Calcium is a major macronutrient for trees and is used in agricultural fertilizers. It is important for preserving membrane and cell wall integrity, as well as intracellular transport (8). At pH 7, calcium bicarbonate lacks a mesoscopic structure and does not exert a cytotoxic effect against microorganisms.

Calcium bicarbonate, whose chemical formula is Ca(HCO₃)₂, does not refer to an actual solid compound, as it exists only in dilute aqueous solutions containing calcium (Ca²⁺), bicarbonate (HCO₃^-), and carbonate (CO₃)^2- ions together with dissolved carbon dioxide (CO₂). The relative concentration of these carbon-containing species depends on the pH, with bicarbonate being predominant at pH 6.36-10.25 in water. These materials are often used in chewing gums with protective action on the oral cavity (9). In clinical practice, calcium bicarbonate water is used to treat patients with gastroesophageal reflux disease (10).

Compared to untreated specimens, CAC-717 treatment has been shown to reduce human norovirus genomic RNA by 3.25 log₁₀ (11). The virucidal effect of CAC-717 was compared to that of a phosphate buffer control at pH 12.4 and calcium bicarbonate without the mesoscopic structure (pH 7.0) (11). A human norovirus (GII.4 Sydney 2012) (12) was used as a target and was purified prior to disinfectant treatment. Phosphate buffer at pH 12.4 and calcium bicarbonate at pH 7.0 failed to inactivate human norovirus (11). Although CAC-717 is an alkaline solution, its pH decreases rapidly (within 1 min) to 8.84 upon contact with human skin (1). Indeed, CAC-717 is classified as non-irritant (Class 0) and skin sensitization tests conducted in rabbits according to the Ministry of Health, Labour and Welfare of Japan Guidelines, Biological Evaluation of Medical Devices-Part 10: Test for irritation and skin sensitization (ISO 10993-10, July 2, 2006) have not revealed any harmful properties (1). Furthermore, rabbit eye toxicity tests using the OECD Guidelines for Testing Chemicals no. 405: Acute Eye Irritation/Corrosion, performed under the same animal welfare requirements (ISO 10993-2, July 2, 2006) confirmed CAC-717 biosafety (1). Concentrated CAC-717 exhibited cytotoxicity in cultivated cell lines (11); but even this effect was lost after a 1:2 dilution in phosphate buffer.

To obtain CAC-717, an electric field is applied to mineral water containing calcium bicarbonate (1,2,13,14). According to the Japanese patent No. 5778328, CAC-717 (Food and Drug Administration/USA Regulation no. 880.6890 Class 1 disinfectant) is produced by applying a voltage of 2x10⁴ V for 48 h using Teflon-coated electrostatic-field electrodes (2). The resulting material has a pH of approximately 12.4 and contains 6.9 mM calcium bicarbonate particles (81,120 mg/l) with a mesoscopic structure (50-500 nm) that can be observed under an electron microscope (Fig. 1). CAC-717 is adsorbed onto a ceramic surface and air-dried for storage in the form of CAC-717 stones (Fig. 2). Prior to use, the CAC-717 stones are placed in fresh distilled water, calcium bicarbonate is dissolved, and CAC-717 is reconstituted. Reconstituted CAC-717 has the same microbicidal properties as original CAC-717.

The CAC-717 suspension, which is a colorless disinfectant, can be sprayed and dried on metal or plastic surfaces, resulting in a white powder coating (1). Scanning electron microscopy samples were obtained from a CAC-717 suspension after drying on a glass slide and used for quality control assessment, together with pH and calcium bicarbonate content measurements (1).

CAC-717 has been shown to be effective in inactivating plant pathogens (14). Black rot is a disease affecting cabbage and other cruciferous plant leaves, and is caused by Xanthomonas campestris pv. campestris (Xcc), a gram-negative seed-borne bacterium (15). The number of viable Xcc cells after treatment with CAC-717 was determined. An Xcc suspension (8.22 log₁₀ colony-forming units/ml) was incubated with an equal volume of CAC-717 at 25˚C for different periods of time (Table I). CAC-717 treatment caused a decrease in the number of Xcc cells after 0.5 min (Fig. 3A). In control experiments, no significant reduction in the number of Xcc cells was observed after treatment with distilled water at 25˚C; whereas hot water (50˚C) treatment significantly decreased the number of Xcc cells (14). In test samples, CAC-717 caused a low incidence of black rot, with only 26.77±3.33% of seeds exhibiting signs of disease after incubation at 25˚C for 5 days, compared with 56.67±8.82% in the distilled water group (Fig. 3B). No significant difference in the germination rate and plant stem length was detected between distilled water (25˚C) and CAC-717 treatment after cultivation for 5 days.

CAC-717 displays a bactericidal effect against Escherichia coli (E. coli) and Salmonella enterica (S. enterica) (13) (Table I). Following treatment with CAC-717 for 2 min, viable E. coli cells decreased by approximately 3.00-2.00 log₁₀ and S. enterica by more than 7.00 log₁₀, indicating that some bacteria are more sensitive than others to CAC-717. When using sodium hypochlorite (4 ppm) as a fungicide for chicken eggs mixed with S. enterica, the average multiple endpoint D value was 0.195 min (16). In contrast, CAC-717 displayed an average multiple-endpoint D value of 0.080 min, indicating a more powerful bactericidal effect than sodium hypochlorite.

