WATER STERILIZATION USING Ag-SUPPORTED SOLIDS WITH IMMOBILIZED SILVER NANOPARTICLES

Abstract

A silver-based solid material for disinfection at room temperature of micro-organisms from water or wastewater such as ship's ballast water. The material includes an aluminum oxide support material with nano-silver particles adhered to a surface of the aluminum oxide support material. The nano-silver particles are formed by decomposition of a silver precursor impregnating the aluminum oxide support in an oxygen-containing atmosphere at a temperature of at least 600°C followed by a reduction in a hydrogen-containing atmosphere at a temperature of at least 300°C. The nano-silver particles are present in an amount between 0.01 and 5 wt% on the aluminum oxide support material. Ships ballast or wastewater reactors including the silver-based solid materials are provided. Advantageously, the disinfection occurs without the use of ozone, microwaves, electro-ionization, ultrasound, UV radiation, or thermal treatment.

Description

WATER STERILIZATION USING Ag-SUPPORTED SOLIDS WITH IMMOBILIZED SILVER NANOPARTICLES

[0001] Field of the Invention:

[0002] The field of the invention relates to sterilization of ship ballast water, and, more particularly, to the sterilization of ship ballast water using silver immobilized on solid supports.

[0003] Background:

[0004] In a ballast system, water moves in and out of a ballast tank in order to balance a ship. Ballast water (BW) is used by ships to provide stability by ensuring hull integrity, stability and maneuverability; ballast water further compensates cargo load during cargo loading and unloading. Ballast water may also compensate for a ship’s fuel and water consumption. According to statistical data, ballast water used in shipping results in about 10 billion tons of water being transferred into and out of ballast tanks; this ballast water is discharged in various ports across the globe. As a result, the biota contained in ballast water, e.g., micro-algae, small invertebrates, larvae of aquatic plants and fish, seeds, spores, eggs, viruses and bacteria may disrupt an aquatic or non- aquatic ecosystem in the region where the ballast water is discharged. These biota may include numerous non-indigenous species. As a result, natural barriers between different global marine species are breached.

[0005] Thus, through the essential use of ballast water a considerable environmental issue arises with potential devastating consequences for both the world economy and global public health. Concerning public health, many microorganisms such as bacterial pathogens which are introduced to new marine environments are responsible for the spread of serious diseases and pandemics in numerous regions of the world. In addition, the high density of those bacteria in ballast water combined with their extremely rapid reproduction rates, their broad resistance to a wide range of environmental conditions, and their ability to survive under harsh conditions, prompted the international community to recognize the treatment of ballast water bacteria and viruses as a priority issue, along with the invasive phytoplankton and zooplankton species. Therefore, in order to reduce the transportation of pathogenic species by ships, strict regulations have been enforced by the International Maritime Organization (IMO) regarding discharge of water including human pathogens: Vibrio cholera (less than 1 cfu / 100 mL), Escherichia coli (less than 250 cfu / 100 mL) and the bacterial group intestinal enterococci (less than 100 cfu / 100 mL) These regulations along with other ballast water management guidelines are included in the International Convention forthe Control and Management of Ships’ Ballast Water and Sediments (BWM Convention) which was enacted by IMO on 13 February 2004.

[0006] Several techniques have been applied to remove pathogens and invasive species from ballast water. Ballast Water Treatment (BWT) systems include UV radiation, electrochlorination, and ozonation. In the UV radiation treatment, UV sterilizes the water during the ballasting and deballasting processes (i.e., when taking in or releasing back the water to / from the sea respectively), using UV lamps. As a technology, it may be used for the eradication of any threatening invasive species, without any risks / hazards for the environment since there is no use of dangerous chemicals. Additionally, there is not any risk of corrosion for ships’ ballast tanks.

[0007] Electrochlorination includes the creation of reactive chlorine compounds by electric current passing through (electrolysis) ballast water with high salt content. Besides the concentration of chlorine in the water, the efficiency of the process also depends on the temperature, reaction time, and amount of any residual chlorine and its compounds. However, the reaction of chlorine with organic material creates hydrogen and chlorine gases, which are toxic and highly flammable when mixed, so the process requires strict safety measures for chemical storage and handling. Further, electrochlorination also causes the creation of dangerous by-products (measured as Treatment Residual Oxidants - TRO) such as chloroform, trichloronitromethane, and dichloroacetic and trichloroacetic acid. For this reason, the neutralization of by-products is considered necessary prior to the discharging of ballast water back into the sea. This may be achieved through the addition of sodium bisulfite or sodium sulfite to the treated water to avoid unauthorised excessive TRO discharges.

[0008] Ozonation uses ozone gas (oxidizing biocide) which is bubbled into the ballast water using an ozone generator. The ozone gas reacts with other chemicals to kill organisms in the water. Ozone is a very powerful but unstable, oxidizing agent which destroys viruses and bacteria. It has been used for over a century as a disinfectant widely in Europe and less widely in the USA. As in the case of electrochlorination, ozonation also causes the creation of dangerous by-products which pose a serious environmental or health concern as these compounds are probable carcinogens.

