36614 Nanotechnology in Biological Agent Decontamination
14.2
Standard Methods for Chemical Decontamination of Biological Agents
There are several approaches generally used for chemical decontamination of biological agents. The most widely used one is when the disinfectant agent is applied as a liquid or solution. Examples of such disinfectants are bleach solutions, chlorine solutions, chloramine T, glutaraldehyde solutions, phenolic solutions, and ethanol/ water mixtures. This approach has the advantage of being relatively quick and very efficient for many bacteria and viruses. Some of the disinfecting compositions, such as bleach, are able to detoxify certain chemical warfare agents as well (e.g., mustard gas). A major disadvantage of this approach is that the solutions deteriorate most surfaces rather rapidly. Although bleach is well suited for situations which require frequent disinfection, like hospitals, public areas, and others, it is not very useful for certain cases, such as industrial spills or terrorist attacks, where many types of surfaces as well as air and water are simultaneously exposed to the biological agent. The application of solutions or liquids is not desirable in the case of sensitive surfaces like those of electronic gear, paper documents, etc. Residues from the solutions are usually corrosive to many surfaces and difficult to remove once the disinfection process is over. Another significant problem of this approach is the need to use relatively concentrated solutions for disinfection of spore cells. Aging of these solutions is a serious issue which is often overlooked – the activity of most disinfecting solutions diminishes drastically with time [1, 2]. Eventually this could lead to overestimation of the efficiency of the disinfection.
An advantage in the liquid-based technology is the development of the so-called nanoemulsions developed by Baker Jr. et al. [3–5]. The nanoemulsions are based on small oil droplets in water stabilized by surfactant (diameter of the order of hundreds of nanometers). They have an advantage over standard liquid treatments in their ability to induce spore proliferation and subsequent death of the generated vegetative cells, which makes these nanoemulsions more effective against sporeforming microorganisms [4]. Although this approach improves the biocidal efficiency and is based on harmless chemicals, it is still limited to the type of surfaces to which it can be applied. It could not be used for air cleaning and would be difficult to remove once the disinfection is over.
Disinfection can also be achieved with gases such as chlorine or chlorine dioxide. Gases are applicable to closed environments only, and they are also very corrosive to practically all surfaces. Gases usually damage sensitive materials and surfaces like those of electronics, art artifacts, paper documents, and many others. Their application also requires specially trained personnel and a well-controlled environment (temperature, humidity, etc.) for the disinfection to be successful.
In general the “traditional” methods of disinfection are not suitable for decontamination from both biological and chemical agents. In these cases separate treatment has to be used for each potential threat, bringing up chemical incompatibility issues.
14.4 Magnesium Oxide 367
14.3
Nanomaterials for Decontamination
The most serious disadvantage of common solid materials as biological decontaminants is lack of good contact with the biological agent, whether virus particles or bacterial cells. Nanomaterials can compensate this disadvantage since they can be fabricated and dispersed as very small particles or aggregates. Additionally, solid nanomaterials can be dispersed either in a liquid or as a relatively stable aerosol, which increases their contact with potential biological hazards. Besides alleviating the contact problem, the nanomaterials introduce much higher chemical and higher biological activity.
The surface of solid materials is a very important factor concerning their overall chemical activity. When a material is broken down to smaller and smaller particles, more and more building elements are exposed on the surface. These exposed molecules have significantly higher energy than those inside the crystal, which translates into higher chemical reactivity. The activity of specific crystal morphologies such as edges, corners, defects, and high index planes is even higher. This is evident in the case of nanomaterials, where the concentration of these highly active sites is much higher compared to the corresponding bulk material. The higher surface reactivity translates into the capability of nanomaterials to adsorb many more molecules per unit surface area and bind them more strongly than a regular solid material can. Besides much higher chemical reactivity, the nanomaterials can be dispersed in much finer form [as airborne particles (aerosol) or dispersed in a liquid (sol)] than other solid materials, which significantly enhances their application and efficiency.
