N95Decon Research Document. Not Peer Reviewed.
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Technical Report for H​2​O​2​-Based N95 Reuse Risk Management
1. Overview
Hydrogen peroxide (H​2​O​2​) vapor and plasma decontamination is an establishedindustrial decontamination method used in research settings, pharmaceutical and medical
industries, and by police and fire departments (​Mickelsen et al., EPA Report, 2017​). Many
hospitals use hydrogen peroxide vapor (wet HPV or dry VHP​TM​) or hydrogen peroxide gas
plasma (HPGP) for decontamination. HPV, VHP​TM​ and HPGP inactivate highly resistantpathogens, including nosocomial bacterial spores and viruses. A number of studies (​Battelle,
2016, 2020; Bergman, 2010, Viscusi, 2009​) have demonstrated that certain N95 Filtering
Facepiece Respirators (N95 FFRs or respirators, often colloquially called ‘N95 masks’) can besafely decontaminated with proper use of HPV. Most of these studies used the Bioquell HPV
system.
For the purpose of bulk decontamination of N95 masks, a whole-room decontamination
system with controlled air-flow will allow carts with trays filled with N95 masks to be wheeled inand out, similar to a protocol developed by Duke University Medical Center. This would provide
for a capacity of 700 N95 masks in a 12 ft x 12 ft room per cycle. Alternatively, an air-flow
controlled BSC could be temporarily outfitted with a HPV system, which would then havecapacity for 100–120 N95 masks, depending on BSC size per cycle. Typically, a
decontamination plus off-gassing cycle takes 6 to 8 hours.
The Battelle study demonstrated that up to 20 cycles of HPV treatments on N95 FFRs
will not affect filter performance, pressure drop, fit, or elastic band quality.
H​2​O​2​ gas plasma (HPGP) reduces filter quality if applied for 3 cycles at high dose.
However, at low dose, filter quality was reported to be preserved for 2 cycles (​personalcommunication, J. Yarwood, ASP​), but this has not yet been independently confirmed andexact protocol might be a major issue.

MethodAbbreviation

Description

Example Provider

Hydrogen Peroxide Vapor

HPV

Wet H​2​O​2​ Vapor, >3 hr.
aeration

BioquellHydrogen Peroxide Gas
PlasmaHPGP

Ionized H​2​O​2 plasma​(not confirmed), 1 hr.
aeration

ASP (STERRAD)

Vaporized Hydrogen
Peroxide

VHP​TM

Dry H​2​O​2​ Vapor, >3 hr.
aeration

Steris

2. Status of Federal Guidance
CDC released guidance on the decontamination and reuse of N95s on March 31, 2020,
including the use of vaporized H​2​O​2​ (​Decontamination and Reuse of Filtering FacepieceN95Decon Research Document. Not Peer Reviewed.
Version 1.2, 4/2/2020.

Respirators, 2020​). The Battelle Decontamination System, an HPV system for decontaminating
N95 masks, received emergency use authorization from the FDA on March 28, 2020. They
report that the system can process at least 80,000 N95 masks per day (​Battelle, 2020​). “Inpreliminary studies, H​2​O​2​ vapor decontamination has been found to be a highly effective
method of eradicating MRSA, ​Serratia marcescens, Clostridium botulinum spores andClostridium difficile​ from rooms, furniture, surfaces and/or equipment; however, further
investigation of this method to demonstrate both safety and effectiveness in reducing infection
rates are required.” (​Rutala et al. 2019​). An FDA-supported study by Battelle (​2016​) found that
Bioquell HPV, with the established decontamination cycle parameters, achieved a 6-log
reduction in organism viability and maintained high filtration and low resistance to air-flow of
N95 masks following exposure to up to 50 cycles of HPV decontamination. Mask users should
perform a seal check after they don the mask. Please refer to current CDC guidelines that are
updated regularly as well as ​N95Decon’s Full Legal Disclaimer​ (​Decontamination and Reuse ofFiltering Facepiece Respirators, 2020​).