[0009] Table 1 summarizes the current methods for ballast water treatment and their shortcomings, demonstrating that these methods are not sufficiently effective for the microbial treatment of ballast water.Table 1 : Evaluation of the current ballast water treatment technologies

[0010] Silver (Ag), along with certain other metals such as mercury and tellurium, is highly toxic to cells, even at extremely low concentrations; the defense mechanisms used by cells are overcome by the activity of metal ions. Although the mechanism of action of silver is not yet fully elucidated, its antimicrobial activity lies mainly in the creation of silver ions, the generation of excess reactive oxygen species (ROS), and the inhibition of gene expression. Silver ions resulting from the oxidation of Ag° may interact with key enzymes and proteins, causing inhibition of metabolic pathways, or induce the formation of pits in the extracellular membrane of microorganisms, resulting in cell lysis. Regarding the generation of excess ROS, penetration of ROS inside the cell can cause oxidative stress or DNA damage leading to cell death; ROS may also give rise to the observation of severe morphological changes in the outer cell membranes. Thus, the present invention determined that the antimicrobial properties of silver could be used for treatment of ballast water.

[0011] Silver nanoparticles (AgNPs) have been shown to cleave the bacterial extracellular membrane, causing the alteration of cell potential and the inhibition of cell respiration by expansion. Although the toxicity of silver is mostly attributed to the release of silver ions, someforms of AgNPs or nanoclusters may exhibit additional capabilities which are not attributed to bulk / ionic silver. However, due to their prevailing mechanism of action, AgNPs do not meet the standards for certain environmental applications, such as water, wastewater and ballast water disinfection. This is because of the need for metal (Ag) recovery. Therefore, the present invention turned to immobilized silver nanoparticles (Ag-iNPs) as an alternative for ballast water treatment. Using silver-supported catalysts; Escherichia coli (gram-negative) and Escherichia faecalis (grampositive) were selected as indicator microbes, according to the regulation D2 of IMO’s BWM Convention.

[0012] Therefore, there is a need for more efficient, affordable and environmentally friendly techniques for ballast water microbial treatment. The present invention addresses this need.

[0013] Summary of the Invention:

[0014] The present invention provides a silver-based solid material that exhibits excellent disinfection activity, at room temperature, towards the elimination of micro-organisms from water or wastewater, including the human pathogens Vibrio cholera, Escherichia coli and the bacterial group intestinal enterococci. The solid material consists of silver (Ag) crystallites in contact with a metal-oxide support (such as AI2O3), in the form of powder or particles. The lwt% Ag / AbO, solid can be prepared by any means known to practitioners of this art, including the technique of wet-impregnation of the support with an aqueous solution of a Ag precursor (e.g., silver nitrate solution). At least 2 hrs of calcination with air at 600°C is needed, following the preparation of supported Ag solid, for complete conversion of the silver precursor crystallites to silver oxide ones. Finally, a reduction with hydrogen at 300°C is necessary for at least 2 hrs for complete conversion of silver oxide to silver in the metallic form. The resulting solid surface then becomes highly stable, exhibiting no deactivation during 72 hrs of reaction time or more. From now on the abovedescribed catalyst will be referred as Ag / AhCh. Almost complete inhibition (destruction) of A. coli is obtained at 20°C over this solid material at a rate of 600 cfu.cm’3.gcat’1.niin’1. Similar results were also obtained towards V. parahaemolyticus and I. enterococci. It is noted that other commercial disinfection methods used for e.g., ballast water treatment, have similar inhibition rates. Upon the application of this Ag-based solid (e.g., lwt% Ag / AbCh) the disinfection of water (e.g., ballast water) at a larger scale may be practically considered.

[0015] Brief Description of the Drawings:

[0016] FIG. 1 presents the E. coli bacterial inhibition profile as a function of Ag metal loading. Reaction conditions: ms= 1.0 g; P= 1.1 atm; T = 25 °C; stirring rate = 150 rpm.

[0017] FIG. 2 presents the effect of support chemical composition (y-ALCh, CeCh-AbCh and QSG) on the inhibition activity of Ag / support against E. coli. Reaction conditions: ms= 1.0 g; P= 1.1 atm; T = 25 °C; stirring rate = 150 rpm.

[0018] FIG. 3 shows the inhibition profile of E. coli, E.faecalis and V. parahaemolyticus obtained on 1 wt% / AhO3, with time on stream. ws= 1 gr, Vsoi=180 mL , T =25 °C , p =1 atm, stirring rate =150 rpm.

[0019] FIG. 4 shows the effect of the presence of 5 ppb of oxalic acid on the antimicrobial activity of 1 g of 1 wt% Ag / y-AbCF solid compared to the antimicrobial activity of 0.3 ppb free Ag+ ions, regarding the treatment of E. coli. Reaction conditions: P = 1.3 atm; T = 25°C; Vsoi. = 200 mL; stirring = 150 rpm.

