Research Progress on the Application of Graphene in the Field of Biomedicine

Yuting YANG Ke PAN Yuanjian ZHONG Jing LENG Lichun ZHAO

Abstract Graphene is the thinest nanomaterial known in the world， which has unique electronic mobility， super specific surface area， high mechanical strength， excellent corrosion resistance and surface chemical structure. Due to its special nanostructure and excellent physical and chemical properties， graphene has a broad application prospect in the fields of electronics， optics， magnetism， biomedicine， catalysis， energy storage and sensors. In order to better develop and utilize graphene data， this paper reviewed the structural characteristics of graphene， as well as its research progress in biosensors， bio-imaging， aerogel and other biomedical fields， hoping to provide scientific basis for better development of graphene and the development of graphene pharmaceutical products.

Key words Graphene; Biological medicine; Biosensor; Bio-imaging; Graphene aerogel

Graphene belonging to fullerene is a kind of two-dimensional carbon nanosheet material with hexagonal honeycomb lattice formed by carbon atoms hybridized with SP2. It is the thinnest nanomaterial known in the world， with a thickness around 0.335 nm[1-2]. In 2004， physicists Andre Geim and Konstantin Novoselov from University of Manchester in the UK successfully obtained a single-layer two-dimensional atomic crystal——graphite， for the first time using mechanical peeling[3]. Graphene can be converted into zero-dimensional fullerene， curled to form one-dimensional carbon nanotubes， and stacked into three-dimensional graphite， so it can be regarded as the basic unit constituting other graphite materials[4]. Graphene is the material known to man with the highest strength， the best toughness， the lightest weight， the highest light transmittance， and the best conductivity[2]. Furthermore， due to its large specific surface area， excellent corrosion resistance， easy adjustment of surface chemical structure and other characteristics， it has been widely used in such seven major fields as new energy， enlarged health， electronic information， energy conservation and environmental protection， biomedicine， chemical industry， aerospace and so on. Graphene has the highest strength among tested materials， reaching 130 GPa[5]. Its carrier mobility is 1.5×104 cm2/（V·s）[6]， which is twice that of the currently known indium antimonide material with the highest mobility， and more than 10 times that of commercial silicon wafers， and the value can even reach 2.5×105 cm2/（V·s） under specific conditions[7].

Graphene oxide is produced from graphene oxidation and has an aromatic structure. Compared with graphene， graphene oxide has larger specific surface area and layered structure. And it has a large number of oxygen-containing groups such as carbonyl， hydroxyl， carboxyl and epoxy groups[8-9]. Graphene oxide also has the advantages of large specific surface area， good hydrophilicity， excellent biological characteristics， good biocompatibility and low cytotoxicity[10].

The Chinese government attaches great importance to the development of the graphene industry， has issued a series of related policies for systematic layout， and included graphene into the 165 major projects of the "Thirteenth Five-Year Plan" in China. The implementation of relevant policies is of great strategic significance for promoting technological innovation， supporting the development of strategic emerging industries and accelerating the path of new industrialization with Chinese characteristics[2]. Graphenes special nanostructures and excellent physical and chemical properties make it promising for applications in electronics， optics， magnetism， biomedicine， catalysis， energy storage， and sensors. Graphene is regarded as the "future material" and "revolutionary material" of the 21st century， which has led to a boom in graphene research.

This paper reviewed the research progress of graphene in the medical field in recent years， and its application is mainly reflected in biosensors， biological imaging and graphene aerogel， in the hope of providing scientific basis for better development of graphene and promoting the research and development of graphene pharmaceutical products.

Application of Graphene in the Field of Biosensors

Graphene materials are one of the most promising materials for next-generation biosensors. Graphenes electron mobility at room temperature is greater than 15 000 cm2/（V·s）， while its electric resistivity is only 10-6 Ω·cm. Its excellent conductivity makes it widely used in sensors[7]. In living organisms， graphene and graphene oxide have good water dispersibility， biocompatibility， and affinity for specific biomolecules[10]. Graphene has excellent electron migration ability for some enzymes， and has good catalytic performance for some small biological molecules （H2O2， caffeine， morphine， etc.）， making it suitable for enzyme-based biosensors[11]. Graphene currently used in gas sensors is generally produced by chemical vapor deposition method， and the products have a complete structure and a large specific surface area， which is conducive to gas adsorption[12]. However，  graphene used in electrochemical sensors is generally produced by redox， and the products usually have more structural defects， and contain some unreduced functional groups， which is conducive to their application in the field of electrochemistry[13].