The high surface area and higher chemical reactivity which are inherent in nanoparticles make them suitable for a dual purpose. Besides playing a role as a biological decontamination agent, they could serve as excellent adsorbents for toxic chemicals and chemical warfare agents and convert them to benign compounds by breaking the labile bond responsible for their toxicity [6–12]. Other toxins are retained by physical adsorption only [13].
An important requirement for the solid material is that it should be practically nontoxic. Because of their small particle size, the nanoparticles and their aggregates can bypass the aerosol-gathering systems of the organism and penetrate to the lungs if they are applied as aerosol. Examples of harmless materials are asbestos and soot, which become dangerous only in the form of tiny aerosol particles. This restricts the choice of nanomaterials as disinfectants to a very small number of compounds. This review concentrates on the two which have the greatest potential and have been most researched in their nanoparticulate form: magnesium oxide and titanium dioxide.
14.4
Magnesium Oxide
Magnesium oxide is a highly ionic compound with high lattice energy. This means that magnesium oxide has a strong driving force to crystallize and a probable amor-
368 14 Nanotechnology in Biological Agent Decontamination
phous state would be unstable. However once “fixed” in nanocrystalline form, the MgO nanoparticles are stable and can endure heating to temperatures as high as 550 C without considerable sintering. Magnesium oxide has the advantage of being benign to organisms as well as practically harmless to most surfaces. One of its major advantages is that when left in contact with the atmosphere it converts to harmless magnesium carbonate.
Magnesium oxide nanocrystals can be prepared in sizes of the order of 4 nm and very high surface area (exceeding 500 m2 g–1) [14]. The particles arrange themselves in the form of a voluminous network made of separate nanoparticles (Fig. 14.1). For comparison, the commercially available magnesium oxide (comprised of large, mi- crometer-sized crystallites) has surface area in the range of 5–80 m2 g–1. The small size of the particles and the porous aggregates is of importance for the stability of the aerosol. Such an aerosol of magnesium oxide nanoparticles is stable for as long as 1 h [11, 15]. This allows the application of nanoparticle materials as aerosols for air volume decontamination combined with surface decontamination.
Figure 14.1. TEM image of magnesium oxide nanoparticles.
The nanoparticles are interconnected in a very porous network.
Magnesium oxide in the form of nanoparticles is a good bactericidal agent [15, 16] capable of killing more than 90% of gram-positive bacteria (B. cereus), gram-neg- ative bacteria (E. coli), and spores (B. globigii) in minutes. This is in contrast with the fact that “normal” magnesium oxide comprised of micrometer-sized particles does not exhibit appreciable bactericidal activity. The mechanism of the bactericidal activity is described later.
The increase of the surface area has been used to adsorb strong disinfectants such as chlorine, bromine, or interhalogen compounds. The resulting solid materials (MgO/X2, where X= Cl, Br, I) have excellent bactericidal and sporicidal properties [17], while at the same time they are not as harsh to the surfaces as most disinfectants. These halogen-loaded nanoparticle compositions contain large amounts of very active halogen (up to 43 wt% in the case of IBr), while at the same time the halogen is “activated” compared to its free state, enhancing its biocidal activity [17].
14.5 Mechanism of Action 369
Nanoparticulate magnesium oxide successfully removed traces of toxins from solution such as aflatoxins at parts per million levels [15]. This capability was observed earlier for both microand nanoparticles and is expected to be due to physical adsorption. However, the capacity to retain toxin molecules is expected to be higher for the nanoparticles due to their higher surface area and higher activity surface.
14.5
Mechanism of Action
It is of fundamental as well as of practical interest to understand the mechanism, of the bactericidal activity of nanoparticles.