3. Mode of Action
HPV methods are used for terminal decontamination of hospital rooms, biosafetycabinets, and medical equipment and materials that are intolerant to heat or havediffusion-restricted space. Sterilizing units use liquid H​2​O​2​ that is vaporized by heat andreleased into the room until the effective concentration is detected by the HPV system.
Individual, unwrapped, contaminated objects are placed in a room or biosafety cabinet.
Typically, the HPV treatment involves a conditioning phase to change room humidity (dry or
wet treatment); a gassing phase to saturate the room (approximately 15 min); a dwell phase tomaintain a certain concentration (approximately 125 min); and an aeration or clearance phasefor off-gassing and breakdown of HPV into oxygen and water vapor (approx. 4–6 h).
Inactivation of microorganisms and viruses is achieved primarily by the combined actions ofH​2​O​2​ gas and the generation of hydroxyl and hydroperoxyl free radicals (Finnegan et al. 2010).
HPGP machines are often used in hospitals for rapid sterilization of surgical tools thatare wrapped or in Tyvek pouches. The plasma penetrates the material even when bagged, andalso more rapidly eliminates any condensed H​2​O​2​.

4. Potential for SARS-CoV-2 Inactivation
HPV, VPH​TM​, and HPGP destroy influenza viruses and other viruses and pathogens thatare more resistant than SARS-CoVs, such as spores from ​G. stearothermophilus​, nosocomial
C. difficile​, and ​mycobacterium tuberculosis (​ ​Battelle, 2016​; ​EPA, 2004​; ​Heckert et al. 1997​;
Kenny et al. 2020; Rudnick et al. 2009; Hall et al. 2007​).
Bergman et al. (​2010​) studied 6 different N95 masks (industrial N95s: 3M 8210; 3M
8000; Moldex 2200 and surgical N95: KC PFR95-270; 3M 1870; 3M 1860; N=6 for each of themodels tested) and applied 3 cycles of decontamination using HPGP, HPV and other methods.
The HPV decontamination method (Clarus R™, Bioquell) involved a gassing phase of 15 min, adwell phase of 125 min in a 64 m​3​ room to achieve a room concentration of 8 g/m​3​ (5700 ppm).
Biological indicators containing ​Geobacillus stearothermophilus​ spores were placed in fivelocations inside the room and a 6-log spore reduction was measured after each treatment.

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The 2016 Battelle Report prepared for the FDA summarized a study on the effects of
HPV on N95 filter quality and microorganism attenuation. An HPV treatment (Clarus C™,
Bioquell) of a 20 min gassing phase (2 g/min) followed by a 150 min dwell phase (0.5 g/min)completely inactivated ​Geobacillus stearothermophilus ​spores inoculated by droplet or aerosol
onto N95 masks (3M 1860). The authors further showed that decontamination was achieved
across 50 cycles of repeated treatments. Although the N95 masks were shown stacked up
against each other in the exposure chamber (Figure 11, Battelle 2016), this resulted in variable
sensor readings and is not recommended (personal communication, B. Heimbuch).
Kenny et al. (​2020​), in a non-peer reviewed report, evaluated viral decontamination of
HPV (BQ-50, Bioquell) after inoculating N95 masks (3M 1870) with 3 different types ofaerosolized bacteriophages. A single cycle of HPV completely eradicated phages from the N95
mask. N95 masks were suspended by their elastic strap on racks in a 33 m​3​ room for a 30–40
min gas phase at 16 g/min, a 25 min dwell phase, and a 150 min aeration phase.
Heckert et al. (​1997​) inoculated glass and stainless steel with 9 exotic animal viruses.
After HPV treatment, virus titer was reduced to 0 (except for hog cholera virus in whole blood).
A VH​TM ​P1000 Steris machine was used to generate a gas phase of 2 g/min for 30 min tomaintain a H​2​O​2​ concentration of 1.73 mg/L (1211 ppm).

