Hydrogen peroxide (H₂O₂) is a highly reactive chemical compound as part of a family of compounds known as “reactive oxygen species” (ROS). It can be used as an oxidiser, removing electrons from another chemical species, and becoming reduced in what is known as a redox reaction. This oxidising property of H₂O₂ is what promotes its biological toxicity, and high concentrations of H₂O₂ are known to cause cell damage through the oxidation of proteins [1] and DNA via the formation of hydroxyl radicals from a Fenton type reaction between H₂O₂ and iron [2] [3] [4].

Hydrogen peroxide was first studied as a disinfectant in 1891 by B. W. Richardson and has since become a popular biocide used for antisepsis, the disinfection of hard surfaces and medical devices, and for room fumigation [4] [5]. The importance of decontaminating surfaces within healthcare settings has only recently been realised with studies showing that transmission of nosocomial pathogens via contaminated surfaces has indeed contributed to an increase in health-care-acquire infections [6]. Disinfecting rooms that had been occupied by infected patients reduces the risk of subsequent patients acquiring the same pathogens, however conventional disinfection/cleaning methods rely on human operation which comes with its own drawbacks. The development of decontamination technologies that are automated is a step towards addressing this problem and eliminates the reliance on a human operator correctly selecting, formulating, distributing, and deciding the contact time of the disinfectant agent [7]. Decontamination systems most used are vapourised H₂O₂ (VHP), aerosolised hydrogen peroxide (AHP), and ultra-violet (UV) light. In this study, technologies utilising VHP were considered.

Vapourised H₂O₂ was first considered for use in sterilisation technologies in the 1980s, whereby liquid H₂O₂ is converted into vapour by either heat or changes in pressure and then delivered into the room to decontaminate and sterilize [8]. There are two types of systems that use H₂O₂ vapour: condensing and non-condensing technologies. Technologies that use condensing systems work by injecting H₂O₂ into the enclosed room until the air is saturated and the H₂O₂ begins condensing onto the surfaces. With non-condensing technologies, the air in the room is first dehumidified by the generators before H₂O₂ that has been vaporised is released and circulated for a set period, and then finally the dry vapour is returned to the generator where it is broken down into water and oxygen [9]. The use of H₂O₂ vapour technologies has become a well-established form of bio-decontamination, with many studies showing effective efficacy against bacterial spores [10] and other microbes, generally achieving 6-log reduction [11] [12] [13]. As well as being highly effective in killing microbes, the decomposition of H₂O₂ into water and oxygen make it environmentally safer in comparison to other biocides like formaldehyde, ethylene oxide, and glutaraldehyde [14].

One of the limitations of using H₂O₂ vapour for decontamination of an enclosed space is ensuring the room is safe for re-entry and the operator does not inhale any vapour. The concentration of VHP is considered safe at <1 ppm, and immediately dangerous to life or health in humans at a concentration of 75 ppm [15] [16]. Therefore, it is essential that an accurate form of monitoring of the H₂O₂ level is achieved, which is typically checked using sensors. In this study the calibration of externally manufactured sensors to a higher degree of accuracy is developed. In order to do this, a H₂O₂ vapour of around 1 ppm [17] was created and exposed to the sensors to perform the calibration.

The method described in the paper is based on the concentration of hydrogen peroxide in saturated vapour over a solution in water at a defined temperature. The saturated vapour is generated by bubbling dry air into a solution of hydrogen peroxide at a defined concentration and temperature as shown in Figure 1.

The results from the 3-day validation study are shown in Table 1. These results show that the concentration of the freshly prepared Hydrogen Peroxide solutions used for the study were between 0.43% and 0.45%. Table 2 shows the concentrations of the hydrogen peroxide vapours formed by the solutions in Table 1.

An average of triplicate samples showed that the concentration of the vapour was between 0.7 and 0.8 ppm.

This variation is expected due to the nature of how vapours are formed. The vapour formed is dependent on the hydrogen peroxide solution and its concentration. Although a 0.42% solution was made up, results show that the exact concentrations of the solutions were between 0.43% and 0.45%. This error is due to the balance used for weighing, making up to volume using diluent, as well as the titration procedure and other human errors. All validation testing and reporting takes into consideration the exact concentration of solution rather than the theoretical concentration to ensure accurate reporting.

Over the 3 days of testing, Table 2 shows that results of 0.7 and 0.8 ppm were obtained. An average of the 9 readings taken gave a result of 0.78 ppm which would give a reading of 0.8 ppm on the gas leak detector gun due to rounding. These standard deviation for these datasets was calculated to be 0.04 which shows low variability and reliable results. Overall, the slight difference in validation results have no suspected impact on calibration of Hydrogen Peroxide cells as a titration test will always be performed prior to calibrating the cell to ensure the cell is being calibrated to the exact concentration of the vapour. Therefore, these validation results are sufficient to prove robustness of the method and confidence in calibrating the Hydrogen peroxide sensors.

Table 3 shows results from the first stage of the verification of the method. Cells 2443, 2444 and 2445 were all calibrated externally and a significant variation in results was observed. Cell 2443 produced readings almost double that of 2444 and 2445. The internally calibrated cell (2447) however, produced more reliable and repeatable results. This cell produced consistent results of 0.8 ppm at each of the 3 readings which showed no variation. A reading of 0.8 ppm is also the closest value to the theoretical and titration concentration of the hydrogen peroxide vapour produced.

The results from the second stage of the verification process are shown in Table 4 and Table 5. In both the small and the large test chamber, the difference of results showed the variability of externally calibrated hydrogen peroxide cells. The use of the small and large test chamber showed the real-life comparison to using the cells in hospitals or other healthcare environments. The two initial decontamination cycles in the small test chamber showed a reading of 0.4 ppm on both occasions using the internally calibrated cell. On the other hand, the other 3

cells showed a difference of results between both runs with readings varying from 0.4 ppm to 0.8 ppm.

A method for calibrating hydrogen peroxide vapour sensors at around 1 ppm can be achieved by the evaporation of aqueous hydrogen peroxide at 0.42% w/w solution. This method was validated by a well-known quantitative iodometric analysis technique to confirm the concentration of the vapour produced. The method was then verified by comparing manufacturer calibrated cells to cells that had been calibrated by the method described in this paper.

In principle, validation and verification processes showed that the calibration procedure developed internally to calibrate Hydrogen Peroxide cells in gas leak detector guns was successfully implemented.

The authors declare no conflicts of interest regarding the publication of this paper.

Zaib, A., Begum, S.S. and Allegra, E. (2022) Calibration of Hydrogen Peroxide Vapour Sensor. Advances in Chemical Engineering and Science, 12, 163-171. https://doi.org/10.4236/aces.2022.123012