**1. Introduction**

For more than a century, chlorination has been the most common method used worldwide for drinking water disinfection. Chlorine and chlorine-based compounds are widely used for the control of waterborne pathogens because of their high oxidizing potential, low cost, and

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residual disinfectant properties that prevent microbial recontamination. Unfortunately, the chemical reaction between chlorine and organic compounds in water generates carcinogenic agents such as trihalomethanes and halogenic acetic acids [1, 2]. Furthermore, some resistant microorganisms may only be inactivated with very high chlorine doses, which can exacerbate the formation of disinfection by-products (DBPs) [3]. Presently, growing concerns about the potential hazards associated with DBPs have boosted efforts to develop chlorination alternatives. Ozonation is effective at inhibiting a variety of pathogens; however, its disadvantages include the high cost and the potential formation of DBPs such as bromate in seawater [4, 5]. Other water treatment methods such as ultraviolet (UV) radiation, ultrasound, cavitation, or heat application can be used for the inactivation of organisms. Although these methods do not produce DBPs or other problematic chemical residues, they require substantial energy consumption and have high operational costs [5]. Besides, the efficiency of UV disinfection is greatly dependent on water quality because the activity of UV light is substantially decreased by turbidity or organic matter present in water [5].

mechanism is still unknown for this process. Pressure cycling is defined as a repetitive pro-

*Escherichia coli* Inactivation Using Pressurized Carbon Dioxide as an Innovative Method for Water Disinfection

suggests that the decompression process may lead to mechanically induced explosive cell

membranes [11–13]. In previous works, the pressure cycling procedure has been conducted

decompression and compression [8, 10, 13, 15]. Despite the good bactericidal performance of

(<1.0 MPa) will enhance the bactericidal activity. Therefore, in this study, we examined the

tion applications such as ballast water treatment. Comparisons of *E. coli* inactivation caused

ined for various conditions of pressure, temperature, flow rates, and working volume ratios (WVRs). In particular, the influence of pressure cycling on *E. coli* inactivation was evaluated.

tron microscopy (SEM). The research objective was to evaluate the bactericidal effectiveness

ambient temperatures, and consequently, high-pressure (4–50 MPa) or ultra-high-pressure (200–700 MPa) conditions are vital for sufficient inactivation. However, to be more attractive

at lower pressures. In this study, we employed the use of a liquid-film-forming apparatus, which enabled improvements in the interaction efficiency but with lower pressures (<1 MPa)

The experimental apparatus for disinfection was a stainless steel chamber with an internal volume of 10 L and pressure tolerance up to 1.0 MPa. The device was designed with a solid stream nozzle and shield to enable vigorous agitation of the influent in such a way that produced liquid films along with fine bubbles (**Figures 1**–**3**). The device was supplemented

high speed through a small nozzle and directed onto the shield. The highly pressurized fluid stream thus collided with the bubble-generating shield. Subsequently, numerous gas bubbles,

pressure prior to the treatments. Sample water was then pumped into the device at

**2. Novel idea: apparatus for forming highly dissolved gas in water**

release requirements are drawbacks owing to the costly and complex operating proce-

technology enhanced by pressure cycling [11–13, 15], the high pressure and

and pressurized air were evaluated in both natural seawater and arti-

for disinfecting water, with the goal of addressing the abovementioned

methods in the field of food preservation, the interaction efficiency

and pathogens in the foodstuffs is probably limited at low pressures and

ruptures [14], while the compression process may intensify the mass transfer of CO2

dures. Presently, it is not clear whether pressure cycling with low-pressure CO2

between each cycle of raised/lowered pressure.

[9, 10]. Evidence so far

discharges between each cycle of

http://dx.doi.org/10.5772/intechopen.68310

at low pressures and with no

against *E. coli* was exam-

at less than 1.0 MPa for seawater disinfec-

treatment were assessed by scanning elec-

technology needs to be implemented

across cell

209

treatments

cedure that involves the decompression and compression of CO2

with high-pressure operations (8–550 MPa) and with CO2

effect of pressure cycling on the bactericidal performance of CO<sup>2</sup>

ficial seawater. The inactivation performance of pressurized CO<sup>2</sup>

emerging problems associated with water disinfection technology.