5. Integrity of N95 Filtering Facepiece Respirators
Viscusi et al. (​2009​) evaluated 6 different N95 masks (​same as Bergman et al. 2011​) andapplied 1 cycle of treatment of 5 decontamination methods including high dose HPGP. There
was no effect of HPGP on filter quality. However, 3 cycles of the same high dose HPGP
decontamination process reduced filtering efficiency by > 5% for 4 of 6 different masks tested,
bringing the efficiency below the FDA-advised 95% threshold (Bergman et al. 2010).
Three cycles of HPV treatment in the Bergman et al. (​2010​) study did not reduce filter
quality (filter efficiency > 98% and no change in airflow resistance) nor were there observable
physical changes to the N95 masks.
The 2016 Battelle Report phase 2 study evaluated filter quality and fit. The same
exposure was applied as described in the SARS-CoV-2 Inactivation section above, with theaddition of 300 min of aeration, to eighty-five N95 masks for 10, 20, 30, 40 and 50 cycles ofdecontamination, 15 N95 masks per cycle set. After decontamination, both inert andbioaerosol collection efficiency remained >99% and no degradation of airflow resistance wasfound for all eighty-five of the N95 masks. After 30 cycles, strap degradation was observed
through strap length elongation and loss of elasticity, which could negatively impact respirator
fit. There was no degradation in mannequin fit testing up to 20 cycles of decontamination (no fit
testing was done beyond 20 cycles). Only the 3M 1860 N95 model was tested.
Duke University & Health HPV system (​Schwartz et al., 2020​) incorporates results fromthe Battelle study. N95 masks (3M 1860) were either suspended by their elastic straps or layed
individually onto stainless steel racks in wheeled carts. An existing disinfection system wasused in a room (12 ft X 12 ft ) of their NIAID Regional Biocontainment Laboratory. The room
was treated (Bioquell Clarus™ C system with a 35% H​2​O​2​ solution and distribution system todisperse HPV uniformly) to attain a 480+ ppm concentration of HPV with a gas time of 25 min

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and dwell time of 20 minutes. One hundred N95 masks (3M 1860) were treated with HPV for 1
cycle. Air concentration near the N95 masks was measured during the aeration period todetermine time until the concentration was below the OSHA Permissible Exposure Limit (1ppm, 1.4 mg/m​3​). At 4 hours, the concentration was below the limit of detection of the device(below detectable limit measured with a PortaSens II™ sensor). A qualitative test was
conducted on the N95 masks by 3 individuals who detected no noticeable odors. There was no
physical nor performance (not described) degradation of the N95 masks. They are currently
evaluating the Bioquell Z‐2 and Bioquell ProteQ™ system with >10 repeated treatment cycles
for fit.
Although no measurements of filter performance were made in the non-peer reviewed
Kenny et al. (​2020​) study, after 5 cycles of HPV treatment, the 3M 1870 N95 masks “appeared
similar to new with no deformity.”
On March 16, 2020, the Dutch National Institute for Public Health reported that 3M
8822 masks with up to 4 cycles of HPGP (STERRAD NX100, Express cycle with AllClear™) andfound that 2 cycles of treatment did not deform masks or compromise fit, but 3 or 4 cycles
compromised the fit. There was no test of filter efficiency. They also noted that used masks
sterilized with this process did not support SARS-CoV-2 growth when in medium for 72 hours.
Data are suggested to show that Sterrad units for HPGP, when used with the proper
settings, can decontaminate 10 individually Tyvek-wrapped N95s in a 25 or 27 min. cycle
without compromising filtration or airflow for 2 cycles (personal communication, J. Yarwood,
ASP). Treatment is followed by 60 min aeration (after opening Tyvek bag) to allow outgassingof residual H​2​O​2​ gas. HPGP can reduce filter quality if applied with intensive treatments(Bergman et al., 2010) and so the exact protocol for this treatment used is a concern. Thismethod is not recommended in the updated CDC report (2020).
It is important to note that HPV, VHP​TM​ and HPGP are not compatible with cellulose,
which is not a component listed in 3M model 1860 N95 masks (3M 1860 Data Sheets), but may
be present in other PPE. The presence of cellulose in PPE is an important consideration in the
adoption of H​2​O​2​-based strategies.

6. Data Summary Tables
Table 1. Impact of HPV on ​G stearothermophilus s​ pores and viruses
Author

Media

Dose

Time (min)Strain(s)Effectiveness(log reduction)

A

Biological indicators inroom (N=5)

8 g/m​3

gas 15;dwell 120

G stearothermophilus spores≥6B

3M 1860 N95 masks
inoculated withaerosol (N=15)

2 g/min then gas 20;
0.5 g/mindwell 150

G stearothermophilus spores≥6C

3M 1860 N95 masks
inoculated withdroplets (N=3)

2 g/min then gas 20;
0.5 g/mindwell 150

G stearothermophilus spores≥6

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D

3M 1860 N95 masks
inoculated withaerosol then treatedwith VHP​TM​ for 50
cycles (N=5)

2 g/min then gas 20;
0.5 g/mindwell 150

G stearothermophilus spores≥6 (50 cycles)E

3M 1860 inoculated
N95 masks (N=3 foreach phage)