[0020] FIG. 5 presents the bacterial inhibition obtained after the long-term exposure of an E. coli bacterial solution to the solids 1 w.t.% Ag CeCh-AbCh and CeCh-AhCh, in continuous flow reactor. Reaction conditions: ws=3 gr, Vsoi. = 180 mL, Qiiquid =10 mL / min, P = 1.3 atm, T = 25°C, stirring rate = 150 rpm.

[0021] FIG. 6 shows the bacterial inhibition of E. coli and Tot. coliforms obtained during the treatment of a ship’s ballast water (pilot test on a real ship) over the 1 w.t.% Ag / QSG solid, using the reactor design depicted in Fig. 8. Reaction conditions: ws=25 kg, P = 1 atm, T = 20°C, air flow rate = 10m3 / min.

[0022] FIG. 7 shows 3D drawings of a typical, simple reactor system that can be used for the process proposed in the present work.

[0023] FIG. 8 presents the effect of the feed composition on the bacterial inhibition vs. time obtained over 1 w.t.% Ag / CeCh-y-AFO,, in a continuous mode reactor system. Reaction conditions: T =25 °C, P =1 atm, Qiiquid =10 mL / min, Qgas=100 mL / min, stirring rate =150 rpm.

[0024] Detailed Description of Specific Embodiments

[0025] The present invention provides a cost effective and efficient treatment method for ships’ ballast water without ecological risk. The present invention uses silver (Ag) supported alumina (AI2O3) spheres, where silver is immobilized on the surface of the alumina (AI2O3) spheres forming silver nano- clusters. This silver-based solid exhibits excellent disinfection activity towards the elimination (inhibition) of A. coli, V. cholera and / , enterococci, at room temperature (20 °C). The solid material was prepared by a wet-impregnation method described in further detail in the Examples below. Identical solid materials can be prepared using other preparation techniques, known to practitioners of this art, having other metal (Ag) loadings and using other metal precursors such as silver acetate, silver acetylacetonate, silver citrate, etc. However, it was found in this work that the preparation of the above-mentioned solid material using the silver nitrate precursor and a lwt% silver loading resulted in better results, in terms of inhibition activity for the bacteria examined (see Fig. 1 and Example 2). Five different silver loadings (wt.%) supported on AI2O3 spheres were prepared and examined within this work, while three different support chemical compositions were evaluated.

[0026] The silver loading on Ag / AbCh solid appears to be a crucial factor that affects its sterilizing performance. As shown in Fig. 1 (see also Example 2), the catalyst with the highest metal loading (1 wt%) exhibits the best disinfection performance compared to the ones with lower metal loadings. The inhibition activity of the Ag / AhCh solid decreases with decreasing silver loading. Since the catalysts with high metal loadings have a higher metal surface area, it can be concluded that the inhibition of bacteria proceeds over the surface of the metal particles. The latter results come in a good agreement with the existing bibliographic data, since silver (Ag) was found to be highly toxic to cells, even at extremely low concentrations; the defence mechanisms used by cells are overcome by the activity of silver metal (in the metallic or ionic form) [1-2],

[0027] It was also found in this work that the nature (chemical composition) of the support has some effect on the activity of the corresponding supported- Ag solid, although this effect cannot be considered as drastic. On one hand, it was found that using CeO2-A12O3 (CeO2-coated AI2O3 spheres) as a support of silver, can lead to increased bacterial inhibition values, in the range of 10- 15% (Example 3, Fig. 2). On the other hand, the use of SiO2-based particles (Quarzo Sferico Griggio, QSG) as a support for Ag, was found to lead to approximately 20% lower bacterial inhibition values compared to those obtained on the Ag / AFO-, solid (Example 3, Fig. 2). However,it should be noted that the QSG support is a much cheaper material, compared to AI2O3 spheres, which is readily available in the market, since it is already being used in many industrial applications. Consequently, the use of Ag supported solids on QSG might be a favourable, cost- effective solution for real industrial applications.

[0028] The Ag / CeCh-AbCh solid was found to present excellent stability with reaction (inhibition of E. coll) time on stream (Example 6, Fig. 5). Practically constant bacterial inhibition values, even after 72 h of continuous reaction on stream, were obtained on the Ag / CeCh-AbCh solid. The fact that no bacterial inhibition was observed when using the support alone, indicates that the observed bacterial inhibition on Ag / CeCh-AhCh solid is due to the silver metal nano-crystallites (clusters), a fact which is in accordance with the literature, as discussed above [1-3], Similar results were also obtained over Ag / AbCh and Ag / QSG solids, as well as for the rest of the bacteria examined in the present work (E parahaemolyticus and I. enterococci). Moreover, the latter Ag-supported solids were also found to retain their stability, even when used for the disinfection of seawater (Example 7, Figure 6). It is noted that, no significant leaching of Ag was observed, even after 72 hrs of continuous reaction over the Ag / AhCh solid. Stability of materials is a major prerequisite when considering real, practical applications and thus, the stability of the Ag-supported solids (under examined reaction conditions) makes them eligible candidates for possible industrial applications.

