Applications of CARBON NANOTUBES in Biotechnology
Creators: name: Hussain, Zahid
Publisher: name: Zenodo
Year: 2024
Subjects: Applications
CARBON
NANOTUBES
Biotechnology

description:  

 

 

Abstract

Carbon nanotubes (CNTs), first discovered by Iijima in 1991, are a remarkable class of nanomaterials within the fullerene family. These elongated, hollow cylindrical structures, often described as "nano test tubes," possess extraordinary mechanical strength, electrical conductivity, and thermal stability. Their nanoscale dimensions and unique properties make them indispensable in various scientific and industrial applications. Among the synthesis methods available, Chemical Vapor Deposition (CVD) is the most widely used and efficient. This process involves the application of a high electric current between graphite electrodes in a helium atmosphere, leading to the controlled deposition of CNTs on vessel walls and electrodes.

The exceptional properties of CNTs have unlocked their potential in a wide range of fields. In electronics, they are utilized for developing high-performance transistors, sensors, and conductive materials. Their lightweight and high-strength characteristics make them ideal for reinforcing composite materials used in aerospace and automotive industries. Additionally, their high surface area and conductivity enable their use in energy storage devices, such as batteries and supercapacitors. The discovery of CNTs has revolutionized material science, paving the way for advanced technologies in medicine, energy, and environmental sustainability.

 

Introduction:

The third allotropic form of carbon buckminsterfullerene also known as buckyballs discovered in 1985 has ball form structure. There are 60 atoms of carbon arranged in icosahedral geometry in its structure. Smalley and kroto vaporize graphite with laser they got symmetrical closed cluster in which carbon atom are bonded to each other in spherical symmetry known as buckyballs which is greatly stable. Then for its commercial applications Wolfgang from max plank institute and Donald Huffman of the University of Arizona used slightly modified technique then described first. They vaporize graphite using carbon arc in a helium atmosphere. As a result of this experiment fullerenes were produced. [1][10]

Carbon nanotube discovered by Iijima belongs to a class of fullerenes rich family that is elongated having hollow structure inside. They were discovered in 1991 at NEC laboratories in Japan. They are nanometers sized ropes and test tubes. Many methods are used for its synthesis but carbon vapor Deposition (CVD) is the best one, in which two graphite electrodes are used and current of 50 A is applied between them in the helium atmosphere. It causes the evaporation of the graphite. Some part of it is deposited on the wall of the vessel and some part is deposited on the cathode, which are mainly the carbon nanotubes. [1][8][10]

 

 They are of two types multi-walled tubes that were discovered first and single walled tubes that were discovered 2 years after the discovery of multi-walled tubes.

 

 

 

 

Structure of CNTs:

Carbon nanotubes are of two types:

1.      Single-walled carbon nanotubes

2.      Multi-walled carbon nanotubes

Structure of SWCNTs:

Single-walled carbon nanotubes are formed by the wrapping of single sheet of graphite to assume the shape of the cylinder. Their wrapping can be of three types:

        I.            Arm chair nanotube

     II.            Zigzag nanotube

  III.            Chiral nanotube

 

 

 

 

Structure of MWCNTs:

MWCNTs are made up of multiple layers of graphite that rolled upon them to form a tube shaped with an interlayer spacing of 3.4 Å. The outer diameter of MWCNTs may range from 1 to 50 nm while the inner diameter can be varied. Two models are used to describe the structures of MWCNTs

 

        I.            Russian Doll model where the sheets of graphite are arranged in concentric cylinders.

     II.            Parchment model in which a single sheet of graphite is rolled around itself like a toilet tissue paper [1][3].

 

Applications:

Properties like small sizes, high conductivity, high mechanical strength and high flexibility make nanotubes use in nanoprobes. These probes can be used in high resolution imaging, nano-lithography, nanoelectrodes, drug delivery, and sensors etc.