Exosomes are microcapsules with a double phospholipid membrane structure. They are rich in proteins， DNA and miRNA， have extremely high homology， and play an important role in intercellular communication， metastasis and angiogenesis[14-16]. In some diseases or cancers， the concentration of exosomes is related to the disease， so the quantitative detection of exosomes is helpful for the early analysis and diagnosis of diseases. Yi et al.[17] developed a reduced graphene oxide field effect transistor （FET） biosensor that can detect exosomes with high sensitivity. Their innovation is to quantify exosomes directly without labeling.

Electrochemical biosensors

Graphene has the ability to quickly transfer electrons， so it can not only effectively promote the redox reaction process near electrodes， but also can be used as a new electrode modification material to improve the repeatability， selectivity and sensitivity of sensors. Liu et al.[18] constructed a graphene-chitosan-horseradish peroxidase biosensor mainly by mixing horseradish peroxidase with graphene-chitosan nanocomposites， and the results showed that the sensor has high sensitivity to hydrogen peroxide and wide detection range， and shows high selectivity and good stability. Wang et al.[19] prepared a molecularly imprinted electrochemical sensor with high sensitivity and selectivity for dopamine， which can be used for the determination of dopamine hydrochloride injection. It uses graphene as an electrode sensitizing material and dopamine imprinted polymer as a specific identification material. Mohammad Hossein Ghanbari et al.[20] prepared a nanocomposite using reduced graphene oxide， gold nanoparticles and 2-amino-5-mercapto-1，3，4-thiadiazole electropolymerization membranes to construct an adriamycin electrochemical sensor. They modified glassy carbon electrode （GCE） with the nanocomposite material， and the results showed that the modified GCE can achieve high sensitivity detection of doxorubicin. This sensor has good application prospects， and can be used as an important tool for pharmacokinetic analysis， quality control， and rutin clinical trials.

Every year， more than 20 million people in the world are infected with Hepatitis E virus （HEV）， and more than 3 million people are infected with Hepatitis E virus[21]. The World Health Organization estimated that 44 000 people died of HEV infection in 2015[22]. However， a sensitive and reliable method for detecting HEV antigens has not yet been established. Therefore， there is an urgent need to develop sensitive and reliable technology for detecting HEV in food and patient samples to ensure health and safety. Ankan Dutta Chowdhury et al.[23] used graphene quantum dots and gold-embedded polyaniline nanowires to prepare pulse-triggered ultrasensitive electrochemical sensors by interfacial polymerization and self-assembly methods. The sensor is applied to various HEV genotypes， such as G1， G3， G7 and ferret HEV obtained from cell culture supernatants， and a series of fecal samples collected from G7-infected monkeys.

Graphene oxide-chitosan nanocomposites can also be used as electrochemical biosensors in the diagnosis of typhoid fever[24]. Zhang et al.[25] developed an efficient hydrogen peroxide biosensor with a gold/graphene-chitosan nanocomposite as a new electrode using electrochemical methods to fix redox proteins. He et al.[26] achieved high-sensitivity detection of proteins using graphene oxide as a substrate based on the characteristic of terminal protection. Wang et al.[27] synthesized by electrochemical method a grapheme/gold nanocomposite， which has high electrocatalytic activity， good stability， selectivity and anti-interference ability and can be used as glucose biosensors for the quantitative detection of glucose of diseases such as diabetes and hyperglycemia. Wang et al.[28] developed a sensor using graphene oxide nanosheets. They labeled ATP aptamers with fluorescent dyes to serve as molecular probes， thereby realizing the detection of ATP in living cells.

Human chorionic gonadotropin （hCG） is a very important protein that can be used to detect pregnancy and related diseases such as ectopic pregnancy， abortion， fetal abnormalities， and testicular tumors[29-32]. Nan-Fu Chiu et al.[33] developed by the unlabeled and non-immunological method of modifying bound peptides with functional carboxy-graphene oxide， a surface plasmon resonance （SPR） sensor， which has ultra-high sensitivity. The experimental results showed that the SPR-type sensor has higher affinity and stronger peptide binding ability， posing a greater impact on the non-immune unlabeled mechanism. In addition， it can detect low concentrations of hCG.