[0029] Maintenance of the stability of the Ag-supported solids requires the continuous feeding of air in the reaction vessel, otherwise the solid materials quickly start to deactivate and become practically completely inactive after 2 hrs of continuous reaction (Example 9, Fig. 8). It was found that the presence of oxygen in the reaction feed is critical for (a) the regulation of the Ag clusters surface charge and (b) the cleaning (through oxidation) of the surface of the Ag clusters from the organic residues formed during bacterial lysis (destruction). Moreover, aeration (feeding of air) is essential for the reduction of mass transport phenomena which may significantly affect (decrease) the Ag-solid’s disinfection activity.

[0030] The silver-based disinfecting solids may be used in a fixed bed reactor, a fluidized bed reactor, a stirred tank reactor or a monolith-type reactor. Typically, these reactors are provided with one or more oxygen gas-introducing inlets to provide an oxygen-containing gas to enhance the disinfection.

[0031] The present invention, i.e., the use of Ag-supported solids (immobilized silver nanoparticles, Ag-iNPs) for the disinfection of water (including seawater and wastewater), can beconsidered a novel solution where the main differences from all the other technologies reported for the same application are the following:

[0032] The proposed technology uses Ag-supported solids which are characterized by low cost and high availability. Consequently, such a technology is expected to have considerably lower manufacturing cost as compared to existing / commercial solutions already available in the marker, such as cyclonic separation, electro-ionization magnetic separation, electric pulse, ultrasound, etc.

[0033] Most of the existing / commercial technologies (cyclonic separation, UV radiation, thermal treatment, microwaves, ultrasound, electric pulse, electro-ionization and ozonation) require significant amounts of electrical energy to operate. To the contrary, the proposed technology (use of Ag-supported solids) requires significantly lower amounts of electrical energy, which correspond to the energy needed for the circulation of water and air through the reaction vessel. That is, the present invention does not require the use of ozone, microwaves, electro-ionization, ultrasound, UV radiation, or thermal treatment, providing considerable energy savings as well as reducing the equipment costs.

[0034] Some of the existing technologies for water or seawater treatment present several environmental issues which are mainly related to the use of certain chemicals (biocides) such as chlorine and ozone. The proposed technology, however, poses no threat to the environment since no such chemicals are required for its operation (only air is needed).

[0035] Metal-based solids are reported to be prone to deactivation, in the presence of water and, in particular, in the presence of seawater [6-7], However, the Ag-supported solids of the present invention (Ag / AhCh, Ag / SQG and Ag / CeCh-AbCh) appear to be highly stable under actual reaction conditions (Examples 6 and 7, Figs. 5 and 6).

[0036] Most of the existing technologies (see Table 1), used for ballast water treatment, require the use of complex equipment, while the proposed Ag-based solution requires a very simple design such as the one shown in Fig. 7.

[0037] Some of the existing technologies used for ballast water treatment are quite bulky and this is a major disadvantage when considering applications where space is limited (e.g., on a ship). The present Ag-based technology though, is expected to require much less space compared to technologies such as cyclonic separation and electro-ionization. In fact, the main part of the proposed technology (reaction chamber) can be fitted within existing water tanks (e.g., ballast water tanks).

[0038] The disinfection activity of the immobilized silver nanoparticles (Ag-iNPs) is not significantly affected by water temperature or chemical composition, as opposed to some of the existing, commercial technologies used for water and seawater treatment.

[0039] As opposed to existing technologies, the proposed Ag-iNPs materials retain high bacterial inhibition activity and stability with reaction time. Moreover, the disinfection activity of the Ag- supported solids is not significantly affected by the concentration of bacteria in the water medium. On the contrary, the disinfection performance of most of the commercial technologies strongly depends on the concentration of microorganisms (level of bacterial pollution).

[0040] The following working examples represent a more detailed description of the invention. It is understood that this detailed description is given merely by way of illustration and does not limit the extent of the invention as many variations may be made therein without departing from the spirit of this invention.

[0041] EXAMPLES

[0042] 1. Immobilized Ag-nanoparticles (Ag-iNPs) synthesis

[0043] 1 wt% Ag / y-AhCh spheres (iNPs) were prepared via a modified wet impregnation method.Commercial y-alumina spheres (Sasol, code 604130), were initially immersed in a solution which contained specific amount of the Ag metal precursor (AgNCh, Panreac, code 131459), dissolved in deionized water which was used as a solvent, in order to obtain the desired loading for each solid. Alumina spheres were left in the above solution for 24 h under mild stirring, followed by evaporation of the solution within 6 h (pH=8). Finally, the nano-Ag supported spheres were mildly dried at 120 °C before their calcination in air at 400 °C for 2 h. A similar process was followed for the preparation of the 1 wt% Ag / QSG solid, where QSG particles were used instead of y-AhO, spheres. For the preparation of 1 wt% Ag / CeCh-AhCh, y-AECh spheres were initially immersed in a 0.12 M aquatic solution of Ce(NO3)3.6H2O (Sigma Aldrich) for 24 h at room temperature. The resulting solid was calcined in air at 600°C for 4 h (5 wt%CeO2 / AhO3) prior the impregnation of the metal (Ag) following the procedure described above.