 

High resolution imaging:

Multi-walled CNTs are attached at the tip of the scanning probe microscope used for the purpose of imaging. These nanotubes tips can be used in SEM, AFM etc. The advantage of using CNTs tips is that they are conducting and can make image of extremely small size particle or structure, deep cracks which are impossible for the probe using silicon or other metal tips. Biological molecules, such as DNA can also be imaged using nanotube tips for example imaging of amyloid-b-protofibrils that is related to Alzheimer’s disease whose resolution was never achieved before. Due to the elasticity of the CNTs they don’t suffer crash on contacting with the substrate. To detect different things CNTs can be chemically modified. They can also be used as molecular probe. [6]

 

DNA-CNT nano wire networks for DNA detection:

DNA in a solution can be detected in the presence of oligo-nucleotide functionalization enzyme.  The reactive site on the CNT is created by treating it with an acid for introducing a carboxyl group at its end. An ss-DNA is bridged between two SWCNTs through its amide groups in such a way that the two tubes are joined to each other at their terminus via covalent linkage through ssDNA. DNA analyte and oligo-nucleotide enzyme acting as a probe is immersed in a solution. DNA-CNT nano wire is kept between two electrodes in that solution. DNA to be analyzed has two recognition sites: one with the probe and the other with the bridging DNA strand. Thus in the presence of analyte DNA and probe the gap between two CNTs is filled by the complementary base pairing of probe. In the presence of suitable reagents, by enzymatic metallization conductive connections take place between two tubes upon the detection of DNA. So this technique can be helpful for detecting and study of various genetic diseases [5].

 

This method is also significant in the detection of matched and mismatched bases. When there is mismatching between the two strands, detection signal falls down slowly and becomes in sensitive. For just a single mismatch, there is a detection decline of about 70%. This device can be used for the detection of DNA at femto molar level and can be used for detection of DNA viruses such as Ebola virus [13].

CNT based electrodes with immobilized enzymes:

CNT-modified electrodes are improved for their selectivity and sensitivity when enzymes are immobilized on their surfaces. The CNT then acts as the transducer that communicates the signal between the enzyme active sites and their specific substrates. Enzymatic interaction can be covalent or non covalent. Non covalent methods do not affect the activity and efficiency of enzyme. Non covalent methods include adsorption, entrapment and encapsulation. In first method, enzyme is attached with help of binder or ion exchange resin. In second technique, (electro) polymerization reaction is carried out and enzyme is dissolved in resulting matrix of polymer. In encapsulation, enzyme is immobilized with help of sol-gels or hydro gels. Non covalent method distributes the enzymes randomly. However, equal distribution takes place through covalent attachment method. It also helps in direct electron transfer to active center of enzyme. This technique is very helpful in sequencing of human genome project. SWCNT modified GCE and MWCNT can detect smaller sequence of about 24 basepairs.

Immobilized enzymes as biosensors:

Detection of glucose:

For detection of glucose, glucose biosensors are used that incorporate an enzyme glucose oxidase immobilized on the surface of CNT. To stabilize the enzyme on CNT electrodes, phosphate buffer is used in presence of Argon conditions. It also prevents the denaturation of enzyme and contamination of biomaterial. Glucose oxidase (GOx) oxidizes β-D glucose to D-glucono 1,5-lectone using oxygen as electron acceptor and producing  H2O2. The amount of this H2O2 produced is detected electrochemically at the electrodes and can be estimated for the level of glucose present inside the body. So it has beneficial application for diabetes diagnostics [14].

 

 

Detection of phenolic compounds:

Some phenolic compounds and cresols are toxic, so their determination in food products and environment is very necessary. MWCNTs layered with Nafions are used for their detection. The enzyme tyrosinase is adsorbed on the surface of CNT modified electrode. In presence of oxygen, the oxidation of phenolic compounds in presence of tyrosinase converts them to quinone. The quinone produced at electrodes is detected electrochemically and its amount is then estimated for maintaining safe level. A CNT paste electrode is also modified with enzyme polyphenyloxidase (PPOx). It detects various phenolic compounds including phenols, catequine and catechols. These compounds are detected in various pharmaceutical products for maintaining their quality [12].

 

Dopamine sensing:

Sensing of dopamine is very important in medical diagnoses. The first biosensor utilizing CNT electrode was the dopamine sensor. Dopamine sensor consists of a multi walled carbon nano tube paste electrode. This electrode is covered by Nafion membrane sometimes, which blocks invading ascorbates that exhibit the same properties as dopamine. MWCNT electrode was used for the detection of dopamine in goat by making its brain tissue homogenate. When a microelectrode of carbon fiber is modified with an MWCNT, this device gives a linear response by increasing concentrations of dopamine in brain. The detection of dopamine is also performed by PPOx-modified CNT paste electrodes, which is sensitive to about 2 µA/mmol/L [9].