Perhaps the most important disadvantage, as emphasized earlier, of using solid materials as disinfectants is the contact issue. Some of the cells are not touched by particles and survive the treatment. Electrostatic interaction is an important parameter of how colloidal particles such as nanoparticles could interact with other colloidal particles (bacteria cells or viruses). In aqueous solution it is described by the f-potential. In water the magnesium oxide nanoparticles are slowly converted to hydroxide. Upon dissociation of hydroxide anions the nanoparticles become positively charged. As determined by f-potential measurement, all magnesium nanoparticles composites (MgO/X2) are positively charged. This is favorable for an interaction with bacterial cells as they are negatively charged at biological pH values [18]. Confocal and optical microscopy studies show that mixing of magnesium oxide nanoparticles in suspension and bacterial cells causes spontaneous coagulation into large aggregates composed of nanoparticle aggregates and bacterial cells. This phenomenon was observed with all types microorganisms tested [16] (Fig. 14.2). The sticking effect was observed with atomic force microscopy, showing that upon contact the bacterial cells suffered considerable cell wall damage [13]. Upon contact with bacterial cells the aggregates of nanoparticles can fall apart into smaller aggregates [16].
Several other effects are of importance for the bactericidal activity of the nanoparticles in the dry state or as a suspension: their charge attraction with bacteria; their enhanced abrasive action; the oxidation capability (in cases where halogen or interhalogens are present); and their basicity.
The bactericidal activity of the nanoparticles is not limited to suspension; they are also bactericidal in the dry state. The important factors for the activity in dry states are explained by the abrasive properties of the particles, which can mechanically damage the cells. Since the nanoparticles adsorb relatively large amounts of water, they are desiccants (in the particles’ vicinity) and generate a high pH. Both factors are considered responsible for the bactericidal activity of the nanoparticles against vegetative bacterial cells.
This is supported by the fact that spores, which have mechanically stronger cell walls and are not sensitive to the amount of water or high pH, are not influenced significantly by the magnesium oxide nanoparticles. However, it is observed that halogen-loaded nanoparticles (nanoparticle compositions prepared by adsorbing free halogens on MgO nanoparticles, MgO/X2, X=Cl, Br, I) have excellent sporicidal
14.7 Summary 371
compared to gas-phase free halogen [17], combined with the high local halogen concentration. The sporicidal activity of the halogen-loaded nanoparticles is schematically shown in Fig. 14.3.
The advantage of a solid disinfectant is apparent in this example: the nanoparticle brings to the spore cell a high concentration of adsorbed active halogen, and its oxidizing activity is promoted by the nanoparticle carrier, which removes the most resistant layer of the spore protection. At the same time, the overall concentration of the halogen in the environment is low and the halogen is released primarily upon contact with a cell and secondarily due to desorption.
14.6
Titanium Dioxide
Another nontoxic metal oxide of interest is titanium dioxide (titania). It becomes a very potent oxidation photocatalyst when illuminated with UV light. Nanoparticulate titania was found to be an active bactericide in water suspensions. Compared to magnesium oxide, its biocidal activity is optimal only in the presence of UV light [22, 23]. Films of titanium dioxide were found to have detoxification abilities by oxidizing biological toxins such as the E. coli endotoxin under UV light in aqueous solutions [24]. The conclusion from the literature reports is that as a very powerful photocatalyst the titania nanoparticles catalyze the catalytic oxidation and destruction of the cell wall by generating high concentrations of highly active inorganic radicals [22]. It was observed that smaller particles and higher anatase content perform better than other titania nanoparticulate composites [23]. An advantage of the nanoparticles versus bulk titania is again easier dispersibility and higher biocidal activity.
14.7
Summary
Nanomaterials have properties different from those of the corresponding bulk material they are made of. They have different chemical and physical properties and sometimes exhibit properties never observed in bulk matter. One direction which still hides many unknowns is how the change in size and properties influences their biological activity. Some materials, as discussed in this chapter, have been found to interact with biological materials in a manner different from the bulk. These new properties and behavior in biosystems can be put to good use as a new line of defense against biological threats.