Optical biosensors

Graphene has photoluminescence properties， and the degree of oxidation of graphene oxide affects its fluorescence quenching properties. Graphene oxide can be used as a quencher and has a higher quenching efficiency than traditional quenchers. Using graphene oxide in a fluorescence resonance energy transfer （FRET） sensor endows the FRET sensor with low background interference， high sensitivity and the significant characteristic of multiple detection[10]. Zhang et al.[34] constructed a label-free graphene FRET sensor that can be used to detect specific sequences of DNA. Liu et al.[35] constructed a FRET sensor  using graphene oxide as a donor and gold nanoparticles as a fluorescent acceptor based on the immune response to DNA hybridization. The sensor can be widely used for detection of DNA， ions and small protein molecules.

Gas sensors

The large surface area of graphene makes it very sensitive to the surrounding environment， and even the adsorption or release of a gas molecule can be detected.  Wei et al.[36] developed a gas sensor that can improve the sensitivity of gas detection by the innovative application of N-doped graphene nanoribbon-modified electrodes. They showed through the investigation of electron migration rate and sensitivity analysis on NH3， CO and N2 that the sensitivity of the doped graphene sensor is much higher than that of non-doped graphene sensors. Wu et al.[37] prepared a sensor for detecting NH3. Their innovation lies in the use of grapheme/polyaniline nanocomposite as sensitive elements. The experimental results showed that the sensor using the nanocomposite as sensitive elements has high sensitivity and wide detection range.

Yuting YANG et al. Research Progress on the Application of Graphene in the Field of Biomedicine

Application of Graphene in the Field of Bio-imaging

As a clinical diagnostic technology， bio-imaging requires its core fluorescent probes to have good solubility， light stability and specificity in buffer， cell culture or body fluids[38]. Traditional biofluorescent probes contain heavy metal elements which are degraded in vivo to produce toxicity[39]. Since graphene oxide contains a large number of oxygen-containing functional groups and is easy to be functionally modified， using it as a fluorescent probe could realize good water solubility and biocompatibility. Peng et al.[40] connected graphene oxide and fluorescent dye via polyethylene glycol and used it for intracellular imaging. Hu et al.[41] produced a quantum dot-reduced graphene oxide nanocomposite. They used quantum dots （QD） to label reduced graphene oxide （rGO） to form a non-toxic QD-rGO nanocomposite with strong fluorescence and carried out bio-imaging experiments in tumor cells.

Application of Graphene in the Field of Aerogel

Aerogel is a three-dimensional nanoporous material with high porosity， large specific surface area and low density. After it is made， it will form a skeleton structure that maintains its original shape and generate nano-scale pores. Therefore， it has important research significance in many fields such as sensing， energy storage， adsorption and chemical catalysis[42]. In order to better develop and utilize graphene， researchers have focused their attention on graphene aerogels with three-dimensional structure. Because they have advantages of both graphene and aerogel， they have attracted widespread attention in academic research and application[43]. Cheng et al.[44] prepared a three-dimensional graphene aerogel with CH4 as the carbon source and nickel foam as the substrate. The experiment results showed that it has the advantages of high specific surface area， high porosity， high conductivity and self-supporting. Li et al.[45] prepared a microcapsule graphene aerogel with high conductivity and large specific surface area using organic cross-linking agents.

Conclusions

Graphene raw materials are cheap， easy to obtain， and simple to prepare， and are considered to be new materials with great application value. Graphenes application develops rapidly， and its research results are widely used in various aspects of the biomedical field. In general， graphene has good biocompatibility and bio-imaging characteristics and can fill some of the gaps in biomedical technology when applied in biosensors， cell imaging and aerogels， thereby playing an important role in greatly promoting the development of biomedical industry. The research on the application of graphene in the field of biomedicine is still at an early stage， but the special structure and excellent properties of graphene provide new possibilities for disease diagnosis and clinical medicine. Therefore， grapheme has great application prospects in biomedicine， and it is significant to further study the action mechanism of graphene.

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