16 g/mingas 30–40;
dwell 25

Phage phi-6
Phage T7
Phage T1

≥6

F

Glass and stainlesssteel inoculated with 9viruses

2 g/mingas 30Avian influenza
African swine fever virus
Bluetongue virus
Hog cholera virus
Newcastle disease virus
Pseudorabies virus
Swine vesicular disease virus
Vesicular exanthema virus
Vesicular stomatitis virus≥6

A: (​Bergman et al., 2010​), B: (​Battelle, 2016​) Phase 1 aerosol inoculated filters, C: (​Battelle, 2016​) Phase 1 droplet inoculated filters,
D: (​Battelle, 2016​) Phase 3 aerosol inoculated filters, E: (​Kenny et al., 2020​), F: (​Heckert et al. 1997​).

Table 2. Impact of HPV on N95 FFRs
Author

N95 masks

DoseTime (min)# cyclesFiltration
efficiency

Respirator damage

G

6 different N95
models (N=6 foreach model)

8 g/m​3

gas 15;dwell 120

3

>97%

None notedH

3M 1860 (N=85
total)

2 g/minthen 0.5
g/min

gas 20;dwell 150;
aeration 300

10, 20,>99% for all
30, 40, 50 N95 masks> 30 cycles straps
fragmented when stretchedI

3M 1860(N=100)

Goal: 480+ppm

gas 25;dwell 20;
aeration 2401None noted

N/A

G: (​Bergman et al., 2010​), H: (​Battelle, 2016​), I: (​Schwartz et al., 2020​).

7. Strategies
Commercial systems are available from companies such as Bioquell, ASP (STERRAD),
Steris, Battelle, and Halosil, though they differ in the method of delivering and sustaining H​2​O​2concentrations in a room/cabinet and in the solvents used. [Note: Bioquell uses the term H​2​O​2vapour (HPV) while Steris uses the term vaporized H​2​O​2​ (VHP​TM​).] Bioquell HPV includes a
generator to produce HPV, a module to measure the concentration of HPV, temperature, andrelative humidity in the enclosure, and an aeration unit to catalyse the breakdown of HPV into
oxygen and water vapour after HPV exposure. A control pedestal is set outside the enclosure
to provide remote control. HPV is delivered until the air in the enclosure becomes saturated
and H​2​O​2​ begins to condense on surfaces (​Hall et al., 2007; Ray et al., 2010​). Steris VHP​TM
systems have a generator inside the room with an integral aeration unit and dehumidifier
designed to achieve a set humidity level prior to the start of the cycle. Alternatively, Steris units

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may be connected to an existing biosafety cabinet in the hospital and operated by
Steris-trained personnel remotely from outside the enclosure. The system delivers‘non-condensing’ VHP​TM​ by drying the vapour stream as it is returned to the generator. Bioquell
systems do not control the H​2​O​2​ air concentration while the Steris systems hold a steady H​2​O​2
air concentration throughout the exposure period.
The Halosil system uses the HaloMist solution which contains a low percentage (5%)
peroxide with a biocidal silver nitrate additive at a low dose (0.01%) that converts to low ppb’s
once the product is aerosolized. Halosil foggers generate a 100-120 ppm H​2​O​2​ vapor content
through initial water evaporation from microdroplets that concentrate both the peroxide andthe silver. This effectively increases the peroxide concentration in the vapor phase above theinitial 5%. While this very low silver concentration is unlikely to impact theelectrostatically-charged membrane of an N95 mask, the filtering function of the N95 masks
should be determined after multiple cycles of decontamination.
The Bioquell systems [used by Duke, Bergman (​2010​) and Battelle (​2016​)] uses a 35%
peroxide solution without additives. It is important that such high concentrations of peroxide behandled safely; high concentrations are toxic and can be explosive. The manufacturer uses
RFID-chipped bottles so use of third party peroxide is not an option.
The Duke Medical Center has developed a standard operating procedure (SOP) for
decontamination of N95 masks using the Bioquell Clarus™ C system and has performed
qualitative testing on more than one hundred N95 masks (see above; ​Schwartz et al., 2020​).
Masks are checked before and after decontamination and prior to each use. Soiled N95 maskswith visible blood, hair or damage are discarded and not decontaminated. N95 masks are
separated into 4 streams based on size of N95 mask (common: 3M 1860 or small: 3M 1860s)
and the visible presence or absence of facial cosmetics. After decontamination, the integrity ofthe straps (evaluate for elongation), nose bridge, and nose foam are checked for integrity. The
Duke Medical Center protocol does not return the mask back to the same user while theBattelle protocol does. A reason for returning the mask to the same user is because the mask
was initially fit and fit tested to that user, and also in case the former user applied any
non-compatible wipe (alcohol or surfactant; ​Viscusi et al., 2007 ​but see​ Heimbuch et al. 2014
for efficacy of bleach wipe option) that could disrupt the hydrophobic coating or electrostatic
charge. In either case, users should perform a seal check before reuse.