[0044] 2. Effect of metal (Ag) loading

[0045] In this example the influence of silver (Ag) loading on the bacterial inhibition profile of E. coli with time on stream, was investigated over the Ag / AFCT solid at room temperature. 1 gr of solid (x wt.% Ag / AFCF) was used in all experiments, while the pressure was kept constant between 1.0- 1.1 atm. The stirring rate was also kept constant at 150 rpm in all experiments. All experimentswhere conducted in a batch-mode reactor. Figure 1 presents the E. coli bacterial inhibition profile as a function of reaction time, at room temperature, obtained over the x wt.% Ag / AbO, solids (Ag- iNPs), for five different Ag loadings, i.e., 0.05, 0.1, 0.2, 0.5 and 1.0 wt.%. As seen in Fig. 1, the percentage of bacterial inhibition increases significantly by increasing Ag loading at all reaction times. Thus, it can be said that the inhibition of E. coli over the x wt.% Ag / AbCh solids is favoured at high loading values of silver (Ag).

[0046] 3. Effect of support chemical composition

[0047] This example examines the effect of the chemical composition of the support on the bacterial inhibition activity of 1 wt.% Ag / support solids, using three different solid supports, i.e., AI2O3, CeCh-AbCh and Quarzo Sferico Griggio (QSG). The reaction conditions used in this example were the same to those described in Example 2. Figure 2 presents the E. coli bacterial inhibition profile as a function of reaction time, at room temperature, obtained over the 1% Ag / AbCE, 1% Ag / CeCh-AbCh and 1% Ag / QSG solids (Ag-iNPs). As seen in Fig. 2 “Comparison of E. coli inhibition depending of the Ag-supported solid used”.

[0048] 4. Disinfection Performance of Ag-iNPs

[0049] In this example the antimicrobial activity of Ag / AhCh is examined, against three different bacteria, i.e., E. coli, E. faecalis and V. parahaemolyticus. The reaction conditions used in this example were the same to those described in Example 2. Figure 3 shows comparatively the bacterial inhibition (as a percentage of initial bacterial concentration) profile of E. coli, E. faecalis and V. parahaemolyticus with reaction time, at room temperature, obtained over 1 wt.% Ag / AbCE As seen in Fig. 3, high bacterial inhibition values are obtained for all examined bacteria. Higher inhibition values are obtained in the case of V. parahaemolyticus, compared to the rest of the bacteria examined, at all reaction times. In particular, practically complete inhibition of V. parahaemolyticus is achieved at about 30 min after the initiation of the reaction (contact with the Ag / AhCh solid), while E. coli is completely inhibited (100% inhibition) 80 min after the initiation of the reaction. In the case of E. faecalis, a value of 75 % bacterial inhibition is achieved 2 hrs after the initiation of reaction. The results of Fig. 3 clearly indicate that the Ag / AbCh solid presents high disinfection activity against all three bacteria examined within the present work.

[0050] It is noted at this point that all antimicrobial experiments were conducted in semi-batch mode using a three-phase continuous stirred tank reactor (Autoclave Engineers, USA, and PID Eng. & Tech., Spain) equipped with a Mahoney -Robinson catalyst basket, which was designed inorder to ensure the maximum contact area between the three phases and to reduce as much as possible, mass transport phenomena. The gas phase medium (20% 02 / 80% He) was kept at a continuous flow while solid and liquid phases were stationary. In the case where 1 wt% immobilized silver nanoparticles were used for E. coli treatment, a pre- weighed (1 g) amount of Ag / AbO-, was placed in the reactor’s basket where in-situ pre- treatment in terms of calcination in air at 400 °C for 2 h, was performed. After temperature decrease to 25 °C (under He flow), the appropriate microbial solution was inserted in the reactor with a constant flow of 24 mL / min for 8 min, under continuous flow of He gas.

[0051] The desired homogenized microbial solution for each experiment was prepared through the dissolution of E. coli commercial lyophilized pellet (Microbiologies, Inc. - LYFO DISK (ATCC®), 200 Cooper Avenue North St. Cloud, MN 56303 USA) using a sterile swab, in 0.5 mL deionized and sterilized water of which, 0.4 pL was inserted in the reactor for each time. Additionally, aiming at the optimum conditions for the survival of bacteria, eliminating at the same time bacterial inhibition because of starvation or due to the lack of vital nutrients, 4,8 mL of Tryptic Soy Broth (TSB) was added in every prepared microbial solution. The ideal quantity of TSB was determined after several experiments were conducted. No dilutions of the initial solution were performed in order to achieve the desired amount of bacterial cells required (ca. 109 cells). Every treated sample was taken from the outlet of the reactor for the time intervals of 0, 5 and 25 min in each case and transferred on a commercial E. coli selective medium (E.C.C. Chrom Agar ACC. ISO 9308-2014) which was able to develop only E. coli bacteria contained in the treated solution.