This study investigated the use of pressurized CO2

Changes in cell morphology after pressurized CO2

in terms of its economic feasibility, pressurized CO2

for the water disinfection purposes.

pressurized CO2

release of CO2

by pressurized CO2

of pressurized CO2

For pressurized CO2

between CO2

with CO2

CO2

Sterilization by using pressurized CO2 has been an active research field for decades [6, 7]. CO2 has been used extensively to sterilize dried food and liquid products via a nonthermal sterilization method [8] because of its effectiveness in inactivating microbes, nontoxicity, and low cost [9]. Prior research on high-pressure CO2 treatments has investigated the effects of several factors such as pressure, temperature, type of microorganisms, agitation speed, decompression rate, and pressure cycling on the inactivation capacity of this method [6, 8, 10–15]. Most studies have reported that high-pressure operating conditions (4–50 MPa) are required to inactivate significant numbers of pathogens [7, 9]. Subsequently, certain concerns involving high-pressure operations (i.e., the need for heavy-duty pressure equipment, high initial investment costs, energy consumption concerns, and pressure control and management issues) have hampered the implementation of high pressurized CO2 preservation technology at a large scale within the food industry.

In recent years, pressurized CO2 has shown great potential as a sustainable disinfection technology in water and wastewater treatment applications [16–22] largely because this method does not generate DBPs [9, 22]. Kobayashi et al. [16, 17] employed CO2 microbubbles in the treatment of drinking water and succeeded in inhibiting *Escherichia coli* within 13.3 min. However, the pressure (10 MPa) and temperature (35–55°C) requirements for effective inactivation [16, 17] are still high from a practical standpoint. Our research group has developed a novel method that uses low-pressure CO2 treatments (0.2–1.0 MPa) based on technology that produces high amounts of dissolved gas in water to inactive bacteria and bacteriophages in freshwater [19–21] and seawater [23, 24]. Cheng et al. [19] suggested that the sudden discharge and resulting reduction of pressure could cause cells to rupture via a mechanical mechanism, and further, that this would be lethal to cells at high levels of dissolved CO2 at 0.3–0.6 MPa and room temperature. Vo et al. [20, 21] demonstrated that acidified water and cellular lipid extraction caused by pressurized CO2 at 0.7 MPa and room temperature were major factors for efficient disinfection within a treatment time of 25 min.

Previous research has shown that pressure cycling is a potential means to improve bacterial inactivation during pressurized CO2 treatments [8–10, 13, 15]; nevertheless, the inactivation mechanism is still unknown for this process. Pressure cycling is defined as a repetitive procedure that involves the decompression and compression of CO2 [9, 10]. Evidence so far suggests that the decompression process may lead to mechanically induced explosive cell ruptures [14], while the compression process may intensify the mass transfer of CO2 across cell membranes [11–13]. In previous works, the pressure cycling procedure has been conducted with high-pressure operations (8–550 MPa) and with CO2 discharges between each cycle of decompression and compression [8, 10, 13, 15]. Despite the good bactericidal performance of pressurized CO2 technology enhanced by pressure cycling [11–13, 15], the high pressure and CO2 release requirements are drawbacks owing to the costly and complex operating procedures. Presently, it is not clear whether pressure cycling with low-pressure CO2 treatments (<1.0 MPa) will enhance the bactericidal activity. Therefore, in this study, we examined the effect of pressure cycling on the bactericidal performance of CO<sup>2</sup> at low pressures and with no release of CO2 between each cycle of raised/lowered pressure.