8. Primary Risks and Unknowns
Dosing protocol is complex and could result in incomplete decontamination or
explosion risk. Therefore only trained personnel should operate HVP, VHP​TM​ or HPGP
equipment. H​2​O​2​ gas is a strong oxidizer that may cause fire or explosions, and is a corrosive
irritant that may cause skin, eye or lung damage. H​2​O​2​ gas may interact with N95 mask
components to form a toxic residue - analytical chemistry tests for H​2​O​2​ can test for this. The
OSHA permissible exposure limit is 1 ppm over an 8-hour Time Weighted Average (TWA).
During the decontamination process, room concentrations may be higher than 100 ppm.
Detection of odor does not provide adequate warning of hazardous residual
concentrations in the N95 mask. Quantitative tests for sufficient decontamination (​G.

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stearothermophilus ​spore growth) and aeration should be done on sentinel N95 masks(PortaSens-II test). Probability of N95 mask straps failure increases with more than 20 cycles ofdecontamination; this will vary with the decontamination method and N95 mask model. Straps
should be examined after decontamination and prior to each use.

9. Conclusions
Multiple studies have confirmed that N95 masks contaminated with aerosol or droplets
containing ​G. stearothermophilus​ spores were successfully decontaminated with H​2​O​2​ with a6-log reduction in spore level. Furthermore, N95 mask filter efficiency did not degrade with up
to 50 cycles of decontamination. However, after 20 cycles of HPV decontamination the N95
mask (3M 1860) straps showed degradation and were permanently deformed when stretched.
Disadvantages are that N95 masks must be individually placed or hung and cannot be bagged
during or after HPV treatment in order to attain complete exposure and aeration. Typicaldecontamination durations, including aeration, are 4 to 8 hours.
Some hospitals have HPGP systems (H​2​O​2​ gas plasma; Sterrad, Irvine, CA). Threecycles of high dose treatments (Sterrad 100S) reduces N95 filter efficiency to an unacceptablelevel (​Bergman, 2010​). According to ASP (​personal communication, J. Yarwood​), 3 cycles oftreatment with lower dose (STERRAD 100NX, AllClear, Express cycle) does not reduce filter
efficiency, but this awaits independent confirmation. H​2​O​2​ gas plasma provides rapid clearing
of toxic H​2​O​2​ vapor (55 minute total processing time) and individual tyvek bagging of masks.
Many hospitals already have HPV systems in-house for use in full room terminal
decontamination. These could be deployed in dedicated decontamination rooms. Processing
carts filled with trays of N95 masks could be wheeled in and out of the room. Alternatively,
existing biosafety cabinets within the hospital can be connected to a VHP​TM​ (pending data for
dry H​2​O​2​ vapor efficacy and safety) or HPV unit. Another solution is to send the respirators toan outside service-provider (​e.g., Battelle​) for decontamination. To achieve an appropriate
concentration of H​2​O​2​ vapor during the gas and dwell phase the specification of the HPV
system should be matched to the treated volume. The long (300 min) aeration phase could be
conducted in an adjacent room where H​2​O​2​ vapor is converted to inert O​2​ and water vapor. The
concentration of HPV should be measured during decontamination to confirm adequate
treatment level and that at the end of aeration workers who enter the room are protected. The
OSHA Permissible Exposure Limit is 1 ppm (1.4 mg/m​3​). HPV can be detected by smell andirritation, but may pose respiratory dangers at levels below user detection.
The Content provided by N95DECON is for INFORMATIONAL PURPOSES ONLY, DOESNOT CONSTITUTE THE PROVIDING OF MEDICAL ADVICE and IS NOT INTENDED TO BE
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Battelle (2020). FDA letter of approval (3/28/2020) for Emergency Use Authorization for the Battelle
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