[0052] The calculation of the bacterial inhibition caused by Ag-iNPs was feasible through the counting of the developed colonies within 24 h and 48 h respectively. The volume of the liquid phase (200 mL), the flow rate of gas feed stream (100 mL / min), the feed stream composition and the stirrer’s rotation speed (150 rpm) were kept constant in all catalytic experiments. In addition, most of the important experimental parameters such as the temperature (25 °C), the pressure (1 atm), the flow of the gaseous feed mixture and the volume of the solution in the reactor, were monitored through a control panel.

[0053] 5. Mode of action of Ag-iNPsThis example examines the possible mode (mechanism) of action of Ag-iNPs against E. coli. Within this example it will be proven that the antibacterial action of Ag-iNPs is due to the biocidal properties of immobilized Ag clusters and not to the properties of free silver ions. In particular,specific experiments were conducted in order to eliminate the possible presence of free Ag+ions in the reaction solution (e.g., due to possible surface dissolution) during the treatment of E. coli with Ag / AbCh. The elimination of free Ag ions was achieved by complexation with oxalic acid. Oxalate anions (C2O42-), due to the full dissociation of oxalic acid under the present experimental conditions, are expected to act as Ag+scavengers in the microbial medium, reducing the bioavailability of any possibly leached silver ions, because the resulting silver oxalate salt is practically completely insoluble in water (Ksp= 1.0 x 10-11) [4], Figure 4 compares the antimicrobial activity (bacterial inhibition) of Ag / AbO, and 0.3 ppb Ag+(for comparison), both in the presence of 5 ppb oxalic acid. As shown in Fig. 4, the antimicrobial activity of Ag / AbCh is not significantly affected by the presence of oxalic acid, because the Ag / AbCh solid practically retains its original activity, with complete bacterial inhibition achieved after just 70 min of treatment (note that a high density of the population of E. coli was used in the initial microbial solution, in order to achieve a minimum microbial concentration that will allow a molecular analysis). However, a completely different effect is observed in the case of 0.3 ppb free Ag+ions, which become practically inactive in the presence of oxalic acid, as expected, revealing the complete complexation of the free Ag+ions. These results further reinforce the notion that the contribution of free Ag+ions to the overall bacterial inhibition profile of Ag / AbO, is rather insignificant and that the strong antimicrobial activity of Ag-iNPs is mainly due to the biocidal activity of immobilized Ag clusters. It also is suggested that oxalic acid does not affect the availability of partially oxidized surface Ag sites on the surface of the solid, probably due to steric effects and charge incompatibility, a fact which is in accordance with the literature [5],

[0054] 6. Stability of Ag-iNPs with time

[0055] In this example the stability of Ag-iNPs (Ag / CeCb-AbCh) with time is examined under continuous reaction conditions. Experiments within tis example were conducted in a continuous flow reactor which has described in Example 4. 3 gr of solid (Ag / CeCh-AbCh) were used, while the flow rate of the bacterial solution (E. coli) that was fed to the reactor was kept constant at 10 mL / min. The rest of the reaction conditions used in this example were the same to those used in Example 2. Figure 5 presents comparatively the inhibition profile of E. coli with continuous reaction time on stream, obtained at room temperature, over 1 wt.% Ag / CeCb-AbCh and CeCh- AI2O3 (for comparison). As seen in Fig. 5 a steady-state (point where stable performance is observed) is achieved after 24 hrs of continuous reaction. In particular, at steady-state conditionsthe very high inhibition percentage of 90% is achieved. As depicted in Fig. 5, the 1 wt.% Ag / CeCh- AI2O3 solid retains its excellent antimicrobial activity (90% inhibition), even after 72 hrs of continuous reaction. Finally, it is noted that no significant activity was observed over the CeCh- AI2O3 support (or the blanc experiment), indicating, once again, that the antimicrobial activity observed over Ag / CeCh-AhCh is due to the presence of immobilized Ag nanoparticles (Ag-iNPs). The stability of Ag-iNPs with time, under continuous reaction conditions, is considered to be a major advantage for a possible practical application.

[0056] 7. Performance of Ag-iNPs in seawater

[0057] This example examines the performance of Ag-iNPs, under the actual conditions that are found in a real practical application (i.e., treatment of ship’s ballast water). For this pilot experiment, a very simple apparatus design was used, which is described in Example 8. Ag-iNPs on QSG were used for this experiment, since QSG is a much more inexpensive material compared to AI2O3 and might be favourable in real applications. In particular, 25 kg of 1 wt.% Ag / QSG were used, while the air flow rate was kept constant at about 1.5 m3 / min. The Ag / QSG solid was loaded in the apparatus (see Example 8) and then submersed in the ship’s ballast water tank (capacity: 100 m3of seawater). The duration of the pilot experiment was 48 hrs. Figure 6 presents the inhibition profiles of E. coli and Tot. coliforms with time, obtained during the pilot experiment described above. As clearly seen in Fig. 6, the system reaches at very high inhibition values of E. coli (>95%), after just 4 hrs of reaction. Similarly, an 80% inhibition of Tot. coliforms is observed after 4 hrs of process initiation. Practically complete inhibition of E. coli is observed after 12 hrs of reaction, while the final inhibition of Tot. coliforms was found to be almost 90%. The results presented in this example, are of utmost importance, since they prove that Ag-iNPs can perform equally well under actual industrial conditions. It is also important to note that, the latter results were achieved by using: (a) a very simple and inexpensive apparatus (Example 8) and (b) an inexpensive route for the preparation of Ag-iNPs (use of QSG instead of AI2O3 as a support).