residual disinfectant properties that prevent microbial recontamination. Unfortunately, the chemical reaction between chlorine and organic compounds in water generates carcinogenic agents such as trihalomethanes and halogenic acetic acids [1, 2]. Furthermore, some resistant microorganisms may only be inactivated with very high chlorine doses, which can exacerbate the formation of disinfection by-products (DBPs) [3]. Presently, growing concerns about the potential hazards associated with DBPs have boosted efforts to develop chlorination alternatives. Ozonation is effective at inhibiting a variety of pathogens; however, its disadvantages include the high cost and the potential formation of DBPs such as bromate in seawater [4, 5]. Other water treatment methods such as ultraviolet (UV) radiation, ultrasound, cavitation, or heat application can be used for the inactivation of organisms. Although these methods do not produce DBPs or other problematic chemical residues, they require substantial energy consumption and have high operational costs [5]. Besides, the efficiency of UV disinfection is greatly dependent on water quality because the activity of UV light is substantially decreased

208 *Escherichia coli* Escherichia coli - Recent Advances on Physiology, Pathogenesis and Biotechnological Applications - Recent Advances on Physiology, Pathogenesis and Biotechnological Applications

 has been used extensively to sterilize dried food and liquid products via a nonthermal sterilization method [8] because of its effectiveness in inactivating microbes, nontoxicity,

of several factors such as pressure, temperature, type of microorganisms, agitation speed, decompression rate, and pressure cycling on the inactivation capacity of this method [6, 8, 10–15]. Most studies have reported that high-pressure operating conditions (4–50 MPa) are required to inactivate significant numbers of pathogens [7, 9]. Subsequently, certain concerns involving high-pressure operations (i.e., the need for heavy-duty pressure equipment, high initial investment costs, energy consumption concerns, and pressure control and manage-

nology in water and wastewater treatment applications [16–22] largely because this method

treatment of drinking water and succeeded in inhibiting *Escherichia coli* within 13.3 min. However, the pressure (10 MPa) and temperature (35–55°C) requirements for effective inactivation [16, 17] are still high from a practical standpoint. Our research group has developed a

produces high amounts of dissolved gas in water to inactive bacteria and bacteriophages in freshwater [19–21] and seawater [23, 24]. Cheng et al. [19] suggested that the sudden discharge and resulting reduction of pressure could cause cells to rupture via a mechanical mechanism,

and room temperature. Vo et al. [20, 21] demonstrated that acidified water and cellular lipid

Previous research has shown that pressure cycling is a potential means to improve bacterial

ment issues) have hampered the implementation of high pressurized CO2

does not generate DBPs [9, 22]. Kobayashi et al. [16, 17] employed CO2

and further, that this would be lethal to cells at high levels of dissolved CO2

has been an active research field for decades [6, 7].

has shown great potential as a sustainable disinfection tech-

treatments (0.2–1.0 MPa) based on technology that

at 0.7 MPa and room temperature were major factors

treatments [8–10, 13, 15]; nevertheless, the inactivation

treatments has investigated the effects

preservation tech-

microbubbles in the

at 0.3–0.6 MPa

by turbidity or organic matter present in water [5].

and low cost [9]. Prior research on high-pressure CO2

nology at a large scale within the food industry.

novel method that uses low-pressure CO2

extraction caused by pressurized CO2

inactivation during pressurized CO2

for efficient disinfection within a treatment time of 25 min.

In recent years, pressurized CO2

Sterilization by using pressurized CO2

CO2

This study investigated the use of pressurized CO2 at less than 1.0 MPa for seawater disinfection applications such as ballast water treatment. Comparisons of *E. coli* inactivation caused by pressurized CO2 and pressurized air were evaluated in both natural seawater and artificial seawater. The inactivation performance of pressurized CO<sup>2</sup> against *E. coli* was examined for various conditions of pressure, temperature, flow rates, and working volume ratios (WVRs). In particular, the influence of pressure cycling on *E. coli* inactivation was evaluated. Changes in cell morphology after pressurized CO2 treatment were assessed by scanning electron microscopy (SEM). The research objective was to evaluate the bactericidal effectiveness of pressurized CO2 for disinfecting water, with the goal of addressing the abovementioned emerging problems associated with water disinfection technology.