Extreme autoclaving conditions (134˚C, 18 min) are required to inactivate prions, although some prions retain transmissibility even after dry-heating at 400˚C (16). Alternatively, prions are inactivated upon treatment with sodium dodecyl sulfate, sodium hydroxide, and sodium hypochlorite (17), but these chemicals are generally impractical due to their corrosive properties.

In another recent study, the authors reported that CAC-717 inactivated prions (2) (Table I). Western blotting was used for scrapie prion (PrP^Sc) analysis (18). To examine the seeding activity of PrP^Sc, protein misfolding cyclic amplification (PMCA) was used, which mimics the in vivo reaction and amplifies PrP^Sc in vitro (19). Western blotting and PMCA were applied to follow proteinase K-resistant PrP^Sc, while a transgenic mouse bioassay was used to test the transmissibility of prions. Western blot analysis showed that the levels of proteinase K-resistant PrP^Sc in lysates obtained from prion-infected N2a cells were markedly reduced after CAC-717 treatment. Next, PMCA was used to assess the conversion activity of any remaining PrP^Sc after CAC-717 treatment. The seeding activity of PrP^Sc was also clearly reduced in CAC-717-treated samples (2) (Table I). In addition, mice injected with CAC-717-treated samples survived longer than those injected with PBS-treated controls. These findings suggest that CAC-717 reduced both PrP^Sc conversion activity and prion transmissibility (Fig. 4).

Further studies are required to determine the mechanism by which CAC-717 inactivates prions. Given that CAC-717 does not cause irritation and is not corrosive, it offers an alternative to other prion disinfectants such as hypochlorous acid (20); however, detailed comparison with the latter is still needed.

Recently, CAC-717 was tested against SARS-CoV-2(21). CAC-717 exhibited a strong virucidal effect against all the examined mutant forms of SARS-CoV-2 isolated in Japan. Viral infectivity decreased by 3.8 to 4.2 log₁₀ within 15 sec (Table II), implying a virucidal activity similar to that of 80% ethanol. This strong microbicidal effect of CAC-717 has been linked to its elevated alkalinity, which may be due to the relative decrease in H⁺ and concomitant increase in OH^-. However, given that SARS-CoV-2 retains its infectivity for up to 30 sec at pH 12.4 (pH-adjusted Dulbecco's modified Eagle's medium) (21), it is more likely that an unidentified mechanism controls the virucidal activity of CAC-717. SARS-CoV-2 is known for the emergence of numerous strains. In addition to the conventional original virus (WK-521), the virucidal activity of CAC-717 was confirmed against α, β, γ, and δ variants, with the log₁₀ reduction in infectivity ranging from 3.6 to 4.4 (Table II). The effectiveness of CAC-717 in the presence of organic matter was analyzed to confirm its beneficial properties in the general environment (21). The tissue culture infectious dose of the virus mixed with 5% bovine serum albumin was 10^4.8 and the virucidal effect of CAC-717 corresponded to >4.3 log₁₀ (21). A lower virucidal effect of sodium hypochlorite has been observed in the presence of organic material; whereas the value obtained for CAC-717 indicates only a minor reduction. Numerous disinfectants have been used for COVID-19 control; hence, their impact on human health and the environment should be taken into account. As CAC-717 is harmless and does not cause skin or eye toxicity in rabbits (1), its future use as an anti-COVID-19 disinfectant is less problematic. Moreover, because the calcium bicarbonate component of CAC-717 is derived from plant material, it is non-flammable and can be used in a variety of applications for which ethanol is not suitable (21).

In addition to SARS-CoV-2, a recent study showed that CAC-717 possessed virucidal activity against 22 different types of DNA or RNA viruses. Destruction of viral nucleic acids in RNA viruses has been observed (22), but the exact effect of CAC-717 against viral DNA remains to be determined (22). In nature, human norovirus is typically associated with the fecal matrix. To investigate the microbiological properties of CAC-717, four fecal specimens were evaluated. Compared to the untreated specimen, CAC-717 resulted in log₁₀ reductions of 1.36, 2.78, 1.64, and 3.52 in the GI.2, GII.4 Sydney 2012, GII.5, and GII.7 specimens, respectively (Table I). Thus, CAC-717 could successfully inactivate human norovirus under clinical conditions. A similar effect was reported with heat treatment, but not with 1,000-ppm sodium hypochlorite in a stool suspension (except for human norovirus GII.17) (11) or 70% (v/v) ethanol.

CAC-717 has been proven effective against influenza virus (1) and feline calicivirus (FCV) (19) (Table I). To investigate the mechanistic effect of CAC-717 against viruses, real-time PCR revealed a progressive decline in the integrity of the FCV genome with increasing CAC-717 treatment time, as opposed to only intact RNA from untreated FCV samples (13).

Hypochlorous acid solution is also effective against several viruses (23,24); however, it is less stable in the presence of ultraviolet irradiation or in contact with air or organic compounds. Consequently, hypochlorous acid must be stored under cool and dark conditions to maintain its microbicidal activity (23).

Recently, concentrated bioshell calcium oxide (BiSCaO) water has been reported as an effective skin disinfectant (7,25). Although BiSCaO is strongly alkaline (pH 12.8), the pH of BiSCaO water decreased to 8.5 within 5 min after spraying on the back skin of hairless rats. The generation of CaCO₃ following the interaction between Ca²⁺ ions in BiSCaO water and CO₂ in air was suspected to be the cause of this rapid drop in pH (25). This finding echoes the rapid decrease in pH of CAC-717 when in contact with rabbit skin (1).