[0058] 8. Indicative design of reactor system

[0059] This example presents an indicative reactor design that can be used for the proposed antimicrobial (disinfection) process with Ag-iNPs. Figure 7 presents 3D images of the very simple design that has been used in this work for the treatment of a ship’s ballast water with Ag-iNPs (see Example 7). The proposed indicative system is comprised of a UPVC tube, having a diameter of 250 mm, which encloses the Ag-iNPs solid’s bed (1% K.0. Ag / QSG). A second UPVC tube, havinga diameter of 200 mm, passes through larger UPVC tube, reaching to the % of its total height. The smaller diameter tube ends to a ceramic diffuser which forces air diffusion in the liquid medium. Air is passed through the smaller diameter tube. The complete system is immersed into the water tank (e.g., ballast water tank). The flow of air (coming out from the diffuser) induces the circular movement of water (draft) which passes through the solid’s bed (Ag / QSG). It is noted that variations of this design or completely different designs can be used with Ag-iNPs, having the same or even better antimicrobial performance.

[0060] 9. Effect of gas feed composition

[0061] Within this example, the effect of the gas feed composition on the antimicrobial performance of Ag-iNPs is examined. The experimental conditions used in this example are described in detail in Example 6 (continuous flow experiment). Figure 8 presents comparatively the inhibition profiles with time, obtained when using air (or 20%O2 / He), He or H2 as a gas feed for the treatment of a bacterial solution (E. coli) with 1 wt.% Ag / CeCh-AECh. As observed in Fig. 8, a quick decline in the antimicrobial activity is observed after just 20 min from the initiation of reaction, when using He or H2 as a gas feed. Such a result is not observed in the case where 20%O2 / He was used as a feed (Fig. 8). These results clearly indicate that oxygen (O2) is essential for the antimicrobial process and for the conservation of a stable disinfection performance over Ag-iNPs. In fact, it was found that the presence of oxygen in the reaction feed is critical for (a) the regulation of the Ag clusters surface charge (need to be partially oxidized), and (b) the cleaning (through oxidation) of the surface of the Ag clusters from the organic residues formed during bacterial lysis (destruction). Moreover, aeration (feeding of air) is essential for the reduction of mass transport phenomena which may significantly affect (decrease) the Ag-solid’s disinfection activity.

[0062] 10. Overall antimicrobial performance of Ag-iNPs - Compliance with IMO regulation

[0063] This example summarizes the antimicrobial performance of Ag-iNPs, as revealed from the numerous tests (lab- and pilot-scale) conducted within this work. Table 2 presents comparative results regarding the main experimental parameters, inhibition performance (for Gram- and Gram+ bacteria) and compliance with IMO regulation, of selected experiments that have been performed within the present work. As reported in Table 2, practically complete inhibition of Gram- bacteria (E. coli) is obtained in all experiments, while high to very high values of Gram+ bacteria inhibition is observed, ranging from 62 to 97%. It is important to note that no significant leaching of silverwas detected in any of the experiments conducted in the present work (Table 2). Moreover, it is of utmost importance to point out that in the cases of pilot experiments (conducted on a real / running ship), compliance with Annex D2 regulation of the IMO Convention (2004) has been achieved.

[0064] Based on the results reported in Table 2, it is safe to claim that the proposed antimicrobial process with Ag-iNPs is a promising technology for the disinfection of water, wastewater or seawater. The results obtained within this work, particularly the ones obtained under simulated or real industrial conditions, further reinforce the latter notion.Table 2: Comparative Results obtained during Lab- and Pilot-scale tests over Ag-iNPs

[0065] INDUSTRIAL APPLICABILITY

[0066] Novel Ag-supported nanoparticles with limited metal surface dissolution but strong antimicrobial activity are applied to confront particular environmental challenges, such as the disinfection of ships’ ballast water (SBW) and numerous types of industrial wastewater. The ballast water treatment system does not require the use of UV radiation or dangerous chemicals which may harm the ecosystem in regions where they are discharged. The present system is a simple solution in that the housing with silver supported catalyst may be placed anywhere within a ballast tank or at the inlet where ballast water is taken up from the sea. Further, multiple units may be placed with multiple tank regions in the ship in order to maintain the antibacterial quality of the water, leading to cleaner hull interiors. The silver treatment units may be readily added to ballast tanks without installation cost as it may be used as an autonomous stand-alone solution. The system is low maintenance and can be renewed through periodic heat treatment, typically overa period of years such as 5 years. The system requires only minimal consumption of electricity in the case where turbulence inside the ballast tank is needed. Further, the system does not require an drydock time as the system is continuous, in situ treatment of ballast water.