Organo phosphorus compounds:

Organo phosphorus agents are used as chemical warfare agents as they are neurotoxins. They are also used as common pesticides. Because of their neurotoxicity, their detection is very necessary. Two of these compounds are paraoxon and methyl parathion. Water resources and food supplies must be monitored for proper detoxification processes. The attractive method is the use of carbon nano tubes. Enzyme organo phosphorus hydrolase converts these organophosphorus compounds in to p-Nitrophenol. This enzyme is immobilized onto MWCNT at SWCNT and produced p-nitrophrnol is oxidized at electrodes. This electrode yields a stable electronic signal for p-nitrophenol over a period of about 60 minutes [11].

Cholesterol:

The clinical diagnosis of cholesterol is very necessary as it causes chronic cardiovascular diseases. Its daily intake must also be regulated. Cholesterol oxidase oxidizes cholesterol and generates H2O2 in the presence of oxygen. The biosensor used for cholesterol utilizes layer by layer technique in which enzyme ChOx is immobilized on gold electrode that is modified with MWCNT. These MWCNT are oxidized by nitric acid before their deposition on gold electrode. This device is sensitive to about 559µA/ (cm2 mol/L) and gives a linear response in range of 0.6-2 mmol/L [12].

NAD+:

Nicotinamide dinucleotide is present inside the living cell that is an important cofactor of redox reactions. NADH generation is detected when it is electrochemically oxidized at the electrodes. Bare electrodes present surface fouling by the formation of reaction intermediates and show high oxidation potential of NADH. Glassy carbon electrodes modified with carbon nano tubes offer advantages for the removal of above two problems. It gives a stable response with >90% activity that remains the same even after 60 minutes of initial use [9].

Nitric oxide:

Nitric oxide is an essential molecular messenger in organisms. It controls blood pressure and blood clotting and is also a neurotransmitter. Glassy carbon electrode which is modified with MWCNT is used for its detection amperometrically [11].

Cystein:

Cystein is sulphur containing amino acid that is frequently used in food and pharmaceutical industry. It is detected by GCE modified with carbon nanotube that is coated with platinum. It can detect in the range between 0.5µmol/L to 0.1mmol/L. It can be used only for 1 month. After this time it loses its activity. It is selective against tryptophan and tyrosine [14].

Cytochrome c:

Cytochrome c is heme protein presnt in Electron Transport chain in the mitochondria of plants and animals cell. It transfers electron through ETC. It can be detected by using GCE modified with SWCNT electrodes.  It can detect the concentration range from 30 μmol/L to 0.7mmol/L with a sensitivity of 3 μA/mmol/ L. Colloidal gold nanoparticles with MWCNT can detect electron transfer more frequently in comparison with using other electrodes [14].

Indole acetic acid (IAA):

Indole acetic acid regulates developmental processes of life cycle of many plants. Their growth conditions can be modified by its proper monitoring. Its concentration is detected by glassy carbon electrode modified with MWCNT that is stabilized by DHP. IAA is detected amperometrically. The oxidation peak is extremely enhanced by using this electrode instead of a bare GCE [12].

Detection of gases:

Nanotubes can be used as advanced chemical sensors. The electrical resistivity of SWNTs was change sensitively on exposure to gases containing molecules of NO2, NH3 and O2. By control monitoring in the change of the conductance of nanotubes, the presence of gases could be accurately monitored. Due to small dimensions and high surface area nanotube sensors could be operated at room temperature or at higher temperatures for sensing applications [9].

Food industry:

Many kinds of sensors and biosensors can be used in the food industry in order to provide safe and pathogenic free food. Contamination of foods caused by bacterial pathogens is an issue of great consumer concern. For instance, CNT- based biosensors can be used to check meat freshness. CNT-based chemical sensors can be used to check harmful chemical residues that may be added by food additives, insecticide, pesticides, herbicides, and other environmental contaminants in raw and processed foods [15].

 

Agriculture and fishing industry

In green house for the growth of plants humidity as well as CO2 concentration is very important. To measure humidity in green house MWCNT quartz crystal microbalance humidity sensor is used that can detect the humidity level from 5-97% and it has recovery time 60-70seconds. For detection of CO2   level carbon nano-tube based biosensor is used that detect the CO2   concentration in green house as well as in garden for the plant growth. CNT-based pressure sensors can be used for uniform spraying of liquid fertilizer, like insecticides, pesticides, and herbicides required for plant growth.

