Toilet research looks not only inside the bowl, but also at the floor and the gap around the toilet’s base. A 2021 study investigated where bacteria were abundant in household toilets and, drawing on those findings, examined a test method in which volatile air-freshener ingredients deposited on materials to inhibit bacterial growth.
The paper directly measured bacteria that could be cultured from household surfaces and the antibacterial properties of materials exposed to ingredients in the laboratory. The authors aimed to prevent odors arising from dirt left on floors and other surfaces, but the study did not include tests measuring improvements in odors perceived by people or reductions in malodorous compounds.
Finding where bacteria remain in household toilets
The first survey involved 10 households in Hyogo Prefecture, Japan. Households were selected that had at least one boy who used the toilet independently. After residents cleaned with their usual products and methods, researchers swabbed the floor, side wall, surface of the toilet rim, and gap between the toilet and floor. Samples were collected again after seven days of normal use without cleaning.
Bacterial counts were determined from colonies formed after spreading the swab fluid on culture medium and incubating it at 35°C for three days. The measurement therefore covered bacteria able to grow under those conditions.
Both immediately after cleaning and seven days later, counts were high in the gap between the toilet and floor compared with the other sampled locations. Floor counts increased significantly after seven days, whereas no significant change over time was found at the other sites.
A subsequent survey examined the floor, walls, underside of the toilet rim, and other locations in greater detail in 12 households in the same prefecture, where toilets had not been cleaned for a week. Here, counts were highest beneath the rim, followed by the floor and walls. Comparisons of floor positions 10 cm and 20 cm from the toilet, at the front and sides, found no significant differences in counts.
The results thus do not establish a single ranking in which the floor always has the most bacteria. They also show bacteria remaining in hard-to-clean locations such as beneath the rim and in the gap at the base. The survey was limited to particular regional and household conditions and does not represent an average for all household toilets.
Delivering ingredients to materials through the air
The next experiment investigated whether volatile ingredients could be taken up by flooring materials and fabrics and leave behind properties that inhibit bacterial growth. Four materials were tested: plain-woven cotton fabric, cotton toweling, a cotton floor mat, and vinyl cushioning material.
After being cut and sterilized, the materials were placed in a sealed chamber with an air-freshener sample for 24 or 72 hours at 23°C. The chamber measured approximately 100 × 63.5 × 54.5 cm, making it an experimental space smaller than a residential toilet room.
The sample combined two groups of alkenals, aldehydes containing a double bond, with different carbon-chain lengths. Specific examples given in the original paper are (E)-2-hexenal and (E)-2-decenal. The intended design combined ingredients with different evaporation rates to deliver them to materials over time.
This was not a household trial of spraying essential oils themselves. It concerned a product formulation containing particular volatile ingredients and controlled material tests.
Exposing the materials first, then inoculating them with bacteria
After exposure, the materials were removed from the chamber and transferred to separate containers. They were then inoculated with five stock bacterial strains, Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, and Enterococcus hirae, and incubated at 37°C for 24 hours.
This was therefore neither a test of spraying air freshener directly onto bacteria nor a household intervention tracking whether floor bacteria decreased. The design measured how much ingredients previously delivered to the materials affected subsequent bacterial growth.
Antibacterial activity values were calculated by comparing, on a logarithmic scale, the changes in bacterial counts from immediately after inoculation to 24 hours later on control and treated materials. Higher values indicate greater growth inhibition relative to the control. They are not simple killing percentages or the percentage of initially present bacteria removed.
Growth inhibition on materials exposed for 72 hours
For the four materials exposed for 72 hours, antibacterial activity values against the five bacterial species exceeded 3 in almost every combination. The exception was the cushioning material with K. pneumoniae, at 2.91. The authors evaluated these findings as growth-inhibitory effects against the criteria of the textile-testing standard they adopted.
Floor mats and cushioning material exposed for 24 hours also had activity values above 2 against the tested S. aureus, E. coli, and E. hirae. An effect remaining on the materials was thus observed even after the shorter exposure.
However, this index registers an effect not only when bacterial counts decline on treated materials, but also when they increase less than on controls. Omitting the specific material–bacterium combinations and interpreting the findings as eliminating all bacteria would be incorrect.
Chemical analysis also confirmed adsorption onto cotton
To investigate whether ingredients actually transferred to the material, cotton fabric was exposed for 15, 24, or 72 hours, and compounds extracted from it were analyzed by GC/MS. After 72 hours, the amount of volatile ingredients adsorbed was approximately 2.7 µg per cm², which the authors interpreted as approaching equilibrium.
This supported a sequence in which ingredients evaporate from the product, transfer to the fabric, and inhibit bacterial growth on it. The authors consider this approach potentially useful for preventing odors originating from toilet surfaces.
In actual toilets, ventilation, soiling, materials, and distance from the product vary. How far adsorption and bacterial growth inhibition in a sealed chamber translate into household odor reduction requires separate verification. The results also do not demonstrate infection prevention or inhalation safety.
Summary
Household toilet surveys found high bacterial counts beneath the rim and in the gap above the floor, while floor counts increased after seven days of use. Against this background, the researchers explored delivering volatile ingredients to materials to inhibit bacterial growth on surfaces.
In the laboratory, growth inhibition was observed on fabrics and cushioning material exposed to the product, and adsorption of ingredients onto cotton was confirmed. The study shows that research into odor-prevention technologies considers not only fragrance in the air, but also how ingredients reach floors and fabrics.
At the time of the study, four of the five authors were affiliated with Procter & Gamble. No separate funding or conflict-of-interest statement was found in the original paper examined. Patents related to the product technology are listed among its references.
Reference
Ishida Y, Vyas R, Chauhan G, Delgado L, Kourai H. Bacterial Contamination Analysis of Residential Toilet Room Environment and Delivery of Residual Bacterial Growth Prevention Efficacy on Residential Toilet Permeable Surfaces Using Toilet Air Freshener Product. Biocontrol Science. 2021;26(2):85–93. doi:10.4265/bio.26.85.
