The Study Of Novel Electrochemiluminescent Biosensors Based On Luminol And Luminol Functional Nanomaterials

Electrochemiluminescence?EC L?is a new analytical method developed on chemiluminescence and electrochemistry,which provides a new method for the detection of biological molecules.EC L technology has the characte ristics both chemiluminescence and electrochemistry,such as high sensitivity,good selectivity,lower background signal,wide linear range,simple equipment,easy control and so on.In particularly,the EC L biosensor based on Luminol expresses great potential and advantages in the detection of bioactive molecules.The EC L intensity of luminol can be strongly enhanced by hydrogen peroxide?H₂O₂?,which is the mainly product in the life system that many bioactive molecules were catalyzed to produce H₂O₂ by the corresponding oxidase.Based on this principle,an EC L biosensor could be easily constructed to ultrasensitively detect the bioactive molecules for example glucose,lactic acid,choline and so on.With the rapid development of life science,the analysis of bioactive molecules has become an important research content of biologists and analysts.It is one of the means to uncover the secrets of life,monitor vital signs and conquer the disease.So the quantitative analysis of biomolecules by electrochemilum inescence has become a research hotspot.However,because of the short development history of EC L biosensors,the service life,stability and sensitivity of the EC L enzyme biosensor based on luminol need to be further improved.Furthermore,in conventio nal fabrication of luminol-enzyme based biosensor,luminol as luminophore was often added into the sample solution.The model is simple but brings some shortcomings: luminol could contaminate sample,is toxic to living sample,increase the error of detection and so on.Therefore with the help of development of nano technology and nano materials,developing more excellent EC L biosensors or novel luminol-based EC L materials for solid-state EC L biosensor to fast,sensitive,non polluting detection become an important subject.Based on the above reasons,the main research contents in this paer are as follows:?1?Construction of glucose EC L biosensor based on a three dimensional response platform.Glucose is an important biomolecule in metabolism and disease diagnosis.It is of great significance to precisely measure the content of glucose in serum for human health.In recent years,the fabrication of glucose biosensor used of EC L method has made great achievements.But in the construction of the traditional EC L sensor of glucose,glucose oxidase?GOD?was closed by Nafion or chitosan polymer film,which was high resistant,packaging enzyme active center.So the sensitivity of those biosensors was not high.In order to improve the sensitivity and to construct a more effective response platform,polyaniline?PANI?and Au nanoparticles?AuNPs?with high conductivity were used.Herein,we built a unique smart three dimensional?3D?network bio-platform by electro-polymerizating PANI nanowires and sequentially self-assembling AuNPs and GO D to sensitively detect glucose.Due to the large specific surface area,excellent conductivity of AuNPs/PAN i and large amount of loading of GOD,the 3D bio-platform shows excellent ECL performance for glucose detection.The sensor also demonstrates a satisfactory result in serum samples detection.?2?Synthesis of novel EC L material of Lu-AuNPs/rGO and applying in detection of H₂O₂ released from living cells.Although the EC L biosensors based on luminol obtained wide application,there are some defects to limit the further use of traditional luminol EC L sensor because of the addition of luminol into the sample solution,which would bring pollution to the sample and electrode,interfer detection,furthermore may have the toxicity to living object.So immobilization of luminol on electrode for developping solid-state EC L biosensor is an important means to overcome the problems.In this experiment,we used reduced graphene oxide?rGO?as a carrier,using luminol directly to reduce gold chloride acid?HAuCl₄?,to successfully synthesized a new EC L material,luminuol reduced gold nanoparticles/reduced graphene oxide composite?Lu-AuNPs/r GO?in hydrothermal conditions.The composite exhibits excellent EC L properties and is easy to fix on electrode.O n this basis,we constructed a transparent EC L sensor using this composite and indium tin oxide glass to realize in situ detection of H₂O₂ from human hepatocellular carcinoma cel s?HepG2?.?3?Controllable synthesis of popcorn-like Au@Polyluminol nanoflowers and its application in solid-state uric acid EC L biosensors.Although the works of Luminol immobilization making some progress,including the synthesis of Lu-AuNPs/rGO,there are still some defect to be improved for example the long-acting and stability of these materials.It is very attractive to look for more durable and stable luminescence materials.Luminol has a similar structure with aniline,previous works have synthesized polyluminol by electropolymerization and demonstrated polyluminol still has EC L characteristic,and more stable and long-term.In this work,by using hydrothermal method,we synthesized a new kind of EC L materials: polyluminolcoated gold nanoparticles with core shell structure?Au@polyluminol?by adjusting the ratio of the reaction between luminol and gold chloride.The product displays highly uniform size,good dispersion and strong ECL response towards trace hydrogen peroxide?H₂O₂?.The synthetic method is fast,high yielding and good repetitive.When it was combined with uricase to fabricate uric acid?UA?EC L sensor for detection of uric acid in serum sample,a satisfactory result also obtained.?4?Synthesis of rambutan-like Au/PLUM/GOD nanoball and its application in detection of glucose.In the process of fabrication of enzyme EC L biosensor based on Au@polyluminol,the amount ratio between enzyme and Au@polyluminol must be optimized for strongest signal,which increases the sensor preparation procedure and error.If Au@polyluminol could directly bind enzyme in the synthesis process,the product would be directly used for the detection of biomolecule,which greatly simplifies the manufacturing process of the EC L biosensor.In this experiment,we successfully synthesized glucose oxidase/Au nanoparticle/polyluminol composite?Au/PLUM/GOD?in room temperature.By adjusting the proportion of reactants,after 48 h reaction time,Au/PLUM/GOD nanoballs with rambutan and uniform shape,good dispersion,400 nm diameter could be produced.The nanoballs were added on the glassy carbon electrode to realize direct detection of glucose and showed good sensitivity and wide linear range.The synthesis method also provides an effective method to construct other enzyme ECL sensor for the detection of corresponding biological molecule.