Optimal pH requires for the growth of fishes in fish industry. CNT-based pH sensors are used to measure the pH of the water. MWCNT based electrodes dipped in ethanol are used for the detection of hypoxanthine in fishes. Hypoxanthine is related with freshness of the fishes [15].

Detection of bacteria by carbon nanotubes:

Sticky nanotubes can detect infectious bacteria in seconds instead of days. Salmonella typhi, the typhus causing bacteria can be detected by sticky electrodes. If the process is modified or revolutionized, it can be carried out for a variety of bacterial detection. Bacterial detection is very laborious process that may take place about two or more days.

Aptamers are the genetic materials used for the detection of these bacteria. These aptamers are attached on surfaces of nanotubes and these sticky tubes are then pasted on the surfaces of electrodes. Then these sticky electrodes are dipped in to a test liquid. Bacteria present in the solution stick to the aptamers attached on nanotubes. Aptamers stick off from the surface of nanotube when bacteria stick on their surfaces. It changes the electrical conductivity of the electrodes. Number of bacteria can be detected in the solution by measuring this change. This technique is very precise; it can detect even a single bacterium in 5mL of solution. Important thing with this technique is that it is cheap and uncomplicated [4].

 

 

 

 

 

 

Detection of viruses by their interaction with antiviral antibodies:

Carbon nanotube biosensors are made by layer by layer technique. Electrodes are chemically functionalized for detection of their interactions with virus and antiviral antibodies. Layer by layer film is made by adsorption of polystyrene sulfonate and SWCNTs are prepared. Viral antibodies are immobilized on specific electrodes, and binding of these antibodies is enhanced by poly L-lysine. The specific antigen binds with these antibodies, and this binding is detected by the conductance of the device. The change in the conductance is directly proportional to the concentration of antigen. The length between the electrodes and the layers upon them affect the performance of this device [7].

 

 

 

Detection of water toxins:

Cyanobacteria and blue green algae produce Microcystin-LR that is a toxic chemical agent in the fresh water reservoirs like Northern Thailand. A large population of Thailand drinks untreated surface water. Microcystin is very lethal agent even in small quantities. It causes severe liver damage and liver cancer. The scientists of Michigan University were capable of detecting and removing microcystin with a small and efficient device, a paper strip infused with carbon nanotubes. Nanotubes are mixed with antibodies against MC-LR and then impregnated in the paper. When paper strips come in contact with contaminated water, antibodies starts squeezing between the nanotubes for making bonds with MC-LR. The detachment between these tubes changes their electrical conductivity. An external monitor connected outside measures the electrical conductivity and the device is 28 times faster than other complicated devices to detect the toxins. Nicholas Kotov who was the professor in the departments of chemical engineering, Material sciences and engineering, and biomedical engineering lead the project for making this device. Water treatment techniques even developed in US cannot work efficiently as this device works for detecting Microcystin LR. This device developed by Kotov provides quick, portable, cheap and sensitive test for allowing water treatment plants to detect multiple toxins, verifying the safety level of water [2].

Ultra sensitive biosensors for proteins detection:

A variety of illnesses can be finding out by a thin layer of protein-recognizing polymer that is coated on the surface of carbon nanotubes. This forms a crucial new bio diagnostic tool that utilizes electrochemical signals for detection of minute amounts of proteins. Boston College researchers reported these biosensors in the journal Nature Nanotechnology.

The primary iron storing protein of humans is ferritin which is detected by this method. Human papilloma virus encoded by E7 oncoprotein is also detected. The sensor that uses calmodulin discriminates between different proteins that adopt a variety of shapes in different solutions. Polymer structures are used for developing biosensors that are capable of recognizing various organic compounds. Subpicogram per liter sensitivity can be achieved by coating a non conducting polymer on surface of wire like nanotubes of approximately one 300th the size of a human hair [16].

Within the non conducting polymer coating, are the imprints of protein molecule that are central to the biosensor function? These imprints reduce the thickness of coating and during the contact between the polymer and the charges on protein surfaces; impedance level lowers down than the rest of the polymer. It signals the presence of proteins.

As compared to the common diagnostics techniques for the viral diseases, the nanotube molecular imprinting technique paves the way for biosensors capable of detecting human papillomavirus or other viruses many weeks before. This sensor can track HPV protein directly. Besides, this label free technique does not require any marker.