sensor exhibited good linear response to ALP over the range from 0.1 to 250 m U/m L with a detection limit of 0.07 m U/m L.Additionally,the method was successfully applied in the detection of ALP in actual human serum,and could clearly distinguish the concentration level of ALP in normal human serum and patient serum.The ALP colorimetric sensing method established in this work has simple operation,response rapid,high sensitivity and excellent selectivity.Thus,it shows a promising prospect for practical applications.