[0067] Most importantly, the present system does not create any dangerous by-products since the use of harmful chemicals is excluded. There is also no residual effect unlike the electrochlorination and ozonation processes since there is no silver leaching from the surface of the catalyst.

[0068] While the present disclosure has been described and illustrated with reference to specific embodiments thereof, these descriptions and illustrations are not limiting. It should be understood by those skilled in the art that various changes may be made and equivalents may be substituted without departing from the true spirit and scope of the present disclosure as defined by the appended claims. The illustrations may not necessarily be drawn to scale. There may be distinctions between the artistic renditions in the present disclosure and the actual apparatus due to manufacturing processes and tolerances. There may be other embodiments of the present disclosure which are not specifically illustrated. The specification and the drawings are to be regarded as illustrative rather than restrictive. Modifications may be made to adapt a particular situation, material, composition of matter, method, or process to the objective, spirit and scope of the present disclosure. All such modifications are intended to be within the scope of the claims appended hereto. While the methods disclosed herein have been described with reference to particular operations performed in a particular order, it will be understood that these operations may be combined, sub-divided, or re-ordered to form an equivalent method without departing from the teachings of the present disclosure. Accordingly, unless specifically indicated herein, the order and grouping of the operations are not limitations.References:1) J. A. Lemire, J. J. Harrison, R.J. Turner, Antimicrobial activity of metals: mechanisms, molecular targets and applications, Nat. Rev. Microbiol. 11 (2013) 371-384.2) C. Marambio-Jones, E.M.V. Hoek, A review of the antibacterial effects of nanomaterials and potential implications for human health and the environment, J. Nanopart. Res. 12 (2010) 1531-1551.3) S. Saha, B. Gupta, K. Gupta, M.G Chaudhuri, Production of putrescine-capped stable silver nanoparticle: its characterization and antibacterial activity against multidrugresistant bacterial strains, Appl. Nanosci. 6 (2016) 1137-1147.) S.P. Theofilou, B.K. Constantinou, V.K. Chatziiona, N. Pantelidou, S. Plyastsov, K. Kapnisis, P.G. Sawa, I. Meshkovsky, A. Anayiotos, C.N. Costa, New insights into the antimicrobial treatment of water on ag-supported solids, J. Chem. Technol. Biotechnol. 94 (2018) 1134-1143. ) Velazquez- Jimenez LH, Hurt RH, Matos J and Rangel-Mendez JR, Zirconium-carbon hybrid sorbent for removal of fluoride from water: oxalic acid mediated Zr(IV) assembly and adsorption mechanism. Environ Sci Technol 48:1166-1174 (2014). ) J. Hu, Q. Yang, L. Yang, Z. Zhang, B. Su, Z. Bao, Q. Ren, H. Xing, S. Dai, Confining Noble Metal (Pd, Au, Pt) Nanoparticles in Surfactant Ionic Liquids: Active Non-Mercury Catalysts for Hydrochlorination of Acetylene, ACS Catal., 5 (2015), 11, 6724-6731.) K. Gold, B. Slay, M. Knackstedt, A.K. Gaharwar, Antimicrobial Activity of Metal and Metal-Oxide Based Nanoparticles, Adv. Therap., 1 (2018), 1700033.Y1

Claims

CLAIMS:

1. A silver-based solid material for disinfection at room temperature of micro-organisms from water or wastewater, comprising: an aluminum oxide support material; nano-silver particles adhered to a surface of the aluminum oxide support material; the nano-silver particles being present in an amount between 0.01 and 5 wt% on the aluminum oxide support material.

2. The silver-based solid material of claim 1, where in the aluminum oxide support material comprises particles or spheres.

3. The silver-based solid material of claim 1, wherein the material further includes one or mor of Ag2O, AI2O3, CeCh, SiCh or Ce2C>3.

4. The silver-based solid material of claim 1, wherein the nano-silver particles are formed by impregnating the aluminum oxide support with a silver precursor, drying the impregnated support, decomposition of the silver precursor in an oxygen-containing atmosphere at a temperature of 400- 600°C.

5. The silver-based solid material of claim 4, wherein the impregnated support is further subjected to a reduction in a hydrogen- containing atmosphere at a temperature of at least 300°C.

6. A method for disinfection of ship ballast water or waste water, comprising: providing the silver-based solid material of claim 1 ; circulating ship ballast water or waste water over the silver-based solid material of claim 1; discharging treated ballast water from the ship into a surrounding body of water.

7. The method for disinfection of ship ballast water or waste water of claim 6, further comprising providing an oxygen-containing gas flow to the circulation of ship ballast water or waster water during the disinfection.

8. The method for disinfection of ship ballast water or waste water of claim 7, wherein the disinfection occurs without the use of ozone, microwaves, electro- ionization, ultrasound, UV radiation, or thermal treatment.

9. A reactor for disinfection of ship ballast water or waste water comprising; a reactor selected from a fixed bed reactor, a fluidized bed reactor, a stirred tank reactor or a monolith-type reactor; the silver-based solid material of claim 1 positioned with the reactor.

10. The reactor of claim 8 further including a gas inlet for introducing an oxygen- containing gas during a reaction.

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